Gene Literature Dashboard

The objective of this study was to investigate the molecular response to damage at the blood brain barrier (BBB) and to elucidate critical pathways that might lead to effective treatment in central nervous system (CNS) pathologies in which the BBB is compromised. We have used a human, stem-cell derived in-vitro BBB injury model to gain a better understanding of the mechanisms controlling BBB integrity. Chemical injury induced by exposure to an organophosphate resulted in rapid lipid peroxidation, initiating a ferroptosis-like process. Additionally, mitochondrial ROS formation (MRF) and increase in mitochondrial membrane permeability were induced, leading to apoptotic cell death. Yet, these processes did not directly result in damage to barrier functionality, since blocking them did not reverse the increased permeability. We found that the iron chelator, Desferal© significantly decreased MRF and apoptosis subsequent to barrier insult, while also rescuing barrier integrity by inhibiting the labile iron pool increase, inducing HIF2α expression and preventing the degradation of Ve-cadherin specifically on the endothelial cell surface. Moreover, the novel nitroxide JP4-039 significantly rescued both injury-induced endothelium cell toxicity and barrier functionality. Elucidating a regulatory pathway that maintains BBB integrity illuminates a potential therapeutic approach to protect the BBB degradation that is evident in many neurological diseases.

The complex network of malfunctioning pathways occurring in the pathogenesis of neurodegenerative diseases (NDDs) represents a huge hurdle in the development of new effective drugs to be used in therapy. In this context, redox reactions act as crucial regulators in the maintenance of neuronal microenvironment homeostasis. Particularly, their imbalance results in the severe compromising of organism's natural defense systems and subsequently, in the instauration of deleterious OS, that plays a fundamental role in the insurgence and progress of NDDs. Despite the huge efforts in drug discovery programs, the identification process of new therapeutic agents able to counteract the relentless progress of neurodegenerative processes has produced low or no effective therapies. Consequently, a paradigm-shift in the drug discovery approach for these diseases is gradually occurring, paving the way for innovative therapeutical approaches, such as polypharmacology. The aim of this review is to provide an overview of the main pharmacological features of most promising nature-based scaffolds for a possible application in drug discovery, especially for NDDs, highlighting their multifaceted effects against OS and neuronal disorders.

No abstract available.

<h4>Background</h4>Little is known about the association among skin color, sunlight exposure. and vitamin D status in Southeast Asian population.<h4>Objective</h4>To investigate the association between skin color measured by von Luschan chromatic scale (VLCS) and vitamin D status in Thai medical ambulatory patients.<h4>Methods</h4>Medical ambulatory patients were enrolled. The eligibility criteria were as follows: aged >18 years, stable medical conditions, and no conditions directly affecting vitamin D status. Serum 25-hydroxyvitamin D [25(OH)D] levels were assessed. Skin color at the outer forearm was assessed using VLCS which grades skin color from the lightest score of 1 to the darkest score of 36. Patients were systematically interviewed to estimate daily sunlight exposure time.<h4>Results</h4>A total of 334 patients were enrolled. Data were expressed as mean ± SD. The mean serum 25(OH)D was 25.21 ± 10.06 ng/mL. There were 17 (5.1%), 217 (65.0%), and 100 (29.9%) patients who had light brown (VLCS score 18-20), medium brown (VLCS score 21-24), and dark brown (VLCS score 25-27) skin colors, respectively. The mean serum 25(OH)D level was higher in patients with dark brown skin than in patients with medium brown and light brown skin (28.31 ± 10.34 <i>vs.</i> 24.28 ± 9.57 and 19.43 ± 9.92 ng/mL, respectively, both <i>p</i> < 0.05). Multivariate analysis showed that darker skin color and increased sunlight exposure time were independently associated with decreased odds of vitamin D deficiency (dark brown vs. light brown: odds ratio, 0.263, 95% CI: 0.081-0.851, <i>p</i>=0.026; medium brown vs. light brown: odds ratio, 0.369, 95% CI: 0.987-1.003, <i>p</i>=0.067; sunlight exposure time odds ratio per 1 minute/day increase 0.955, 95% CI: 0.991-1.000, <i>p</i>=0.037), after adjusting for possible confounders.<h4>Conclusions</h4>We found that darker skin color at sunlight exposure area and increased sunlight exposure time were independently associated with decreased odds of vitamin D deficiency in Thai medical ambulatory patients.

<h4>Background</h4>Periventricular nodular heterotopia (PNH) is a malformation of cortical development characterized by nodules of abnormally migrated neurons. The cause of posteriorly placed PNH is not well characterised and we present a case that provides insights into the cause of posterior PNH.<h4>Case presentation</h4>We report a fetus with extensive posterior PNH in association with biallelic variants in LAMC3. LAMC3 mutations have previously been shown to cause polymicrogyria and pachygyria in the occipital cortex, but not PNH. The occipital location of PNH in our case and the proposed function of LAMC3 in cortical development suggest that the identified LAMC3 variants may be causal of PNH in this fetus.<h4>Conclusion</h4>We hypothesise that this finding extends the cortical phenotype associated with LAMC3 and provides valuable insight into genetic cause of posterior PNH.

The high deposition of intramuscular fat and the content of oleic fatty acid are characteristic of the Iberian pig. These two parameters present great variability and are differentiated amongst the varieties that make up the Iberian pig population. Although previous studies generated evidence for causal genes and polymorphisms associated to the adipogenic potential of the Iberian pig, there is little information about how genetic expression influences this trait's variability. The aim of this study was to analyses the expression profile between two varieties of Iberian pig (Torbiscal and Retinto) and their reciprocal crosses differentiated in their intramuscular fat (IMF) content and fatty acid (FA) composition in the <i>Longissimus thoracis</i> muscle using an RNA-seq approach. Our results corroborate that the Retinto variety is the fattiest amongst all studied varieties as its upregulated genes, such as <i>FABP3</i> and <i>FABP5</i>, <i>SLC27A1</i> and <i>VEGFA</i> among others, contribute to increasing adiposity. In its turn, Torbiscal pigs showed an upregulation of genes associated with the inhibition of fat deposition such as <i>ADIPOQ</i> and <i>CPT1A</i>. Further genetic variation analysis in these Iberian varieties showed relevant associations for SNP located within the differentially expressed genes with IMF and FA content. Thus, the differences found in the genetic architecture and the muscle transcriptome of these Iberian varieties might explain the variability in their fat content and composition and hence, their meat quality.

Advances in understanding disease pathogenesis correlates to modifications in gene expression within different tissues and organ systems. In depth knowledge about the dysregulation of gene expression profiles is fundamental to fully uncover mechanisms in disease development and changes in host homeostasis. The body of knowledge surrounding mammalian regulatory elements, specifically regulators of chromatin structure, transcriptional and translational activation, has considerably surged within the past decade. A set of key regulators whose function still needs to be fully elucidated are small non-coding RNAs (sncRNAs). Due to their broad range of unfolding functions in the regulation of gene expression during transcription and translation, sncRNAs are becoming vital to many cellular processes. Within the past decade, a novel class of sncRNAs called PIWI-interacting RNAs (piRNAs) have been implicated in various diseases, and understanding their complete function is of vital importance. Historically, piRNAs have been shown to be indispensable in germline integrity and stem cell development. Accumulating research evidence continue to reveal the many arms of piRNA function. Although piRNA function and biogenesis has been extensively studied in <i>Drosophila</i>, it is thought that they play similar roles in vertebrate species, including humans. Compounding evidence suggests that piRNAs encompass a wider functional range than small interfering RNAs (siRNAs) and microRNAs (miRNAs), which have been studied more in terms of cellular homeostasis and disease. This review aims to summarize contemporary knowledge regarding biogenesis, and homeostatic function of piRNAs and their emerging roles in the development of pathologies related to cardiomyopathies, cancer, and infectious diseases.

Best known as a transcriptional factor, CCCTC-binding factor (CTCF) is a highly conserved multifunctional DNA-binding protein with 11 zinc fingers. It functions in diverse genomic processes, including transcriptional activation/repression, insulation, genome imprinting and three-dimensional genome organization. A big surprise has recently emerged with the identification of CTCF engaging in the repair of DNA double-strand breaks (DSBs) and in the maintenance of genome fidelity. This discovery now adds a new dimension to the multifaceted attributes of this protein. CTCF facilitates the most accurate DSB repair via homologous recombination (HR) that occurs through an elaborate pathway, which entails a chain of timely assembly/disassembly of various HR-repair complexes and chromatin modifications and coordinates multistep HR processes to faithfully restore the original DNA sequences of broken DNA sites. Understanding the functional crosstalks between CTCF and other HR factors will illuminate the molecular basis of various human diseases that range from developmental disorders to cancer and arise from impaired repair. Such knowledge will also help understand the molecular mechanisms underlying the diverse functions of CTCF in genome biology. In this review, we discuss the recent advances regarding this newly assigned versatile role of CTCF and the mechanism whereby CTCF functions in DSB repair.

Due to the extremely high incidence of lesions and diseases in aging population, it is critical to put all efforts into developing a successful implant for osteochondral tissue regeneration. Many of the patients undergoing surgery present osteochondral fissure extending until the subchondral bone (corresponding to a IV grade according to the conventional radiographic classification by Berndt and Harty). Therefore, strategies for functional tissue regeneration should also aim at healing the subchondral bone and joint interface, besides hyaline cartilage. With the ambition of contributing to solving this problem, several research groups have been working intensively on the development of tailored implants that could promote that complex osteochondral regeneration. These implants may be manufactured through a wide variety of processes and use a wide variety of (bio)materials. This review aimed to examine the state of the art regarding the challenges, advantages, and drawbacks of the current strategies for osteochondral regeneration. One of the most promising approaches relies on the principles of additive manufacturing, where technologies are used that allow for the production of complex 3D structures with a high level of control, intended and predefined geometry, size, and interconnected pores, in a reproducible way. However, not all materials are suitable for these processes, and their features should be examined, targeting a successful regeneration.

Alphaviruses are arthropod-borne RNA viruses which can cause either mild to severe febrile arthritis which may persist for months, or encephalitis which can lead to death or lifelong cognitive impairments. The non-assembly molecular role(s), functions, and protein-protein interactions of the alphavirus capsid proteins have been largely overlooked. Here we detail the use of a BioID2 biotin ligase system to identify the protein-protein interactions of the Sindbis virus capsid protein. These efforts led to the discovery of a series of novel host-pathogen interactions, including the identification of an interaction between the alphaviral capsid protein and the host IRAK1 protein. Importantly, this capsid-IRAK1 interaction is conserved across multiple alphavirus species, including arthritogenic alphaviruses SINV, Ross River virus, and Chikungunya virus; and encephalitic alphaviruses Eastern Equine Encephalitis virus, and Venezuelan Equine Encephalitis virus. The impact of the capsid-IRAK1 interaction was evaluated using a robust set of cellular model systems, leading to the realization that the alphaviral capsid protein specifically inhibits IRAK1-dependent signaling. This inhibition represents a means by which alphaviruses may evade innate immune detection and activation prior to viral gene expression. Altogether, these data identify novel capsid protein-protein interactions, establish the capsid-IRAK1 interaction as a common alphavirus host-pathogen interface, and delineate the molecular consequences of the capsid-IRAK1 interaction on IRAK1-dependent signaling.

The demand for organs to be transplanted increases pressure on procurement centers, to the detriment of organ quality, increasing complications. New preservation protocols are urgently needed, requiring an in-depth understanding of ischemia-reperfusion mechanisms. We performed a proteomic analysis using LC-MS/MS-TOF data analyzed through R software and Cytoscape's ClueGO application, comparing the proteome of kidney endothelial cells, key cell type, subjected to 3, 6, 12, 19, and 24 h of cold ischemia and 6 h reperfusion. Critical pathways such as energy metabolism, cytoskeleton structure/transport system, and gene transcription/translation were modulated. Important time windows were revealed: a-during the first 3 h, central proteins were upregulated within these pathways; b-the majority of these upregulations were maintained until 12 h cold ischemia time (CIT); c-after that time, the overall decrease in protein expression was observed; d-at reperfusion, proteins expressed in response to cold ischemia were all downregulated. This shows that cold ischemia is not a simple slowing down of metabolism, as deep changes take place within the proteome on major pathways. Time-sensitive expression of key protein reveals possible quality biomarkers as well as potential targets for new strategies to maintain or optimize organ quality.

Colorectal cancer (CRC) initiation is believed to result from the conversion of normal intestinal stem cells (ISCs) into cancer stem cells (CSCs), also known as tumor-initiating cells (TICs). Hence, CRC evolves through the multiple acquisition of well-established genetic and epigenetic alterations with an adenoma-carcinoma sequence progression. Unlike other stem cells elsewhere in the body, ISCs cohabit with the intestinal microbiota, which consists of a diverse community of microorganisms, including bacteria, fungi, and viruses. The gut microbiota communicates closely with ISCs and mounting evidence suggests that there is significant crosstalk between host and microbiota at the ISC niche level. Metagenomic analyses have demonstrated that the host-microbiota mutually beneficial symbiosis existing under physiologic conditions is lost during a state of pathological microbial imbalance due to the alteration of microbiota composition (dysbiosis) and/or the genetic susceptibility of the host. The complex interaction between CRC and microbiota is at the forefront of the current CRC research, and there is growing attention on a possible role of the gut microbiome in the pathogenesis of CRC through ISC niche impairment. Here we primarily review the most recent findings on the molecular mechanism underlying the complex interplay between gut microbiota and ISCs, revealing a possible key role of microbiota in the aberrant reprogramming of CSCs in the initiation of CRC. We also discuss recent advances in OMICS approaches and single-cell analyses to explore the relationship between gut microbiota and ISC/CSC niche biology leading to a desirable implementation of the current precision medicine approaches.

Ectodomain shedding is a key mechanism of several biological processes, including cell-communication. Disintegrin and metalloproteinases (ADAMs), together with the membrane-type matrix metalloproteinases, play a pivotal role in shedding transmembrane proteins. Aberrant shedding is associated to several pathological conditions, including arthritis. Tissue inhibitor of metalloproteases 3 (TIMP-3), an endogenous inhibitor of ADAMs and matrix metalloproteases (MMPs), has been proven to be beneficial in such diseases. Thus, strategies to increase TIMP-3 bioavailability in the tissue have been sought for development of therapeutics. Nevertheless, high levels of TIMP-3 may lead to mechanism-based side-effects, as its overall effects on cell behavior are still unknown. In this study, we used a high-resolution mass-spectrometry-based workflow to analyze alterations induced by sustained expression of TIMP-3 in the cell surfaceome. In agreement with its multifunctional properties, TIMP-3 induced changes on the protein composition of the cell surface. We found that TIMP-3 had differential effects on metalloproteinase substrates, with several that accumulated in TIMP-3-overexpressing cells. In addition, our study identified potentially novel ADAM substrates, including ADAM15, whose levels at the cell surface are regulated by the inhibitor. In conclusion, our study reveals that high levels of TIMP-3 induce modifications in the cell surfaceome and identifies molecular pathways that can be deregulated via TIMP-3-based therapies.

The intestinal epithelium fulfils pleiotropic functions in nutrient uptake, waste elimination, and immune surveillance while also forming a barrier against luminal toxins and gut-resident microbiota. Incessantly barraged by extraneous stresses, the intestine must continuously replenish its epithelial lining and regenerate the full gamut of specialized cell types that underpin its functions. Homeostatic remodelling is orchestrated by the intestinal stem cell (ISC) niche: a convergence of epithelial- and stromal-derived cues, which maintains ISCs in a multipotent state. Following demise of homeostatic ISCs post injury, plasticity is pervasive among multiple populations of reserve stem-like cells, lineage-committed progenitors, and/or fully differentiated cell types, all of which can contribute to regeneration and repair. Failure to restore the epithelial barrier risks seepage of toxic luminal contents, resulting in inflammation and likely predisposing to tumour formation. Here, we explore how homeostatic niche-signalling pathways are subverted in tumorigenesis, enabling ISCs to gain autonomy from niche restraints ("ISC emancipation") and transform into cancer stem cells capable of driving tumour initiation, progression, and therapy resistance. We further consider the implications of the pervasive plasticity of the intestinal epithelium for the trajectory of colorectal cancer, the emergence of distinct molecular subtypes, the propensity to metastasize, and the development of effective therapeutic strategies.

Asthma is one of the most common respiratory disease that affects both children and adults worldwide, with diverse phenotypes and underlying pathogenetic mechanisms poorly understood. As technology in genome sequencing progressed, scientific efforts were made to explain and predict asthma's complexity and heterogeneity, and genome-wide association studies (GWAS) quickly became the preferred study method. Several gene markers and loci associated with asthma susceptibility, atopic and childhood-onset asthma were identified during the last few decades. Markers near the <i>ORMDL3/GSDMB</i> genes were associated with childhood-onset asthma, interleukin (IL)33 and <i>IL1RL1</i> SNPs were associated with atopic asthma, and the <i>Thymic Stromal Lymphopoietin (TSLP)</i> gene was identified as protective against the risk to TH2-asthma. The latest efforts and advances in identifying and decoding asthma susceptibility are focused on epigenetics, heritable characteristics that affect gene expression without altering DNA sequence, with DNA methylation being the most described mechanism. Other less studied epigenetic mechanisms include histone modifications and alterations of miR expression. Recent findings suggest that the DNA methylation pattern is tissue and cell-specific. Several studies attempt to describe DNA methylation of different types of cells and tissues of asthmatic patients that regulate airway remodeling, phagocytosis, and other lung functions in asthma. In this review, we attempt to briefly present the latest advancements in the field of genetics and mainly epigenetics concerning asthma susceptibility.

Cortical interneurons (cINs) are substantially affected in Schizophrenia (SCZ) and enriched for SCZ heritability during development. To understand SCZ-specific changes in these cells during development, we isolated migratory cINs from cIN spheres derived from 5 healthy control (HC) and 5 SCZ induced pluripotent stem cell lines (iPSCs). Transcriptome analyses show dysregulation in extracellular matrix pathways as the major disturbances in SCZ migratory cINs, whereas sphere cINs show dysregulation in immune pathways. This result suggests the importance of using homogeneous cell populations to identify stage-specific abnormalities and provides a platform to further study the biology of schizophrenia pathogenesis during early development.

The expression patterns of some cytokines were compared by RT-qPCR between lambs with and without Taenia hydatigena larvae vesicular concentrate (ThLVC) administration and subsequent infection with Haemonchus contortus. Lambs that received ThLVC prior to infection with H. contortus showed lower (p < 0.03) cumulative FEC (AUC = 18450 ± 3384) than infected lambs who did not receive ThLVC (AUC = 31081 ± 3277). Lambs infected with H. contortus, in general, overexpressed Th1 and Th2 cytokines in abomasal mucosa and abomasal lymph nodes, which seems to indicate a generalized and nonpolarized activation of the immune response by H. contortus. The main immunomodulatory effects of ThLVC were observed in the abomasal fundic region. The lambs that were given ThLVC prior to infection strongly overexpressed most of the studied cytokines representing the Th1 (IFNγ and IL2) and Th2 profiles (IL4, IL5, IL6 and IL10), proinflammatory cytokines (SOD1 and PRDX6) and IgE receptor; in contrast, lambs that were infected but did not receive ThLVC only moderately overexpressed IFNγ, IL4 and IL6. The absence of the significant overexpression of cytokines in lambs that only received ThLVC suggests that this derived from T. hydatigena does not have a stimulating effect per se; however, the presence of H. contortus did produce the highest expression (p < 0.01) cytokine profile among lambs that received ThLVC prior to infection compared to those who did not receive it, so its effect seems to be immunomodulatory and not only immunostimulatory.

<h4>Background</h4>Primary osteoporosis is a common complication of aging and menopause. The negative effects of osteoporosis in the coming years will increase by increasing life expectancy and population aging. The purpose of this research was to compare health-promoting lifestyle and quality of life in postmenopausal women with and without primary osteoporosis.<h4>Materials and methods</h4>This cross-sectional analytical research was conducted on 445 postmenopausal women aged 50-65 selected by simple random sampling in Tabriz health centers from September 2018 to July 2019. Data collection instruments included demographic, midwifery, anthropometric, health-promoting lifestyle profile II and menopausal quality-of-life questionnaire questionnaires, and serum test checklist (25-hydroxy vitamin D, complete blood count/diff, thyroid-stimulating hormone, fasting blood sugar, Calcium, and Phosphor). Dual-energy X-ray absorptiometry method was used to measure bone density. Data were analyzed using SPSS/23 through descriptive and inferential statistics such as Chi-square, independent <i>t</i>-test, Mann-Whitney, and multiple regression.<h4>Results</h4>The mean score of lifestyle was 141.2 ± 21.9 in normal and 127.2 ± 25.4 in osteoporosis group, and differences were statistically significant in total score (<i>P</i> < 0.001) and all subdomains. The mean score of quality of life was 3.9 ± 1.2 in the normal and 4.5 ± 1.4 in the osteoporotic group. The differences were significant in total score (<i>P</i> < 0.001) and all subdomains except for sexual function subdomain (<i>P</i> = 0.064). Logistic regression adjusted for confounders indicated by one unit increase in total lifestyle score, the odds of primary osteoporosis reduced by 2.2% (adjusted odds ratio [0.95% confidence interval]: 0.978 [0.963-0.994], <i>P</i> = 0.006).<h4>Conclusion</h4>To prevent of primary osteoporosis and improve the quality of life of postmenopausal women, it seems that education and implementation of health-promoting lifestyle are essential. The research findings can be used to plan for health care in middle and old ages.

<h4>Background</h4>Nonalcoholic fatty liver disease (NAFLD) is becoming the most common cause of cirrhosis. Although magnetic resonance spectroscopy (MRS) is considered the gold standard, it has a few limitations. The role of diffusion-weighted imaging (DWI), which is a simpler sequence, in the diagnosis and grading of fatty liver is not well studied. The aim of the study was to investigate the value of DWI in the diagnosis and grading of hepatic steatosis in patients with NAFLD.<h4>Materials and methods</h4>Fifty-one adults (mean age: 38 years; 28 men, 23 women) with NAFLD, diagnosed clinically and by ultrasonography (USG), were included in the study after obtaining informed consent and approval from the institute ethics committee. USG was performed for grading of hepatic steatosis in all patients, followed by magnetic resonance imaging with DWI and MRS, on a 1.5T scanner. The mean apparent diffusion coefficient (ADC) values and proton density fat fraction (PDFF) were calculated, and MRS was used as the gold standard. The mean ADC values were compared with the PDFF and USG grades.<h4>Results</h4>There was a weak correlation between ADC values and PDFF (r = -0.36; <i>P</i> < 0.05). In addition, there was a weak correlation between the ADC values of the liver and USG grade (r = -0.34; <i>P</i> < 0.05). However, an overall increase in USG grades and PDFF was associated with decrease in the mean ADC value (<i>P</i> < 0.001).<h4>Conclusion</h4>DWI is not accurate in the diagnosis and grading of hepatic steatosis in patients with NAFLD. However, a significant increase in fat deposition in the liver lowers the ADC values.

Non-alcoholic fatty liver disease (NAFLD) has become the most common chronic liver disorder worldwide. Murine models of NAFLD have been widely used to explore its pathogenesis. In this study, we performed a systematic evaluation of hepatic genome-wide mRNA expression by RNA-Sequencing using three mouse models of NAFLD: leptin receptor deficient db/db mice, high-fat high-sugar diet (HSHF)-induced obese mice, and dexamethasone (DEX)-induced NAFLD mice. As a result, we found both distinct and common pathways in the regulation of lipid metabolism from transcriptomes of three mouse models. Moreover, only a total of 12 differentially expressed genes (DEGs) were commonly detected among all three mouse groups, indicating very little overlap among all three models. Therefore, our results suggest that NAFLD is a heterogeneous disease with highly variable molecular mechanisms.

<h4>Background</h4>Lymphopenia is the most important criterion of mortality and discharging feature for patients infected with coronavirus disease 2019 (COVID-19). This study aimed to investigate the clinical impact of a low molecular weight heparin (LMWH) treatment on the clinical course of COVID-19.<h4>Materials and methods</h4>Patients’ clinical symptoms, radiologic outcomes, hematologic, biochemical, D-dimer, and C-reactive protein (CRP) results were obtained from their medical records. Participants were separated into 2 groups: one was treated with LMWH and the other was not. Improvement in the patients was compared before and after treatment.<h4>Results</h4>Ninety-six patients who were diagnosed with COVID-19 between April and May 2020 were retrospectively analyzed. The multivariable analysis showed that the count of lymphocytes, D-dimer, and CRP levels were significantly improved in the LMWH group, as compared to the control group (OR, (95% CI) 0.628 (0.248–0.965), P < 0.001); OR, (95% CI) 0.356 (0.089–0.674), P < 0.001, respectively). The area under the receiver operating characteristic (ROC) curve analysis was AUC: 0.679 ± 0.055, 0.615 ± 0.058, and 0.633 ± 0.057, respectively; the β-value was found to be –1.032, –0.026, and –0.465, respectively.<h4>Conclusion</h4>The LMWH treatment group demonstrated better laboratory findings, including recovery in the lymphocyte count, CRP, and D-dimer results.

<h4>Background</h4>To support patient-centred care, healthcare organisations increasingly offer patients access to data stored in the institutional electronic health record (EHR).<h4>Objectives</h4>Primary objective 1. To assess the effects of providing adult patients with access to electronic health records (EHRs) alone or with additional functionalities on a range of patient, patient-provider, and health resource consumption outcomes, including patient knowledge and understanding, patient empowerment, patient adherence, patient satisfaction with care, adverse events, health-related quality of life, health-related outcomes, psychosocial health outcomes, health resource consumption, and patient-provider communication. Secondary objective 1. To assess whether effects of providing adult patients with EHR access alone versus EHR access with additional functionalities differ among patient groups according to age, educational level, or different status of disease (chronic or acute).<h4>Search methods</h4>We searched CENTRAL, MEDLINE, Embase, PsycINFO, CINAHL, and Scopus in June 2017 and in April 2020.<h4>Selection criteria</h4>Randomised controlled trials and cluster-randomised trials of EHR access with or without additional functionalities for adults with any medical condition.<h4>Data collection and analysis</h4>We used standard Cochrane methodological procedures.<h4>Main results</h4>We included 10 studies with 78 to 4500 participants and follow-up from 3 to 24 months. Nine studies assessed the effects of EHR with additional functionalities, each addressing a subset of outcomes sought by this review. Five studies focused on patients with diabetes mellitus, four on patients with specific diseases, and one on all patients. All studies compared EHR access alone or with additional functionalities plus usual care versus usual care only. No studies assessing the effects of EHR access alone versus EHR access with additional functionalities were identified. Interventions required a variety of data within the EHR, such as patient history, problem list, medication, allergies, and lab results. In addition to EHR access, eight studies allowed patients to share self-documented data, seven offered individualised disease management functions, seven offered educational disease-related information, six supported secure communication, and one offered preventive reminders. Only two studies were at low or unclear risk of bias across domains. Meta-analysis could not be performed, as participants, interventions, and outcomes were too heterogeneous, and most studies presented results based on different adjustment methods or variables. The quality of evidence was rated as low or very low across outcomes. Overall differences between intervention and control groups, if any, were small. The relevance of any small effects remains unclear for most outcomes because in most cases, trial authors did not define a minimal clinically important difference. Overall, results suggest that the effects of EHR access alone and with additional functionalities are mostly uncertain when compared with usual care. Patient knowledge and understanding: very low-quality evidence is available from one study, so we are uncertain about effects of the intervention on patient knowledge about diabetes and blood glucose testing. Patient empowerment: low-quality evidence from three studies suggests that the intervention may have little or no effect on patient empowerment measures. Patient adherence: low-quality evidence from two studies suggests that the intervention may slightly improve adherence to the process of monitoring risk factors and preventive services. Effects on medication adherence are conflicting in two studies; this may or may not improve to a clinically relevant degree. Patient satisfaction with care: low-quality evidence from three studies suggests that the intervention may have little or no effect on patient satisfaction, with conflicting results. Adverse events: two small studies reported on mortality; one of these also reported on serious and other adverse events, but sample sizes were too small for small differences to be detected. Therefore, low-quality evidence suggests that the intervention may have little to no effect on mortality and other adverse events. Health-related quality of life: only very low-quality evidence from one study is available. We are uncertain whether the intervention improves disease-specific quality of life of patients with asthma. Health-related outcomes: low-quality evidence from eight studies suggests that the intervention may have little to no effect on asthma control, glycosylated haemoglobin (HbA1c) levels, blood pressure, low-density lipoprotein or total cholesterol levels, body mass index or weight, or 10-year Framingham risk scores. Low-quality evidence from one study suggests that the composite scores of risk factors for diabetes mellitus may improve slightly with the intervention, but there is uncertainty about effects on ophthalmic medications or intraocular pressure. Psychosocial health outcomes: no study investigated psychosocial health outcomes in a more than anecdotal way. Health resource consumption: low-quality evidence for adult patients in three studies suggests that there may be little to no effect of the intervention on different measures of healthcare use. Patient-provider communication: very low-quality evidence is available from a single small study, and we are uncertain whether the intervention improves communication measures, such as the number of messages sent.<h4>Authors' conclusions</h4>The effects of EHR access with additional functionalities in comparison with usual care for the most part are uncertain. Only adherence to the process of monitoring risk factors and providing preventive services as well as a composite score of risk factors for diabetes mellitus may improve slightly with EHR access with additional functionalities. Due to inconsistent terminology in this area, our search may have missed relevant studies. As the overall quality of evidence is very low to low, future research is likely to change these results. Further trials should investigate the impact of EHR access in a broader range of countries and clinical settings, including more patients over a longer period of follow-up, as this may increase the likelihood of detecting effects of the intervention, should these exist. More studies should focus on assessing outcomes such as patient empowerment and behavioural outcomes, rather than concentrating on health-related outcomes alone. Future studies should distinguish between effects of EHR access only and effects of additional functionalities, and investigate the impact of mobile EHR tools. Future studies should include information on usage patterns, and consider the potential for widening health inequalities with implementation of EHR access. A taxonomy for EHR access and additional functionalities should be developed to promote consistency and comparability of outcome measures, and facilitate future reviews by better enabling cross-study comparisons.

The miniaturization of polymerase chain reaction (PCR) using drop-based microfluidics allows for amplification of single nucleic acids in aqueous picoliter-sized drops. Accurate data collection during PCR requires that drops remain stable to coalescence during thermocycling and drop contents are retained. Following systematic testing of known PCR additives, we identified an optimized formulation of 1% w/v Tween-20, 0.8 μg/μL bovine serum albumin, 1 M betaine in the aqueous phase, and 3 wt % (w/w) of the polyethylene glycol-perfluoropolyether₂ surfactant in the oil phase of 50 μm diameter drops that maintains drop stability and prevents dye transport. This formulation enables a method we call off-chip drop reverse transcription quantitative PCR (OCD RT-qPCR) in which drops are thermocycled in a qPCR machine and sampled at various cycle numbers "off-chip", or outside of a microfluidic chip. qPCR amplification curves constructed from hundreds of individual drops using OCD RT-qPCR and imaged using epifluorescence microscopy correlate with amplification curves of ≈300,000 drops thermocycled using a qPCR machine. To demonstrate the utility of OCD RT-qPCR, influenza A virus (IAV) RNA was detected down to a single viral genome copy per drop, or 0.320 cpd. This work was extended to perform multiplexed detection of IAV M gene RNA and cellular β-actin DNA in drops, and direct amplification of IAV genomes from infected cells without a separate RNA extraction step. The optimized additive formulation and the OCD-qPCR method allow for drop-based RT-qPCR without complex devices and demonstrate the ability to quantify individual or rare nucleic acid species within drops with minimal processing.

The ubiquitin-proteasome system (UPS) and autophagy are two major quality control processes whose impairment is linked to a wide variety of diseases. The coordination between UPS and autophagy remains incompletely understood. Here, we show that ubiquitin ligase UBE3C and deubiquitinating enzyme TRABID reciprocally regulate K29/K48-branched ubiquitination of VPS34. We find that this ubiquitination enhances the binding of VPS34 to proteasomes for degradation, thereby suppressing autophagosome formation and maturation. Under ER and proteotoxic stresses, UBE3C recruitment to phagophores is compromised with a concomitant increase of its association with proteasomes. This switch attenuates the action of UBE3C on VPS34, thereby elevating autophagy activity to facilitate proteostasis, ER quality control and cell survival. Specifically in the liver, we show that TRABID-mediated VPS34 stabilization is critical for lipid metabolism and is downregulated during the pathogenesis of steatosis. This study identifies a ubiquitination type on VPS34 and elucidates its cellular fate and physiological functions in proteostasis and liver metabolism.

Despite the immense importance of enzyme-substrate reactions, there is a lack of general and unbiased tools for identifying and prioritizing substrate proteins that are modified by the enzyme on the structural level. Here we describe a high-throughput unbiased proteomics method called System-wide Identification and prioritization of Enzyme Substrates by Thermal Analysis (SIESTA). The approach assumes that the enzymatic post-translational modification of substrate proteins is likely to change their thermal stability. In our proof-of-concept studies, SIESTA successfully identifies several known and novel substrate candidates for selenoprotein thioredoxin reductase 1, protein kinase B (AKT1) and poly-(ADP-ribose) polymerase-10 systems. Wider application of SIESTA can enhance our understanding of the role of enzymes in homeostasis and disease, opening opportunities to investigate the effect of post-translational modifications on signal transduction and facilitate drug discovery.

Osteoarthritis causes pain and functional disability for over 500 million people worldwide. To develop disease-stratifying tools and modifying therapies, we need a better understanding of the molecular basis of the disease in relevant tissue and cell types. Here, we study primary cartilage and synovium from 115 patients with osteoarthritis to construct a deep molecular signature map of the disease. By integrating genetics with transcriptomics and proteomics, we discover molecular trait loci in each tissue type and omics level, identify likely effector genes for osteoarthritis-associated genetic signals and highlight high-value targets for drug development and repurposing. These findings provide insights into disease aetiopathology, and offer translational opportunities in response to the global clinical challenge of osteoarthritis.

Methionine (Met) is an essential amino acid and critical precursor to the cellular methyl donor S-adenosylmethionine. Unlike nontransformed cells, cancer cells have a unique metabolic requirement for Met and are unable to proliferate in growth media where Met is replaced with its metabolic precursor, homocysteine. This metabolic vulnerability is common among cancer cells regardless of tissue origin and is known as "methionine dependence", "methionine stress sensitivity", or the Hoffman effect. The response of lipids to Met stress, however, is not well-understood. Using mass spectroscopy, label-free vibrational microscopy, and next-generation sequencing, we characterize the response of lipids to Met stress in the triple-negative breast cancer cell line MDA-MB-468 and its Met stress insensitive derivative, MDA-MB-468res-R8. Lipidome analysis identified an immediate, global decrease in lipid abundances with the exception of triglycerides and an increase in lipid droplets in response to Met stress specifically in MDA-MB-468 cells. Furthermore, specific gene expression changes were observed as a secondary response to Met stress in MDA-MB-468, resulting in a downregulation of fatty acid metabolic genes and an upregulation of genes in the unfolded protein response pathway. We conclude that the extensive changes in lipid abundance during Met stress is a direct consequence of the modified metabolic profile previously described in Met stress-sensitive cells. The changes in lipid abundance likely results in changes in membrane composition inducing the unfolded protein response we observe.

The translocator protein (TSPO) is a 18kDa transmembrane protein, ubiquitously present in human mitochondria. It is overexpressed in tumor cells and at the sites of neuroinflammation, thus representing an important biomarker, as well as a promising drug target. In mammalian TSPO, there are cholesterol-binding motifs, as well as a binding cavity able to accommodate different chemical compounds. Given the lack of structural information for the human protein, we built a model of human (<i>h</i>) TSPO in the apo state and in complex with PK11195, a molecule routinely used in positron emission tomography (PET) for imaging of neuroinflammatory sites. To better understand the interactions of PK11195 and cholesterol with this pharmacologically relevant protein, we ran molecular dynamics simulations of the apo and holo proteins embedded in a model membrane. We found that: (i) PK11195 stabilizes <i>h</i>TSPO structural fold; (ii) PK11195 might enter in the binding site through transmembrane helices I and II of <i>h</i>TSPO; (iii) PK11195 reduces the frequency of cholesterol binding to the lower, N-terminal part of <i>h</i>TSPO in the inner membrane leaflet, while this impact is less pronounced for the upper, C-terminal part in the outer membrane leaflet, where the ligand binding site is located; (iv) very interestingly, cholesterol most frequently binds <i>simultaneously</i> to the so-called CRAC and CARC regions in TM V in the free form (residues L150-X-Y152-X(3)-R156 and R135-X(2)-Y138-X(2)-L141, respectively). However, when the protein is in complex with PK11195, cholesterol binds equally frequently to the CRAC-resembling motif that we observed in TM I (residues L17-X(2)-F20-X(3)-R24) and to CRAC in TM V. We expect that the CRAC-like motif in TM I will be of interest in future experimental investigations. Thus, our MD simulations provide insight into the structural features of <i>h</i>TSPO and the previously unknown interplay between PK11195 and cholesterol interactions with this pharmacologically relevant protein.

Teneurins have been identified in vertebrates as four different genes (TENM1-4), coding for membrane proteins that are mainly involved in embryonic and neuronal development. Genetic studies have correlated them with various diseases, including developmental problems, neurological disorders and congenital general anosmia. There is some evidence to suggest their possible involvement in cancer initiation and progression, and drug resistance. Indeed, mutations, chromosomal alterations and the deregulation of teneurins expression have been associated with several tumor types and patient survival. However, the role of teneurins in cancer-related regulatory networks is not fully understood, as both a tumor-suppressor role and pro-tumoral functions have been proposed, depending on tumor histotype. Here, we summarize and discuss the literature data on teneurins expression and their potential role in different tumor types, while highlighting the possibility of using teneurins as novel molecular diagnostic and prognostic biomarkers and as targets for cancer treatments, such as immunotherapy, in some tumors.

High-dose intravenously administered vitamin C (IVC) is widely used in cancer patients by complementary and alternative medicine practitioners. The most frequent indications for IVC therapy result from the belief in its effectiveness as a potent anti-cancer agent which additionally enhances chemosensitivity of cancer cells and reduces chemotherapy-related toxicities and fatigue intensity. In this narrative review, we decided to deal with this issue, trying to answer the question whether there is any scientific evidence supporting the rationale for application of high-dose IVC therapy in advanced-stage cancer patients. Although results obtained from preclinical studies demonstrated that millimolar ascorbate plasma concentrations achievable only after IVC administration were cytotoxic to fast-growing malignant cells and inhibited tumor growth as well as prolonged the survival of laboratory animals, such positive effects were not found in human studies with advanced-stage cancer patients. We also have not found the rationale for the use of IVC to increase the effectiveness of chemotherapy and to reduce the chemotherapy-induced toxicity in the above mentioned group. Nevertheless, in palliative care, high-dose IVC might be considered as a therapy improving the quality of life and reducing cancer-related symptoms, such as fatigue and bone pain. However, because of the absence of placebo-controlled randomized trials on IVC efficacy in advanced-stage cancer patients, the placebo effect cannot be excluded.

<i>Cannabis sativa</i> (<i>Cannabis</i>) is one of the world's most well-known, yet maligned plant species. However, significant recent research is starting to unveil the potential of <i>Cannabis</i> to produce secondary compounds that may offer a suite of medical benefits, elevating this unique plant species from its illicit narcotic status into a genuine biopharmaceutical. This review summarises the lengthy history of <i>Cannabis</i> and details the molecular pathways that underpin the production of key secondary metabolites that may confer medical efficacy. We also provide an up-to-date summary of the molecular targets and potential of the relatively unknown minor compounds offered by the <i>Cannabis</i> plant. Furthermore, we detail the recent advances in plant science, as well as synthetic biology, and the pharmacology surrounding <i>Cannabis.</i> Given the relative infancy of <i>Cannabis</i> research, we go on to highlight the parallels to previous research conducted in another medically relevant and versatile plant, <i>Papaver somniferum</i> (opium poppy), as an indicator of the possible future direction of <i>Cannabis</i> plant biology. Overall, this review highlights the future directions of cannabis research outside of the medical biology aspects of its well-characterised constituents and explores additional avenues for the potential improvement of the medical potential of the <i>Cannabis</i> plant.

With the extension of life span in recent decades, there is an increasing burden of late-onset neurodegenerative diseases, for which effective treatments are lacking. Neurodegenerative diseases include the widespread Alzheimer's disease (AD) and Parkinson's disease (PD), the less frequent Huntington's disease (HD) and Amyotrophic Lateral Sclerosis (ALS) and also rare early-onset diseases linked to mutations that cause protein aggregation or loss of function in genes that maintain protein homeostasis. The difficulties in applying gene therapy approaches to tackle these diseases is drawing increasing attention to strategies that aim to inhibit cellular toxicity and restore homeostasis by intervening in cellular pathways. These include the unfolded protein response (UPR), activated in response to endoplasmic reticulum (ER) stress, a cellular affliction that is shared by these diseases. Special focus is turned to the PKR-like ER kinase (PERK) pathway of the UPR as a target for intervention. However, the complexity of the pathway and its ability to promote cell survival or death, depending on ER stress resolution, has led to some confusion in conflicting studies. Both inhibition and activation of the PERK pathway have been reported to be beneficial in disease models, although there are also some reports where they are counterproductive. Although with the current knowledge a definitive answer cannot be given on whether it is better to activate or to inhibit the pathway, the most encouraging strategies appear to rely on boosting some steps without compromising downstream recovery.

Smith-Lemli-Opitz Syndrome (SLOS) results from mutations in the gene encoding the enzyme DHCR7, which catalyzes conversion of 7-dehydrocholesterol (7DHC) to cholesterol (CHOL). Rats treated with a DHCR7 inhibitor serve as a SLOS animal model, and exhibit progressive photoreceptor-specific cell death, with accumulation of 7DHC and oxidized sterols. To understand the basis of this cell type specificity, we performed transcriptomic analyses on a photoreceptor-derived cell line (661W), treating cells with two 7DHC-derived oxysterols, which accumulate in tissues and bodily fluids of SLOS patients and in the rat SLOS model, as well as with CHOL (negative control), and evaluated differentially expressed genes (DEGs) for each treatment. Gene enrichment analysis and compilation of DEG sets indicated that endoplasmic reticulum stress, oxidative stress, DNA damage and repair, and autophagy were all highly up-regulated pathways in oxysterol-treated cells. Detailed analysis indicated that the two oxysterols exert their effects via different molecular mechanisms. Changes in expression of key genes in highlighted pathways (<i>Hmox1</i>, <i>Ddit3</i>, <i>Trib3</i>, and <i>Herpud1</i>) were validated by immunofluorescence confocal microscopy. The results extend our understanding of the pathobiology of retinal degeneration and SLOS, identifying potential new druggable targets for therapeutic intervention into these and other related orphan diseases.

In search of anti-inflammatory compounds, novel scaffolds containing isonicotinoyl motif were synthesized via an efficient strategy. The compounds were screened for their in vitro anti-inflammatory activity. Remarkably high activities were observed for isonicotinates <b>5</b>-<b>6</b> and <b>8a</b>-<b>8b</b>. The compound <b>5</b> exhibits an exceptional IC₅₀ value (1.42 ± 0.1 µg/mL) with 95.9% inhibition at 25 µg/mL, which is eight folds better than the standard drug ibuprofen (11.2 ± 1.9 µg/mL). To gain an insight into the mode of action of anti-inflammatory compounds, molecular docking studies were also performed. Decisively, further development and fine tuning of these isonicotinates based scaffolds for the treatment of various aberrations is still a wide-open field of research.

Various bone graft products are commercially available worldwide. However, there is no clear consensus regarding the appropriate bone graft products in different clinical situations. This review is intended to summarize bone graft products, especially alloplastic bone substitutes that are available in multiple countries. It also provides dental clinicians with detailed and accurate information concerning these products. Furthermore, it discusses the prospects of alloplastic bone substitutes based on an analysis of the current market status, as well as a comparison of trends among countries. In this review, we focus on alloplastic bone substitutes approved in the United States, Japan, and Korea for use in periodontal and bone regeneration. According to the Food and Drug Administration database, 87 alloplastic bone graft products have been approved in the United States since 1996. According to the Pharmaceuticals and Medical Devices Agency database, 10 alloplastic bone graft products have been approved in Japan since 2004. According to the Ministry of Health and Welfare database, 36 alloplastic bone graft products have been approved in Korea since 1980. The approved products are mainly hydroxyapatite, β-tricalcium phosphate, and biphasic calcium phosphate. The formulations of the products differed among countries. The development of new alloplastic bone products has been remarkable. In the near future, alloplastic bone substitutes with safety and standardized quality may be the first choice instead of autologous bone; they may offer new osteoconductive and osteoinductive products with easier handling form and an adequate resorption rate, which can be used with growth factors and/or cell transplantation. Careful selection of alloplastic bone graft products is necessary to achieve predictable outcomes according to each clinical situation.

Skeletal muscle possesses remarkable plasticity that permits functional adaptations to a wide range of signals such as motor input, exercise, and disease. Small animal models have been pivotal in elucidating the molecular mechanisms regulating skeletal muscle adaptation and plasticity. However, these small animal models fail to accurately model human muscle disease resulting in poor clinical success of therapies. Here, we review the potential of <i>in vitro</i> three-dimensional tissue-engineered skeletal muscle models to study muscle function, plasticity, and disease. First, we discuss the generation and function of <i>in vitro</i> skeletal muscle models. We then discuss the genetic, neural, and hormonal factors regulating skeletal muscle fiber-type <i>in vivo</i> and the ability of current <i>in vitro</i> models to study muscle fiber-type regulation. We also evaluate the potential of these systems to be utilized in a patient-specific manner to accurately model and gain novel insights into diseases such as Duchenne muscular dystrophy (DMD) and volumetric muscle loss. We conclude with a discussion on future developments required for tissue-engineered skeletal muscle models to become more mature, biomimetic, and widely utilized for studying muscle physiology, disease, and clinical use.

Vertebrates have evolved a complex immune system required for the identification of and coordinated response to harmful pathogens. Migratory species spend periods of their life-cycle in more than one environment, and their immune system consequently faces a greater diversity of pathogens residing in different environments. In facultatively anadromous salmonids, individuals may spend parts of their life-cycle in freshwater and marine environments. For species such as the brown trout <i>Salmo trutta</i>, sexes differ in their life-histories with females more likely to migrate to sea while males are more likely to stay and complete their life-cycle in their natal river. Salmonids have also undergone a lineage-specific whole genome duplication event, which may provide novel immune innovations but our current understanding of the differences in salmonid immune expression between the sexes is limited. We characterized the brown trout immune gene repertoire, identifying a number of canonical immune genes in non-salmonid teleosts to be duplicated in <i>S. trutta</i>, with genes involved in innate and adaptive immunity. Through genome-wide transcriptional profiling ("RNA-seq") of male and female livers to investigate sex differences in gene expression amplitude and alternative splicing, we identified immune genes as being generally male-biased in expression. Our study provides important insights into the evolutionary consequences of whole genome duplication events on the salmonid immune gene repertoire and how the sexes differ in constitutive immune expression.

The development of a non-sputum-based, point-of-care diagnostic test for tuberculosis (TB) is a priority in the global effort to combat this disease, particularly in resource-constrained settings. Previous studies have identified host biomarker signatures which showed potential, but there is a need to validate and refine these for development as a test. We recruited 1,403 adults presenting with symptoms suggestive of pulmonary TB at primary healthcare clinics in six countries from West, East and Southern Africa. Of the study cohort, 326 were diagnosed with TB and 787 with other respiratory diseases, from whom we randomly selected 1005 participants. Using Luminex^® technology, we measured the levels of 20 host biomarkers in serum samples which we used to evaluate the diagnostic accuracy of previously identified and novel bio-signatures. Our previously identified seven-marker bio-signature did not perform well (sensitivity: 89%, specificity: 60%). We also identified an optimal, two-marker bio-signature with a sensitivity of 94% and specificity of 69% in patients with no history of previous TB. This signature performed slightly better than C-reactive protein (CRP) alone. The cut-off value for a positive diagnosis differed for human immuno-deficiency virus (HIV)-positive and -negative individuals. Notably, we also found that no signature was able to diagnose TB adequately in patients with a prior history of the disease. We have identified a two-marker, pan-African bio-signature which is more robust than CRP alone and meets the World Health Organization (WHO) target product profile requirements for a triage test in both HIV-negative and HIV-positive individuals. This signature could be incorporated into a point-of-care device, greatly reducing the necessity for expensive confirmatory diagnostics and potentially reducing the number of cases currently lost to follow-up. It might also potentially be useful with individuals unable to provide sputum or with paucibacillary disease. We suggest that the performance of TB diagnostic signatures can be improved by incorporating the HIV-status of the patient. We further suggest that only patients who have never had TB be subjected to a triage test and that those with a history of previous TB be evaluated using more direct diagnostic techniques.

During bone resorption, the osteoclast must sustain an extraordinarily low pH environment, withstand immense ionic pressures, and coordinate nutrient and waste exchange across its membrane to sustain its unique structural and functional polarity. To achieve this, osteoclasts are equipped with an elaborate set of membrane transport proteins (pumps, transporters and channels) that serve as molecular 'gatekeepers' to regulate the bilateral exchange of ions, amino acids, metabolites and macromolecules across the ruffled border and basolateral domains. Whereas the importance of the vacuolar-ATPase proton pump and chloride voltage-gated channel 7 in osteoclasts has long been established, comparatively little is known about the contributions of other membrane transport proteins, including those categorized as secondary active transporters. In this Special Issue review, we provide a contemporary update on the 'ins and outs' of membrane transport proteins implicated in osteoclast differentiation, function and bone homeostasis and discuss their therapeutic potential for the treatment of metabolic bone diseases.

Small bowel neuroendocrine tumors are rare tumors with an increasing incidence over the last several decades. Early detection remains challenging because patients commonly develop symptoms late in the disease course, often after the tumors have metastasized. Although these tumors were thought to arise from sporadic genetic mutations, large epidemiological studies strongly support genetic predisposition and increased risk of disease in affected families. Recent studies of familial small bowel neuroendocrine tumors have identified several novel genetic mutations. Screening for familial small bowel neuroendocrine tumors can lead to earlier diagnosis and improved patient outcomes. This review aims to summarize the current knowledge of molecular changes seen in familial small bowel neuroendocrine tumors, identify clinical features specific to familial disease, and provide strategies for screening and treatment.

The development of genome-editing technologies in 1970s has discerned a new beginning in the field of science. Out of different genome-editing approaches such as Zing-finger nucleases, TALENs, and meganucleases, clustered regularly interspaced short palindromic repeats-CRISPR-associated protein 9 (CRISPR/Cas9) is a recent and versatile technology that has the ability of making changes to the genome of different organisms with high specificity. Cancer is a complex process that is characterized by multiple genetic and epigenetic changes resulting in abnormal cell growth and proliferation. As cancer is one of the leading causes of deaths worldwide, a large number of studies are done to understand the molecular mechanisms underlying the development of cancer. Because of its high efficiency and specificity, CRISPR/Cas9 has emerged as a novel and powerful tool in the field of cancer research. CRISPR/Cas9 has the potential to accelerate cancer research by dissecting tumorigenesis process, generating animal and cellular models, and identify drug targets for chemotherapeutic approaches. However, despite having tremendous potential, there are certain challenges associated with CRISPR/Cas9 such as safe delivery to the target, potential off-target effects and its efficacy which needs to be addressed prior to its clinical application. In this review, we give a gist of different genome-editing technologies with a special focus on CRISPR/Cas9 development, its mechanism of action and its applications, especially in different type of cancers. We also highlight the importance of CRISPR/Cas9 in generating animal models of different cancers. Finally, we present an overview of the clinical trials and discuss the challenges associated with translating CRISPR/Cas9 in clinical use.

Polycystic ovary syndrome (PCOS) and Hashimoto's thyroiditis (HT) are endocrine disorders that commonly occur among young women. A higher prevalence of HT in women with PCOS, relative to healthy individuals, is observed consistently. Combined occurrence of both diseases is associated with a higher risk of severe metabolic and reproductive complications. Genetic factors strongly impact the pathogenesis of both PCOS and HT and several susceptibility loci associated with a higher risk of both disorders have been identified. Furthermore, some candidate gene polymorphisms are thought to be functionally relevant; however, few genetic variants are proposed to be causally associated with the incidence of both disorders together.

The livestock sector is one of the most important sectors of the agricultural economy due to an increase in the demand for animal protein. This increase generates serious waste disposal concerns and has negative environmental consequences. Furthermore, the food production chain needs phosphorus (P), which is listed as a critical raw material due to its high demand and limited availability in Europe. Manure contains large amounts of P and other elements that may be recycled, in the frame of circular economy and "zero waste" principles, and reused as a by-product for fertilizer production and other applications. This paper focuses on the extraction and recovery of amorphous silica from rice husk poultry litter ash. Two different extraction procedures are proposed and compared, and the obtained silica is characterized. This work shows that amorphous silica can be recovered as an almost pure material rendering the residual ash free of P. It also addresses the possibility of more specific phosphorous extraction procedures <i>via</i> acid leaching.

Condensins are best known for their role in shaping chromosomes. Other functions such as organizing interphase chromatin and transcriptional control have been reported in yeasts and animals, but little is known about their function in plants. To elucidate the specific composition of condensin complexes and the expression of CAP-D2 (condensin I) and CAP-D3 (condensin II), we performed biochemical analyses in Arabidopsis. The role of CAP-D3 in interphase chromatin organization and function was evaluated using cytogenetic and transcriptome analysis in cap-d3 T-DNA insertion mutants. CAP-D2 and CAP-D3 are highly expressed in mitotically active tissues. In silico and pull-down experiments indicate that both CAP-D proteins interact with the other condensin I and II subunits. In cap-d3 mutants, an association of heterochromatic sequences occurs, but the nuclear size and the general histone and DNA methylation patterns remain unchanged. Also, CAP-D3 influences the expression of genes affecting the response to water, chemicals, and stress. The expression and composition of the condensin complexes in Arabidopsis are similar to those in other higher eukaryotes. We propose a model for the CAP-D3 function during interphase in which CAP-D3 localizes in euchromatin loops to stiffen them and consequently separates centromeric regions and 45S rDNA repeats.

Human coronaviruses (HCoVs) cause mild to severe respiratory infection. Most of the common cold illnesses are caused by one of four HCoVs, namely HCoV-229E, HCoV-NL63, HCoV-HKU1 and HCoV-OC43. Several studies have applied global transcriptomic methods to understand host responses to HCoV infection, with most studies focusing on the pandemic severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome CoV (MERS-CoV) and the newly emerging SARS-CoV-2. In this study, Next Generation Sequencing was used to gain new insights into cellular transcriptomic changes elicited by alphacoronavirus HCoV-229E. HCoV-229E-infected MRC-5 cells showed marked downregulation of superpathway of cholesterol biosynthesis and eIF2 signaling pathways. Moreover, upregulation of cyclins, cell cycle control of chromosomal replication, and the role of BRCA1 in DNA damage response, alongside downregulation of the cell cycle G1/S checkpoint, suggest that HCoV-229E may favors S phase for viral infection. Intriguingly, a significant portion of key factors of cell innate immunity, interferon-stimulated genes (ISGs) and other transcripts of early antiviral response genes were downregulated early in HCoV-229E infection. On the other hand, early upregulation of the antiviral response factor Apolipoprotein B mRNA editing enzyme catalytic subunit 3B (APOBEC3B) was observed. APOBEC3B cytidine deaminase signature (C-to-T) was previously observed in genomic analysis of SARS-CoV-2 but not HCoV-229E. Higher levels of C-to-T mutations were found in countries with high mortality rates caused by SARS-CoV-2. APOBEC activity could be a marker for new emerging CoVs. This study will enhance our understanding of commonly circulating HCoVs and hopefully provide critical information about still-emerging coronaviruses.

<h4>Background</h4>A substantial number of infants infected with RSV develop severe symptoms requiring hospitalization. We currently lack accurate biomarkers that are associated with severe illness.<h4>Method</h4>We defined airway gene expression profiles based on RNA sequencing from nasal brush samples from 106 full-tem previously healthy RSV infected subjects during acute infection (day 1-10 of illness) and convalescence stage (day 28 of illness). All subjects were assigned a clinical illness severity score (GRSS). Using AIC-based model selection, we built a sparse linear correlate of GRSS based on 41 genes (NGSS1). We also built an alternate model based upon 13 genes associated with severe infection acutely but displaying stable expression over time (NGSS2).<h4>Results</h4>NGSS1 is strongly correlated with the disease severity, demonstrating a naïve correlation (ρ) of ρ = 0.935 and cross-validated correlation of 0.813. As a binary classifier (mild versus severe), NGSS1 correctly classifies disease severity in 89.6% of the subjects following cross-validation. NGSS2 has slightly less, but comparable, accuracy with a cross-validated correlation of 0.741 and classification accuracy of 84.0%.<h4>Conclusion</h4>Airway gene expression patterns, obtained following a minimally-invasive procedure, have potential utility for development of clinically useful biomarkers that correlate with disease severity in primary RSV infection.

DNA and RNA can fold into a variety of alternative conformations. In recent years, a particular nucleic acid structure was discussed to play a role in malignant transformation and cancer development. This structure is called a G-quadruplex (G4). G4 structure formation can drive genome instability by creating mutations, deletions and stimulating recombination events. The importance of G4 structures in the characterization of malignant cells was currently demonstrated in breast cancer samples. In this analysis a correlation between G4 structure formation and an increased intratumor heterogeneity was identified. This suggests that G4 structures might allow breast cancer stratification and supports the identification of new personalized treatment options. Because of the stability of G4 structures and their presence within most human oncogenic promoters and at telomeres, G4 structures are currently tested as a therapeutic target to downregulate transcription or to block telomere elongation in cancer cells. To date, different chemical molecules (G4 ligands) have been developed that aim to target G4 structures. In this review we discuss and compare G4 function and relevance for therapeutic approaches and their impact on cancer development for three cancer entities, which differ significantly in their amount and type of mutations: pancreatic cancer, leukemia and malignant melanoma. G4 structures might present a promising new strategy to individually target tumor cells and could support personalized treatment approaches in the future.

<h4>Background</h4>Triple negative breast cancer (TNBC) is one of the most biologically aggressive breast cancers and lacks effective treatment options, resulting in a poor prognosis. Therefore, studies aiming to explore new therapeutic strategies for advanced TNBC are urgently needed. According to recent studies, microRNA-124 (miR124) not only inhibits tumour growth but also increases the sensitivity of TNBC to paclitaxel (PTX), suggesting that a platform combining PTX and miR124 may be an advanced solution for TNBC.<h4>Results</h4>Herein, we constructed a stepped cleavable calcium phosphate composite lipid nanosystem (CaP/LNS) to codeliver PTX and miR124 (PTX/miR124-NP). PTX/miR124-NP exhibited superior tumor microenvironment responsive ability, in which the surface PEG layer was shed in the mildly acidic environment of tumor tissues and exposed oligomeric hyaluronic acid (o-HA) facilitated the cellular uptake of CaP/LNS by targeting the CD44 receptor on the surface of tumor cells. Inside tumour cells, o-HA detached from CaP/LNS due to the reduction of disulfide bonds by glutathione (GSH) and inhibited tumour metastasis. Then, PTX and miR124 were sequentially released from CaP/LNS and exerted synergistic antitumour effects by reversing the Epithelial-Mesenchymal Transition (EMT) process in MDA-MB-231 cells. Moreover, PTX/miR124-NP showed significant antitumour efficiency and excellent safety in mice bearing MDA-MB-231 tumours.<h4>Conclusion</h4>Based on these results, the codelivery of PTX and miR124 by the CaP/LNS nanosystem might be a promising therapeutic strategy for TNBC.

<h4>Background</h4>Clinical-grade whole-genome sequencing (cWGS) has the potential to become the standard of care within the clinic because of its breadth of coverage and lack of bias towards certain regions of the genome. Colorectal cancer presents a difficult treatment paradigm, with over 40% of patients presenting at diagnosis with metastatic disease. We hypothesised that cWGS coupled with 3' transcriptome analysis would give new insights into colorectal cancer.<h4>Methods</h4>Patients underwent PCR-free whole-genome sequencing and alignment and variant calling using a standardised pipeline to output SNVs, indels, SVs and CNAs. Additional insights into the mutational signatures and tumour biology were gained by the use of 3' RNA-seq.<h4>Results</h4>Fifty-four patients were studied in total. Driver analysis identified the Wnt pathway gene APC as the only consistently mutated driver in colorectal cancer. Alterations in the PI3K/mTOR pathways were seen as previously observed in CRC. Multiple private CNAs, SVs and gene fusions were unique to individual tumours. Approximately 30% of patients had a tumour mutational burden of > 10 mutations/Mb of DNA, suggesting suitability for immunotherapy.<h4>Conclusions</h4>Clinical whole-genome sequencing offers a potential avenue for the identification of private genomic variation that may confer sensitivity to targeted agents and offer patients new options for targeted therapies.

Fear dysregulation is one of the symptoms found in post-traumatic stress disorder (PTSD) patients. The functional abnormality of the hippocampus is known to be implicated in the development of such pathology. Peroxiredoxin 6 (PRDX6) belongs to the peroxiredoxin family. This antioxidant enzyme is expressed throughout the brain, including the hippocampus. Recent evidence reveals that PRDX6 plays an important role in redox regulation and the modulation of several signaling molecules involved in fear regulation. Thus, we hypothesized that PRDX6 plays a role in the regulation of fear memory. We subjected a systemic Prdx6 knockout (Prdx6^-/-) mice to trace fear conditioning and observed enhanced fear response after training. Intraventricular injection of lentivirus-carried mouse Prdx6 into the 3rd ventricle reduced the enhanced fear response in these knockout mice. Proteomic analysis followed by validation of western blot analysis revealed that several proteins in the MAPK pathway, such as NTRK2, AKT, and phospho-ERK1/2, cPLA2 were significantly upregulated in the hippocampus of Prdx6^-/- mice during the retrieval stage of contextual fear memory. The distribution of PRDX6 found in the astrocytes was also observed throughout the hippocampus. This study identifies PRDX6 as a participant in the regulation of fear response. It suggests that PRDX6 and related molecules may have important implications for understanding fear-dysregulation associated disorders like PTSD.

<h4>Background</h4>Epigenetic alterations may contribute to early detection of cancer. We evaluated the association of blood DNA methylation with lymphatic-hematopoietic cancers and, for comparison, with solid cancers. We also evaluated the predictive ability of DNA methylation for lymphatic-hematopoietic cancers.<h4>Methods</h4>Blood DNA methylation was measured using the Illumina Infinium methylationEPIC array in 2324 Strong Heart Study participants (41.4% men, mean age 56 years). 788,368 CpG sites were available for differential DNA methylation analysis for lymphatic-hematopoietic, solid and overall cancers using elastic-net and Cox regression models. We conducted replication in an independent population: the Framingham Heart Study. We also analyzed differential variability and conducted bioinformatic analyses to assess for potential biological mechanisms.<h4>Results</h4>Over a follow-up of up to 28 years (mean 15), we identified 41 lymphatic-hematopoietic and 394 solid cancer cases. A total of 126 CpGs for lymphatic-hematopoietic cancers, 396 for solid cancers, and 414 for overall cancers were selected as predictors by the elastic-net model. For lymphatic-hematopoietic cancers, the predictive ability (C index) increased from 0.58 to 0.87 when adding these 126 CpGs to the risk factor model in the discovery set. The association was replicated with hazard ratios in the same direction in 28 CpGs in the Framingham Heart Study. When considering the association of variability, rather than mean differences, we found 432 differentially variable regions for lymphatic-hematopoietic cancers.<h4>Conclusions</h4>This study suggests that differential methylation and differential variability in blood DNA methylation are associated with lymphatic-hematopoietic cancer risk. DNA methylation data may contribute to early detection of lymphatic-hematopoietic cancers.

Long-read and strand-specific sequencing technologies together facilitate the de novo assembly of high-quality haplotype-resolved human genomes without parent-child trio data. We present 64 assembled haplotypes from 32 diverse human genomes. These highly contiguous haplotype assemblies (average minimum contig length needed to cover 50% of the genome: 26 million base pairs) integrate all forms of genetic variation, even across complex loci. We identified 107,590 structural variants (SVs), of which 68% were not discovered with short-read sequencing, and 278 SV hotspots (spanning megabases of gene-rich sequence). We characterized 130 of the most active mobile element source elements and found that 63% of all SVs arise through homology-mediated mechanisms. This resource enables reliable graph-based genotyping from short reads of up to 50,340 SVs, resulting in the identification of 1526 expression quantitative trait loci as well as SV candidates for adaptive selection within the human population.

<h4>Purpose</h4>Perineural invasion (PNI) is associated with aggressive tumor behavior, recurrence, and metastasis, and can influence the administration of adjuvant treatment. However, standard histopathologic examination has limited sensitivity in detecting PNI and does not provide insights into its mechanistic underpinnings.<h4>Experimental design</h4>A multivariate Cox regression was performed to validate associations between PNI and survival in 2,029 patients across 12 cancer types. Differential expression and gene set enrichment analysis were used to learn PNI-associated programs. Machine learning models were applied to build a PNI gene expression classifier. A blinded re-review of hematoxylin and eosin (H&E) slides by a board-certified pathologist helped determine whether the classifier could improve occult histopathologic detection of PNI.<h4>Results</h4>PNI associated with both poor overall survival [HR, 1.73; 95% confidence interval (CI), 1.27-2.36; <i>P</i> < 0.001] and disease-free survival (HR, 1.79; 95% CI, 1.38-2.32; <i>P</i> < 0.001). Neural-like, prosurvival, and invasive programs were enriched in PNI-positive tumors (<i>P</i> _adj < 0.001). Although PNI-associated features likely reflect in part the increased presence of nerves, many differentially expressed genes mapped specifically to malignant cells from single-cell atlases. A PNI gene expression classifier was derived using random forest and evaluated as a tool for occult histopathologic detection. On a blinded H&E re-review of sections initially described as PNI negative, more specimens were reannotated as PNI positive in the high classifier score cohort compared with the low-scoring cohort (<i>P</i> = 0.03, Fisher exact test).<h4>Conclusions</h4>This study provides salient biological insights regarding PNI and demonstrates a role for gene expression classifiers to augment detection of histopathologic features.

Trisomy of human chromosome 21 (Down syndrome, DS) alters development of multiple organ systems, including the face and underlying skeleton. Besides causing stigmata, these facial dysmorphologies can impair vital functions such as hearing, breathing, mastication, and health. To investigate the therapeutic potential of green tea extracts containing epigallocatechin-3-gallate (GTE-EGCG) for alleviating facial dysmorphologies associated with DS, we performed an experimental study with continued pre- and postnatal treatment with two doses of GTE-EGCG supplementation in a mouse model of DS, and an observational study of children with DS whose parents administered EGCG as a green tea supplement. We evaluated the effect of high (100 mg/kg/day) or low doses (30 mg/kg/day) of GTE-EGCG, administered from embryonic day 9 to post-natal day 29, on the facial skeletal development in the Ts65Dn mouse model. In a cross-sectional observational study, we assessed the facial shape in DS and evaluated the effects of self-medication with green tea extracts in children from 0 to 18 years old. The main outcomes are 3D quantitative morphometric measures of the face, acquired either with micro-computed tomography (animal study) or photogrammetry (human study). The lowest experimentally tested GTE-EGCG dose improved the facial skeleton morphology in a mouse model of DS. In humans, GTE-EGCG supplementation was associated with reduced facial dysmorphology in children with DS when treatment was administered during the first 3 years of life. However, higher GTE-EGCG dosing disrupted normal development and increased facial dysmorphology in both trisomic and euploid mice. We conclude that GTE-EGCG modulates facial development with dose-dependent effects. Considering the potentially detrimental effects observed in mice, the therapeutic relevance of controlled GTE-EGCG administration towards reducing facial dysmorphology in young children with Down syndrome has yet to be confirmed by clinical studies.

This study presents a non-invasive, automated, clinical diagnostic system for early diagnosis of lung cancer that integrates imaging data from a single computed tomography scan and breath bio-markers obtained from a single exhaled breath to quickly and accurately classify lung nodules. CT imaging and breath volatile organic compounds data were collected from 47 patients. Spherical Harmonics-based shape features to quantify the shape complexity of the pulmonary nodules, 7th-Order Markov Gibbs Random Field based appearance model to describe the spatial non-homogeneities in the pulmonary nodule, and volumetric features (size) of pulmonary nodules were calculated from CT images. 27 VOCs in exhaled breath were captured by a micro-reactor approach and quantied using mass spectrometry. CT and breath markers were input into a deep-learning autoencoder classifier with a leave-one-subject-out cross validation for nodule classification. To mitigate the limitation of a small sample size and validate the methodology for individual markers, retrospective CT scans from 467 patients with 727 pulmonary nodules, and breath samples from 504 patients were analyzed. The CAD system achieved 97.8% accuracy, 97.3% sensitivity, 100% specificity, and 99.1% area under curve in classifying pulmonary nodules.

Within the oocytes of chicken preovulatory follicles, the engulfed yolk constitutes 99% of the oocyte content, while the small germinal disc (GD) (which contains the nucleus and 99% ooplasm) occupies only less than 1%. Relative to the position of the GD, the single granulosa cell layer surrounding the oocyte can be sub-divided into two sub-populations: granulosa cells proximal (named Gp cells) and distal (Gd cells) to the GD. It was reported that Gp cells and Gd cells differ in their morphology, proliferative rate and steroidogenic capacity, however, the underlying mechanism controlling granulosa cell heterogeneity remains unclear. Here we analyzed the transcriptomes of Gd and Gp cells of preovulatory (F5 and F1) follicles in chicken ovaries. We found that: (1) genes associated with cell cycle and DNA replication (CDK1, CCNB3 etc.) have comparatively higher expression levels in Gp cells than in Gd cells, while genes associated with steroidogenesis (CYP51A1, DHCR24) are highly expressed in Gd cells, indicating that Gp cells are likely more mitotic and less steroidogenic than Gd cells; (2) genes associated with extracellular matrix remodeling, cell adhesion and sperm binding (ZP3, ZP2) are differentially expressed in Gp and Gd cells; (3) Furthermore, signaling molecules (WNT4/IHH) and receptors for NGF (NGFR), epidermal growth factor (EGFR), gonadotropins (FSHR/LHR) and prostaglandin (PTGER3) are abundantly but differentially expressed in Gp and Gd cells. Taken together, our data strongly supports the notion that Gp and Gd cells of preovulatory follicles differ in their proliferation rate, steroidogenic activity, ECM organization and sperm binding capacity, which are likely controlled by gonadotropins and local ovarian factors, such as GD-derived factors.

Trim-Away is a recently developed technology that exploits off-the-shelf antibodies and the RING E3 ligase and cytosolic antibody receptor TRIM21 to carry out rapid protein depletion. How TRIM21 is catalytically activated upon target engagement, either during its normal immune function or when repurposed for targeted protein degradation, is unknown. Here we show that a mechanism of target-induced clustering triggers intermolecular dimerization of the RING domain to switch on the ubiquitination activity of TRIM21 and induce virus neutralization or drive Trim-Away. We harness this mechanism for selective degradation of disease-causing huntingtin protein containing long polyglutamine tracts and expand the Trim-Away toolbox with highly active TRIM21-nanobody chimeras that can also be controlled optogenetically. This work provides a mechanism for cellular activation of TRIM RING ligases and has implications for targeted protein degradation technologies.

Rare monogenic disorders of the primary cilium, termed ciliopathies, are characterized by extreme presentations of otherwise common diseases, such as diabetes, hepatic fibrosis, and kidney failure. However, despite a recent revolution in our understanding of the cilium's role in rare disease pathogenesis, the organelle's contribution to common disease remains largely unknown. Hypothesizing that common genetic variants within Mendelian ciliopathy genes might contribute to common complex diseases pathogenesis, we performed association studies of 16,874 common genetic variants across 122 ciliary genes with 12 quantitative laboratory traits characteristic of ciliopathy syndromes in 452,593 individuals in the UK Biobank. We incorporated tissue-specific gene expression analysis, expression quantitative trait loci, and Mendelian disease phenotype information into our analysis and replicated our findings in meta-analysis. 101 statistically significant associations were identified across 42 of the 122 examined ciliary genes (including eight novel replicating associations). These ciliary genes were widely expressed in tissues relevant to the phenotypes being studied, and eQTL analysis revealed strong evidence for correlation between ciliary gene expression levels and laboratory traits. Perhaps most interestingly, our analysis identified different ciliary subcompartments as being specifically associated with distinct sets of phenotypes. Taken together, our data demonstrate the utility of a Mendelian pathway-based approach to genomic association studies, challenge the widely held belief that the cilium is an organelle important mainly in development and in rare syndromic disease pathogenesis, and provide a framework for the continued integration of common and rare disease genetics to provide insight into the pathophysiology of human diseases of immense public health burden.

Synchrotron serial crystallography (SSX) is an emerging data-collection method for micro-crystallography on synchrotron macromolecular (MX) crystallography beamlines. At SPring-8, the feasibility of the fixed-target approach was examined by collecting data using a 2D raster scan combined with goniometer rotation. Results at cryogenic temperatures demonstrated that rotation is effective for efficient data collection in SSX and the method was named serial synchrotron rotation crystallography (SS-ROX). To use this method for room-temperature (RT) data collection, a humid air and glue-coating (HAG) method was developed in which data were collected from polyvinyl alcohol-coated microcrystals fixed on a loop under humidity-controlled air. The performance and the RT data-collection strategy for micro-crystallography were evaluated using microcrystals of lysozyme. Although a change in unit-cell dimensions of up to 1% was observed during data collection, the impact on data quality was marginal. A comparison of data obtained at various absorbed doses revealed that absorbed doses of up to 210 kGy were tolerable in both global and local damage. Although this limits the number of photons deposited on each crystal, increasing the number of merged images improved the resolution. On the basis of these results, an equation was proposed that relates the achievable resolution to the total photon flux used to obtain a data set.

<h4>Introduction</h4>A functional tandem repeat polymorphism in the promoter of the serotonin transporter (SERT) gene (SLC6A4) has been studied for association to neuropsychiatric conditions, including substance use disorders. Short (S) forms of this repeat result in reduced transcription, and presumably greater synaptic levels of serotonin, which are involved in opioid and cocaine-induced reward. Dual exposure to heroin and cocaine is a common pattern of poly-drug use and is associated with considerable morbidity. We hypothesize that SLC6A4 variants are associated with cocaine exposure in subjects with an opioid dependence diagnosis (OD), and also in non-dependent opioid users (NOD). Other single nucleotide polymorphisms (SNPs) of SLC6A4 may also be likewise associated.<h4>Materials and methods</h4>This study determined whether variants of the SLC6A4 promoter repeats and two intronic SNPs, rs16965628 and rs2066713, are associated with categorical diagnoses of opioid dependence (DSM-IV criteria) and with dimensional aspects of cocaine use, in a Caucasian cohort (n=591). Three groups of subjects were examined: (1) 276 subjects with opioid dependence diagnosis (OD); (2) 163 subjects who had used opioids for non-medical reasons but never had an opioid dependence diagnosis (NOD); (3) 152 healthy controls (HC).<h4>Results</h4>Aside from high exposure to heroin in the OD group, relatively high exposure to cocaine was detected in both OD and NOD groups. The SERT repeat genotype (classified as "long-long" [LL] versus "short-long" plus "short-short" [SL+SS]) was not associated with categorical opioid dependence diagnoses. A nominally significant association was identified with the [SL+SS] genotype of SLC6A4 and cocaine KMSK scores ≥"cutpoint" for a cocaine dependence diagnosis (<i>p</i>=0.026). The [SL+SS] genotype was associated with more rapid cocaine escalation than the LL genotype. No significant associations of rs16965628 and rs2066713 SNPs were found overall.<h4>Conclusion</h4>The functional SERT promoter tandem repeat genotype may be associated to heavy cocaine exposure and more rapid escalation of cocaine use, in persons with and without opioid dependence diagnosis.

Biliary tract cancer (BTC) is a leading cause of cancer-related death, due to the limited benefits of current systematic therapies and the heterogeneity of the tumor itself. High heterogeneity means that the clinical and molecular features vary between different subtypes of BTC, while the underlying molecular mechanisms remain unclear. Targeted therapy, where inhibitors are developed to selectively combine with targeted molecules in order to block abnormal signaling pathways in BTC, has shown promise as an emerging form of treatment for various types of cancer. In this article, a comprehensive review is conducted to examine potential molecular targets for BTC targeted therapy and their mechanisms. Furthermore, preliminary data published from clinical trials is utilized to analyze the main drugs used to combat BTC. The collective information presented in this article has provided useful insights into the current understanding of BTC.

Evidence suggests that stem cells exert regenerative potential via the release of extracellular vesicles. Mesenchymal stem cell extracellular vesicles (MSCEVs) offer therapeutic benefits for various pathophysiological ailments by restoring tissues. Facts suggest that MSCEV action can be potentiated by modifying the mesenchymal stem cells culturing methodology and bioengineering EVs. Limited clinical trials of MSCEVs have questioned their superiority, culturing quality, production scale-up and isolation, and administration format. Translation of preclinically successful MSCEVs into a clinical platform requires paying attention to several critical matters, such as the production technique, quantification/characterization, pharmacokinetics/targeting/transfer to the target site, and the safety profile. Keeping these issues as a priority, the present review was designed to highlight the challenges in translating preclinical MSCEV research into clinical platforms and provide evidence for the regenerative potential of MSCEVs in various conditions of the liver, kidney, heart, nervous system, bone, muscle, cartilage, and other organs/tissues.

Hybrid linear-dendritic block copolymers (LDBCs) having dendrons with a precise number of peripheral groups that are able to supramolecular bind functional moieties are challenging materials as versatile polymeric platforms for the preparation of functional polymeric nanocarriers. <b>PEG_2k-<i>b</i>-dxDAP</b> LDBCs that are based on polyethylene glycol (PEG) as hydrophilic blocks and dendrons derived from bis-MPA having 2,6-diacylaminopyridine (DAP) units have been efficiently synthesized by the click coupling of preformed blocks, as was demonstrated by spectroscopic techniques and mass spectrometry. Self-assembly ability was first checked by nanoprecipitation. A reproducible and fast synthesis of aggregates was accomplished by microfluidics optimizing the total flow rate and phase ratio to achieve spherical micelles and/or vesicles depending on dendron generation and experimental parameters. The morphology and size of the self-assemblies were studied by TEM, Cryogenic Transmission Electron Microscopy (cryo-TEM), and Dynamic Light Scattering (DLS). The cytotoxicity of aggregates synthesized by microfluidics and the influence on apoptosis and cell cycle evaluation was studied on four cell lines. The self-assemblies are not cytotoxic at doses below 0.4 mg mL^-1. Supramolecular functionalization using thymine derivatives was explored for reversibly cross-linking the hydrophobic blocks. The results open new possibilities for their use as drug nanocarriers with a dynamic cross-linking to improve nanocarrier stability but without hindering disassembly to release molecular cargoes.

Signal transducer and activator of transcription 3 (STAT3)/Cyclin-dependent kinases are multifunctional proteins that play an important implicative role in cancer initiations, progression, drug resistance, and metastasis, and has been extensively explored in cancer therapy. However, the genetic alterations of STAT3/CDK2/4/6 and its role in predicting immune infiltration and immunotherapeutic response are yet to be well exploited. In this study, we use in silico methods to analyze differential expression, prognostic value, genetic and epigenetic alterations, association with tumor-infiltrating immune cells, and cancer-associated fibroblast (CAF) infiltrations of STAT3/CDK2/4/6 in multiple cancer types. Our results revealed that the expression of STAT3/CDK2/4/6 was altered in various cancers and is associated with poor overall and disease-free survival of the cohorts. Moreover, genetic alterations in STAT3/CDK2/4/6 co-occurred with a number of other genetic alterations and are associated with poorer prognoses of the cohorts. The protein-protein interaction (PPI) network analysis suggests CDK2/4/6/STAT3 may directly interact with factors that promote tumorigenesis and immune response. We found that STAT3/CDK2/4/6 expressions were associated with infiltrations of CAF and the various immune cells in multiple cancers and it's associated with poor response to immunotherapy. Collectively, our study suggested that STAT3/CDK2/4/6 are important onco-immune signatures that play central roles in tumor immune invasion, poor prognoses and, immune therapy response. Findings from the present study may therefore be clinically useful in prognosis assessment and follow-up management of immunotherapy.

A novel ion-selective electrode with membranes based on iron(III) phosphate and silver sulfide integrated into a completely new electrode body design has been developed for the determination of iron(III) cations. The best response characteristics with linear potential change were found in the iron(III) concentration range from 3.97 × 10^-5 to 10^-2 mol L^-1. The detection limit was found to be 2.41 × 10^-5 mol L^-1 with a slope of -20.53 ± 0.63 and regression coefficient of 0.9925, while the quantification limit was 3.97 × 10^-5 M. The potential change per concentration decade ranged from -13.59 ± 0.54 to -20.53 ± 1.56 for Electrode Body 1 (EB1) and from -17.28 ± 1.04 to -24 ± 1.87 for Electrode Body 2 (EB2), which is presented for the first time in this work. The prepared electrode has a long lifetime and the ability to detect changes in the concentration of iron cations within 20 s. Membrane M1 showed high recoveries in the determination of iron cations in iron(III) standard solutions (98.2-101.2%) as well as in two different pharmaceuticals (98.6-106.5%). This proves that this type of sensor is applicable in the determination of ferric cations in unknown samples, and the fact that all sensor parts are completely manufactured in our laboratory proves the simplicity of the method.

3D-printed bone scaffolds hold great promise for the individualized treatment of critical-size bone defects. Among the resorbable polymers available for use as 3D-printable scaffold materials, poly(ε-caprolactone) (PCL) has many benefits. However, its relatively low stiffness and lack of bioactivity limit its use in load-bearing bone scaffolds. This study tests the hypothesis that surface-oxidized cellulose nanocrystals (SO-CNCs), decorated with carboxyl groups, can act as multi-functional scaffold additives that (1) improve the mechanical properties of PCL and (2) induce biomineral formation upon PCL resorption. To this end, an <i>in vitro</i> biomineralization study was performed to assess the ability of SO-CNCs to induce the formation of calcium phosphate minerals. In addition, PCL nanocomposites containing different amounts of SO-CNCs (1, 2, 3, 5, and 10 wt%) were prepared using melt compounding extrusion and characterized in terms of Young's modulus, ultimate tensile strength, crystallinity, thermal transitions, and water contact angle. Neither sulfuric acid-hydrolyzed CNCs (SH-CNCs) nor SO-CNCs were toxic to MC3T3 preosteoblasts during a 24 h exposure at concentrations ranging from 0.25 to 3.0 mg/mL. SO-CNCs were more effective at inducing mineral formation than SH-CNCs in simulated body fluid (1x). An SO-CNC content of 10 wt% in the PCL matrix caused a more than 2-fold increase in Young's modulus (stiffness) and a more than 60% increase in ultimate tensile strength. The matrix glass transition and melting temperatures were not affected by the SO-CNCs but the crystallization temperature increased by about 5.5°C upon addition of 10 wt% SO-CNCs, the matrix crystallinity decreased from about 43 to about 40%, and the water contact angle decreased from 87 to 82.6°. The abilities of SO-CNCs to induce calcium phosphate mineral formation and increase the Young's modulus of PCL render them attractive for applications as multi-functional nanoscale additives in PCL-based bone scaffolds.

Autism spectrum disorder (ASD) is a heterogeneous neuropsychiatric disorder with a complex genetic background. Analysis of altered molecular processes in ASD patients requires linear and nonlinear methods that provide interpretable solutions. Interpretable machine learning provides legible models that allow explaining biological mechanisms and support analysis of clinical subgroups. In this work, we investigated several case-control studies of gene expression measurements of ASD individuals. We constructed a rule-based learning model from three independent datasets that we further visualized as a nonlinear gene-gene co-predictive network. To find dissimilarities between ASD subtypes, we scrutinized a topological structure of the network and estimated a centrality distance. Our analysis revealed that autism is the most severe subtype of ASD, while pervasive developmental disorder-not otherwise specified and Asperger syndrome are closely related and milder ASD subtypes. Furthermore, we analyzed the most important ASD-related features that were described in terms of gene co-predictors. Among others, we found a strong co-predictive mechanism between <i>EMC4</i> and <i>TMEM30A</i>, which may suggest a co-regulation between these genes. The present study demonstrates the potential of applying interpretable machine learning in bioinformatics analyses. Although the proposed methodology was designed for transcriptomics data, it can be applied to other omics disciplines.

<h4>Objectives</h4>Recently long non-coding RNAs (lncRNAs) have emerged as novel gene regulators involved in tumorigenic processes, including oral squamous cell carcinoma (OSCC). Here, we identified a differentiation-related lncRNA, terminal differentiation-induced non-coding RNA (TINCR). However, its biological function and clinicopathological significance in OSCC still remain unclear.<h4>Methods</h4>The lncRNA expression profiles in OSCC tissues and paired adjacent non-tumor tissues (NATs) from 10 patients were detected by lncRNA microarrays. Weighted gene co-expression network analysis (WGCNA) and gene ontology (GO) enrichment were performed to identify the most significant module and module functional annotation, respectively. Potential differentiation-related lncRNAs were screened by differential expression analysis. TINCR was further confirmed in OSCC cell lines and tissues of another patient cohort by using qRT-PCR. The correlation between the TINCR expression level and clinicopathological characteristics was analyzed. The effects of TINCR on cell differentiation, migration and invasion were assessed by knockdown or knock-in <i>in vitro</i> and <i>in vivo</i>.<h4>Results</h4>WGCNA and GO enrichment analysis showed that one co-expression network was significantly enriched for epithelial cell differentiation, among which, TINCR was significantly downregulated. qRT-PCR analyses validated down-regulation of TINCR in tumor tissues compared with paired NATs, and its expression was closely correlated with pathological differentiation and lymph node metastasis in patients with OSCC. Patients with lower TINCR expression levels had worse survival. Cell function experiments showed that TINCR played a crucial role in epithelial differentiation. Both TINCR and epithelial differentiation-associated genes, including IVL and KRT4, were significantly upregulated during OSCC cell calcium-induced differentiation but were reduced when cell dedifferentiation occurred in tumor spheres. Overexpression of TINCR dramatically suppressed cell dedifferentiation, migration and invasion <i>in vitro</i>, while knockdown of TINCR had the opposite effects. Upregulation of TINCR significantly elevated the expression of terminal differentiation genes and repressed tumor growth <i>in vivo</i>. Moreover, TINCR significantly suppressed the activation of JAK2/STAT3 signaling in OSCC cells.<h4>Conclusion</h4>Our study suggests that TINCR functions as a tumor suppressor by inducing cell differentiation through modulating JAK2/STAT3 signaling in OSCC. TINCR may serve as a prognostic biomarker and therapeutic target for OSCC.

<h4>Background</h4>Some colorectal adenocarcinoma (CRC) patients are susceptible to recurrence, and they rapidly progress to advanced cancer stages and have a poor prognosis. There is an urgent need for efficient screening criteria to identify patients who tend to relapse in order to treat them earlier and more systematically.<h4>Methods</h4>We identified two groups of patients with significantly different outcomes by unsupervised cluster analysis of GSE39582 based on 101 significantly differentially expressed immune genes. To develop an accurate and specific signature based on immune-related genes to predict the recurrence of CRC, a multivariate Cox risk regression model was constructed with a training cohort composed of 519 CRC samples. The model was then validated using 129, 292, and 446 samples in the real-time quantitative reverse transcription PCR (qRT-PCR), test, and validation cohorts, respectively.<h4>Results</h4>This classification system can also be used to predict the prognosis in clinical subgroups and patients with different mutation states. Four independent datasets, including qRT-PCR and The Cancer Genome Atlas (TCGA), demonstrated that they can also be used to accurately predict the overall survival of CRC patients. Further analysis suggested that high-risk patients were characterized by worse effects of chemotherapy and immunotherapy, as well as lower immune scores. Ultimately, the signature was identified as an independent prognostic factor.<h4>Conclusion</h4>The signature can accurately predict recurrence and overall survival in patients with CRC and may serve as a powerful prognostic tool to further optimize cancer immunotherapy.

Autosomal recessive mutations in <i>G6PC3</i> cause isolated and syndromic congenital neutropenia which includes congenital heart disease and atypical inflammatory bowel disease (IBD). In a highly consanguineous pedigree with novel mutations in <i>G6PC3</i> and <i>MPL</i>, we performed comprehensive multi-omics analyses. Structural analysis of variant G6PC3 and MPL proteins suggests a damaging effect. A distinct molecular cytokine profile (cytokinome) in the affected proband with IBD was detected. Liquid chromatography-mass spectrometry-based proteomics analysis of the G6PC3-deficient plasma samples identified 460 distinct proteins including 75 upregulated and 73 downregulated proteins. Specifically, the transcription factor GATA4 and LST1 were downregulated while platelet factor 4 (PF4) was upregulated. GATA4 and PF4 have been linked to congenital heart disease and IBD respectively, while LST1 may have perturbed a variety of essential cell functions as it is required for normal cell-cell communication. Together, these studies provide potentially novel insights into the pathogenesis of syndromic congenital G6PC3 deficiency.

Differences in health outcomes and treatment responses within and between global populations have been well documented. There is growing recognition of the need to move beyond simple inventories and descriptions of these differences and our linear explanations for them, and gain a better understanding of the multifaceted systems and networks underlying them in order to develop more precise and effective remedies. Typical targets for such integrative research have been common multifactorial diseases. We propose sickle cell disease, one of the most common monogenic diseases, as an ideal candidate for elucidating the complexity of the influences of endogenous and exogenous factors on disease pathophysiology, phenotypic diversity, and variations in responses to treatments at both the individual and population levels. We provide data-informed representations of diverse contributors to sickle cell disease complications that could guide innovative efforts to advance scientific knowledge, clinical practice, and policy formulation related to the disease; help improve outcomes for people worldwide with sickle cell disease; and inform approaches to studying and addressing other diseases.

<title>Abstract</title> <p>Background: Gossypol (GP) is a polyphenolic compound in cottonseed. In porcine, GP affects female reproduction and the respiratory system. The aim of this study was to identify differentially expressed genes in porcine granulosa cells (GCs) treated with GP (6.25 and 12.5 μM) for 72 h, in vitro . <h4>Results:</h4> In GP-treated groups (6.25 and 12.5 μM), the expressions of a number of genes were found to be significantly changed. Gene ontology analysis indicated that the identified DEGs were primarily related to the mitotic cell cycle, chromosome, centromeric region and protein binding. Kyoto Encyclopedia of Genes and Genomes pathway analysis of the GP6.25 group revealed that pathways related to the cell cycle, oocyte meiosis, progesterone-mediated oocyte maturation and p53 signaling were the most changed, while that of the GP12.5 group revealed that pathways related to the PI3K-Akt signaling, focal adhesion, HIF-1 signaling, cell cycle and ECM-receptor interaction were the most changed. Genes associated with female reproductive function ( CDK1 , CCNB1 , CPEB1 and MMP3 ), cellular component organization ( BIRC5 , CYP1A1 , TGFβ3 and COL1A2 ) and oxidation-reduction processes ( PRDX6 , MGST1 and SOD3 ) were confirmed to be differently expressed in GP-treated groups. <h4>Conclusion:</h4> Our findings provide insight regarding changes in GC gene expression in porcine exposed to GP.</p>

<title>Abstract</title> <p>Amyotrophic lateral sclerosis (ALS) is a rapidly progressive adult-onset neurodegenerative disease that is often diagnosed with a delay due to initial non-specific symptoms. Therefore, reliable and easy-to-obtain biomarkers are in desperate need for earlier and more accurate diagnostics. Circular RNAs (circRNAs) have been already proposed as potential biomarkers for several neurodegenerative diseases. In this study, we further investigated the usefulness of circRNAs as potential biomarkers for ALS. We first performed a microarray analysis of circRNAs on peripheral blood mononuclear cells of a subset of ALS patients and controls. Among the differently expressed circRNA by microarray analysis, we selected only the ones with a host gene that harbors the highest level of conservation and genetic constraints. This selection was based under the hypothesis that genes under selective pressure and genetic constraints could have a major role in determining a trait or disease. Then we performed a linear regression between ALS cases and controls using each circRNA as a predictor variable. With an FDR threshold of 0.1, only six circRNAs passed the filtering and merely one of them remained statistically significant after Bonferroni correction: hsa_circ_0060762 and its host gene CSE1L. Finally, we observed a significant difference in expression levels between larger sets of patients and healthy controls for both hsa_circ_0060762 and CSE1L . In addition, receiver operating characteristics curve analysis showed diagnostic potential for CSE1L and hsa_circ_0060762 . Hsa_circ_0060762 thus represent a novel potential peripheral blood circRNA biomarker for ALS.</p>

The SARS-CoV-2 pandemic has caused over 1 million deaths globally, mostly due to acute lung injury and acute respiratory distress syndrome, or direct complications resulting in multiple-organ failures. Little is known about the host tissue immune and cellular responses associated with COVID-19 infection, symptoms, and lethality. To address this, we collected tissues from 11 organs during the clinical autopsy of 17 individuals who succumbed to COVID-19, resulting in a tissue bank of approximately 420 specimens. We generated comprehensive cellular maps capturing COVID-19 biology related to patients’ demise through single-cell and single-nucleus RNA-Seq of lung, kidney, liver and heart tissues, and further contextualized our findings through spatial RNA profiling of distinct lung regions. We developed a computational framework that incorporates removal of ambient RNA and automated cell type annotation to facilitate comparison with other healthy and diseased tissue atlases. In the lung, we uncovered significantly altered transcriptional programs within the epithelial, immune, and stromal compartments and cell intrinsic changes in multiple cell types relative to lung tissue from healthy controls. We observed evidence of: alveolar type 2 (AT2) differentiation replacing depleted alveolar type 1 (AT1) lung epithelial cells, as previously seen in fibrosis; a concomitant increase in myofibroblasts reflective of defective tissue repair; and, putative TP63 + intrapulmonary basal-like progenitor (IPBLP) cells, similar to cells identified in H1N1 influenza, that may serve as an emergency cellular reserve for severely damaged alveoli. Together, these findings suggest the activation and failure of multiple avenues for regeneration of the epithelium in these terminal lungs. SARS-CoV-2 RNA reads were enriched in lung mononuclear phagocytic cells and endothelial cells, and these cells expressed distinct host response transcriptional programs. We corroborated the compositional and transcriptional changes in lung tissue through spatial analysis of RNA profiles in situ and distinguished unique tissue host responses between regions with and without viral RNA, and in COVID-19 donor tissues relative to healthy lung. Finally, we analyzed genetic regions implicated in COVID-19 GWAS with transcriptomic data to implicate specific cell types and genes associated with disease severity. Overall, our COVID-19 cell atlas is a foundational dataset to better understand the biological impact of SARS-CoV-2 infection across the human body and empowers the identification of new therapeutic interventions and prevention strategies.

Microgels are soft colloids that show responsive behavior and are easy to functionalize for applications. They are considered key components for future smart colloidal material systems. However, so far microgel systems have almost exclusively been studied in classical responsive switching settings using external triggers, while internally organized, autonomous control mechanisms as found in supramolecular chemistry and DNA nanotechnology relying on fuel-driven out-of-equilibrium concepts have not been implemented into microgel systems. Here, we introduce chemically fueled transient volume phase transitions (VPTs) for poly(methacrylic acid) (PMAA) microgels, where the collapsed hydrophobic state can be programmed using the fuel concentration in a cyclic reaction network. We discuss details of the system behavior as a function of pH and fuel amount, unravel kinetically trapped regions and showcase transient encapsulation and time-programmed release as a first application.

Infection with <i>Zaire ebolavirus</i> (EBOV), a member of the <i>Filoviridae</i> family, causes a disease characterized by high levels of viremia, aberrant inflammation, coagulopathy, and lymphopenia. EBOV initially replicates in lymphoid tissues and disseminates via dendritic cells (DCs) and monocytes to liver, spleen, adrenal gland, and other secondary organs. EBOV protein VP35 is a critical immune evasion factor that inhibits type I interferon signaling and DC maturation. Nonhuman primates (NHPs) immunized with a high dose (5 × 10⁵ PFU) of recombinant EBOV containing a mutated VP35 (VP35m) are protected from challenge with wild-type EBOV (wtEBOV). This protection is accompanied by a transcriptional response in the peripheral blood reflecting a regulated innate immune response and a robust induction of adaptive immune genes. However, the host transcriptional response to VP35m in lymphoid tissues has not been evaluated. Therefore, we conducted a transcriptional analysis of axillary and inguinal lymph nodes and spleen tissues of NHPs infected with a low dose (2 × 10⁴ PFU) of VP35m and then back-challenged with a lethal dose of wtEBOV. VP35m induced early transcriptional responses in lymphoid tissues that are distinct from those observed in wtEBOV challenge. Specifically, we detected robust antiviral innate and adaptive responses and fewer transcriptional changes in genes with roles in angiogenesis, apoptosis, and inflammation. Two of three macaques survived wtEBOV back-challenge, with only the nonsurvivor displaying a transcriptional response reflecting Ebola virus disease. These data suggest that VP35 is a key modulator of early host responses in lymphoid tissues, thereby regulating disease progression and severity following EBOV challenge.<b>IMPORTANCE</b> Zaire Ebola virus (EBOV) infection causes a severe and often fatal disease characterized by inflammation, coagulation defects, and organ failure driven by a defective host immune response. Lymphoid tissues are key sites of EBOV pathogenesis and the generation of an effective immune response to infection. A recent study demonstrated that infection with an EBOV encoding a mutant VP35, a viral protein that antagonizes host immunity, can protect nonhuman primates (NHPs) against lethal EBOV challenge. However, no studies have examined the response to this mutant EBOV in lymphoid tissues. Here, we characterize gene expression in lymphoid tissues from NHPs challenged with the mutant EBOV and subsequently with wild-type EBOV to identify signatures of a protective host response. Our findings are critical for elucidating viral pathogenesis, mechanisms of host antagonism, and the role of lymphoid organs in protective responses to EBOV to improve the development of antivirals and vaccines against EBOV.

The phenylalanine-tyrosine-dopa-dopamine pathway provides dopamine to the brain. In this process, tyrosine hydroxylase (TH) is the rate-limiting enzyme that hydroxylates tyrosine and generates levodopa (L-dopa) with tetrahydrobiopterin (BH₄) as a coenzyme. Here, we show that oral berberine (BBR) might supply H^• through dihydroberberine (reduced BBR produced by bacterial nitroreductase) and promote the production of BH₄ from dihydrobiopterin; the increased BH₄ enhances TH activity, which accelerates the production of L-dopa by the gut bacteria. Oral BBR acts in a way similar to vitamins. The L-dopa produced by the intestinal bacteria enters the brain through the circulation and is transformed to dopamine. To verify the gut-brain dialog activated by BBR's effect, Enterococcus faecalis or Enterococcus faecium was transplanted into Parkinson's disease (PD) mice. The bacteria significantly increased brain dopamine and ameliorated PD manifestation in mice; additionally, combination of BBR with bacteria showed better therapeutic effect than that with bacteria alone. Moreover, 2,4,6-trimethyl-pyranylium tetrafluoroborate (TMP-TFB)-derivatized matrix-assisted laser desorption mass spectrometry (MALDI-MS) imaging of dopamine identified elevated striatal dopamine levels in mouse brains with oral Enterococcus, and BBR strengthened the imaging intensity of brain dopamine. These results demonstrated that BBR was an agonist of TH in Enterococcus and could lead to the production of L-dopa in the gut. Furthermore, a study of 28 patients with hyperlipidemia confirmed that oral BBR increased blood/fecal L-dopa by the intestinal bacteria. Hence, BBR might improve the brain function by upregulating the biosynthesis of L-dopa in the gut microbiota through a vitamin-like effect.

PD-1 is a critical therapeutic target in cancer immunotherapy and antibodies blocking PD-1 are approved for multiple types of malignancies. The phosphatase SHP2 is the main effector mediating PD-1 downstream signaling and accordingly attempts have been made to target this enzyme as an alternative approach to treat immunogenic tumors. Unfortunately, small molecule inhibitors of SHP2 do not work as expected, suggesting that the role of SHP2 in T cells is more complex than initially hypothesized. To better understand the perplexing role of SHP2 in T cells, we performed interactome mapping of SAP, an adapter protein that is associated with SHP2 downstream signaling. Using genetic and pharmacological approaches, we discovered that SHP2 dephosphorylates ITK specifically downstream of PD-1 and that this event was associated with PD-1 inhibitory cellular functions. This study suggests that ITK is a unique target in this pathway, and since ITK is a SHP2-dependent specific mediator of PD-1 signaling, the combination of ITK inhibitors with PD-1 blockade may improve upon PD-1 monotherapy in the treatment of cancer.

Managing the pandemic caused by SARS-CoV-2 requires new capabilities in testing, including the possibility of identifying, in minutes, infected individuals as they enter spaces where they must congregate in a functioning society, including workspaces, schools, points of entry, and commercial business establishments. Here, the only useful tests (a) require no sample transport, (b) require minimal sample manipulation, (c) can be performed by unlicensed individuals, (d) return results on the spot in much less than one hour, and (e) cost no more than a few dollars. The sensitivity need not be as high as normally required by the FDA for screening asymptomatic carriers (as few as 10 virions per sample), as these viral loads are almost certainly not high enough for an individual to present a risk for forward infection. This allows tests specifically useful for this pandemic to trade-off unneeded sensitivity for necessary speed, simplicity, and frugality. In some studies, it was shown that viral load that creates forward-infection risk may exceed 105 virions per milliliter, easily within the sensitivity of an RNA amplification architecture, but unattainable by antibody-based architectures that simply target viral antigens. Here, we describe such a test based on a displaceable probe loop amplification architecture.

<h4>Background</h4>The increasing number of genome-wide association studies (GWAS) has revealed several loci that are associated to multiple distinct phenotypes, suggesting the existence of pleiotropic effects. Highlighting these cross-phenotype genetic associations could help to identify and understand common biological mechanisms underlying some diseases. Common approaches test the association between genetic variants and multiple traits at the SNP level. In this paper, we propose a novel gene- and a pathway-level approach in the case where several independent GWAS on independent traits are available. The method is based on a generalization of the sparse group Partial Least Squares (sgPLS) to take into account groups of variables, and a Lasso penalization that links all independent data sets. This method, called joint-sgPLS, is able to convincingly detect signal at the variable level and at the group level.<h4>Results</h4>Our method has the advantage to propose a global readable model while coping with the architecture of data. It can outperform traditional methods and provides a wider insight in terms of a priori information. We compared the performance of the proposed method to other benchmark methods on simulated data and gave an example of application on real data with the aim to highlight common susceptibility variants to breast and thyroid cancers.<h4>Conclusion</h4>The joint-sgPLS shows interesting properties for detecting a signal. As an extension of the PLS, the method is suited for data with a large number of variables. The choice of Lasso penalization copes with architectures of groups of variables and observations sets. Furthermore, although the method has been applied to a genetic study, its formulation is adapted to any data with high number of variables and an exposed a priori architecture in other application fields.

The DEAD-box helicase family member DDX3X (DBX, DDX3) functions in nearly all stages of RNA metabolism and participates in the progression of many diseases, including virus infection, inflammation, intellectual disabilities and cancer. Over two decades, many studies have gradually unveiled the role of DDX3X in tumorigenesis and tumour progression. In fact, DDX3X possesses numerous functions in cancer biology and is closely related to many well-known molecules. In this review, we describe the function of DDX3X in RNA metabolism, cellular stress response, innate immune response, metabolic stress response in pancreatic β cells and embryo development. Then, we focused on the role of DDX3X in cancer biology and systematically demonstrated its functions in various aspects of tumorigenesis and development. To provide a more intuitive understanding of the role of DDX3X in cancer, we summarized its functions and specific mechanisms in various types of cancer and presented its involvement in cancer-related signalling pathways.

<h4>Background</h4>The uterine cervix is a mechanical and immunological barrier against ascending infection during pregnancy. Cervical insufficiency (CI), a painless cervical dilation that occurs in the mid-trimester, is an important cause of extremely preterm birth. We hypothesized that women with CI have a differential transcriptomic profile. Therefore, we compared the transcriptomic profile of peripheral blood in women with CI and that of controls.<h4>Methods</h4>RNA sequencing was used to generate the global gene expression profiles of 11 women with CI and 4 controls, and differential expression analysis was performed to identify genes showing significant expression changes between the CI (n = 11) and control (n = 4) groups as well as between the CI-preterm (n = 7) and CI-term (n = 4) groups. Gene set enrichment was assessed in terms of Gene Ontology processes, and a subset of differentially expressed genes in CI was validated in a different sample-set by qRT-PCR and ELISA.<h4>Results</h4>Thirty genes were differentially expressed between the CI and control groups. Differentially upregulated genes in the CI group included neutrophil-mediated immunity-associated (DEFA3 and ELANE) and bicarbonate transport-related genes. The serum concentration of alpha defensin 3 was significantly higher in women with CI than in controls (P = 0.014). Analysis of differential gene expression according to pregnancy outcomes revealed 338 differentially expressed genes between the CI-term and CI-preterm groups. Immune and defense response to organism-associated genes and influenza A and NOD-like receptor signaling pathways were upregulated in the CI-term group.<h4>Conclusions</h4>Our results revealed significant differences in the whole blood transcriptomic profiles of women with CI compared to those of controls. Different immune responses in women with CI may affect pregnancy outcomes.

<h4>Background</h4>The non-receptor tyrosine kinase Abelson (Abl) is a key player in oncogenesis, with kinase inhibitors serving as paradigms of targeted therapy. Abl also is a critical regulator of normal development, playing conserved roles in regulating cell behavior, brain development and morphogenesis. Drosophila offers a superb model for studying Abl's normal function, because, unlike mammals, there is only a single fly Abl family member. In exploring the mechanism of action of multi-domain scaffolding proteins like Abl, one route is to define the roles of their individual domains. Research into Abl's diverse roles in embryonic morphogenesis revealed many surprises. For instance, kinase activity, while important, is not crucial for all Abl activities, and the C-terminal F-actin binding domain plays a very modest role. This turned our attention to one of Abl's least understood features-the long intrinsically-disordered region (IDR) linking Abl's kinase and F-actin binding domains. The past decade revealed unexpected, important roles for IDRs in diverse cell functions, as sites of posttranslational modifications, mediating multivalent interactions and enabling assembly of biomolecular condensates via phase separation. Previous work deleting conserved regions in Abl's IDR revealed an important role for a PXXP motif, but did not identify any other essential regions.<h4>Methods</h4>Here we extend this analysis by deleting the entire IDR, and asking whether Abl∆IDR rescues the diverse roles of Abl in viability and embryonic morphogenesis in Drosophila.<h4>Results</h4>This revealed that the IDR is essential for embryonic and adult viability, and for cell shape changes and cytoskeletal regulation during embryonic morphogenesis, and, most surprisingly, revealed a role in modulating protein stability.<h4>Conclusion</h4>Our data provide new insights into the role of the IDR in an important signaling protein, the non-receptor kinase Abl, suggesting that it is essential for all aspects of protein function during embryogenesis, and revealing a role in protein stability. These data will stimulate new explorations of the mechanisms by which the IDR regulates Abl stability and function, both in Drosophila and also in mammals. They also will stimulate further interest in the broader roles IDRs play in diverse signaling proteins. Video Abstract.

Memory B cells (MBCs) have enhanced capabilities to differentiate to plasma cells and generate a rapid burst of Abs upon secondary stimulation. To determine if MBCs harbor an epigenetic landscape that contributes to increased differentiation potential, we derived the chromatin accessibility and transcriptomes of influenza-specific IgM and IgG MBCs compared with naive cells. MBCs possessed an accessible chromatin architecture surrounding plasma cell-specific genes, as well as altered expression of transcription factors and genes encoding cell cycle, chemotaxis, and signal transduction processes. Intriguingly, this MBC signature was conserved between humans and mice. MBCs of both species possessed a heightened heme signature compared with naive cells. Differentiation in the presence of hemin enhanced oxidative phosphorylation metabolism and MBC differentiation into Ab-secreting plasma cells. Thus, these data define conserved MBC transcriptional and epigenetic signatures that include a central role for heme and multiple other pathways in augmenting MBC reactivation potential.

Mechanical stimuli on cells and mechanotransduction are essential in many biological and pathological processes. Glucocorticoid is an important hormone, roles, and mechanisms of which in cellular mechanotransduction remain unknown. Here, we report that glucocorticoid counteracted cellular mechanoresponses dependently on a novel long noncoding RNA (lncRNA), <i>LINC01569</i> Further, <i>LINC01569</i> mediated glucocorticoid effects on mechanotransduction by destabilizing messenger RNA (mRNA) of mechanosensors including early growth response protein 1 (<i>EGR1</i>), Cbp/P300-interacting transactivator 2 (<i>CITED2</i>), and bone morphogenic protein 7 (<i>BMP7</i>) in glucocorticoid receptor-mediated mRNA decay (GMD) manner. Mechanistically, <i>LINC01569</i> directly bound to the GMD factor Y-box-binding protein 1 (<i>YBX1</i>). Then, the <i>LINC01569-YBX1</i> complex was guided to the mRNAs of <i>EGR1</i>, <i>CITED2</i>, and <i>BMP7</i> through specific <i>LINC01569</i>-mRNA interaction, thereby contributing to the successful assembly of GMD complex and triggering GMD. Our results uncovered roles of glucocorticoid in cellular mechanotransduction and novel lncRNA-dependent GMD machinery and provided potential strategy for early intervention in mechanical disorder-associated diseases.

Blood circulating proteins are confounded readouts of the biological processes that occur in different tissues and organs. Many proteins have been linked to complex disorders and are also under substantial genetic control. Here, we investigate the associations between over 1000 blood circulating proteins and body mass index (BMI) in three studies including over 4600 participants. We show that BMI is associated with widespread changes in the plasma proteome. We observe 152 replicated protein associations with BMI. 24 proteins also associate with a genome-wide polygenic score (GPS) for BMI. These proteins are involved in lipid metabolism and inflammatory pathways impacting clinically relevant pathways of adiposity. Mendelian randomization suggests a bi-directional causal relationship of BMI with LEPR/LEP, IGFBP1, and WFIKKN2, a protein-to-BMI relationship for AGER, DPT, and CTSA, and a BMI-to-protein relationship for another 21 proteins. Combined with animal model and tissue-specific gene expression data, our findings suggest potential therapeutic targets further elucidating the role of these proteins in obesity associated pathologies.

Primary open-angle glaucoma (POAG), is a heritable common cause of blindness world-wide. To identify risk loci, we conduct a large multi-ethnic meta-analysis of genome-wide association studies on a total of 34,179 cases and 349,321 controls, identifying 44 previously unreported risk loci and confirming 83 loci that were previously known. The majority of loci have broadly consistent effects across European, Asian and African ancestries. Cross-ancestry data improve fine-mapping of causal variants for several loci. Integration of multiple lines of genetic evidence support the functional relevance of the identified POAG risk loci and highlight potential contributions of several genes to POAG pathogenesis, including SVEP1, RERE, VCAM1, ZNF638, CLIC5, SLC2A12, YAP1, MXRA5, and SMAD6. Several drug compounds targeting POAG risk genes may be potential glaucoma therapeutic candidates.

Biphasic calcium phosphate bioceramics (BCP) consist of a mixture of hydroxyapatite (HA) and beta-tricalcium phosphate (β-TCP) within the same particle. Due to their osteoconductive properties, biocompatibility and resemblance to natural bone, these materials have become a promising and suitable alternative to autologous bone grafting. First, the topography characteristics, specific surface area, and total pore volume of BCP were evaluated using scanning electron microscopy and the BET and BJH methods. Next, this study aimed to evaluate the intensity of the inflammatory process and the bone neoformation capacity of various particle sizes of BCP in the repair of critical defects in the calvaria of rats. A xenogeneic biomaterial was used in the control group. After 30, 60, and 90 days, the animals were euthanized, followed by the processing of the samples to measure the intensity of inflammatory infiltrates and the areas of bone neoformation. Our results indicate that no considerable differences were observed in the inflammatory scores in sites treated with distinct BCP grain sizes. A greater area of bone neoformation was measured in the xenogeneic group at all analysis times, with no substantial differences in bone formation between the BCP particle size in the range of 250-500 µm and 500-1000 µm.

Prophylactic antibiotic bone cements are extensively used in orthopaedics. However, the development of antimicrobial resistance to antibiotics, demonstrates a need to find alternative treatments. Herein, an antimicrobial honey (SurgihoneyRO-SHRO) has been successfully incorporated into a calcium sulphate (CS) based cement to produce a hard tissue scaffold with the ability to inhibit bacterial growth. Antimicrobial properties elicited from SHRO are predominantly owed to the water-initiated production of reactive oxygen species (ROS). As an alternative to initially loading CS cement with SHRO, in order to prevent premature activation, SHRO was added into the already developing cement matrix, locking available water into the CS crystal structure before SHRO addition. Promisingly, this methodology produced > 2.5 times (715.0 ± 147.3 μM/mL/g) more ROS over 24 h and exhibited a compressive strength (32.2 ± 5.8 MPa) comparable to trabecular bone after 3 weeks of immersion. In-vitro the SHRO loaded CS scaffolds were shown to inhibit growth of clinically relevant organisms, Staphylococcus aureus and Pseudomonas aeruginosa, with comparable potency to equivalent doses of gentamicin. Encouragingly, formulations did not inhibit wound healing or induce an inflammatory response from osteoblasts. Overall this study highlights the prophylactic potential of CS-SHRO cements as an alternative to traditional antibiotics.

<h4>Objectives</h4>Diagnostic process of mitochondrial disorders (MD) is challenging because of the clinical variability and genetic heterogeneity of these conditions. Next-Generation Sequencing (NGS) technology offers a high-throughput platform for nuclear MD.<h4>Methods</h4>We included 59 of 72 patients that undergone WES and targeted exome sequencing panel suspected to have potential PMDs. Patients who were included in the analysis considering the possible PMD were reviewed retrospectively and scored according to the Mitochondrial Disease Criteria Scale.<h4>Results</h4>Sixty-one percent of the patients were diagnosed with whole-exome sequencing (WES) (36/59) and 15% with targeted exome sequencing (TES) (9/59). Patients with MD-related gene defects were included in the mito group, patients without MD-related gene defects were included in the nonmito group, and patients in whom no etiological cause could be identified were included in the unknown etiology group. In 11 out of 36 patients diagnosed with WES, a TES panel was applied prior to WES. In 47 probands in 39 genes (<i>SURF1, SDHAF1, MTO1, FBXL4, SLC25A12, GLRX5, C19oRF12, NDUFAF6, DARS2, BOLA3, SLC19A3, SCO1, HIBCH, PDHA1, PDHAX, PC, ETFA, TRMU, TUFM, NDUFS6, WWOX, UBCD TREX1, ATL1, VAC14, GFAP, PLA2G6, TPRKB, ATP8A2, PEX13, IGHMBP2, LAMB2, LPIN1, GFPT1, CLN5, DOLK</i>) (20 mito group, 19 nonmito group) 59 variants (31 mito group, 18 nonmito group) were detected. Seven novel variants in the mito group (<i>SLC25A12, GLRX5, DARS2, SCO1, PC, ETFA, NDUFS6</i>), nine novel variants in the nonmito group (<i>IVD, GCDH, COG4, VAC14, GFAP, PLA2G6, ATP8A2, PEX13, LPIN1</i>) were detected.<h4>Conclusions</h4>We explored the feasibility of identifying pathogenic alleles using WES and TES in MD. Our results show that WES is the primary method of choice in the diagnosis of MD until at least all genes responsible for PMD are found and are highly effective in facilitating the diagnosis process.

Point mutations within the histone H3.3 are frequent in aggressive childhood brain tumors known as pediatric high-grade gliomas (pHGGs). Intriguingly, distinct mutations arise in discrete anatomical regions: H3.3-G34R within the forebrain and H3.3-K27M preferentially within the hindbrain. The reasons for this contrasting etiology are unknown. By engineering human fetal neural stem cell cultures from distinct brain regions, we demonstrate here that cell-intrinsic regional identity provides differential responsiveness to each mutant that mirrors the origins of pHGGs. Focusing on H3.3-G34R, we find that the oncohistone supports proliferation of forebrain cells while inducing a cytostatic response in the hindbrain. Mechanistically, H3.3-G34R does not impose widespread transcriptional or epigenetic changes but instead impairs recruitment of ZMYND11, a transcriptional repressor of highly expressed genes. We therefore propose that H3.3-G34R promotes tumorigenesis by focally stabilizing the expression of key progenitor genes, thereby locking initiating forebrain cells into their pre-existing immature state.

Huntington's disease (HD) is a neurodegenerative disorder characterized by accumulation of mutant huntingtin protein and significant loss of neurons in striatum and cortex. Along with motor difficulties, the HD patients also manifest anxiety and loss of cognition. Unfortunately, the clinically approved drugs only offer symptomatic relief and are not free from side effects. This study underlines the importance of glyceryl tribenzoate (GTB), an FDA-approved food flavoring ingredient, in alleviating HD pathology in transgenic N171-82Q mouse model. Oral administration of GTB significantly reduced mutant huntingtin level in striatum, motor cortex as well as hippocampus and increased the integrity of viable neurons. Furthermore, we found the presence of sodium benzoate (NaB), a FDA-approved drug for urea cycle disorders and glycine encephalopathy, in the brain of GTB-fed HD mice. Accordingly, NaB administration also markedly decreased huntingtin level in striatum and cortex. Glial activation is found to coincide with neuronal death in affected regions of HD brains. Interestingly, both GTB and NaB treatment suppressed activation of glial cells and inflammation in the brain. Finally, neuroprotective effect of GTB and NaB resulted in improved motor performance of HD mice. Collectively, these results suggest that GTB and NaB may be repurposed for HD.

The frameshift mutants K192S^fs*7 and R153S^fs*41, of the polyglutamine tract-binding protein 1 (PQBP-1), are stable intrinsically disordered proteins (IDPs). They are each associated with the severe cognitive disorder known as the Renpenning syndrome, a form of X-linked intellectual disability (XLID). Relative to the monomeric wild-type protein, these mutants are dimeric, contain more folded contents, and have higher thermal stabilities. Comparisons can be drawn to the toxic oligomerisation in the "conformational diseases", which collectively describe medical conditions involving a substantial protein structural transition in the pathogenic mechanism. At the molecular level, the end state of these diseases is often cytotoxic protein aggregation. The conformational disease proteins contain varying extents of intrinsic disorder, and the consensus pathogenesis includes an early oligomer formation. We reviewed the experimental characterisation of the toxic oligomers in representative cases. PQBP-1 mutant dimerisation was then compared to the oligomerisation of the conformational disease proteins. The PQBP-1 mutants are unique in behaving as stable soluble dimers, which do not further develop into higher oligomers or aggregates. The toxicity of the PQBP-1 mutant dimers lies in the native functions (in transcription regulation and possibly, RNA splicing) being compromised, rather than proceeding to aggregation. Other examples of stable IDP dimers were discussed and we speculated on the roles of IDP dimerisation in protein evolution.

Alpha-toxin is a major virulence factor of <i>Staphylococcus aureus</i>. Monomer binding to host cell membranes results in the formation of heptameric transmembrane pores. Among human model airway epithelial cell lines, A549 cells were most sensitive toward the toxin followed by 16HBE14o^- and S9 cells. In this study we investigated the processes of internalization of pore-containing plasma membrane areas as well as potential pathways for heptamer degradation (lysosomal, proteasomal) or disposal (formation of exosomes/micro-vesicles). The abundance of toxin heptamers upon applying an alpha-toxin pulse to the cells declined both in extracts of whole cells and of cellular membranes of S9 cells, but not in those of 16HBE14o^- or A549 cells. Comparisons of heptamer degradation rates under inhibition of lysosomal or proteasomal degradation revealed that an important route of heptamer degradation, at least in S9 cells, seems to be the lysosomal pathway, while proteasomal degradation appears to be irrelevant. Exosomes prepared from culture supernatants of toxin-exposed S9 cells contained alpha-toxin as well as low amounts of exosome and micro-vesicle markers. These results indicate that lysosomal degradation of internalized toxin heptamers may be the most important determinant of toxin-resistance of some types of airway epithelial cells.

Cancers are a large group of diseases that mostly emerge because of the uncontrollable action of many different genes in human cells [...].

FOXC1, a transcription factor involved in cell differentiation and embryogenesis, is demonstrated to be a negative regulator of Nanog in this study. FOXC1 is up-regulated in retinoic acid-induced differentiation of F9 Embryonal Carcinoma (EC) cells; furthermore, FOXC1 specifically inhibits the core pluripotency factor Nanog by binding to the proximal promoter. Overexpression of FOXC1 in F9 or knockdown in 3T3 results in the down-regulation or up-regulation of Nanog mRNA and proteins, respectively. In order to explain the mechanism by which FOXC1 inhibits Nanog expression, we identified the co-repressor HDAC2 from the FOXC1 interactome. FOXC1 recruits HDAC2 to Nanog promoter to decrease H3K27ac enrichment, resulting in transcription inhibition of Nanog. To the best of our knowledge, this is the first report that FOXC1 is involved in the epigenetic regulation of gene expression.

Transplantation is essential and crucial for individuals suffering from end-stage organ failure diseases. However, there are still many challenges regarding these procedures, such as high rates of organ rejection, shortage of organ donors, and long waiting lines. Thus, investments and efforts to develop laboratory-grown organs have increased over the past years, and with the recent progress in regenerative medicine, growing organs <i>in vitro</i> might be a reality within the next decades. One of the many different strategies to address this issue relies on organoid technology, a miniaturized and simplified version of an organ. Here, we address recent progress on organoid research, focusing on transplantation of intestine, retina, kidney, liver, pancreas, brain, lung, and heart organoids. Also, we discuss the main outcomes after organoid transplantation, common challenges faced by these promising regenerative medicine approaches, and future perspectives on the field.

Obesity cardiomyopathy increases the risk of heart failure and death. Obesity is curable, leading to the restoration of the heart phenotype, but it is not clear if there are any after-effects of obesity present after weight loss. We characterize the proteomic landscape of obesity cardiomyopathy with an evaluation of whether the cardiac phenotype is still shaped after weight loss. Cardiomyopathy was validated by cardiac hypertrophy, fibrosis, oversized myocytes, and mTOR upregulation in a rat model of cafeteria diet-induced developmental obesity. By global proteomic techniques (LC-MS/MS) a plethora of molecular changes was observed in the heart and circulation of obese animals, suggesting abnormal utilization of metabolic substrates. This was confirmed by increased levels of cardiac ACSL-1, a key enzyme for fatty acid degradation and decreased GLUT-1, a glucose transporter in obese rats. Calorie restriction and weight loss led to the normalization of the heart's size, but fibrosis was still excessive. The proteomic compositions of cardiac tissue and plasma were different after weight loss as compared to control. In addition to morphological consequences, obesity cardiomyopathy involves many proteomic changes. Weight loss provides for a partial repair of the heart's architecture, but the trace of fibrotic deposition and proteomic alterations may occur.

<h4>Introduction</h4>Neutrophils contribute to host defense through different mechanisms, including the formation of neutrophil extracellular traps (NETs). The genetic background and underlying mechanisms contributing to NET formation remain unclear.<h4>Materials and methods</h4>We performed a genome-wide association study (GWAS) and exome-sequencing analysis to identify common and rare genetic variants associated with plasma myeloperoxidase (MPO)-DNA complex levels, a biomarker for NETs, in the population-based Rotterdam Study cohort. GWAS was performed using haplotype reference consortium(HRC)-imputed genotypes of common variants in 3,514 individuals from the first and 2,076 individuals from the second cohort of the Rotterdam Study. We additionally performed exome-sequencing analysis in 960 individuals to investigate rare variants in candidate genes.<h4>Results</h4>The GWAS yielded suggestive associations (p-value < 5.0 × 10^-6) of SNPs annotated to four genes. In the exome-sequencing analysis, a variant in <i>TMPRSS13</i> gene was significantly associated with MPO-DNA complex levels (p-value < 3.06×10^-8). Moreover, gene-based analysis showed ten genes (<i>OR10H1, RP11-461L13.5, RP11-24B19.4, RP11-461L13.3, KHDRBS1, ZNF200, RP11-395I6.1, RP11-696P8.2, RGPD1, AC007036.5</i>) to be associated with MPO-DNA complex levels (p-value between 4.48 × 10^-9 and 1.05 × 10^-6). Pathway analysis of the identified genes showed their involvement in cellular development, molecular transport, RNA trafficking, cell-to-cell signaling and interaction, cellular growth and proliferation. Cancer was the top disease linked to the NET-associated genes.<h4>Conclusion</h4>In this first GWAS and exome-sequencing analysis of NETs levels, we found several genes that were associated with NETs. The precise mechanism of how these genes may contribute to neutrophil function or the formation of NETs remains unclear and should be further investigated in experimental studies.

Glioblastoma is the most common and lethal primary brain malignancy. Despite major investments in research into glioblastoma biology and drug development, treatment remains limited and survival has not substantially improved beyond 1-2 years. Cancer stem cells (CSC) or glioma stem cells (GSC) refer to a population of tumor originating cells capable of self-renewal and differentiation. While controversial and challenging to study, evidence suggests that GCSs may result in glioblastoma tumor recurrence and resistance to treatment. Multiple treatment strategies have been suggested at targeting GCSs, including immunotherapy, posttranscriptional regulation, modulation of the tumor microenvironment, and epigenetic modulation. In this review, we discuss recent advances in glioblastoma treatment specifically focused on targeting of GCSs as well as their potential integration into current clinical pathways and trials.

Herein, we report the enantiospecific synthesis of two artificial glutamate analogs designed based on IKM-159, an antagonist selective to the AMPA-type ionotropic glutamate receptor. The synthesis features the chiral resolution of the carboxylic acid intermediate by the esterification with ʟ-menthol, followed by a configurational analysis by NMR, conformational calculation, and X-ray crystallography. A mice in vivo assay showed that (2<i>R</i>)-MC-27, with a six-membered oxacycle, is neuroactive, whereas the (2<i>S</i>)-counterpart is inactive. It was also found that TKM-38, with an eight-membered azacycle, is neuronally inactive, showing that the activity is controlled by the ring C.

Neurological disorders are becoming a growing burden as society ages, and there is a compelling need to address this spiraling problem. Stem cell-based regenerative medicine is becoming an increasingly attractive approach to designing therapies for such disorders. The unique characteristics of mesenchymal stem cells (MSCs) make them among the most sought after cell sources. Researchers have extensively studied the modulatory properties of MSCs and their engineering, labeling, and delivery methods to the brain. The first part of this review provides an overview of studies on the application of MSCs to various neurological diseases, including stroke, traumatic brain injury, spinal cord injury, multiple sclerosis, amyotrophic lateral sclerosis, Alzheimer's disease, Huntington's disease, Parkinson's disease, and other less frequently studied clinical entities. In the second part, stem cell delivery to the brain is focused. This fundamental but still understudied problem needs to be overcome to apply stem cells to brain diseases successfully. Here the value of cell engineering is also emphasized to facilitate MSC diapedesis, migration, and homing to brain areas affected by the disease to implement precision medicine paradigms into stem cell-based therapies.

Neuroendocrine prostate cancer (NEPC) is the most lethal malignancy of prostate cancer (PCa). Treatment with next-generation androgen receptor (AR) pathway inhibitors (ARPIs) has successfully extended patients' lifespan. However, with the emergence of drug resistance, PCa tumors increasingly adapt to potent ARPI therapies by transitioning to alternative cellular lineage. Such therapy-induced drug resistance is largely driven from the cellular plasticity of PCa cells to alter their phenotypes of AR independence for cell growth and survival. Some of the resistant PCa cells undergo cellular reprogramming to form neuroendocrine phenotypes. Recent evidences suggest that this cellular reprogramming or the lineage plasticity is driven by dysregulation of the epigenome and transcriptional networks. Aberrant DNA methylation and altered expression of epigenetic modifiers, such as enhancer of zeste-homolog 2, transcription factors, histone demethylases, are hallmarks of NEPC. In this review, we discuss the nature of the epigenetic and transcriptional landscapes of PCa cells which lose their AR independence and transition to the neuroendocrine lineage. We also discuss how oncogenic signaling and metabolic reprogramming fuel epigenetic and transcriptional alterations. In addition, the current state of epigenetic therapies for NEPC is addressed.

<h4>Background</h4>Alzheimer's disease (AD) is associated with abnormal tau and amyloid-β accumulation in the brain, leading to neurofibrillary tangles, neuropil threads and extracellular amyloid-β plaques. Treatment is limited to symptom management, a disease-modifying therapy is not available. To advance search of therapy approaches, there is a continued need to identify targets for disease intervention both by confirming existing hypotheses and generating new hypotheses.<h4>Methods</h4>We conducted a mRNA-seq study to identify genes associated with AD in post-mortem brain samples from the superior temporal gyrus (STG, n ​= ​76), and inferior frontal gyrus (IFG, n ​= ​65) brain regions. Differentially expressed genes (DEGs) were identified correcting for gender and surrogate variables to capture hidden variation not accounted for by pre-planned covariates. The results from this study were compared with the transcriptome studies from the Accelerated Medicine Partnership - Alzheimer's Disease (AMP-AD) initiative. Over-representation and gene set enrichment analysis (GSEA) was used to identify disease-associated pathways. Protein-protein interaction (PPI) and weighted gene co-expression network analysis (WGCNA) analyses were carried out and co-expressed gene modules and their hub genes were identified and associated with additional phenotypic traits of interest.<h4>Results</h4>Several hundred mRNAs were differentially expressed between AD cases and cognitively normal controls in the STG, while no and few transcripts met the same criteria (adjusted p less than 0.05 and fold change greater than 1.2) in the IFG. The findings were consistent at the gene set level with two out of three cohorts from AMP-AD. PPI analysis suggested that the DEGs were enriched in protein-protein interactions than expected by random chance. Over-representation and GSEA analysis suggested genes playing roles in neuroinflammation, amyloid-β, autophagy and trafficking being important for the AD disease process. At the gene level, 10 genes from the STG that were consistently differentially expressed in this study and in the MSBB study (one of the three cohorts within the AMP-AD initiative) were enriched in microglial genes (TREM2, C3AR1, ITGAX, OLR1, CD74, and HLA-DRA), but also included genes with a broader cell type expression pattern such as CDK2AP1. Among the DEGs with supporting evidence from an independent study, CDK2AP1 (most abundantly expressed in astrocyte) was the transcript with strongest association with antemortem cognitive measure (last Mini-Mental State Examination score) and neurofibril tangle burden but also associated with amyloid plaque burden, while OLR1 was the transcript with strongest association with amyloid plaque burden. GSEA and over-representation analyses revealed gene sets related to immune processes including neutrophil degranulation, interleukin 10 signaling, and interferon gamma signaling, complement and coagulation cascades, phosphatidylinositol signaling system, phagosome and neurotransmitter receptors and postsynaptic signal transmission were enriched from this study and replicated in an independent study.<h4>Conclusion</h4>This study identified differential gene sets, common with two out of three AMP-AD cohorts (ROSMAP and MSBB) and highlights microglia and astrocyte as the key cell-types with DGEs associated with AD clinical diagnosis, and/or antemortem cognitive measure as well as neuropathological indices. Future meta-analysis and causal inferential analysis will be helpful in pinpointing the most relevant pathways and genes to intervene.

<h4>ABSTRACT</h4> Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the virus responsible for the coronavirus disease 2019 pandemic. ORF6 is known to antagonize the interferon signaling by inhibiting the nuclear translocation of STAT1. Here we show that ORF6 acts as a virulence factor through two distinct strategies. First, ORF6 directly interacts with STAT1 in an IFN-independent manner to inhibit its nuclear translocation. Second, ORF6 directly binds to importin α1, which is a nuclear transport factor encoded by KPNA2 , leading to a significant suppression of importin α1-mediated nuclear transport. Furthermore, we found that KPNA2 knockout enhances the viral replication, suggesting that importin α1 suppresses the viral propagation. Additionally, the analyses of gene expression data revealed that importin α1 levels decreased significantly in the lungs of older individuals. Taken together, SARS-CoV-2 ORF6 disrupts the nucleocytoplasmic trafficking to accelerate the viral replication, resulting in the disease progression, especially in older individuals.

The coronavirus disease (COVID-19) pandemic caused by a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly spread worldwide recently, leading to a global social and economic disruption. Although the emergently approved vaccine programs against SARS-CoV-2 have been rolled out globally, the number of COVID-19 daily cases and deaths has remained significantly high. Here, we attempted to computationally screen for possible medications for COVID-19 via rapidly estimate the highly potential inhibitors from an FDA-approved drug database against the main protease (Mpro) of SARS-CoV-2. The approach combined molecular docking and fast pulling of ligand (FPL) simulations that were demonstrated to be accurate and suitable for quick prediction of SARS-CoV-2 Mpro inhibitors. The results suggested that twentyseven compounds were capable of strongly associating with SARS-CoV-2 Mpro. Among them, the seven top leads are daclatasvir, teniposide, etoposide, levoleucovorin, naldemedine, cabozantinib, and irinotecan. The potential application of these drugs in COVID-19 therapy has thus been discussed.

Loss of cellular homeostasis has been implicated in the etiology of several neurodegenerative diseases (NDs). However, the molecular mechanisms that underlie this loss remain poorly understood on a systems level in each case. Here, using a novel computational approach to integrate dimensional RNA-seq and in vivo neuron survival data, we map the temporal dynamics of homeostatic and pathogenic responses in four striatal cell types of Huntington's disease (HD) model mice. This map shows that most pathogenic responses are mitigated and most homeostatic responses are decreased over time, suggesting that neuronal death in HD is primarily driven by the loss of homeostatic responses. Moreover, different cell types may lose similar homeostatic processes, for example, endosome biogenesis and mitochondrial quality control in <i>Drd1</i>-expressing neurons and astrocytes. HD relevance is validated by human stem cell, genome-wide association study, and post-mortem brain data. These findings provide a new paradigm and framework for therapeutic discovery in HD and other NDs.

<h4>Introduction</h4>Hereditary haemochromatosis (HH) is a disease characterised by the excessive absorption of iron and its deposition in various organs. Late complications of this disease include cirrhosis, hepatocellular carcinoma, and endocrine disorders. Data from the literature on thyroid disorders in patients with HH are inconsistent and ambiguous, and no research has been done to determine the relationship between excessive accumulation of iron and the thyroid morphology. Therefore, the aim of this study was to characterise thyroid function and ultrasound images in patients with clinically overt hereditary haemochromatosis.<h4>Material and methods</h4>We studied 40 patients who were diagnosed with hereditary haemochromatosis with one of the mutations of the HFE gene and iron deposits in liver in specimen from liver biopsies (graded G2 to G4) or in MRI. To assess thyroid function, ultrasound examinations of the thyroid gland were performed and serum TSH concentrations were measured.<h4>Results</h4>We showed in our study that patients with HH have been diagnosed with thyroid focal lesions statistically less frequent than in the control group. We did not reveal any statistically significant difference in TSH concentration between patients with HH and the general population. However, patients with more severe iron deposits in liver showed lower TSH concentration.<h4>Conclusions</h4>Our results indicate lower incidence of focal lesions in thyroid gland in a group of patients with clinically overt hereditary haemochromatosis.

<h4>Background</h4>The clinical implications of SARS-CoV-2 infection are highly variable. Some people with SARS-CoV-2 infection remain asymptomatic, whilst the infection can cause mild to moderate COVID-19 and COVID-19 pneumonia in others. This can lead to some people requiring intensive care support and, in some cases, to death, especially in older adults. Symptoms such as fever, cough, or loss of smell or taste, and signs such as oxygen saturation are the first and most readily available diagnostic information. Such information could be used to either rule out COVID-19, or select patients for further testing. This is an update of this review, the first version of which published in July 2020.<h4>Objectives</h4>To assess the diagnostic accuracy of signs and symptoms to determine if a person presenting in primary care or to hospital outpatient settings, such as the emergency department or dedicated COVID-19 clinics, has COVID-19.<h4>Search methods</h4>For this review iteration we undertook electronic searches up to 15 July 2020 in the Cochrane COVID-19 Study Register and the University of Bern living search database. In addition, we checked repositories of COVID-19 publications. We did not apply any language restrictions.<h4>Selection criteria</h4>Studies were eligible if they included patients with clinically suspected COVID-19, or if they recruited known cases with COVID-19 and controls without COVID-19. Studies were eligible when they recruited patients presenting to primary care or hospital outpatient settings. Studies in hospitalised patients were only included if symptoms and signs were recorded on admission or at presentation. Studies including patients who contracted SARS-CoV-2 infection while admitted to hospital were not eligible. The minimum eligible sample size of studies was 10 participants. All signs and symptoms were eligible for this review, including individual signs and symptoms or combinations. We accepted a range of reference standards.<h4>Data collection and analysis</h4>Pairs of review authors independently selected all studies, at both title and abstract stage and full-text stage. They resolved any disagreements by discussion with a third review author. Two review authors independently extracted data and resolved disagreements by discussion with a third review author. Two review authors independently assessed risk of bias using the Quality Assessment tool for Diagnostic Accuracy Studies (QUADAS-2) checklist. We presented sensitivity and specificity in paired forest plots, in receiver operating characteristic space and in dumbbell plots. We estimated summary parameters using a bivariate random-effects meta-analysis whenever five or more primary studies were available, and whenever heterogeneity across studies was deemed acceptable.<h4>Main results</h4>We identified 44 studies including 26,884 participants in total. Prevalence of COVID-19 varied from 3% to 71% with a median of 21%. There were three studies from primary care settings (1824 participants), nine studies from outpatient testing centres (10,717 participants), 12 studies performed in hospital outpatient wards (5061 participants), seven studies in hospitalised patients (1048 participants), 10 studies in the emergency department (3173 participants), and three studies in which the setting was not specified (5061 participants). The studies did not clearly distinguish mild from severe COVID-19, so we present the results for all disease severities together. Fifteen studies had a high risk of bias for selection of participants because inclusion in the studies depended on the applicable testing and referral protocols, which included many of the signs and symptoms under study in this review. This may have especially influenced the sensitivity of those features used in referral protocols, such as fever and cough. Five studies only included participants with pneumonia on imaging, suggesting that this is a highly selected population. In an additional 12 studies, we were unable to assess the risk for selection bias. This makes it very difficult to judge the validity of the diagnostic accuracy of the signs and symptoms from these included studies. The applicability of the results of this review update improved in comparison with the original review. A greater proportion of studies included participants who presented to outpatient settings, which is where the majority of clinical assessments for COVID-19 take place. However, still none of the studies presented any data on children separately, and only one focused specifically on older adults. We found data on 84 signs and symptoms. Results were highly variable across studies. Most had very low sensitivity and high specificity. Only cough (25 studies) and fever (7 studies) had a pooled sensitivity of at least 50% but specificities were moderate to low. Cough had a sensitivity of 67.4% (95% confidence interval (CI) 59.8% to 74.1%) and specificity of 35.0% (95% CI 28.7% to 41.9%). Fever had a sensitivity of 53.8% (95% CI 35.0% to 71.7%) and a specificity of 67.4% (95% CI 53.3% to 78.9%). The pooled positive likelihood ratio of cough was only 1.04 (95% CI 0.97 to 1.11) and that of fever 1.65 (95% CI 1.41 to 1.93). Anosmia alone (11 studies), ageusia alone (6 studies), and anosmia or ageusia (6 studies) had sensitivities below 50% but specificities over 90%. Anosmia had a pooled sensitivity of 28.0% (95% CI 17.7% to 41.3%) and a specificity of 93.4% (95% CI 88.3% to 96.4%). Ageusia had a pooled sensitivity of 24.8% (95% CI 12.4% to 43.5%) and a specificity of 91.4% (95% CI 81.3% to 96.3%). Anosmia or ageusia had a pooled sensitivity of 41.0% (95% CI 27.0% to 56.6%) and a specificity of 90.5% (95% CI 81.2% to 95.4%). The pooled positive likelihood ratios of anosmia alone and anosmia or ageusia were 4.25 (95% CI 3.17 to 5.71) and 4.31 (95% CI 3.00 to 6.18) respectively, which is just below our arbitrary definition of a 'red flag', that is, a positive likelihood ratio of at least 5. The pooled positive likelihood ratio of ageusia alone was only 2.88 (95% CI 2.02 to 4.09). Only two studies assessed combinations of different signs and symptoms, mostly combining fever and cough with other symptoms. These combinations had a specificity above 80%, but at the cost of very low sensitivity (< 30%).<h4>Authors' conclusions</h4>The majority of individual signs and symptoms included in this review appear to have very poor diagnostic accuracy, although this should be interpreted in the context of selection bias and heterogeneity between studies. Based on currently available data, neither absence nor presence of signs or symptoms are accurate enough to rule in or rule out COVID-19. The presence of anosmia or ageusia may be useful as a red flag for COVID-19. The presence of fever or cough, given their high sensitivities, may also be useful to identify people for further testing. Prospective studies in an unselected population presenting to primary care or hospital outpatient settings, examining combinations of signs and symptoms to evaluate the syndromic presentation of COVID-19, are still urgently needed. Results from such studies could inform subsequent management decisions.

Microgliosis is a prominent pathological feature in many neurological diseases including multiple sclerosis (MS), a progressive auto-immune demyelinating disorder. The precise role of microglia, parenchymal central nervous system (CNS) macrophages, during demyelination, and the relative contributions of peripheral macrophages are incompletely understood. Classical markers used to identify microglia do not reliably discriminate between microglia and peripheral macrophages, confounding analyses. Here, we use a genetic fate mapping strategy to identify microglia as predominant responders and key effectors of demyelination in the cuprizone (CUP) model. Colony-stimulating factor 1 (CSF1), also known as macrophage colony-stimulating factor (M-CSF) - a secreted cytokine that regulates microglia development and survival-is upregulated in demyelinated white matter lesions. Depletion of microglia with the CSF1R inhibitor PLX3397 greatly abrogates the demyelination, loss of oligodendrocytes, and reactive astrocytosis that results from CUP treatment. Electron microscopy (EM) and serial block face imaging show myelin sheaths remain intact in CUP treated mice depleted of microglia. However, these CUP-damaged myelin sheaths are lost and robustly phagocytosed upon-repopulation of microglia. Direct injection of CSF1 into CNS white matter induces focal microgliosis and demyelination indicating active CSF1 signaling can promote demyelination. Finally, mice defective in adopting a toxic astrocyte phenotype that is driven by microglia nevertheless demyelinate normally upon CUP treatment implicating microglia rather than astrocytes as the primary drivers of CUP-mediated demyelination. Together, these studies indicate activated microglia are required for and can drive demyelination directly and implicate CSF1 signaling in these events.

<h4>Background</h4>Individuals undergoing hemodialysis in the United States frequently report pain and receive three-fold more opioid prescriptions than the general population. While opioid use is appropriate for select patients, high-dose utilization may contribute to an increased risk of death due to possible accumulation of opioid metabolites.<h4>Methods</h4>We studied high-dose opioid utilization (≥120 morphine milligram equivalents [MME] per day) among adults initiating hemodialysis in the United States between 2007 and 2014 using national registry data. We calculated the cumulative incidence (%) of high-dose utilization and depicted trends in the average percentage of days individuals were exposed to opioids. We used adjusted Cox proportional hazards models to identify which opioid doses were associated with mortality.<h4>Results</h4>Among 327,344 adults undergoing hemodialysis, the cumulative incidence of high-dose utilization was 14.9% at 2 years after initiating hemodialysis. Among patients with ≥1 opioid prescription during follow-up, the average percentage of days exposed to high-dose utilization increased from 13.9% in 2007 to 26.1% in 2014. Compared to 0MME per day, doses < 60MME were not associated with an increased risk of mortality, but high-dose utilization was associated with a 1.63-fold (95% CI, 1.57, 1.69) increased risk of mortality. The risk of mortality associated with opioid dose was highest in the first year after hemodialysis initiation.<h4>Conclusions</h4>The risk of mortality associated with opioid utilization among individuals on hemodialysis increases as doses exceed 60MME per day and is greatest during periods of high-dose utilization. Patients and clinicians should carefully weigh the risks and benefits of opioid doses exceeding 60MME per day.

<h4>Background</h4>Super-enhancer-associated long noncoding RNAs (SE-lncRNAs) have been reported to play essential roles in tumorigenesis, but the fundamental mechanism of SE-lncRNAs in colorectal cancer (CRC) remains largely unknown.<h4>Methods</h4>A microarray was performed to identify the differentially expressed SE-lncRNAs between CRC tissues and peritumoral tissues. A novel SE-lncRNA, AC005592.2, was selected from these differentially expressed SE-lncRNAs to explore its effects on CRC development. Fluorescence quantitative real-time PCR (qRT-PCR) was used to assay the expression of AC005592.2 in CRC tissues and cell lines. Functional assays were applied to identify the biological effects of AC005592.2 in CRC cells. Furthermore, RNA-seq was employed to predict potential targets of AC005592.2.<h4>Results</h4>AC005592.2 was significantly increased in CRC tissues and cells. High expression of AC005592.2 was significantly associated with TNM stage and tumor differentiation in CRC patients. Knockdown of AC005592.2 suppressed CRC cell proliferation, invasion and migration but promoted apoptosis, while AC005592.2 overexpression exerted the opposite effects on CRC cells. In addition, AC005592.2 positively regulated the expression of olfactomedin 4 (OLFM4), which was also upregulated in CRC tissues.<h4>Conclusion</h4>The findings suggested that AC005592.2 is a crucial promoter of CRC progression and may serve as an attractive therapeutic target for CRC.

To enhance understanding of polycystic ovary syndrome (PCOS) at the molecular level; this investigation intends to examine the genes and pathways associated with PCOS by using an integrated bioinformatics analysis. Based on the expression profiling by high throughput sequencing data GSE84958 derived from the Gene Expression Omnibus (GEO) database, the differentially expressed genes (DEGs) between PCOS samples and normal controls were identified. We performed a functional enrichment analysis. A protein-protein interaction (PPI) network, miRNA- target genes and TF - target gene networks, were constructed and visualized, with which the hub gene nodes were identified. Validation of hub genes was performed by using receiver operating characteristic (ROC) and RT-PCR. Small drug molecules were predicted by using molecular docking. A total of 739 DEGs were identified, of which 360 genes were up regulated and 379 genes were down regulated. GO enrichment analysis revealed that up regulated genes were mainly involved in peptide metabolic process, organelle envelope and RNA binding and the down regulated genes were significantly enriched in plasma membrane bounded cell projection organization, neuron projection and DNA-binding transcription factor activity, RNA polymerase II-specific. REACTOME pathway enrichment analysis revealed that the up regulated genes were mainly enriched in translation and respiratory electron transport and the down regulated genes were mainly enriched in generic transcription pathway and transmembrane transport of small molecules. The top 10 hub genes (SAA1, ADCY6, POLR2K, RPS15, RPS15A, CTNND1, ESR1, NEDD4L, KNTC1 and NGFR) were identified from PPI network, miRNA - target gene network and TF - target gene network. The modules analysis showed that genes in modules were mainly associated with the transport of respiratory electrons and signaling NGF, respectively. We find a series of crucial genes along with the pathways that were most closely related with PCOS initiation and advancement. Our investigations provide a more detailed molecular mechanism for the progression of PCOS, detail information on the potential biomarkers and therapeutic targets.

<h4>Background</h4>Long non-coding RNAs (lncRNA) are reported to influence colorectal cancer (CRC) progression. Currently, the functions of the lncRNA ZNF561 antisense RNA 1 (ZNF561-AS1) in CRC are unknown.<h4>Methods</h4>ZNF561-AS1 and SRSF6 expression in CRC patient samples and CRC cell lines was evaluated through TCGA database analysis, western blot along with real-time PCR. SRSF6 expression in CRC cells was also examined upon ZNF561-AS1 depletion or overexpression. Interaction between miR-26a-3p, miR-128-5p, ZNF561-AS1, and SRSF6 was examined by dual luciferase reporter assay, as well as RNA binding protein immunoprecipitation (RIP) assay. Small interfering RNA (siRNA) mediated knockdown experiments were performed to assess the role of ZNF561-AS1 and SRSF6 in the proliferative actives and apoptosis rate of CRC cells. A mouse xenograft model was employed to assess tumor growth upon ZNF561-AS1 knockdown and SRSF6 rescue.<h4>Results</h4>We find that ZNF561-AS1 and SRSF6 were upregulated in CRC patient tissues. ZNF561-AS1 expression was reduced in tissues from treated CRC patients but upregulated in CRC tissues from relapsed patients. SRSF6 expression was suppressed and enhanced by ZNF561-AS1 depletion and overexpression, respectively. Mechanistically, ZNF561-AS1 regulated SRSF6 expression by sponging miR-26a-3p and miR-128-5p. ZNF561-AS1-miR-26a-3p/miR-128-5p-SRSF6 axis was required for CRC proliferation and survival. ZNF561-AS1 knockdown suppressed CRC cell proliferation and triggered apoptosis. ZNF561-AS1 depletion suppressed the growth of tumors in a model of a nude mouse xenograft. Similar observations were made upon SRSF6 depletion. SRSF6 overexpression reversed the inhibitory activities of ZNF561-AS1 in vivo, as well as in vitro.<h4>Conclusion</h4>In summary, we find that ZNF561-AS1 promotes CRC progression via the miR-26a-3p/miR-128-5p-SRSF6 axis. This study reveals new perspectives into the role of ZNF561-AS1 in CRC.

<h4>Background</h4>Brain energy metabolism is impaired in Alzheimer's disease (AD), which may be mitigated by a ketogenic diet. We conducted a randomized crossover trial to determine whether a 12-week modified ketogenic diet improved cognition, daily function, or quality of life in a hospital clinic of AD patients.<h4>Methods</h4>We randomly assigned patients with clinically confirmed diagnoses of AD to a modified ketogenic diet or usual diet supplemented with low-fat healthy-eating guidelines and enrolled them in a single-phase, assessor-blinded, two-period crossover trial (two 12-week treatment periods, separated by a 10-week washout period). Primary outcomes were mean within-individual changes in the Addenbrookes Cognitive Examination - III (ACE-III) scale, AD Cooperative Study - Activities of Daily Living (ADCS-ADL) inventory, and Quality of Life in AD (QOL-AD) questionnaire over 12 weeks. Secondary outcomes considered changes in cardiovascular risk factors and adverse effects.<h4>Results</h4>We randomized 26 patients, of whom 21 (81%) completed the ketogenic diet; only one withdrawal was attributed to the ketogenic diet. While on the ketogenic diet, patients achieved sustained physiological ketosis (12-week mean beta-hydroxybutyrate level: 0.95 ± 0.34 mmol/L). Compared with usual diet, patients on the ketogenic diet increased their mean within-individual ADCS-ADL (+ 3.13 ± 5.01 points, P = 0.0067) and QOL-AD (+ 3.37 ± 6.86 points, P = 0.023) scores; the ACE-III also increased, but not significantly (+ 2.12 ± 8.70 points, P = 0.24). Changes in cardiovascular risk factors were mostly favourable, and adverse effects were mild.<h4>Conclusions</h4>This is the first randomized trial to investigate the impact of a ketogenic diet in patients with uniform diagnoses of AD. High rates of retention, adherence, and safety appear to be achievable in applying a 12-week modified ketogenic diet to AD patients. Compared with a usual diet supplemented with low-fat healthy-eating guidelines, patients on the ketogenic diet improved in daily function and quality of life, two factors of great importance to people living with dementia.<h4>Trial registration</h4>This trial is registered on the Australia New Zealand Clinical Trials Registry, number ACTRN12618001450202 . The trial was registered on August 28, 2018.

Ribosome biogenesis in eukaryotes requires the coordinated production and assembly of 80 ribosomal proteins and four ribosomal RNAs (rRNAs), and its rate must be synchronized with cellular growth. Here, we showed that the Microprocessor complex, which mediates the first step of microRNA processing, potentiated the transcription of ribosomal protein genes by eliminating DNA/RNA hybrids known as R-loops. Nutrient deprivation triggered the nuclear export of Drosha, a key component of the Microprocessor complex, and its subsequent degradation by the E3 ubiquitin ligase Nedd4, thereby reducing ribosomal protein production and protein synthesis. In mouse erythroid progenitors, conditional deletion of <i>Drosha</i> led to the reduced production of ribosomal proteins, translational inhibition of the mRNA encoding the erythroid transcription factor Gata1, and impaired erythropoiesis. This phenotype mirrored the clinical presentation of human "ribosomopathies." Thus, the Microprocessor complex plays a pivotal role in synchronizing protein synthesis capacity with cellular growth rate and is a potential drug target for anemias caused by ribosomal insufficiency.

Droplet libraries consisting of many reagents encapsulated in separate droplets are necessary for applications of microfluidics, including combinatorial chemical synthesis, DNA-encoded libraries, and massively multiplexed PCR. However, existing approaches for generating them are laborious and impractical. Here, we describe an automated approach using a commercial array spotter. The approach can controllably emulsify hundreds of different reagents in a fraction of the time of manual operation of a microfluidic device, and without any user intervention. We demonstrate that the droplets produced by the spotter are similarly uniform to those produced by microfluidics and automate the generation of a ~ 2 mL emulsion containing 192 different reagents in ~ 4 h. The ease with which it can generate high diversity droplet libraries should make combinatorial applications more feasible in droplet microfluidics. Moreover, the instrument serves as an automated droplet generator, allowing execution of droplet reactions without microfluidic expertise.

Infants with KMT2A-rearranged acute lymphoblastic leukemia (KMT2A-r ALL) have a poor prognosis. KMT2A-r ALL overexpresses FLT3, and the FLT3 inhibitor (FLT3i) lestaurtinib potentiates chemotherapy-induced cytotoxicity in preclinical models. Children's Oncology Group (COG) AALL0631 tested whether adding lestaurtinib to post-induction chemotherapy improved event-free survival (EFS). After chemotherapy induction, KMT2A-r infants received either chemotherapy only or chemotherapy plus lestaurtinib. Correlative assays included FLT3i plasma pharmacodynamics (PD), which categorized patients as inhibited or uninhibited, and FLT3i ex vivo sensitivity (EVS), which categorized leukemic blasts as sensitive or resistant. There was no difference in 3-year EFS between patients treated with chemotherapy plus lestaurtinib (n = 67, 36 ± 6%) vs. chemotherapy only (n = 54, 39 ± 7%, p = 0.67). However, for the lestaurtinib-treated patients, FLT3i PD and FLT3i EVS significantly correlated with EFS. For FLT3i PD, EFS for inhibited/uninhibited was 59 ± 10%/28 ± 7% (p = 0.009) and for FLTi EVS, EFS for sensitive/resistant was 52 ± 8%/5 ± 5% (p < 0.001). Seventeen patients were both inhibited and sensitive, with an EFS of 88 ± 8%. Adding lestaurtinib did not improve EFS overall, but patients achieving potent FLT3 inhibition and those whose leukemia blasts were sensitive FLT3-inhibition ex vivo did benefit from the addition of lestaurtinib. Patient selection and PD-guided dose escalation may enhance the efficacy of FLT3 inhibition for KMT2A-r infant ALL.

Circular RNAs (circRNAs) are involved in various biological processes and disease pathogenesis. However, only a small number of functional circRNAs have been identified among hundreds of thousands of circRNA species, partly because most current methods are based on circular junction counts and overlook the fact that a circRNA is formed from the host gene by back-splicing (BS). To distinguish the expression difference originating from BS or the host gene, we present differentially expressed back-splicing (DEBKS), a software program to streamline the discovery of differential BS events between two rRNA-depleted RNA sequencing (RNA-seq) sample groups. By applying to real and simulated data and employing RT-qPCR for validation, we demonstrate that DEBKS is efficient and accurate in detecting circRNAs with differential BS events between paired and unpaired sample groups. DEBKS is available at https://github.com/yangence/DEBKS as open-source software.

<h4>Background</h4>Cholesterol gallstone (CG) is the most common gallstone disease, which is induced by biliary cholesterol supersaturation. The purpose of this study is to investigate the pathogenesis of CG.<h4>Methods</h4>Sixteen mice were equally and randomly divided into model group and normal control group. The model group was fed with lithogenic diets to induce CG, and then gallbladder bile lipid analysis was performed. After RNA-seq library was constructed, differentially expressed mRNAs (DE-mRNAs) and differentially expressed lncRNAs (DE-lncRNAs) between model group and normal control group were analyzed by DESeq2 package. Using the cluster Profiler package, enrichment analysis for the DE-mRNAs was carried out. Based on Cytoscape software, the protein-protein interaction (PPI) network and competing endogenous RNA (ceRNA) network were built. Using quantitative real-time reverse transcription-PCR (qRT-PCR) analysis, the key RNAs were validated.<h4>Results</h4>The mouse model of CG was suc cessfully established, and then 181 DE-mRNAs and 33 DE-lncRNAs between model and normal groups were obtained. Moreover, KDM4A was selected as a hub node in the PPI network, and lncRNA <i>MEG3</i> was considered as a key lncRNA in the regulatory network. Additionally, the <i>miR-107-5p/miR-149-3p/miR-346-3-MEG3</i> regulatory pairs and <i>MEG3-PABPC4/CEP131/NUMB1</i> co-expression pairs existed in the regulatory network. The qRT-PCR analysis showed that <i>KDM4A</i> expression was increased, and the expressions of <i>MEG3</i>, <i>PABPC4</i>, <i>CEP131</i>, and <i>NUMB1</i> were downregulated.<h4>Conclusion</h4>These RNAs might be related to the pathogenesis of CG.

Pulmonary arterial hypertension (PAH) is a severe cardiovascular disease that is caused by the progressive occlusion of the distal pulmonary arteries, eventually leading to right heart failure and death. Almost 40% of patients with PAH are iron deficient. Although widely studied, the mechanisms linking between PAH and iron deficiency remain unclear. Here we review the mechanisms regulating iron homeostasis and the preclinical and clinical data available on iron deficiency in PAH. Then we discuss the potential implications of iron deficiency on the development and management of PAH.

Although many studies have been conducted on single gene therapies in cancer patients, the reality is that tumor arises from different coordinating protein groups. Unveiling perturbations in protein interactome related to the tumor formation may contribute to the development of effective diagnosis, treatment strategies, and prognosis. In this study, considering the clinical and transcriptome data of three Renal Cell Carcinoma (RCC) subtypes (ccRCC, pRCC, and chRCC) retrieved from The Cancer Genome Atlas (TCGA) and the human protein interactome, the differential protein-protein interactions were identified in each RCC subtype. The approach enabled the identification of differentially interacting proteins (DIPs) indicating prominent changes in their interaction patterns during tumor formation. Further, diagnostic and prognostic performances were generated by taking into account DIP clusters which are specific to the relevant subtypes. Furthermore, considering the mesenchymal epithelial transition (MET) receptor tyrosine kinase (PDB ID: 3DKF) as a potential drug target specific to pRCC, twenty-one lead compounds were identified through virtual screening of ZINC molecules. In this study, we presented remarkable findings in terms of early diagnosis, prognosis, and effective treatment strategies, that deserve further experimental and clinical efforts.

<h4>Background</h4>Incidence of biliary tract cancers (BTCs) in China is high, and surgery is the only curative option. Preoperative CA19-9 has been identified as a predictor for survival in patients with resectable BTCs, but more potential predictors need to be studied. This retrospective study aimed to establish the prognostic significance of CA19-9/TBIL ratio (CTR) in patients with BTCs.<h4>Methods</h4>A retrospective analysis was performed in patients who were diagnosed with BTCs and received surgical resection between 2013 and 2018 at PUMCH. Demographic and clinical parameters were collected. Preoperative CA19-9 and CTR were classified as elevated (>58.6 and >0.83) according to the receiver operating characteristic (ROC) analysis. Demographic and clinical parameters were compared between the groups using Student's <i>t</i>-test, chi-square, or Fisher's exact test. Survival analysis was performed by the Kaplan-Meier methods, and the relationship between variables and survival was assessed by the log-rank test. Cox regression analysis was conducted to identify potential risk factors for overall survival.<h4>Results</h4>In total, 109 participants were involved in the final analysis. The overall survival rate was 18.0% at 5 years, with a median survival duration of 1.58 years. The Kaplan-Meier analysis indicated that higher CTR was associated with shorter OS (15 vs. 50, <i>p</i> < 0.01). Univariate survival analysis identified TNM staging, CA19-9, and CTR as statistically significant prognostic factors. In a multiple Cox analysis, only CTR was proved as a significantly independent prognostic factor.<h4>Conclusion</h4>CTR acts as an independent prognostic predictor for patients with biliary tract cancer.

Extracellular vesicles or EVs are secreted by most, if not all, eukaryote cell types and recaptured by neighboring or distant cells. Their cargo, composed of a vast diversity of proteins, lipids, and nucleic acids, supports the EVs' inter-cellular communication. The role of EVs in many cellular processes is now well documented both in physiological and pathological conditions. In this review, we focus on the role of EVs in the central nervous system (CNS) in physiological as well as pathological conditions such as neurodegenerative diseases or brain cancers. We also discuss the future of EVs in clinical research, in particular, their value as biomarkers as well as innovative therapeutic agents. While an increasing number of studies reveal EV research as a promising field, progress in the standardization of protocols and innovation in analysis as well as in research tools is needed to make a breakthrough in our understanding of their impact in the pathophysiology of the brain.

Normal or "good" inflammation process starts from a local cellular response against injury or any infectious agent, with the activation of neutrophils, macrophages, Langerhans cells, dendritic cells, and innate immune cells. Cytokines and chemokines are produced to amplify the local inflammatory process followed by the migration of immune cells to the regional lymph nodes where adaptive immune response is initiated. Systemic inflammation enhances the biological response to mobilize additional cells from central and peripheral immune/hematopoietic system. Local mechanisms to limit inflammation are initiated and lead to healing. During the normal inflammatory process, there is a balance between the production of inflammatory chemokines/cytokines such as Tumor Necrosis Factor (TNF)-α, interleukin (IL)-6 and IL-1 and the production of compounds that limit inflammation and have an immune suppressive effect, such as IL-10 and Transforming Factor (TGF) β. IL-6 and IL-6/soluble IL-6 Receptor (R) complex stimulate liver cells to produce inflammatory proteins, which represents the systemic inflammation response. The magnitude and the duration of the systemic inflammatory response are linked to the cause, under genetic and epigenetic control. Significant inflammation as seen in septic shock, in severe forms of infections or in certain active cancers, represents the "bad inflammation", correlated with a poor prognosis. In addition, the persistence of a chronic smoldering inflammation may lead to pathological situations which are observed in the majority of inflammatory, degenerative, dysmetabolic, or dysimmune diseases and cancer. Chronic smoldering inflammation is a cross between different pathological situations possibly linked. In addition, within the tumor microenvironment, inflammatory process results from different cellular mechanisms modulated by metabolic and vascular changes. On the contrary, a limited and balanced inflammation initiates the normal immune response, including the adaptive response which amplifies any immunotherapy, including vaccines. Immune checkpoint inhibitors and chimeric antigen receptor (CAR) T-cells are associated with cytokine release syndrome, a clinical risk leading to the use of anti-cytokine drugs. Nowadays, it is time to monitor the dynamic inflammatory process for a better immune precision medicine in both infections and cancer.

<h4>Background</h4>Innate lymphoid cells (ILCs), so far studied mostly in mouse models, are important tissue-resident innate immune cells that play important roles in the colorectal cancer microenvironment and maintain mucosal tissue homeostasis. Plasmacytoid dendritic cells (pDCs) present complexity in various tumor types and are correlated with poor prognosis. pDCs can promote HIV-1-induced group 3 ILC (ILC3) depletion through the CD95 pathway. However, the role of ILC3s in human colon cancer and their correlation with other immune cells, especially pDCs, remain unclear.<h4>Methods</h4>We characterized ILCs and pDCs in the tumor microenvironment of 58 colon cancer patients by flow cytometry and selected three patients for RNA sequencing.<h4>Results</h4>ILC3s were negatively correlated, and pDCs were positively correlated, with cancer pathological stage. There was a negative correlation between the numbers of ILC3s and pDCs in tumor tissues. RNA sequencing confirmed the correlations between ILC3s and pDCs and highlighted the potential function of many ILC- and pDC-associated differentially expressed genes in the regulation of tumor immunity. pDCs can induce apoptosis of ILC3s through the CD95 pathway in the tumor-like microenvironment.<h4>Conclusions</h4>One of the interactions between ILC3s and pDCs is <i>via</i> the CD95 pathway, which may help explain the role of ILC3s in colon cancer.

<b>Background:</b> Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a poorly understood disease involving a high inflammatory status. Neutrophil extracellular traps (NETs) have been described as a new pathway to contain infectious diseases but can also participate in the imbalance of the inflammatory and the coagulation systems. NETs could be a therapeutic target in COVID-19 patients. <b>Methods:</b> Consecutive patients with SARS-CoV2 related pneumonia admitted to the intensive care unit were included in a prospective bicentric study. Neutrophil extracellular trap concentrations were quantified in whole blood samples at day-1 and day-3 by flow cytometry. The primary outcome was the association between the blood NET quantification at ICU admission and the number of days with refractory hypoxemia defined by a PaO₂/FIO₂ ratio ≤100 mmHg. <b>Results:</b> Among 181 patients admitted to the ICUs for acute respiratory failure related to SARS-CoV2 pneumonia, 58 were included in the analysis. Patients were 62 [54, 69] years old in median, mostly male (75.9%). The median number of days with severe hypoxemia was 4 [2, 6] days and day-28 mortality was 27.6% (<i>n</i> = 16). The blood level of NETs significantly decreased between day-1 and day-3 in patients who survived (59.5 [30.5, 116.6] to 47 [33.2, 62.4] <i>p</i> = 0.006; 8.6 [3.4, 18.0] to 4 [1.4, 10.7] <i>p</i> = 0.001 and 7.4 [4.0, 16.7] to 2.6 [1.0, 8.3] <i>p</i> = 0.001 for MPO+, Cit-H3+, and MPO+ Cit-H3+ NETs, respectively) while it remained stable in patients who died (38.4 [26.0, 54.8] to 44.5 [36.4, 77.7] <i>p</i> = 0.542; 4.9 [1.3, 13.0] to 5.5 [2.8, 6.9] <i>p</i> = 0.839 and 4 [1.3, 13.6] to 2.7 [1.4, 4.5] <i>p</i> = 0.421 for MPO+, Cit-H3+, and MPO+ Cit-H3+ NETs, respectively). In multivariable negative binomial regression, the blood level of MPO+ NETs was negatively associated with the number of days with severe hypoxemia within 7 days (0.84 [0.73, 0.97]), while neither Cit-H3+ NETs nor double-positive NETs were significantly associated with the primary outcome. <b>Conclusion:</b> The whole blood level of NETs at day-1 was negatively associated with the number of days with severe hypoxemia in patients admitted to the intensive care unit for SARS-CoV2 related pneumonia. The lack of decrease of the blood level of NETs between day-1 and day-3 discriminated patients who died within day-28.

Dedifferentiation of chondrocyte greatly restricts its function and application, however, it is poorly understood except a small number of canonical markers. The non-cell-adhesive property endows polysaccharide hydrogel with the ability to maintain chondrocyte phenotype, which can serve as a platform to identify new molecular markers and therapeutic targets of chondrocyte dedifferentiation. In this study, the high-throughput RNA sequencing (RNA-seq) was first performed on articular chondrocytes at primary (P0) and passage 1 (P1) stages to explore the global alteration of gene expression along with chondrocyte dedifferentiation. Significantly, several potential marker genes, such as PFKFB3, KDM6B, had been identified via comparatively analyzing their expression in P0 and P1 chondrocytes as well as in 3D constructs (i.e. chondrocyte-laden alginate hydrogel and HA-MA hydrogel) at both mRNA and protein level. Besides, the changes in cellular morphology and enriched pathway of differentially expressed genes during chondrocyte dedifferentiation was studied in detail. This study developed the use of hydrogel as a platform to investigate chondrocyte dedifferentiation; the results provided new molecular markers and potential therapeutic targets of chondrocyte dedifferentiation.

Isoflavonoid phytoalexins (isoflavones: genistein, 2'-hydroxygenistein, and daidzein; isoflavanones: dalbergioidin and kievitone; coumestrol; pterocarpans: phaseollidin and phaseollin; and the isoflavan: phaseollinisoflavan) production in response to the application of eleven 1-oxo-indane-4-carboxylic acid derivatives (indanoyl esters and indanoyl amino acids conjugates), in cotyledons and hypocotyl/root of two common bean (<i>Phaseolus vulgaris</i> L.) cultivars was evaluated. The content of isoflavonoids depended on the cultivar, the treated tissue, the time after induction, the structure and concentration of the elicitor. The highest isoflavonoid contents were found when 1-oxo-indanoyl-amino acids conjugates were used as elicitors. Cotyledons and hypocotyl/root of the anthracnose-resistant cultivar produced significantly higher isoflavonoid contents as compared to the susceptible one. Maximum levels of phaseollin were obtained using 0.66 mM 1-oxo-indanoyl-l-isoleucyl methyl ester and between 72 and 96 h post-induction. So, 1-oxo-indane-4-carboxylic acid derivatives, may be used to enhance the amount of isoflavonoid phytoalexins in common bean and protect crops from phytopathogenic microorganisms.

Coronavirus disease 2019 (COVID-19) is an emerging infectious disease that leads to severe respiratory failure (RF). It is known that host exposure to viral infection triggers an iron-lowering response to mitigate pathogenic load and tissue damage. However, the association between host iron-lowering response and COVID-19 severity is not clear. This two-center observational study of 136 adult hospitalized COVID-19 patients analyzed the association between disease severity and initial serum iron, total iron-binding capacity (TIBC), and transferrin saturation (TSAT) levels. Serum iron levels were significantly lower in patients with mild RF than in the non-RF group; however, there were no significant differences in iron levels between the non-RF and severe RF groups, depicting a U-shaped association between serum iron levels and disease severity. TIBC levels decreased significantly with increasing severity; consequently, TSAT was significantly higher in patients with severe RF than in other patients. Multivariate analysis including only patients with RF adjusted for age and sex demonstrated that higher serum iron and TSAT levels were independently associated with the development of severe RF, indicating that inadequate response to lower serum iron might be an exacerbating factor for COVID-19.

<title>Abstract</title> <p>Expanding the exercise capacity of skeletal muscle is an emerging strategy to combat obesity-related metabolic diseases and this can be achieved by shifting skeletal muscle fibers toward slow-twitch oxidative type. Here, we report that Sirt6, an anti-aging histone deacetylase, is critical in regulating myofiber configuration toward oxidative type and that Sirt6 activator can be an exercise mimetic. Genetic inactivation of Sirt6 in skeletal muscle reduced while its transgenic overexpression increased mitochondrial oxidative capacity and exercise performance in mice. Mechanistically, we show that Sirt6 downregulated Sox6, a key repressor of slow fiber specific gene, by increasing the transcription of CREB. Sirt6 expression is elevated in chronically exercised humans and mice treated with an activator of Sirt6 showed an increase in exercise endurance as compared to exercise-trained controls. Thus, the current study identifies Sirt6 as a new molecular target for reprogramming myofiber composition toward the oxidative type and for improving muscle performance.</p>

Limited data exists regarding the disease course of Huntington's Disease (HD) in children and young adults. Here, we evaluate the trajectory of various cognitive skill development as a function of cytosine-adenine-guanine (CAG) repeat length in children and adolescents that carry the mutation that causes HD. We discovered that the development of verbal skills seems to plateau earlier as CAG repeat length increases. These findings increase our understanding of the relationship between neurodegeneration and neurodevelopment and may have far-reaching implications for future gene-therapy treatment strategies. ANN NEUROL 2021;89:1036-1040.

Critical periods are discrete developmental stages when the nervous system is especially sensitive to stimuli that facilitate circuit maturation. The distinctive landscapes assumed by the developing CNS create analogous periods of susceptibility to pathogenic insults and responsiveness to therapy. Here, we review critical periods in nervous system development and disease, with an emphasis on the neurodevelopmental disorder DYT1 dystonia. We highlight clinical and laboratory observations supporting the existence of a critical period during which the DYT1 mutation is uniquely harmful, and the implications for future therapeutic development.

<h4>Purpose</h4>We aimed to investigate the prognostic value of multiple immune cell markers including programmed death-ligand 1 (PD-L1) and poliovirus receptor (PVR) in head and neck squamous cell carcinoma (HNSCC) using archival tumor tissues METHODS: Patients diagnosed with HNSCC who have undergone surgical resection in 2005-2012 were included. Correlations between PVR and PD-L1 expression and patient characteristics were analyzed by analysis of variance. The Kaplan-Meier method and log-rank test were used to estimate survival. P values < 0.05 were considered statistically significant.<h4>Results</h4>In total, 375 primary tumor tissues were analyzed using immunohistochemistry. High PVR expression was associated with a poor prognosis in terms of overall survival (OS) and recurrence-free survival (RFS), and tumors with high PVR expression were associated with a short OS. PD-L1 tumor expression did not have a prognostic impact on survival. Univariate analysis revealed that OS and RFS were affected by age and p16 and PVR expression; multivariate analysis revealed that age and p16 and PVR expression were the most important determinants of RFS.<h4>Conclusion</h4>PVR overexpression is a poor prognostic factor in patients with HNSCC and co-targeting PVR and PD-L1 may be a promising therapeutic option that needs further investigation.

Isoniazid and rifampicin are crucial for treating tuberculosis (TB); however, they can cause severe hepatotoxicity leading to liver failure. Therapeutic options are limited and ineffective. We hypothesized that prophylaxis with quercetin attenuates isoniazid- and rifampicin-induced liver injury. We randomly divided Wistar rats into seven groups (<i>n</i> = 6). The animals received isoniazid and rifampicin or were co-treated with quercetin or silymarin for 28 days. The protective effect of quercetin was assessed using liver function tests and liver histology. Nuclear factor erythroid 2-related factor 2 (NRF2) and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathways were explored to elucidate the mechanism of action. Quercetin co-administration prevented the elevation of alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP) and bilirubin compared with isoniazid and rifampicin treatment alone. In the histological analysis, we observed that quercetin prophylaxis lessened the severity of hepatic necrosis and inflammation compared with the anti-TB drug-treated group. Quercetin attenuated anti-TB drug-induced oxidative stress by increasing NRF2 activation and expression, boosting endogenous antioxidant levels. Additionally, quercetin blocked inflammatory mediators high mobility group box-1 (HMGB-1) and interferon γ (IFN-γ), inhibiting activation of the NF-κB/ toll like receptor 4 (TLR-4) axis. Quercetin protects against anti-TB liver injury by activating NRF2 and blocking NF-κB/TLR-4.

<h4>Background</h4>Despite the high number of researches on pancreatic adenocarcinoma (PAAD) over past decades, little progress had been made due to lack of effective treatment regimens. We aimed to investigate the expression level, mutation, and clinical significance of the Frizzled (FZD) family in PAAD so as to establish a sufficient scientific evidence for clinical decisions and risk management.<h4>Methods</h4>PAAD samples were extracted from The Cancer Genome Atlas (TCGA). Oncomine, Gene expression profiling interactive analysis (GEPIA), human protein atlas (HPA), Kaplan-Meier Plotter, cBioPortal, LinkedOmics, DAVID database, and R software (× 64 3.6.2) were used to comprehensively analyze the roles of FZDs. p value below to 0.05 was considered as significant difference.<h4>Results</h4>In total, 179 PAAD tissues and 171 paracancerous tissues were included. The expression levels of FZD1, 2, 6, 7, and 8 were higher in PAAD tissues than those in normal pancreatic tissue. The higher the expression levels of FZD2 and FZD7, the higher the clinical stage. The overall survival (OS) time was significantly different between low FZD3, 4, 5, 6, and 9 expression group and high expression group. Multivariable analysis showed that FZD3 and FZD6 were independent prognostic factors. The recurrence free survival (RFS) time was significantly different between low FZD4 and FZD8 expression group and high expression group. The RFS difference between low FZD6 expression group and high expression group had not reached statistical significance (p = 0.067), which might be due to the small sample size. However, multivariable analysis showed that FZD6 was the only independent factor for RFS. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed that FZDs played a critical role in the Wnt signaling pathway, which was further confirmation that FZDs were transmembrane receptors of Wnt signaling pathway.<h4>Conclusions</h4>Our results strongly indicated a crucial role of the FZD family in PAAD. FZD3 and FZD6 could be potential prognostic and predictive markers, and FZD6 might also function as a potential therapeutic target in PAAD by blocking Wnt/β-catenin pathway.

Colorectal cancer is the third leading cause of cancer-related death in the United States. About 15% of colorectal cancers are associated with microsatellite instability (MSI) due to loss of function in the DNA mismatch repair pathway. This subgroup of patients has better survival rates and is more sensitive to immunotherapy. However, it remains unclear whether microsatellite stable (MSS) patients with colorectal cancer can be further stratified into subgroups with differential clinical characteristics. In this study, we analyzed The Cancer Genome Atlas data and found that Chr20q amplification is the most frequent copy number alteration that occurs specifically in colon (46%) and rectum (61%) cancer and is mutually exclusive with MSI. Importantly, MSS patients with Chr20q amplification (MSS-A) were associated with better recurrence-free survival compared with MSS patients without Chr20q amplification (MSS-N; <i>P</i> = 0.03). MSS-A tumors were associated with high level of chromosome instability and low immune infiltrations. In addition, MSS-A and MSS-N tumors were associated with somatic mutations in different driver genes, with high frequencies of mutated <i>TP53</i> in MSS-A and mutated <i>KRAS</i> and <i>BRAF</i> in MSS-N. Our results suggest that MSS-A and MSS-N represent two subtypes of MSS colorectal cancer, and such stratification may be used to improve therapeutic treatment in an individualized manner. SIGNIFICANCE: This study shows that chromosome 20q amplification occurs predominately in microsatellite-stable colorectal cancer and defines a distinct subtype with good prognosis, high chromosomal instability, distinct mutation profiles, and low immune infiltrations.

Induction of host cell autophagy by starvation was shown to enhance lysosomal delivery to mycobacterial phagosomes, resulting in the restriction of Mycobacterium tuberculosis reference strain H37Rv. Our previous study showed that strains belonging to M. tuberculosis Beijing genotype resisted starvation-induced autophagic elimination but the factors involved remained unclear. Here, we conducted RNA-Seq of macrophages infected with the autophagy-resistant Beijing strain (BJN) compared to macrophages infected with H37Rv upon autophagy induction by starvation. Results identified several genes uniquely upregulated in BJN-infected macrophages but not in H37Rv-infected cells, including those encoding Kxd1 and Plekhm2, which function in lysosome positioning towards the cell periphery. Unlike H37Rv, BJN suppressed enhanced lysosome positioning towards the perinuclear region and lysosomal delivery to its phagosome upon autophagy induction by starvation, while depletion of Kxd1 and Plekhm2 reverted such effects, resulting in restriction of BJN intracellular survival upon autophagy induction by starvation. Taken together, these data indicated that Kxd1 and Plekhm2 are important for the BJN strain to suppress lysosome positioning towards the perinuclear region and lysosomal delivery into its phagosome during autophagy induction by starvation to evade starvation-induced autophagic restriction.

Obesity is a major risk factor for cardiometabolic diseases. Nevertheless, a substantial proportion of individuals with obesity do not suffer cardiometabolic comorbidities. The mechanisms that uncouple adiposity from its cardiometabolic complications are not fully understood. Here, we identify 62 loci of which the same allele is significantly associated with both higher adiposity and lower cardiometabolic risk. Functional analyses show that the 62 loci are enriched for genes expressed in adipose tissue, and for regulatory variants that influence nearby genes that affect adipocyte differentiation. Genes prioritized in each locus support a key role of fat distribution (FAM13A, IRS1 and PPARG) and adipocyte function (ALDH2, CCDC92, DNAH10, ESR1, FAM13A, MTOR, PIK3R1 and VEGFB). Several additional mechanisms are involved as well, such as insulin-glucose signalling (ADCY5, ARAP1, CREBBP, FAM13A, MTOR, PEPD, RAC1 and SH2B3), energy expenditure and fatty acid oxidation (IGF2BP2), browning of white adipose tissue (CSK, VEGFA, VEGFB and SLC22A3) and inflammation (SH2B3, DAGLB and ADCY9). Some of these genes may represent therapeutic targets to reduce cardiometabolic risk linked to excess adiposity.

Chromatin conformation capture (3C) methods and fluorescent in situ hybridization (FISH) microscopy have been used to investigate the spatial organization of the genome. Although powerful, both techniques have limitations. Hi-C is challenging for low cell numbers and requires very deep sequencing to achieve its high resolution. In contrast, FISH can be done on small cell numbers and capture rare cell populations, but typically targets pairs of loci at a lower resolution. Here we detail a protocol for optical reconstruction of chromatin architecture (ORCA), a microscopy approach to trace the 3D DNA path within the nuclei of fixed tissues and cultured cells with a genomic resolution as fine as 2 kb and a throughput of ~10,000 cells per experiment. ORCA can identify structural features with comparable resolution to Hi-C while providing single-cell resolution and multimodal measurements characteristic of microscopy. We describe how to use this DNA labeling in parallel with multiplexed labeling of dozens of RNAs to relate chromatin structure and gene expression in the same cells. Oligopaint probe design, primary probe making, sample collection, cryosectioning and RNA/DNA primary probe hybridization can be completed in 1.5 weeks, while automated RNA/DNA barcode hybridization and RNA/DNA imaging typically takes 2-6 d for data collection and 2-7 d for the automated steps of image analysis.

Human stem-cell-derived models provide the promise of accelerating our understanding of brain disorders, but not knowing whether they possess the ability to mature beyond mid- to late-fetal stages potentially limits their utility. We leveraged a directed differentiation protocol to comprehensively assess maturation in vitro. Based on genome-wide analysis of the epigenetic clock and transcriptomics, as well as RNA editing, we observe that three-dimensional human cortical organoids reach postnatal stages between 250 and 300 days, a timeline paralleling in vivo development. We demonstrate the presence of several known developmental milestones, including switches in the histone deacetylase complex and NMDA receptor subunits, which we confirm at the protein and physiological levels. These results suggest that important components of an intrinsic in vivo developmental program persist in vitro. We further map neurodevelopmental and neurodegenerative disease risk genes onto in vitro gene expression trajectories to provide a resource and webtool (Gene Expression in Cortical Organoids, GECO) to guide disease modeling.

Despite a growing number of investigative studies on pancreatic fat deposition, there remains no clear indication regarding the clinical relevance of fat infiltration in the pancreas, also called fatty pancreas (FP). An individual's body weight is correlated with their pancreatic weight. Moreover, lipid infiltration causes disorders that compromise not only morphology but also metabolic functions. Fat infiltration leads to insulin resistance, type II diabetes mellitus, and pancreatic cancer; however, knowledge about pancreatic fat content and aspects related to the clinical profile remains unclear in the literature. The present review describes the current knowledge of FP, including its pathophysiology and clinical implications, as well as lifestyle changes in FP.

Fatty liver disease is a frequent diagnosis. Rarely, it adopts a multifocal nodular pattern mimicking multiple liver metastases. Multifocal nodular fatty infiltration of the liver entails a challenging problem that must be included as a differential diagnosis when dealing with healthy patients with an incidental finding of multiple liver lesions, even in the absence of obesity or metabolic syndrome. A complete clinical examination and high-quality imaging, including magnetic resonance imaging, might help to confirm diagnosis and to avoid unnecessary liver biopsies.

Neurodegenerative diseases resulting from the progressive loss of structure and/or function of neurons contribute to different paralysis degrees and loss of cognition and sensation. The lack of successful curative therapies for neurodegenerative disorders leads to a considerable burden on society and a high economic impact. Over the past 20 years, regenerative cell therapy, also known as stem cell therapy, has provided an excellent opportunity to investigate potentially powerful innovative strategies for treating neurodegenerative diseases. This is due to stem cells' capability to repair injured neuronal tissue by replacing the damaged or lost cells with differentiated cells, providing a conducive environment that is in favor of regeneration, or protecting the existing healthy neurons and glial cells from further damage. Thus, in this review, the various types of stem cells, the current knowledge of stem-cell-based therapies in neurodegenerative diseases, and the recent advances in this field are summarized. Indeed, a better understanding and further studies of stem cell technologies cause progress into realistic and efficacious treatments of neurodegenerative disorders.

Green synthetic protocol refers to the development of processes for the sustainable production of chemicals and materials. For the synthesis of various biologically active compounds, energy-efficient and environmentally benign processes are applied, such as microwave irradiation technology, ultrasound-mediated synthesis, photo-catalysis (ultraviolet, visible and infrared irradiation), molecular sieving, grinding and milling techniques, etc. Thesemethods are considered sustainable technology and become valuable green protocol to synthesize new drug molecules as theyprovidenumerous benefits over conventional synthetic methods.Based on this concept, oxadiazole derivatives are synthesized under microwave irradiation technique to reduce the formation of byproduct so that the product yield can be increased quantitatively in less reaction time. Hence, the synthesis of drug molecules under microwave irradiation follows a green chemistry approach that employs a set of principles to minimize or remove the utilization and production of hazardous toxic materials during the design, manufacture and application of chemical substances.This approach plays a major role in controlling environmental pollution by utilizing safer solvents, catalysts, suitable reaction conditions and thereby increases the atom economy and energy efficiency. Oxadiazole is a five-membered heterocyclic compound that possesses one oxygen and two nitrogen atoms in the ring system.Oxadiazole moiety is drawing considerable interest for the development of new drug candidates with potential therapeutic activities including antibacterial, antifungal, antiviral, anticonvulsant, anticancer, antimalarial, antitubercular, anti-asthmatic, antidepressant, antidiabetic, antioxidant, antiparkinsonian, analgesic and antiinflammatory, etc. This review focuses on different synthetic approaches of oxadiazole derivatives under microwave heating method and study of their various biological activities.

Attention-deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder with poorly understood molecular mechanisms that results in significant impairment in children. In this study, we sought to assess the role of rare recurrent variants in non-European populations and outside of coding regions. We generated whole genome sequence (WGS) data on 875 individuals, including 205 ADHD cases and 670 non-ADHD controls. The cases included 116 African Americans (AA) and 89 European Americans (EA), and the controls included 408 AA and 262 EA. Multiple novel rare recurrent variants were identified in exonic regions, functionally classified as stop-gains and frameshifts for known ADHD genes. Deletion in introns of the protocadherins families and the ncRNA <i>HGB8P</i> were identified in two independent EA ADHD patients. A meta-analysis of the two ethnicities for differential ADHD recurrent variants compared to controls shows a small number of overlaps. These results suggest that rare recurrent variants in noncoding regions may be involved in the pathogenesis of ADHD in children of both AA and EA ancestry; thus, WGS could be a powerful discovery tool for studying the molecular mechanisms of ADHD.

A complicated clinical course for critically ill patients admitted to the intensive care unit (ICU) usually includes multiorgan dysfunction and subsequent death. Owing to the heterogeneity, complexity, and unpredictability of the disease progression, ICU patient care is challenging. Identifying the predictors of complicated courses and subsequent mortality at the early stages of the disease and recognizing the trajectory of the disease from the vast array of longitudinal quantitative clinical data is difficult. Therefore, we attempted to perform a meta-analysis of previously published gene expression datasets to identify novel early biomarkers and train the artificial intelligence systems to recognize the disease trajectories and subsequent clinical outcomes. Using the gene expression profile of peripheral blood cells obtained within 24 h of pediatric ICU (PICU) admission and numerous clinical data from 228 septic patients from pediatric ICU, we identified 20 differentially expressed genes predictive of complicated course outcomes and developed a new machine learning model. After 5-fold cross-validation with 10 iterations, the overall mean area under the curve reached 0.82. Using a subset of the same set of genes, we further achieved an overall area under the curve of 0.72, 0.96, 0.83, and 0.82, respectively, on four independent external validation sets. This model was highly effective in identifying the clinical trajectories of the patients and mortality. Artificial intelligence systems identified eight out of twenty novel genetic markers (<i>SDC4, CLEC5A, TCN1, MS4A3, HCAR3, OLAH, PLCB1</i>, and <i>NLRP1</i>) that help predict sepsis severity or mortality. While these genes have been previously associated with sepsis mortality, in this work, we show that these genes are also implicated in complex disease courses, even among survivors. The discovery of eight novel genetic biomarkers related to the overactive innate immune system, including neutrophil function, and a new predictive machine learning method provides options to effectively recognize sepsis trajectories, modify real-time treatment options, improve prognosis, and patient survival.

An important manifestation of severe COVID-19 is the ARDS-like lung injury that is associated with vascular endothelialitis, thrombosis, and angiogenesis. The intravascular innate immune system (IIIS), including the complement, contact, coagulation, and fibrinolysis systems, which is crucial for recognizing and eliminating microorganisms and debris in the body, is likely to be involved in the pathogenesis of COVID-19 ARDS. Biomarkers for IIIS activation were studied in the first 66 patients with COVID-19 admitted to the ICU in Uppsala University Hospital, both cross-sectionally on day 1 and in 19 patients longitudinally for up to a month, in a prospective study. IIIS analyses were compared with biochemical parameters and clinical outcome and survival. Blood cascade systems activation leading to an overreactive conjunct thromboinflammation was demonstrated, reflected in consumption of individual cascade system components, e.g., FXII, prekallikrein, and high molecular weight kininogen and in increased levels of activation products, e.g., C4d, C3a, C3d,g, sC5b-9, TAT, and D-dimer. Strong associations were found between the blood cascade systems and organ damage, illness severity scores, and survival. We show that critically ill COVID-19 patients display a conjunct activation of the IIIS that is linked to organ damage of the lung, heart, kidneys, and death. We present evidence that the complement and in particular the kallikrein/kinin system is strongly activated and that both systems are prognostic markers of the outcome of the patients suggesting their role in driving the inflammation. Already licensed kallikrein/kinin inhibitors are potential drugs for treatment of critically ill patients with COVID-19.

Anthocyanins are the primary pigments contributing to the variety of flower colors among angiosperms and are considered essential for survival and reproduction. Anthocyanins are members of the flavonoids, a broader class of secondary metabolites, of which there are numerous structural genes and regulators thereof. In western European populations of <i>Lysimachia arvensis</i>, there are blue- and orange-petaled individuals. The proportion of blue-flowered plants increases with temperature and daylength yet decreases with precipitation. Here, we performed a transcriptome analysis to characterize the coding sequences of a large group of flavonoid biosynthetic genes, examine their expression and compare our results to flavonoid biochemical analysis for blue and orange petals. Among a set of 140 structural and regulatory genes broadly representing the flavonoid biosynthetic pathway, we found 39 genes with significant differential expression including some that have previously been reported to be involved in similar flower color transitions. In particular, <i>F3'5'H</i> and <i>DFR</i>, two genes at a critical branchpoint in the ABP for determining flower color, showed differential expression. The expression results were complemented by careful examination of the SNPs that differentiate the two color types for these two critical genes. The decreased expression of <i>F3'5'H</i> in orange petals and differential expression of two distinct copies of <i>DFR</i>, which also exhibit amino acid changes in the color-determining substrate specificity region, strongly correlate with the blue to orange transition. Our biochemical analysis was consistent with the transcriptome data indicating that the shift from blue to orange petals is caused by a change from primarily malvidin to largely pelargonidin forms of anthocyanins. Overall, we have identified several flavonoid biosynthetic pathway loci likely involved in the shift in flower color in <i>L. arvensis</i> and even more loci that may represent the complex network of genetic and physiological consequences of this flower color polymorphism.

The SALL2 transcription factor, an evolutionarily conserved gene through vertebrates, is involved in normal development and neuronal differentiation. In disease, SALL2 is associated with eye, kidney, and brain disorders, but mainly is related to cancer. Some studies support a tumor suppressor role and others an oncogenic role for SALL2, which seems to depend on the cancer type. An additional consideration is tissue-dependent expression of different SALL2 isoforms. Human and mouse <i>SALL2</i> gene loci contain two promoters, each controlling the expression of a different protein isoform (E1 and E1A). Also, several improvements on the human genome assembly and gene annotation through next-generation sequencing technologies reveal correction and annotation of additional isoforms, obscuring dissection of SALL2 isoform-specific transcriptional targets and functions. We here integrated current data of normal/tumor gene expression databases along with ChIP-seq binding profiles to analyze SALL2 isoforms expression distribution and infer isoform-specific SALL2 targets. We found that the canonical SALL2 E1 isoform is one of the lowest expressed, while the E1A isoform is highly predominant across cell types. To dissect SALL2 isoform-specific targets, we analyzed publicly available ChIP-seq data from Glioblastoma tumor-propagating cells and in-house ChIP-seq datasets performed in SALL2 wild-type and E1A isoform knockout HEK293 cells. Another available ChIP-seq data in HEK293 cells (ENCODE Consortium Phase III) overexpressing a non-canonical SALL2 isoform (short_E1A) was also analyzed. Regardless of cell type, our analysis indicates that the SALL2 long E1 and E1A isoforms, but not short_E1A, are mostly contributing to transcriptional control, and reveals a highly conserved network of brain-specific transcription factors (i.e., SALL3, POU3F2, and NPAS3). Our data integration identified a conserved molecular network in which SALL2 regulates genes associated with neural function, cell differentiation, development, and cell adhesion between others. Also, we identified <i>PODXL</i> as a gene that is likely regulated by SALL2 across tissues. Our study encourages the validation of publicly available ChIP-seq datasets to assess a specific gene/isoform's transcriptional targets. The knowledge of SALL2 isoforms expression and function in different tissue contexts is relevant to understanding its role in disease.

The small intestinal epithelium is the primary route of infection for many protozoan parasites. Understanding the mechanisms of infection, however, has been hindered due to the lack of appropriate models that recapitulate the complexity of the intestinal epithelium. Here, we describe an <i>in vitro</i> platform using stem cell-derived intestinal organoids established for four species that are important hosts of Apicomplexa and other protozoa in a zoonotic context: human, mouse, pig and chicken. The focus was set to create organoid-derived monolayers (ODMs) using the transwell system amenable for infection studies, and we provide straightforward guidelines for their generation and differentiation from organ-derived intestinal crypts. To this end, we reduced medium variations to an absolute minimum, allowing generation and differentiation of three-dimensional organoids for all four species and the subsequent generation of ODMs. Quantitative RT-PCR, immunolabeling with antibodies against marker proteins as well as transepithelial-electrical resistance (TEER) measurements were used to characterize ODM's integrity and functional state. These experiments show an overall uniform generation of monolayers suitable for <i>Toxoplasma gondii</i> infection, although robustness in terms of generation of stable TEER levels and cell differentiation status varies from species to species. Murine duodenal ODMs were then infected with <i>T. gondii</i> and/or <i>Giardia duodenalis</i>, two parasites that temporarily co-inhabit the intestinal niche but have not been studied previously in cellular co-infection models. <i>T. gondii</i> alone did not alter TEER values, integrity and transcriptional abundance of tight junction components. In contrast, in <i>G. duodenalis</i>-infected ODMs all these parameters were altered and <i>T. gondii</i> had no apparent influence on the <i>G. duodenalis</i>-triggered phenotype. In conclusion, we provide robust protocols for the generation, differentiation and characterization of intestinal organoids and ODMs from four species. We show their applications for comparative studies on parasite-host interactions during the early phase of a <i>T. gondii</i> infection but also its use for co-infections with other relevant intestinal protozoans.

SIN3A is the central scaffold protein of the SIN3/histone deacetylase (HDAC) transcriptional repressor complex. SIN3A participates in the mouse preimplantation development by fine-tuning <i>HDAC1</i> expression. However, it remains unresolved if this functional significance of SIN3A was conserved in other mammals. Herein, RNA-seq results show a large amount of <i>SIN3A</i> mRNA is present in oocytes and early embryos prior to embryonic genome activation and a low amount thereafter, suggesting a maternal origin of SIN3A in pigs, cattle, mice, and humans. Interestingly, immunofluorescence data show that SIN3A protein level peaks at four-cell stage in pigs compared with morula stage in cattle. SIN3A depletion in early embryos causes a developmental arrest at two-cell stage in pigs but does not affect bovine early embryonic development. In contrast with mouse data, SIN3A depletion results in only a slight decrease and even no difference in HDAC1 expression in porcine and bovine early embryos, respectively. In addition, HDAC1 knockdown does not cause two-cell block but leads to a reduced blastocyst rate. By using unbiased RNA-seq approach, we found that <i>Cyclin B1</i> (<i>CCNB1</i>) transcript level is dramatically reduced. Moreover, CCNB1 knockdown results in a similar phenotype as SIN3A depletion. Injection of exogenous <i>CCNB1</i> mRNA into SIN3A-depleted embryos could partly rescue embryonic development to pass two-cell stage. In conclusion, our results indicate SIN3A plays an essential role in porcine early embryonic development, which probably involves the regulation of <i>CCNB1</i> expression.

Knowledge of the host response to the novel coronavirus SARS-CoV-2 remains limited, hindering the understanding of COVID-19 pathogenesis and the development of therapeutic strategies. During the course of a viral infection, host cells release exosomes and other extracellular vesicles carrying viral and host components that can modulate the immune response. The present study used a shotgun proteomic approach to map the host circulating exosomes' response to SARS-CoV-2 infection. We investigated how SARS-CoV-2 infection modulates exosome content, exosomes' involvement in disease progression, and the potential use of plasma exosomes as biomarkers of disease severity. A proteomic analysis of patient-derived exosomes identified several molecules involved in the immune response, inflammation, and activation of the coagulation and complement pathways, which are the main mechanisms of COVID-19-associated tissue damage and multiple organ dysfunctions. In addition, several potential biomarkers-such as fibrinogen, fibronectin, complement C1r subcomponent and serum amyloid P-component-were shown to have a diagnostic feature presenting an area under the curve (AUC) of almost 1. Proteins correlating with disease severity were also detected. Moreover, for the first time, we identified the presence of SARS-CoV-2 RNA in the exosomal cargo, which suggests that the virus might use the endocytosis route to spread infection. Our findings indicate circulating exosomes' significant contribution to several processes-such as inflammation, coagulation, and immunomodulation-during SARS-CoV-2 infection. The study's data are available via ProteomeXchange with the identifier PXD021144.

Isothiocyanates (ITCs) are well-known chemopreventive agents that have received significant interest across the nutrition and pharmaceutical industries owing to their anticancer properties, thus it is essential to increase the conversion of glucosinolate to ITCs by myrosinase to maximize their health benefits. In this paper, using broccoli seed meals as a raw material, we comparatively analyzed the outcomes of two extraction methods: (i) hydrolysis followed by extraction (HFE) and (ii) simultaneous hydrolysis and extraction (SHE) in terms of the ITC yield. The results revealed that the SHE method showed a relatively greater erucin production from broccoli seeds and greater antitumor and antioxidant activities. A similar phenomenon was found for the hydrolysates of crude myrosinase and crude glucosinolate separated from broccoli seeds. However, when the crude glucosinolates were hydrolyzed by purified broccoli myrosinase, or when pure glucoraphanin was hydrolyzed by crude myrosinase, no significant effects were noted on the types and yields of ITCs between the SHE and HFE methods.

Ovarian clear cell carcinoma (OCCC) is a special pathological type of epithelial ovarian carcinoma (EOC) and has a high prevalence in Asia without specific molecular subtype classification. Endometriosis is a recognized precancerous lesion that carries 3-fold increased risk of OCCC. Ovarian endometrioid carcinoma, which also originates from endometriosis, shares several features with OCCC, including platinum resistance and younger age at diagnosis. Patients with OCCC have about a 2.5 to 4 times greater risk of having a venous thromboembolism (VTE) compared with other EOC, and OCCC tends to metastasize through lymphatic vesicular and peritoneal spread as opposed to hematogenous metastasis. There is only mild elevation of the conventional biomarker CA125. Staging surgery or optimal cytoreduction combined with chemotherapy is a common therapeutic strategy for OCCC. However, platinum resistance commonly portends a poor prognosis, so novel treatments are urgently needed. Targeted therapy and immunotherapy are currently being studied, including PARP, EZH2, and ATR inhibitors combined with the synthetic lethality of ARID1A-dificiency, and MAPK/PI3K/HER2, VEGF/bFGF/PDGF, HNF1β, and PD-1/PD-L1 inhibitors. Advanced stage, suboptimal cytoreduction, platinum resistance, lymph node metastasis, and VTE are major prognostic predictors for OCCC. We focus on update pathogenesis, diagnostic methods and therapeutic approaches to provide future directions for clinical diagnosis and treatment of OCCC.

<b>Background:</b> Disrupted gene levels are intimately correlated with the occurrence and prognosis of gastric cancer (GC). As genes do not function in isolation, we set out to investigate the possible relationship among mRNA and non-coding RNAs (ncRNAs). <b>Materials and methods:</b> RNA sequencing from 406 cases of GC was acquired through the TCGA database. R packages were utilized to assess differential RNA expression. The competing endogenous RNA (ceRNA) network was predicted using miRcode, miRDB, mirTarBase, Target Scan and constructed by Cytoscape 3.6.1. GO enrichment analysis, KEGG pathway analysis, GSEA, and WGCNA were applied for pathway analysis. The expression of select candidate molecules was confirmed using western blot and RT-PCR in GC cells and tissues. CCK-8, EdU staining, and Transwell assays were conducted to assess the influence of candidate molecules on proliferation and invasion. The gain and loss-of-function were achieved by co-culture with sh-lncRNA, mimics and sh-mRNA. Luciferase reporters were created using the psiCHECK2 vector, and the relative luciferase activity was calculated. <b>Results:</b> Using data from TCGA, we determined differentially expressed RNAs and created a ceRNA regulatory network. Interestingly, we identified a regulatory complex surrounding ANGPT2. We detected that ANGPT2 was highly expressed in GC, which correlated with a worse prognosis. Our findings indicated that ANGPT2 encourages growth, invasion, and epithelial-mesenchymal transition (EMT) in GC. Importantly, miR-145 inhibits ANGPT2 and abrogates its effects. Furthermore, LINC00184, a ceRNA, blocks miR-145, thereby improving ANGPT2-mediated carcinogenesis. <b>Conclusions:</b> Our findings indicate that the LINC00184/miR-145/ANGPT2 pathway has a crucial function in the development of GC and can act as a possible biomarker and targets for GC therapy.

Diabetic gastroparesis (DG) exhibits delayed gastric emptying (GE) due to impaired gastric non-adrenergic, non-cholinergic (NANC) relaxation. These defects are due to loss or reduction of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) that causes reduced expression and/or dimerization of neuronal nitric oxide synthase alpha (nNOSα) gene expression and function. We investigated the effect of potent Nrf2 activators (cinnamaldehyde [CNM] & curcumin [CUR]) on GE in obesity-induced diabetic female mice. We fed adult female homozygous Nfe2l2^-/- (Nrf2 KO) and wild-type (WT) female mice with either a high-fat diet (HFD) or a normal diet (ND) for a period of 16 weeks. Groups of HFD mice were fed with CUR or CNM either at 6th or 10th week respectively. Our results demonstrate that supplementation of CNM or CUR restored impaired nitrergic relaxation and attenuated delayed GE in HFD fed mice. Supplementation of CNM or CUR normalized altered gastric antrum protein expression of (1) p-ERK/p-JNK/MAPK/p-GSK-3β, (2) BH₄ (Cofactor of nNOS) biosynthesis enzyme GCH-1 and the GSH/GSSG ratio, (3) nNOSα protein & dimerization and soluble guanylate cyclase (sGC), (4) AhR and ER expression, (5) inflammatory cytokines (TNF α, IL-1β, IL-6), (6)TLR-4, as well as (7) reduced oxidative stress markers in WT but not in Nrf2 KO obesity-induced chronic diabetic female mice. Immunoprecipitation experiments revealed an interaction between nNOS and Nrf2 proteins. Our results conclude that Nrf2 activation restores nitrergic-mediated gastric motility and GE by normalizing inflammation and oxidative stress induced by obesity-induced chronic diabetes.

Using the radicals generated during pH oscillations, a semibatch pH oscillator is used as the chemical fuel and engine to drive polymerization induced self-assembly (PISA) for the one-pot autonomous synthesis of functional giant vesicles. Vesicles with diameters ranging from sub-micron to ∼5 µm are generated. Radical formation is found to be switched ON/OFF and be autonomously controlled by the pH oscillator itself, inducing a periodic polymerization process. The mechanism underlying these complex processes is studied and compared to conventional (non-oscillatory) initiation by the same redox pair. The pH oscillations along with the continuous increase in salt concentration in the semibatch reactor make the self-assembled objects undergo morphological evolution. This process provides a self-regulated means for the synthesis of soft giant polymersomes and opens the door for new applications of pH oscillators in a variety of contexts, from the exploration of new geochemical scenarios for the origin of life and the autonomous emergence of the necessary free-energy and proton gradients, to the creation of active functional microreactors and programmable release of cargo molecules for pH-responsive materials.

Many types of elective ancillary tests may be required to support the cytopathologic interpretations. Most of these tests can be performed on cell-blocks of different cytology specimens. The cell-block sections can be used for almost any special stains including various <i>histochemistry stains</i> and for <i>special stains for different microorganisms</i> including fungi, Pneumocystis jirovecii (carinii), and various organisms including acid-fast organisms similar to the surgical biopsy specimens. Similarly, in addition to <i>immunochemistry</i>, different <i>molecular tests</i> can be performed on cell-blocks. Molecular tests broadly can be divided into two main types <i>Molecular genetic tests</i> and <i>Proteomics</i>.

Abstract Objective: Pregnant women may develop disseminated intravascular coagulation (DIC), possibly resulting in massive maternal haemorrhage and perinatal death. The Japan guideline recommends use of antithrombin Ⅲ (ATⅢ) for DIC in obstetrics; however, its effect remains uncertain. The present study therefore aimed to investigate the effect of ATⅢ for DIC patients in obstetrics, using a national inpatient database in Japan. <h4>Design:</h4> Nationwide observational study Setting: Japan Population: We used the Diagnosis Procedure Combination inpatient database to identify patients who delivered at hospital and were diagnosed with DIC from July 2010 to March 2018. <h4>Methods:</h4> Propensity score matching analyses were performed to compare in-hospital maternal mortality and hysterectomy during hospitalization between users and non-users of ATⅢ on the day of delivery. Main Outcome Measures: In-hospital mortality, hysterectomy Results: A total of 9,920 patients were enrolled, including 4,329 patients (44%) who used ATⅢ and 5,511 patients (56%) who did not use ATⅢ. One-to-one propensity score matching created 3290 pairs. In-hospital maternal mortality did not differ significantly between the propensity-matched groups (0.3% in the ATⅢ group vs. 0.5% in the control group; odds ratio, 0.73; 95% confidence interval, 0.35–1.54). Patients in the ATⅢ group, compared with those in the control group, had a significantly lower proportion of receiving hysterectomy during hospitalization (5.3% vs. 8.7%; difference, -2.9%; 95% confidence interval, -4.2 to -1.6%). <h4>Conclusions:</h4> The present study did not show mortality-reducing effect of ATIII for patients with DIC in obstetrics. ATⅢ may have clinical benefit in terms of reduction in receiving hysterectomy.

We aim to establish a comprehensive COVID-19 autoantigen atlas in order to understand autoimmune diseases caused by SARS-CoV-2 infection. Based on the unique affinity between dermatan sulfate and autoantigens, we identified 348 proteins from human lung A549 cells, of which 198 are known targets of autoantibodies. Comparison with current COVID data identified 291 proteins that are altered at protein or transcript level in SARS-CoV-2 infection, with 191 being known autoantigens. These known and putative autoantigens are significantly associated with viral replication and trafficking processes, including gene expression, ribonucleoprotein biogenesis, mRNA metabolism, translation, vesicle and vesicle-mediated transport, and apoptosis. They are also associated with cytoskeleton, platelet degranulation, IL-12 signaling, and smooth muscle contraction. Host proteins that interact with and that are perturbed by viral proteins are a major source of autoantigens. Orf3 induces the largest number of protein alterations, Orf9 affects the mitochondrial ribosome, and they and E, M, N, and Nsp proteins affect protein localization to membrane, immune responses, and apoptosis. Phosphorylation and ubiquitination alterations by viral infection define major molecular changes in autoantigen origination. This study provides a large list of autoantigens as well as new targets for future investigation, e.g., UBA1, UCHL1, USP7, CDK11A, PRKDC, PLD3, PSAT1, RAB1A, SLC2A1, platelet activating factor acetylhydrolase, and mitochondrial ribosomal proteins. This study illustrates how viral infection can modify host cellular proteins extensively, yield diverse autoantigens, and trigger a myriad of autoimmune sequelae.

<title>Abstract</title> <p><bold>Background:</bold> Gut ecosystem has profound effects on host physiology and health. Gastrointestinal (GI) symptoms were frequently observed in patients with COVID-19. Compared with other organs, gut antiviral response can result in more complicated immune responses because of the interactions between the gut microbiota and host immunity. However, there are still large knowledge gaps in the impact of COVID-19 on gut molecular profiles and commensal microbiome, hindering our comprehensive understanding of the pathogenesis of SARS-CoV-2 and the treatment of COVID-19.<bold>Results: </bold>We performed longitudinal stool multi-omics profiling to systemically investigate the molecular phenomics alterations of gut ecosystem in COVID-19. Gut proteomes of COVID-19 were characterized by disturbed immune, proteolysis and redox homeostasis. The expression and glycosylation of proteins involved in neutrophil degranulation and migration were suppressed, while those of proteases were upregulated. The variable domains of Ig heavy chains were downregulated and the overall glycosylation of IgA heavy chain constant regions, IgGFc-binding protein, and J chain were suppressed with glycan-specific variations. There was a reduction of beneficial gut bacteria and an enrichment of bacteria derived deleterious metabolites potentially associated with multiple types of diseases (such as ethyl glucuronide). The reduction of Ig heave chain variable domains may contribute to the increase of some <italic>Bacteroidetes </italic>species. Many bacteria ceramide lipids with a C17-sphingoid based were downregulated in COVID-19. In many cases, the gut phenome did not restore two months after symptom onset.<bold>Conclusions:</bold> Our study indicates widely disturbed gut molecular profiles which may play a role in the development of symptoms in COVID-19. Our findings also emphasis the need for ongoing investigation of the long-term gut molecular and microbial alterations during COVID-19 recovery process. Considering the gut ecosystem as a potential target could offer a valuable approach in managing the disease.</p>

AMPA receptors are tetrameric glutamate-gated ion channels that mediate a majority of fast excitatory neurotransmission in the brain. They exist as calcium-impermeable (CI-) and calcium-permeable (CP-) subtypes, the latter of which lacks the GluA2 subunit. CP-AMPARs display an array of distinctive biophysical and pharmacological properties that allow them to be functionally identified. This has revealed that they play crucial roles in diverse forms of central synaptic plasticity. Here we summarise the functional hallmarks of CP-AMPARs and describe how these are modified by the presence of auxiliary subunits that have emerged as pivotal regulators of AMPARs. A lasting change in the prevalence of GluA2-containing AMPARs, and hence in the fraction of CP-AMPARs, is a feature in many maladaptive forms of synaptic plasticity and neurological disorders. These include modifications of glutamatergic transmission induced by inflammatory pain, fear conditioning, cocaine exposure, and anoxia-induced damage in neurons and glia. Furthermore, defective RNA editing of GluA2 can cause altered expression of CP-AMPARs and is implicated in motor neuron damage (amyotrophic lateral sclerosis) and the proliferation of cells in malignant gliomas. A number of the players involved in CP-AMPAR regulation have been identified, providing useful insight into interventions that may prevent the aberrant CP-AMPAR expression. Furthermore, recent molecular and pharmacological developments, particularly the discovery of TARP subtype-selective drugs, offer the exciting potential to modify some of the harmful effects of increased CP-AMPAR prevalence in a brain region-specific manner.

Lumpy skin disease (LSD) is a transboundary, systemic, viral disease of cattle. The first outbreaks of LSD were reported in Lang Son Province of Vietnam (bordered to China), and an official document has been submitted to OIE on 1 November 2020. Here, we described first the genetic profiles of this pathogen based on four well-known marker regions. The LSD virus isolated in these first outbreaks was 100% identical to viruses isolated in China (2019) based on the p32 and RP030 genes. Additionally, it is very close to the virus isolated in Russia (2017) based on the p32, RP030, thymidine kinase and ORF103 genes (100%, 99.01%, 99.08% and 99.47% identities). This finding is new, and a success in LSD virus isolation using MDBK cells from first outbreaks is important for vaccine development to control and eradicate LSD in Vietnam.

<h4>Background</h4>Analyses of gut microbiome composition in livestock species have shown its potential to contribute to the regulation of complex phenotypes. However, little is known about the host genetic control over the gut microbial communities. In pigs, previous studies are based on classical "single-gene-single-trait" approaches and have evaluated the role of host genome controlling gut prokaryote and eukaryote communities separately.<h4>Results</h4>In order to determine the ability of the host genome to control the diversity and composition of microbial communities in healthy pigs, we undertook genome-wide association studies (GWAS) for 39 microbial phenotypes that included 2 diversity indexes, and the relative abundance of 31 bacterial and six commensal protist genera in 390 pigs genotyped for 70 K SNPs. The GWAS results were processed through a 3-step analytical pipeline comprised of (1) association weight matrix; (2) regulatory impact factor; and (3) partial correlation and information theory. The inferred gene regulatory network comprised 3561 genes (within a 5 kb distance from a relevant SNP-P < 0.05) and 738,913 connections (SNP-to-SNP co-associations). Our findings highlight the complexity and polygenic nature of the pig gut microbial ecosystem. Prominent within the network were 5 regulators, PRDM15, STAT1, ssc-mir-371, SOX9 and RUNX2 which gathered 942, 607, 588, 284 and 273 connections, respectively. PRDM15 modulates the transcription of upstream regulators of WNT and MAPK-ERK signaling to safeguard naive pluripotency and regulates the production of Th1- and Th2-type immune response. The signal transducer STAT1 has long been associated with immune processes and was recently identified as a potential regulator of vaccine response to porcine reproductive and respiratory syndrome. The list of regulators was enriched for immune-related pathways, and the list of predicted targets includes candidate genes previously reported as associated with microbiota profile in pigs, mice and human, such as SLIT3, SLC39A8, NOS1, IL1R2, DAB1, TOX3, SPP1, THSD7B, ELF2, PIANP, A2ML1, and IFNAR1. Moreover, we show the existence of host-genetic variants jointly associated with the relative abundance of butyrate producer bacteria and host performance.<h4>Conclusions</h4>Taken together, our results identified regulators, candidate genes, and mechanisms linked with microbiome modulation by the host. They further highlight the value of the proposed analytical pipeline to exploit pleiotropy and the crosstalk between bacteria and protists as significant contributors to host-microbiome interactions and identify genetic markers and candidate genes that can be incorporated in breeding program to improve host-performance and microbial traits. Video Abstract.

Metastatic spread in breast cancer patients is the major driver of cancer-related deaths. A unique subset of cells disseminated from pre-invasive or primary tumor lesions are recognized as the main seeds for metastatic outgrowth. Disseminated cancer cells (DCCs) can migrate to distant organs and settle in a dormant state for a prolonged period until they emerge to overt metastases. Understanding the biology of breast cancer cells dissemination, dormancy and reactivation to form overt metastases has become an important focus. In this review, we discuss the recent advancements of molecular pathways involving breast cancer cell dissemination, role of chemokine-chemokine receptor networks in DCCs migration, DCCs phenotypic heterogeneity and unique genes signatures in tumor dormancy, microenvironmental regulation and specific niches that favors DCCs homing and dormancy. In addition, we also discuss recent findings relating to the role of immune response on DCC dissemination and dormancy. With recent advances in the field of immunotherapy/targeted therapy and its beneficial effects in cancer treatment, this review will focus on their impact on DCCs, reversal of stemness, tumor dormancy and metastatic relapse.

<b>Objective:</b> The present study aimed to determine the prognostic value of HOXA cluster antisense RNA2 (HOXA-AS2) in acute myeloid leukemia (AML), and to explore its potential molecular mechanisms. We also screening of potential drugs targeting HOXA-AS2 in AML. <b>Methods:</b> The level 3 raw genome-wide RNA sequencing dataset of AML was download from The Cancer Genome Atlas (TCGA) Data Portal, and the potential molecular mechanisms and drugs prediction of HOXA-AS2 in AML were explored using multiple bioinformatics analysis approaches. <b>Results:</b> TCGA AML cohort dataset indicated that HOXA-AS2 was significantly up-regulated in AML bone marrow tissues, and high HOXA-AS2 expression was related to poor overall survival (log-rank <i>P</i>=0.0284, hazard ratio 1.640, 95% confidence interval 1.046-2.573). Functional enrichment of differentially expressed genes (DEGs) suggested that the difference in prognosis between AML patients with high- and low-HOXA-AS2 expression may be due to differences in biological processes and pathways, including cell adhesion, angiogenesis, mitogen-activated protein kinase, cell differentiation, and other biological processes, and phosphatidylinositol 3 kinase-protein kinase B and Wnt signaling pathways. We also screened out three potential HOXA-AS2-targeted therapeutic drugs for AML, megestrol, carmustine, and cefoxitin, based on these DEGs. Functional enrichment analysis of HOXA-AS2-co-expressed genes revealed that HOXA-AS2 may act a part in AML by regulating nuclear factor-κB transcription factor activity, DNA methylation, angiogenesis, apoptosis, cell migration, Toll-like receptor 4, and Wnt signaling pathways. <b>Conclusion:</b> Our findings suggest that HOXA-AS2 is up-regulated in the bone marrow in patients with AML, and may serve as a novel prognostic biomarker for AML.

Post-translational modifications (PTMs) within the first 17 amino acids (Nt17) of the Huntingtin protein (Htt) have been shown to inhibit the aggregation and attenuate the toxicity of mutant Htt proteins in vitro and in various models of Huntington’s disease. Our group’s previous studies suggested that the Nt17 PTM code is a combinatorial code that involves a complex interplay between different PTMs. Here, we expand on these studies by investigating the effect of methionine 8 oxidation (oxM8) and crosstalk between this PTM and either lysine 6 acetylation (AcK6) or threonine 3 phosphorylation (pT3) on the aggregation of mutant Httex1. We show that M8 oxidation delays but does not inhibit the aggregation and has no effect on the final morphologies of mutant Httex1 aggregates. This delay in aggregation kinetics could be attributed to the transient accumulation of oligomeric aggregates, which disappear upon the formation of Httex1 oxM8 fibrils. Interestingly, the presence of both oxM8 and AcK6 resulted in dramatic inhibition of Httex1 fibrillization, whereas the presence of oxM8 did not influence the aggregation inhibitory effect of pT3. To gain insight into the structural basis underlying these proteins’ aggregation properties, we investigated the impact of each PTM and the combination of these PTMs on the conformational properties of the Nt17 peptide by circular dichroism spectroscopy and molecular dynamics simulation. These studies show that M8 oxidation decreases the helicity of the Nt17 in the presence or absence of PTMs and provides novel insight into the structural basis underlying the effects of different PTMs on mutant Httex1 aggregation. PTMs that lower the mutant Httex1 aggregation rate (oxM8, AcK6/oxM8, pT3, pT3/oxM8, and phosphorylation at Serine 13) result in stabilization and increased population of a short N-terminal helix (first eight residues) in Nt17 or decreased abundance of other helical forms, including long helix and short C-terminal helix. PTMs that did not alter the aggregation of mutant Httex1 exhibit a similar distribution of helical conformation as the unmodified peptides. These results show that the relative abundance of N- vs. C-terminal helical conformations and long helices, rather than the overall helicity of Nt17, better explains the effect of different Nt17 PTMs on mutant Httex1; thus, explaining the lack of correlation between the effect of PTMs on the overall helicity of Nt17 and mutant Httex1 aggregation in vitro . Taken together, our results provide novel structural insight into the differential effects of single PTMs and crosstalk between different PTMs in regulating mutant Httex1 aggregation. <h4>TOC Figure</h4>

<h4>Purpose</h4>The expansion of CAG (glutamine; Q) trinucleotide repeats (TNRs) predominantly occurs through male lineage in Huntington's disease (HD). As a result, offspring will have larger CAG repeats compared to their fathers, which causes an earlier onset of the disease called genetic anticipation. This study aims to develop a novel in vitro model to replicate CAG repeat instability in early spermatogenesis and demonstrate the biological process of genetic anticipation by using the HD stem cell model for the first time.<h4>Methods</h4>HD rhesus monkey embryonic stem cells (rESCs) were cultured in vitro for an extended period. Male rESCs were used to derive spermatogenic cells in vitro with a 10-day differentiation. The assessment of CAG repeat instability was performed by GeneScan and curve fit analysis.<h4>Results</h4>Spermatogenic cells derived from rESCs exhibit progressive expansion of CAG repeats with high daily expansion rates compared to the extended culture of rESCs. The expansion of CAG repeats is cell type-specific and size-dependent.<h4>Conclusions</h4>Here, we report a novel stem cell model that replicates genome instability and CAG repeat expansion in in vitro derived HD monkey spermatogenic cells. The in vitro spermatogenic cell model opens a new opportunity for studying TNR instability and the underlying mechanism of genetic anticipation, not only in HD but also in other TNR diseases.

Linker histone H1 is an essential regulatory protein for many critical biological processes, such as eukaryotic chromatin packaging and gene expression. Mis-regulation of H1s is commonly observed in tumor cells, where the balance between different H1 subtypes has been shown to alter the cancer phenotype. Consisting of a rigid globular domain and two highly charged terminal domains, H1 can bind to multiple sites on a nucleosomal particle to alter chromatin hierarchical condensation levels. In particular, the disordered H1 amino- and carboxyl-terminal domains (NTD/CTD) are believed to enhance this binding affinity, but their detailed dynamics and functions remain unclear. In this work, we used a coarse-grained computational model, AWSEM-DNA, to simulate the H1.0b-nucleosome complex, namely chromatosome. Our results demonstrate that H1 disordered domains restrict the dynamics and conformation of both globular H1 and linker DNA arms, resulting in a more compact and rigid chromatosome particle. Furthermore, we identified regions of H1 disordered domains that are tightly tethered to DNA near the entry-exit site. Overall, our study elucidates at near-atomic resolution the way the disordered linker histone H1 modulates nucleosome's structural preferences and conformational dynamics.

Serial measurement of a large panel of protein biomarkers near the bedside could provide a promising pathway to transform the critical care of acutely ill patients. However, attaining the combination of high sensitivity and multiplexity with a short assay turnaround poses a formidable technological challenge. Here, the authors develop a rapid, accurate, and highly multiplexed microfluidic digital immunoassay by incorporating machine learning-based autonomous image analysis. The assay has achieved 12-plexed biomarker detection in sample volume <15 μL at concentrations < 5 pg/mL while only requiring a 5-min assay incubation, allowing for all processes from sampling to result to be completed within 40 min. The assay procedure applies both a spatial-spectral microfluidic encoding scheme and an image data analysis algorithm based on machine learning with a convolutional neural network (CNN) for pre-equilibrated single-molecule protein digital counting. This unique approach remarkably reduces errors facing the high-capacity multiplexing of digital immunoassay at low protein concentrations. Longitudinal data obtained for a panel of 12 serum cytokines in human patients receiving chimeric antigen receptor-T (CAR-T) cell therapy reveals the powerful biomarker profiling capability. The assay could also be deployed for near-real-time immune status monitoring of critically ill COVID-19 patients developing cytokine storm syndrome.

Palmitoylethanolamide (PEA) is an endogenous anti-inflammatory lipid mediator and a widely used nutraceutical. In this study, we designed, realized, and tested a drug-carrier conjugate between PEA (the active drug) and glucuronic acid (the carrier). The conjugate, named GLUPEA, was characterized for its capability of increasing PEA levels and exerting anti-inflammatory activity both in vitro and in vivo. GLUPEA treatment, compared to the same concentration of PEA, resulted in higher cellular amounts of PEA and the endocannabinoid 2-arachidonoyl glycerol (2-AG), and increased 2-AG-induced transient receptor potential vanilloid type 1 (TRPV1) channel desensitization to capsaicin. GLUPEA inhibited pro-inflammatory monocyte chemoattractant protein 2 (MCP-2) release from stimulated keratinocytes, and it was almost as efficacious as ultra-micronized PEA at reducing colitis in dinitrobenzene sulfonic acid (DNBS)-injected mice when using the same dose. GLUPEA is a novel pro-drug able to efficiently mimic the anti-inflammatory and endocannabinoid enhancing actions of PEA.

Prostate cancer (PCa) mortality remains a significant public health problem, as advanced disease has poor survivability due to the development of resistance in response to both standard and novel therapeutic interventions. Therapeutic resistance is a multifaceted problem involving the interplay of a number of biological mechanisms including genetic, signaling, and phenotypic alterations, compounded by the contributions of a tumor microenvironment that supports tumor growth, invasiveness, and metastasis. The androgen receptor (AR) is a primary regulator of prostate cell growth, response and maintenance, and the target of most standard PCa therapies designed to inhibit AR from interacting with androgens, its native ligands. As such, AR remains the main driver of therapeutic response in patients with metastatic castration-resistant prostate cancer (mCRPC). While androgen deprivation therapy (ADT), in combination with microtubule-targeting taxane chemotherapy, offers survival benefits in patients with mCRPC, therapeutic resistance invariably develops, leading to lethal disease. Understanding the mechanisms underlying resistance is critical to improving therapeutic outcomes and also to the development of biomarker signatures of predictive value. The interconversions between epithelial-to-mesenchymal transition (EMT) and mesenchymal-to-epithelial transition (MET) navigate the prostate tumor therapeutic response, and provide a novel targeting platform in overcoming therapeutic resistance. Both microRNA (miRNA)- and long non-coding RNA (lncRNA)-mediated mechanisms have been associated with epigenetic changes in prostate cancer. This review discusses the current evidence-based knowledge of the role of the phenotypic transitions and novel molecular determinants (non-coding RNAs) as contributors to the emergence of therapeutic resistance and metastasis and their integrated predictive value in prostate cancer progression to advanced disease.

As leiomyosarcoma patients are challenged by the development of metastatic disease, effective systemic therapies are the cornerstone of outcome. However, the overall activity of the currently available conventional systemic treatments and the prognosis of patients with advanced or metastatic disease are still poor, making the treatment of this patient group challenging. Therefore, in a joint effort together with patient networks and organizations, namely Sarcoma Patients EuroNet (SPAEN), the international network of sarcoma patients organizations, and the National LeioMyoSarcoma Foundation (NLMSF) in the United States, we aim to summarize state-of-the-art treatments for leiomyosarcoma patients in order to identify knowledge gaps and current unmet needs, thereby guiding the community to design innovative clinical trials and basic research and close these research gaps. This position paper arose from a leiomyosarcoma research meeting in October 2020 hosted by the NLMSF and SPAEN.

Varicella-zoster virus (VZV), a common and ubiquitous human-restricted pathogen, causes a primary infection (varicella or chickenpox) followed by establishment of latency in sensory ganglia. The virus can reactivate, causing herpes zoster (HZ, shingles) and leading to significant morbidity but rarely mortality, although in immunocompromised hosts, VZV can cause severe disseminated and occasionally fatal disease. We discuss VZV diseases and the decrease in their incidence due to the introduction of live-attenuated vaccines to prevent varicella or HZ. We also focus on acyclovir, valacyclovir, and famciclovir (FDA approved drugs to treat VZV infections), brivudine (used in some European countries) and amenamevir (a helicase-primase inhibitor, approved in Japan) that augur the beginning of a new era of anti-VZV therapy. Valnivudine hydrochloride (FV-100) and valomaciclovir stearate (in advanced stage of development) and several new molecules potentially good as anti-VZV candidates described during the last year are examined. We reflect on the role of antiviral agents in the treatment of VZV-associated diseases, as a large percentage of the at-risk population is not immunized, and on the limitations of currently FDA-approved anti-VZV drugs. Their low efficacy in controlling HZ pain and post-herpetic neuralgia development, and the need of multiple dosing regimens requiring daily dose adaptation for patients with renal failure urges the development of novel anti-VZV drugs.

Inactivation of thrombin by the endogenous inhibitor antithrombin (AT) is a central mechanism in the regulation of hemostasis. This makes hereditary AT deficiency, which is caused by SERPINC1 gene mutations, a major thrombophilic risk factor. Aim of this study was to assess to what extent AT mutations impair thrombin inhibition kinetics. The study population included 36 thrombophilic patients with 19 different mutations and mean AT levels of 65% in a thrombin-based functional assay, and 26 healthy controls. To assess thrombin inhibition kinetics, thrombin (3.94 mU/mL final concentration) was added to citrated plasma. Subsequently, endogenous thrombin inhibition was stopped by addition of the reversible thrombin inhibitor argatroban and the amount of argatroban-complexed thrombin quantified using an oligonucleotide-based enzyme capture assay. The plasma half-life of human thrombin was significantly longer in patients with AT mutations than in the controls (119.9 versus 55.9 s). Moreover, it was disproportionately prolonged when compared with preparations of wild type AT in plasma, in whom a comparable thrombin half-life of 120.8 s was reached at a distinctly lower AT level of 20%. These findings may help to better understand the increased thrombotic risk of SERPINC1 mutations with near normal AT plasma levels in functional assays.

Identified as the second leading cause of cancer-related deaths among American women after lung cancer, breast cancer of all types has been the focus of numerous research studies. Even though triple-negative breast cancer (TNBC) represents 15-20% of the number of breast cancer cases worldwide, its existing therapeutic options are fairly limited. Due to the pivotal role of the presence/absence of specific receptors to luminal A, luminal B, HER-2+, and TNBC in the molecular classification of breast cancer, the lack of these receptors has accounted for the aforementioned limitation. Thereupon, in an attempt to participate in the ongoing research endeavors to overcome such a limitation, the conducted study adopts a combination strategy as a therapeutic paradigm for TNBC, which has proven notable results with respect to both: improving patient outcomes and survivability rates. The study hinges upon an investigation of a promising NPs platform for CD44 mediated theranostic that can be combined with JAK/STAT inhibitors for the treatment of TNBC. The ability of momelotinib (MMB), which is a JAK/STAT inhibitor, to sensitize the TNBC to apoptosis inducer (CFM-4.16) has been evaluated in MDA-MB-231 and MDA-MB-468. MMB + CFM-4.16 combination with a combination index (CI) ≤0.5, has been selected for in vitro and in vivo studies. MMB has been combined with CD44 directed polymeric nanoparticles (PNPs) loaded with CFM-4.16, namely CD44-T-PNPs, which selectively delivered the payload to CD44 overexpressing TNBC with a significant decrease in cell viability associated with a high dose reduction index (DRI). The mechanism underlying their synergism is based on the simultaneous downregulation of P-STAT3 and the up-regulation of CARP-1, which has induced ROS-dependent apoptosis leading to caspase 3/7 elevation, cell shrinkage, DNA damage, and suppressed migration. CD44-T-PNPs showed a remarkable cellular internalization, demonstrated by uptake of a Rhodamine B dye in vitro and S0456 (NIR dye) in vivo. S0456 was conjugated to PNPs to form CD44-T-PNPs/S0456 that simultaneously delivered CFM-4.16 and S0456 parenterally with selective tumor targeting, prolonged circulation, minimized off-target distribution.

Mitochondrial dysfunction is associated with a wide range of chronic human disorders, including atherosclerosis and diabetes mellitus. Mitochondria are dynamic organelles that undergo constant turnover in living cells. Through the processes of mitochondrial fission and fusion, a functional population of mitochondria is maintained, that responds to the energy needs of the cell. Damaged or excessive mitochondria are degraded by mitophagy, a specialized type of autophagy. These processes are orchestrated by a number of proteins and genes, and are tightly regulated. When one or several of these processes are affected, it can lead to the accumulation of dysfunctional mitochondria, deficient energy production, increased oxidative stress and cell death-features that are described in many human disorders. While severe mitochondrial dysfunction is known to cause specific and mitochondrial disorders in humans, progressing damage of the mitochondria is also observed in a wide range of other chronic diseases, including cancer and atherosclerosis, and appears to play an important role in disease development. Therefore, correction of mitochondrial dynamics can help in developing new therapies for the treatment of these conditions. In this review, we summarize the recent knowledge on the processes of mitochondrial turnover and the proteins and genes involved in it. We provide a list of known mutations that affect mitochondrial function, and discuss the emerging therapeutic approaches.

No abstract available.

<b>Rationale:</b> Combinations of two or more therapeutic agents targeting different signaling pathways involved in tumor progression can have synergistic anticancer effects. However, combination chemotherapies are greatly limited by the different pharmacokinetics, tumor targeting, and cellular uptake capacities of the combined drugs. We have previously demonstrated the potential synergistic efficacy of paclitaxel (PTX) and the natural anti-angiogenic agent tetramethylpyrazine (TMP) for suppressing ovarian carcinoma growth. An efficient, facile, and smart nanosystem to deliver PTX and TMP simultaneously <i>in vivo</i> is greatly desired. <b>Methods:</b> We constructed a redox-sensitive nanosystem based on the amphiphilic PTX-ss-TMP conjugate, in which PTX and TMP are linked by a disulfide bond. We characterized the structure of the drug conjugate by ¹H NMR and LC-MS, and then prepared PTX-ss-TMP NPs by a one-step nanoprecipitation method. We investigated the redox sensitivity, tumor-targeting ability, anticancer efficacy, and anti-angiogenesis activity of PTX-ss-TMP NPs <i>in vitro</i> and <i>in vivo</i>. <b>Results:</b> The amphiphilic PTX-ss-TMP conjugate readily self-assembled into stable nanoparticles in aqueous solution with a low critical association concentration of 1.35 µg/mL, well-defined spherical structure, small particle size (152 nm), high drug loading, redox-responsive drug release, high biocompatibility, and high storage stability. In cancer cells pretreated with GSH-OEt, PTX-ss-TMP NPs exhibited higher cytotoxicity, apoptosis rate, and cell-cycle arrest than monotherapy or combination therapy with free drugs, which was attributed to their improved cellular uptake and rapid intracellular drug release. Additionally, PTX-ss-TMP NPs also had a stronger anti-angiogenesis effect in HUVECs than free drug, which was mediated by VEGFR2-involved downstream signals. Finally, PTX-ss-TMP NPs showed tumor-specific accumulation and excellent antitumor activity in A2780 xenograft mice compared with free drug. <b>Conclusions:</b> These <i>in vitro</i> and <i>in vivo</i> results provide clear evidence that this redox-responsive carrier-free nanosystem with intrinsic amphiphilicity has great potential for combination cancer chemotherapy.

Salmonella enterica and Escherichia coli are bacteria of concern to veterinary public health and poultry health. Our research aimed to determine the factors associated with S. enterica and E. coli in commercial broiler chicken barns during the rest period between flocks to identify the best methods of sanitation for bacterial load reduction. This involved collecting samples from September 2015 to July 2016 from the floors of 36 barns before sanitation (baseline) and at 2 time intervals after sanitation, followed by microbiological and molecular analysis. A priori variables of interest included sanitation procedure (dry cleaning, wet cleaning, disinfection), sampling point (baseline, 2 d after sanitation, 6 d after sanitation), and flooring type (concrete, wood). The odds of detecting S. enterica were higher on wooden floors that were wet-cleaned than on concrete floors that were dry-cleaned, lower in the winter and spring than in the fall, and lower when samples were collected 2 d and 6 d after sanitation than at baseline. For E. coli, the concentration was higher on wooden floors than on concrete floors and in the summer than in the fall, and it was lower in postsanitation samples from disinfected barns than in presanitation samples from dry-cleaned barns and in the winter than in the fall. Among E. coli isolates, factors associated with the presence of qacEΔ1, a gene associated with resistance to quaternary ammonium compounds, included sanitation procedure, flooring type, cycle length, and the number of times per yr the barn is disinfected. Our findings highlight the importance of cleaning after litter removal, although the sanitation procedure chosen might differ depending on which pathogen is present and causing disease issues; dry cleaning appears to be preferable for S. enterica control, especially in barns with wooden floors, whereas disinfection appears to be preferable for E. coli reduction.

<h4>Objective</h4>Arthropathy is a major clinical problem in patients with hemochromatosis, the most common genetic disorder of iron overload. The pathological features of hemochromatosis arthropathy (HA) are heterogeneous and its specific nature remains unknown. One important drawback is the lack of proper <i>in vitro</i> models. The aim of the present study was to set up a model to investigate the biological response of cartilage to iron exposure.<h4>Design</h4>Bovine articular cartilage explants were incubated with ferric citrate for up to 9 days. We evaluated chondrocyte viability, iron deposition, and biomarkers of cartilage degradation in the conditioned medium.<h4>Results</h4>Iron accumulated within chondrocytes, which was associated with programmed cell death through chondroptosis. Iron treatment increased the release of sulfated glycosaminoglycans (sGAG), a component of the extracellular matrix, into the medium (p=0.0189). This was dependent on the presence of viable chondrocytes and was associated with increased activity of matrix-degrading metalloproteinases (MMP) (pro/active MMP-9, p=0.0317; pro MMP-2, p=0.0092; active MMP-2, p=0.0288). Co-treatment with the broad MMP/aggrecanase inhibitor prinomastat reduced iron-mediated sGAG release (0.02 ​μM, p=0.0425; 2 ​μM, p=0.0014), confirming that iron induces sGAG release via the activation of catabolic enzymes. Notably, iron-treated cartilage continued to release an increased amount of sGAG into the medium for 6 days after termination of the ferric citrate treatment (p=0.0259).<h4>Conclusions</h4>Iron triggers the early stages of cartilage degeneration. Removal of iron exposure does not prevent further damage to the cartilage, thus providing a possible explanation why HA is not prevented after iron depletion by phlebotomy treatment.

The current article aims to summarize all possible spectrum of protein-protein interactions for thromboxane A synthase (CYP5A1) and prostacyclin synthase (CYP8A1). These enzymes metabolize the same substrate (prostaglandin H₂ ) and can participate in cardiovascular, inflammatory, immune processes, and apoptosis modulation, as well as significantly influence the risk of cancers. Binary protein-protein and multiprotein complexes are of great importance in enzyme-regulating and signal-transduction pathways. However, protein partners of CYP5A1 and CYP8A1 are not yet fully identified, although both synthases are considered as prospective drug targets. At least 36 novel protein partners of CYP5A1 and CYP8A1 were revealed from different tissue types using an approach based on affinity isolation and mass spectrometry. Enrichment analysis showed that these proteins have different molecular functions: folding (refolding), unfolded protein and chaperon binding, protein transport (export/import), posttranslational modification, protein domain-specific binding, antioxidant activity, and glutathione homeostasis. A significant part of them, belonging to molecular chaperones, were common partners for CYP5A1 and CYP8A1, while other proteins were unique with the tissue-dependent distribution. New aspects of CYP5A1 and CYP8A1 interactomics and hetero-complex formation with different protein partners, including cytochrome P450s are discussed.

The serotonin transporter (5-HTT) is a key molecule of serotoninergic neurotransmission and target of many anxiolytics and antidepressants. In humans, 5-HTT gene variants resulting in lower expression levels are associated with behavioral traits of anxiety. Furthermore, functional magnetic resonance imaging (fMRI) studies reported increased cerebral blood flow (CBF) during resting state (RS) and amygdala hyperreactivity. 5-HTT deficient mice as an established animal model for anxiety disorders seem to be well suited for investigating amygdala (re-)activity in an fMRI study. We investigated wildtype (5-HTT+/+), heterozygous (5-HTT+/-), and homozygous 5-HTT-knockout mice (5-HTT-/-) of both sexes in an ultra-high-field 17.6 Tesla magnetic resonance scanner. CBF was measured with continuous arterial spin labeling during RS, stimulation state (SS; with odor of rats as aversive stimulus), and post-stimulation state (PS). Subsequently, post mortem c-Fos immunohistochemistry elucidated neural activation on cellular level. The results showed that in reaction to the aversive odor CBF in total brain and amygdala of all mice significantly increased. In male 5-HTT+/+ mice amygdala RS CBF levels were found to be significantly lower than in 5-HTT+/- mice. From RS to SS 5-HTT+/+ amygdala perfusion significantly increased compared to both 5-HTT+/- and 5-HTT-/- mice. Perfusion level changes of male mice correlated with the density of c-Fos-immunoreactive cells in the amygdaloid nuclei. In female mice the perfusion was not modulated by the 5-Htt-genotype, but by estrous cycle stages. We conclude that amygdala reactivity is modulated by the 5-Htt genotype in males. In females, gonadal hormones have an impact which might have obscured genotype effects. Furthermore, our results demonstrate experimental support for the tonic model of 5-HTTLPR function.

Ferroptosis is a newly described form of regulated cell death triggered by oxidative stresses and characterized by extensive lipid peroxidation and membrane damages. The name of ferroptosis indicates that the ferroptotic death process depends on iron, but not other metals, as one of its canonical features. Here, we reported that zinc is also essential for ferroptosis in breast and renal cancer cells. Zinc chelator suppressed ferroptosis, and zinc addition promoted ferroptosis, even during iron chelation. By interrogating zinc-related genes in a genome-wide RNAi screen of ferroptosis, we identified SLC39A7, encoding ZIP7 that controls zinc transport from endoplasmic reticulum (ER) to cytosol, as a novel genetic determinant of ferroptosis. Genetic and chemical inhibition of the ZIP7 protected cells against ferroptosis, and the ferroptosis protection upon ZIP7 knockdown can be abolished by zinc supplementation. We found that the genetic and chemical inhibition of ZIP7 triggered ER stresses, including the induction of the expression of HERPUD1 and ATF3. Importantly, the knockdown of HERPUD1 abolished the ferroptosis protection phenotypes of ZIP7 inhibition. Together, we have uncovered an unexpected role of ZIP7 in ferroptosis by maintaining ER homeostasis. These findings may have therapeutic implications for human diseases involving ferroptosis and zinc dysregulations.

Somites arising from paraxial mesoderm are a hallmark of the segmented vertebrate body plan. They form sequentially during axis extension and generate musculoskeletal cell lineages. How paraxial mesoderm becomes regionalised along the axis and how this correlates with dynamic changes of chromatin accessibility and the transcriptome remains unknown. Here, we report a spatiotemporal series of ATAC-seq and RNA-seq along the chick embryonic axis. Footprint analysis shows differential coverage of binding sites for several key transcription factors, including CDX2, LEF1 and members of HOX clusters. Associating accessible chromatin with nearby expressed genes identifies cis-regulatory elements (CRE) for TCF15 and MEOX1. We determine their spatiotemporal activity and evolutionary conservation in Xenopus and human. Epigenome silencing of endogenous CREs disrupts TCF15 and MEOX1 gene expression and recapitulates phenotypic abnormalities of anterior-posterior axis extension. Our integrated approach allows dissection of paraxial mesoderm regulatory circuits in vivo and has implications for investigating gene regulatory networks.

Two large-scale Florida manatee (Trichechus manatus latirostris) mortality episodes were reported on separate coasts of Florida in 2013. The east coast mortality episode was associated with an unknown etiology in the Indian River Lagoon (IRL). The west coast mortality episode was attributed to a persistent Karenia brevis algal bloom or 'red tide' centered in Southwest Florida. Manatees from the IRL also had signs of cold stress. To investigate these two mortality episodes, two proteomic experiments were performed, using two-dimensional difference in gel electrophoresis (2D-DIGE) and isobaric tags for relative and absolute quantification (iTRAQ) LC-MS/MS. Manatees from the IRL displayed increased levels of several proteins in their serum samples compared to controls, including kininogen-1 isoform 1, alpha-1-microglobulin/bikunen precursor, histidine-rich glycoprotein, properdin, and complement C4-A isoform 1. In the red tide group, the following proteins were increased: ceruloplasmin, pyruvate kinase isozymes M1/M2 isoform 3, angiotensinogen, complement C4-A isoform 1, and complement C3. These proteins are associated with acute-phase response, amyloid formation and accumulation, copper and iron homeostasis, the complement cascade pathway, and other important cellular functions. The increased level of complement C4 protein observed in the red tide group was confirmed through the use of Western Blot.

Timely transition to flowering, maturity and plant height are important for agronomic adaptation and productivity of Indian mustard (B. juncea), which is a major edible oilseed crop of low input ecologies in Indian subcontinent. Breeding manipulation for these traits is difficult because of the involvement of multiple interacting genetic and environmental factors. Here, we report a genetic analysis of these traits using a population comprising 92 diverse genotypes of mustard. These genotypes were evaluated under deficient (N75), normal (N100) or excess (N125) conditions of nitrogen (N) application. Lower N availability induced early flowering and maturity in most genotypes, while high N conditions delayed both. A genotyping-by-sequencing approach helped to identify 406,888 SNP markers and undertake genome wide association studies (GWAS). 282 significant marker-trait associations (MTA's) were identified. We detected strong interactions between GWAS loci and nitrogen levels. Though some trait associated SNPs were detected repeatedly across fertility gradients, majority were identified under deficient or normal levels of N applications. Annotation of the genomic region (s) within ± 50 kb of the peak SNPs facilitated prediction of 30 candidate genes belonging to light perception, circadian, floral meristem identity, flowering regulation, gibberellic acid pathways and plant development. These included over one copy each of AGL24, AP1, FVE, FRI, GID1A and GNC. FLC and CO were predicted on chromosomes A02 and B08 respectively. CDF1, CO, FLC, AGL24, GNC and FAF2 appeared to influence the variation for plant height. Our findings may help in improving phenotypic plasticity of mustard across fertility gradients through marker-assisted breeding strategies.

<h4>Objective</h4>Cartilage in joints such as the hip and knee experiences repeated phases of heavy loading and low load recovery during the 24-h day/night cycle. Our previous work has shown 24 h rhythmic changes in gene expression at transcript level between night and day in wild type mouse cartilage which is lost in a circadian clock knock-out mouse model. However, it remains unknown to what extent circadian rhythms also regulate protein level gene expression in this matrix rich tissue.<h4>Methods</h4>We investigated daily changes of protein abundance in mouse femoral head articular cartilage by performing a 48-h time-series LC-MS/MS analysis.<h4>Results</h4>Out of the 1,177 proteins we identified across all time points, 145 proteins showed rhythmic changes in their abundance within the femoral head cartilage. Among these were molecules that have been implicated in key cartilage functions, including CTGF, MATN1, PAI-1 and PLOD1 & 2. Pathway analysis revealed that protein synthesis, cytoskeleton and glucose metabolism exhibited time-of-day dependent functions. Analysis of published cartilage proteomics datasets revealed that a significant portion of rhythmic proteins were dysregulated in osteoarthritis and/or ageing.<h4>Conclusions</h4>Our circadian proteomics study reveals that articular cartilage is a much more dynamic tissue than previously thought, with chondrocytes driving circadian rhythms not only in gene transcription but also in protein abundance. Our results clearly call for the consideration of circadian timing mechanisms not only in cartilage biology, but also in the pathogenesis, treatment strategies and biomarker detection in osteoarthritis.

<i>Background</i>: Setleis syndrome (SS) is a focal facial dermal dysplasia presenting with bilateral temporal skin lesions, eyelash abnormalities and absent meibomian glands. SS is a rare autosomal recessive disorder caused by mutations in the TWIST2 gene, which codes for a transcription factor of the bHLH family known to be involved in skin and facial development. <i>Methods</i>: We obtained gene expression profiles by microarray analyses from control and SS patient primary skin fibroblast and lymphoblastoid cell lines. <i>Results</i>: Out of 983 differentially regulated genes in fibroblasts (fold change ≥ 2.0), 479 were down-regulated and 509 were up-regulated, while in lymphoblasts, 1248 genes were down-regulated and 73 up-regulated. RT-PCR reactions confirmed altered expression of selected genes. <i>Conclusions</i>: TWIST2 is described as a repressor, but expression profiling suggests an important role in gene activation as well, as evidenced by the number of genes that are down-regulated, with a much higher proportion of down-regulated genes found in lymphoblastoid cells from an SS patient. As expected, both types of cell types showed dysregulation of cytokine genes. These results identify potential TWIST2 target genes in two important cell types relevant to rare disorders caused by mutations in this bHLH gene.

<b>Background and objectives</b>: Exposure to ionizing radiation (IR) has increased immensely over the past years, owing to diagnostic and therapeutic reasons. However, certain radiosensitive individuals show toxic enhanced reaction to IR, and it is necessary to specifically protect them from unwanted exposure. Although predicting radiosensitivity is the way forward in the field of personalised medicine, there is limited information on the potential biomarkers. The aim of this systematic review is to identify evidence from a range of literature in order to present the status quo of our knowledge of IR-induced changes in protein expression in normal tissues, which can be correlated to radiosensitivity. <b>Methods</b>: Studies were searched in NCBI Pubmed and in ISI Web of Science databases and field experts were consulted for relevant studies. Primary peer-reviewed studies in English language within the time-frame of 2011 to 2020 were considered. Human non-tumour tissues and human-derived non-tumour model systems that have been exposed to IR were considered if they reported changes in protein levels, which could be correlated to radiosensitivity. At least two reviewers screened the titles, keywords, and abstracts of the studies against the eligibility criteria at the first phase and full texts of potential studies at the second phase. Similarly, at least two reviewers manually extracted the data and accessed the risk of bias (National Toxicology Program/Office for Health Assessment and Translation-NTP/OHAT) for the included studies. Finally, the data were synthesised narratively in accordance to synthesis without meta analyses (SWiM) method. <b>Results</b>: In total, 28 studies were included in this review. Most of the records (16) demonstrated increased residual DNA damage in radiosensitive individuals compared to normo-sensitive individuals based on γH2AX and TP53BP1. Overall, 15 studies included proteins other than DNA repair foci, of which five proteins were selected, Vascular endothelial growth factor (VEGF), Caspase 3, p16^INK4A (Cyclin-dependent kinase inhibitor 2A, CDKN2A), Interleukin-6, and Interleukin-1β, that were connected to radiosensitivity in normal tissue and were reported at least in two independent studies. <b>Conclusions and implication of key findings</b>: A majority of studies used repair foci as a tool to predict radiosensitivity. However, its correlation to outcome parameters such as repair deficient cell lines and patients, as well as an association to moderate and severe clinical radiation reactions, still remain contradictory. When IR-induced proteins reported in at least two studies were considered, a protein network was discovered, which provides a direction for further studies to elucidate the mechanisms of radiosensitivity. Although the identification of only a few of the commonly reported proteins might raise a concern, this could be because (i) our eligibility criteria were strict and (ii) radiosensitivity is influenced by multiple factors. <b>Registration</b>: PROSPERO (CRD42020220064).

Inflammatory bowel diseases (IBD) are chronic conditions that primarily affect the gastrointestinal tract, with a complex pathogenesis; they are characterized by a significant heterogeneity of clinical presentations and of inflammatory pathways that sustain intestinal damage. After the introduction of the first biological therapies, the pipeline of therapies for IBD has been constantly expanding, and a significant number of new molecules is expected in the next few years. Evidence from clinical trials and real-life experiences has taught us that up to 40% of patients do not respond to a specific drug. Unfortunately, to date, clinicians lack a valid tool that can predict each patient's response to therapies and that could help them in choosing what drug to administer. Several candidate biomarkers have been investigated so far, with conflicting results: clinical, genetic, immunological, pharmacokinetic and microbial markers have been tested, but no ideal marker has been identified so far. Based on recent evidence, multiparametric models seemingly hold the greatest potential for predicting response to therapy. In this narrative review, we aim to summarize the current knowledge on predictors and early markers of response to biological therapies in IBD.

Genomic integrity is maintained by DNA repair and the DNA damage response (DDR). Defects in certain DNA repair genes give rise to many rare progressive neurodegenerative diseases (NDDs), such as ocular motor ataxia, Huntington disease (HD), and spinocerebellar ataxias (SCA). Dysregulation or dysfunction of DDR is also proposed to contribute to more common NDDs, such as Parkinson's disease (PD), Alzheimer's disease (AD), and Amyotrophic Lateral Sclerosis (ALS). Here, we present mechanisms that link DDR with neurodegeneration in rare NDDs caused by defects in the DDR and discuss the relevance for more common age-related neurodegenerative diseases. Moreover, we highlight recent insight into the crosstalk between the DDR and other cellular processes known to be disturbed during NDDs. We compare the strengths and limitations of established model systems to model human NDDs, ranging from <i>C. elegans</i> and mouse models towards advanced stem cell-based 3D models.

Triclocarban (TCC), a broad-spectrum lipophilic antimicrobial agent, is a diarylurea derivative that has been used for more than 60 years as a major ingredient of toys, clothing, food packaging materials, food industry floors, medical supplies and especially of personal care products, such as soaps, toothpaste and shampoo. In September 2016, the U.S. FDA banned nineteen antimicrobial ingredients, including TCC, in over-the-counter consumer antiseptic wash products, due to their toxicity. Withdrawal of TCC has prompted efforts to search for new antimicrobial compounds. In this paper, we present the synthesis and biological evaluation, as antibiotic and non-cytotoxic agents, of a series of diarylureas, analogues of TCC. These compounds are characterized by an intriguingly simple chemistry and can be easily synthesized. Among the synthesized compounds, <b>1ab</b> and <b>1bc</b> emerge as the most interesting compounds as they show the same activity of TCC (MIC = 16 µg/mL) against <i>Staphylococcus aureus,</i> and a higher activity than TCC against <i>Enterococcus faecalis</i> (MIC = 32 µg/mL versus MIC = 64 µg/mL). Moreover, <b>1ab</b> and <b>1bc</b> show no cytotoxicity towards the human mammary epithelial cells MCF-10A and embryonic kidney epithelial cells Hek-293, in opposition to TCC, which exhibits a marked cytotoxicity on the same cell lines and shows a good antitumor activity on a panel of cell lines tested.

This research is focused on a synthesis of copper-cellulose phosphates antimicrobial complexes. Vapor-phase phosphorylations of cellulose were achieved by exposing microcrystalline cellulose to phosphorus trichloride (PCl₃) vapors. The cellulose-<i>O</i>-dichlorophosphines (Cell-<i>O</i>-PCl₂) formed were hydrolyzed to cellulose-<i>O</i>-hydrogenphosphate (P(III)) (Cell-<i>O</i>-P(O)(H)(OH)), which, in turn, were converted into corresponding copper(II) complexes (Cell-<i>O</i>-P(O)(H)(OH)∙Cu²⁺). The analysis of the complexes Cell-<i>O</i>-P(O)(H)(OH)∙Cu²⁺ covered: scanning electron microscopy (SEM), attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), atomic absorption spectrometry with flame excitation (FAAS), and bioactivity tests against representative Gram-negative bacteria (<i>Escherichia coli</i>) and Gram-positive bacteria (<i>Staphylococcus aureus</i>). The antimicrobial tests of synthesized Cell-<i>O</i>-P(O)(H)(OH)∙Cu²⁺ revealed their potential applications as an antibacterial material.

Despite the improvements in life expectancy, neurodegenerative conditions have arguably become the most dreaded maladies of older people. The neuroprotective and anti-ageing potentials of essential oils (EOs) are widely evaluated around the globe. The objective of this review is to analyse the effectiveness of EOs as neuroprotective remedies among the four common age-related neurodegenerative diseases. The literature was extracted from three databases (PubMed, Web of Science and Google Scholar) between the years of 2010 to 2020 using the medical subject heading (MeSH) terms "essential oil", crossed with "Alzheimer's disease (AD)", "Huntington's disease (HD)", "Parkinson's disease (PD)" or "amyotrophic lateral sclerosis (ALS)". Eighty three percent (83%) of the studies were focused on AD, while another 12% focused on PD. No classifiable study was recorded on HD or ALS. EO from <i>Salvia officinalis</i> has been recorded as one of the most effective acetylcholinesterase and butyrylcholinesterase inhibitors. However, only <i>Cinnamomum</i> sp. has been assessed for its effectiveness in both AD and PD. Our review provided useful evidence on EOs as potential neuroprotective remedies for age-related neurodegenerative diseases.

The endoplasmic reticulum (ER) is an interconnected organelle that plays fundamental roles in the biosynthesis, folding, stabilization, maturation, and trafficking of secretory and transmembrane proteins. It is the largest organelle and critically modulates nearly all aspects of life. Therefore, in the endoplasmic reticulum, an enormous investment of resources, including chaperones and protein folding facilitators, is dedicated to adequate protein maturation and delivery to final destinations. Unfortunately, the folding and assembly of proteins can be quite error-prone, which leads to the generation of misfolded proteins. Notably, protein homeostasis, referred to as proteostasis, is constantly exposed to danger by flows of misfolded proteins and subsequent protein aggregates. To maintain proteostasis, the ER triages and eliminates terminally misfolded proteins by delivering substrates to the ubiquitin-proteasome system (UPS) or to the lysosome, which is termed ER-associated degradation (ERAD) or ER-phagy, respectively. ERAD not only eliminates misfolded or unassembled proteins via protein quality control but also fine-tunes correctly folded proteins via protein quantity control. Intriguingly, the diversity and distinctive nature of E3 ubiquitin ligases determine efficiency, complexity, and specificity of ubiquitination during ERAD. ER-phagy utilizes the core autophagy machinery and eliminates ERAD-resistant misfolded proteins. Here, we conceptually outline not only ubiquitination machinery but also catalytic mechanisms of E3 ubiquitin ligases. Further, we discuss the mechanistic insights into E3 ubiquitin ligases involved in the two guardian pathways in the ER, ERAD and ER-phagy. Finally, we provide the molecular mechanisms by which ERAD and ER-phagy conduct not only protein quality control but also protein quantity control to ensure proteostasis and subsequent organismal homeostasis.

Cell adhesion molecules and their extracellular ligands control morphogenetic events such as directed cell migration. The migration of neuroblasts and neural crest cells establishes the structure of the central and peripheral nervous systems. In <i>C. elegans</i>, the bilateral Q neuroblasts and their descendants undergo long-range migrations with left/right asymmetry. QR and its descendants on the right migrate anteriorly, and QL and its descendants on the left migrate posteriorly, despite identical patterns of cell division, cell death, and neuronal generation. The initial direction of protrusion of the Q cells relies on the left/right asymmetric functions of the transmembrane receptors UNC-40/DCC and PTP-3/LAR in the Q cells. Here, we show that Q cell left/right asymmetry of migration is independent of the GPA-16/Ga pathway which regulates other left/right asymmetries, including nervous system L/R asymmetry. No extracellular cue has been identified that guides initial Q anterior versus posterior migrations. We show that collagens DPY-17 and SQT-3 control initial Q direction of protrusion. Genetic interactions with UNC-40/DCC and PTP-3/LAR suggest that DPY-17 and SQT-3 drive posterior migration and might act with both receptors or in a parallel pathway. Analysis of mutants in other collagens and extracellular matrix components indicated that general perturbation of collagens and the extracellular matrix (ECM) did not result in directional defects, and that the effect of DPY-17 and SQT-3 on Q direction is specific. DPY-17 and SQT-3 are components of the cuticle, but a role in the basement membrane cannot be excluded. Possibly, DPY-17 and SQT-3 are part of a pattern in the cuticle and/or basement membrane that is oriented to the anterior-posterior axis of the animal and that is deciphered by the Q cells in a left-right asymmetric fashion. Alternatively, DPY-17 and SQT-3 might be involved in the production or stabilization of a guidance cue that directs Q migrations. In any case, these results describe a novel role for the DPY-17 and SQT-3 collagens in directing posterior Q neuroblast migration.

Serotonin (5-HT) is member of a family of indolamine molecules that participate in a wide variety of biological processes. Despite its important role in the regulation of local blood systems, little is known about the physiological function of 5-HT in reproductive organs, its functional implications, and its role in the reproduction of mammals. In the present work, we evaluated the localization and distribution of 5-HT (using histochemical analysis of indolamines) and different components of the serotoninergic system in rat testes. We detected local synthesis and degradation through immunofluorescence and western blot analyses against the TPH1, MAO_A, 5-HT_T, and VMAT1 serotonin transporters. We also identified the localization and distribution of the 5-HT_1B, 5-HT_2A, and 5-HT_3A receptors. RT-PCR results showed the presence of the Tph1, Maoa, Slc6a4, and Htr3a genes in testes and in the brain stem (Tph1 was used as a negative control). High-performance liquid chromatography was used to determine the presence of 5-HT and the activity of tryptophan hydroxylase in testes homogenates <i>in vitro</i>. Our observations suggest that TPH1 activity and local 5-HT synthesis befall in rat testes. We propose that 5-HT could participate in the regulation of testosterone synthesis and in the spermatogenesis process via local serotoninergic system. However, more studies are needed before concluding that rat testes, or those of other mammals, contain an active form of tryptophan hydroxylase and produce 5-HT.

Huntington's disease (HD) is a life-threatening neurodegenerative disorder. Altered levels and functions of the purinergic ionotropic P2X7 receptors (P2X7Rs) have been found in animal and cellular models of HD, suggesting their possible role in the pathogenesis of the disease; accordingly, the therapeutic potential of P2X7R antagonists in HD has been proposed. Here we further investigated the effects of P2X7R ligands in <i>in vitro</i> and <i>ex vivo</i> HD experimental models. In ST14A/Q120 rat striatal cells, we found a reduction of P2X7R expression; however, the P2X7R agonist 2'(3')-O-(4-benzoylbenzoyl)adenosine-5'-triphosphate (BzATP) induced cellular death, and this effect was fully reversed by the antagonist periodate-oxidized adenosine 5'-triphosphate (OxATP). Moreover, in corticostriatal slices from symptomatic R6/2 mice, BzATP reduced the synaptic transmission to a larger extent than in wild-type (WT) mice. Such an effect was accompanied by a concomitant increase of the paired-pulse ratio, suggesting a presynaptic inhibitory action. This was confirmed to be the case, since while the effects of BzATP were unaffected by the P2X7R antagonist OxATP, they were blocked by the adenosine A₁ receptor (A₁R) antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), suggesting possible BzATP hydrolysis to 2'(3')-O-(4-benzoylbenzoyl)adenosine (Bz-adenosine) and consequent activation of A₁Rs as a mechanism. Taken together, these data point out that 1) P2X7R expression and activity are confirmed to be altered in the presence of HD mutation; 2) in some experimental settings, such an abnormal functioning can be ascribed to presynaptic A₁Rs activation.

A huge diversification of phospholipids, forming the aqueous interfaces of all biomembranes, cannot be accommodated within a simple concept of their role as membrane building blocks. Indeed, a number of signaling functions of (phospho)lipid molecules has been discovered. Among these signaling lipids, a particular group of oxygenated polyunsaturated fatty acids (PUFA), so called lipid mediators, has been thoroughly investigated over several decades. This group includes oxygenated octadecanoids, eicosanoids, and docosanoids and includes several hundreds of individual species. Oxygenation of PUFA can occur when they are esterified into major classes of phospholipids. Initially, these events have been associated with non-specific oxidative injury of biomembranes. An alternative concept is that these post-synthetically oxidatively modified phospholipids and their adducts with proteins are a part of a redox epiphospholipidome that represents a rich and versatile language for intra- and inter-cellular communications. The redox epiphospholipidome may include hundreds of thousands of individual molecular species acting as meaningful biological signals. This review describes the signaling role of oxygenated phospholipids in programs of regulated cell death. Although phospholipid peroxidation has been associated with almost all known cell death programs, we chose to discuss enzymatic pathways activated during apoptosis and ferroptosis and leading to peroxidation of two phospholipid classes, cardiolipins (CLs) and phosphatidylethanolamines (PEs). This is based on the available LC-MS identification and quantitative information on the respective peroxidation products of CLs and PEs. We focused on molecular mechanisms through which two proteins, a mitochondrial hemoprotein cytochrome <i>c</i> (cyt <i>c</i>), and non-heme Fe lipoxygenase (LOX), change their catalytic properties to fulfill new functions of generating oxygenated CL and PE species. Given the high selectivity and specificity of CL and PE peroxidation we argue that enzymatic reactions catalyzed by cyt <i>c</i>/CL complexes and 15-lipoxygenase/phosphatidylethanolamine binding protein 1 (15LOX/PEBP1) complexes dominate, at least during the initiation stage of peroxidation, in apoptosis and ferroptosis. We contrast cell-autonomous nature of CLox signaling in apoptosis correlating with its anti-inflammatory functions <i>vs.</i> non-cell-autonomous ferroptotic signaling facilitating pro-inflammatory (necro-inflammatory) responses. Finally, we propose that small molecule mechanism-based regulators of enzymatic phospholipid peroxidation may lead to highly specific anti-apoptotic and anti-ferroptotic therapeutic modalities.

Emerging evidence indicates that gut dysbiosis may play a regulatory role in the onset and progression of Huntington's disease (HD). However, any alterations in the fecal microbiome of HD patients and its relation to the host cytokine response remain unknown. The present study investigated alterations and host cytokine responses in patients with HD. We enrolled 33 HD patients and 33 sex- and age- matched healthy controls. Fecal microbiota communities were determined through 16S ribosomal DNA gene sequencing, from which we analyzed fecal microbial richness, evenness, structure, and differential abundance of individual taxa between HD patients and healthy controls. HD patients were evaluated for their clinical characteristics, and the relationships of fecal microbiota with these clinical characteristics were analyzed. Plasma concentrations of interferon gamma (IFN-γ), interleukin 1 beta (IL-1β), IL-2, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, and tumor necrosis factor alpha were measured by Meso Scale Discovery (MSD) assays, and relationships between microbiota and cytokine levels were analyzed in the HD group. HD patients showed increased α-diversity (richness), β-diversity (structure), and altered relative abundances of several taxa compared to those in healthy controls. HD-associated clinical characteristics correlated with the abundances of components of fecal microbiota at the genus level. Genus <i>Intestinimonas</i> was correlated with total functional capacity scores and IL-4 levels. Our present study also revealed that genus <i>Bilophila</i> were negatively correlated with proinflammatory IL-6 levels. Taken together, our present study represents the first to demonstrate alterations in fecal microbiota and inflammatory cytokine responses in HD patients. Further elucidation of interactions between microbial and host immune responses may help to better understand the pathogenesis of HD.

Myelodysplasticsyndrome (MDS) and acute myeloid leukemia (AML) are clonal hematopoietic stem cell diseases leading to an insufficient formation of functional blood cells. Disease-immanent factors as insufficient erythropoiesis and treatment-related factors as recurrent treatment with red blood cell transfusions frequently lead to systemic iron overload in MDS and AML patients. In addition, alterations of function and expression of proteins associated with iron metabolism are increasingly recognized to be pathogenetic factors and potential vulnerabilities of these diseases. Iron is known to be involved in multiple intracellular and extracellular processes. It is essential for cell metabolism as well as for cell proliferation and closely linked to the formation of reactive oxygen species. Therefore, iron can influence the course of clonal myeloid disorders, the leukemic environment and the occurrence as well as the defense of infections. Imbalances of iron homeostasis may induce cell death of normal but also of malignant cells. New potential treatment strategies utilizing the importance of the iron homeostasis include iron chelation, modulation of proteins involved in iron metabolism, induction of leukemic cell death <i>via</i> ferroptosis and exploitation of iron proteins for the delivery of antileukemic drugs. Here, we provide an overview of some of the latest findings about the function, the prognostic impact and potential treatment strategies of iron in patients with MDS and AML.

Recent studies have reported patients who concurrently exhibit conditions of epilepsy and schizophrenia, indicating certain shared pathologies between them. This study aimed to investigate the genetic effects of the schizophrenia-related gene <i>DTNBP1</i> in temporal lobe epilepsy (TLE). A total of 496 TLE patients and 528 healthy individuals were successfully genotyped for six <i>DTNBP1</i> polymorphisms (rs760665, rs1011313, rs2619528, rs2619522, rs909706, and rs2619538), including 335 TLE patients and 325 healthy controls in cohort 1, and 161 TLE patients and 203 healthy controls in cohort 2. The frequency of the TT genotype at rs909706 T > C was lower in TLE patients than in normal controls in the initial cohort (cohort 1), which was confirmed in an independent cohort (cohort 2). However, the intronic T allele failed to be in linkage disequilibrium (LD) with any functional variations nearby; thus, together with the CCAC and TCAT haplotypes (rs1011313-rs2619528-rs2619522-rs909706) observed in the study, this allele acts only as a protective factor against susceptibility to TLE. Meanwhile, a <i>novo</i> mutant allele rs2619538 T > A was exclusively observed in TLE patients, and a dual-luciferase assay revealed that the mutant allele was increased by approximately 22% in the <i>DTNBP2</i> promoter compared with the wild-type allele. Together with the trend of increasing <i>DTNBP1</i> expression in epilepsy patients and animal models in this study, these are the first findings to demonstrate the genetic association of <i>DTNBP1</i> with TLE. Homozygous mutation of rs2619538 T > A likely promotes <i>DTNBP1</i> expression and facilitates subsequent processes in epilepsy pathologies. Thus, the role of <i>DTNBP1</i> in TLE deserves further exploration in the future.

The spread of drug-resistant bacteria via food has contributed to the dissemination of resistant bacteria among humans. However, the status of food contamination with resistant bacteria, particularly the quantitative level of resistant bacteria in food, has not yet been well elucidated. In this study, the abundance of colistin-resistant <i>Escherichia coli</i> in meat samples was quantified to understand the origin of the contamination of meat available in local Vietnamese markets. Fifteen samples each of chicken and pork meat purchased from local Vietnamese markets were assessed for the presence of colistin-resistant <i>E</i>. <i>coli</i> with the mobile colistin resistance gene, <i>mcr</i>. The results showed that 40% (6/15) and 66% (10/15) of the pork and chicken meat samples, respectively, were contaminated with colistin-resistant <i>E</i>. <i>coli</i>. The median quantitative levels of colistin-resistant <i>E</i>. <i>coli</i> in the contaminated pork and chicken samples were 1.8 × 10⁴ and 4.2 × 10³ CFU/g, respectively. The results of phylogenetic analysis of isolates from a chicken meat sample showed that the contaminated colistin-resistant <i>E</i>. <i>coli</i> was a mix of multiple phylogenetical clones of bacteria that may have multiplied during sale. This is the first study to quantify the abundance of colistin-resistant <i>E</i>. <i>coli</i> in meat samples.

<b><i>Aim</i>/<i>Hypothesis</i></b> : Lysine residues are known for the post-translational modifications (PTMs) such as acetylation, ubiquitination, and SUMOylation. In acetylation, histone deacetylase (HDAC) and its interactors cause transcriptional deregulation and cause mitochondrial dysfunction, apoptosis, inflammatory response, and cell-cycle impairment that cause brain homeostasis and neuronal cell death. Other regulatory PTMs involved in the pathogenesis of neurodegenerative diseases (NDDs) are ubiquitination and SUMOylation for the degradation of the misfolded proteins. Thus, we aim to investigate the potential acetylation/ubiquitination/SUMOylation crosstalk sites in the HDAC interactors, which cause NDDs. Furthermore, we aim to identify the influence of PTMs on the structural features of proteins and the impact of putative lysine mutation on disease susceptibility. Last, we aim to examine the impact of the putative mutation on acetylated lysine for ubiquitination and SUMOylation. <b><i>Results</i></b> : Herein, we integrate 1455 genes, 3094 genes, and 1940 genes related to HDAC interactors, Alzheimer's disease (AD), and Parkinson's disease (PD), respectively. Furthermore, the protein-protein interaction and PTM integrations from different databases identified 32 proteins that are associated with HDAC, AD, and PD with 1489 potential lysine-modified sites. HDAC interactors poly(ADP-ribose) polymerase 1 (PARP1), nucleophosmin (NPM1), and cyclin-dependent kinase 1 (CDK1) involved in the progression of NDDs and 64 and 75% of PTM sites in PARP1, NPM1, and CDK1 fall into coiled and ordered regions, respectively. Moreover, 15 putative lysine sites have been found in the crosstalk and K148, K249, K528, K637, K700, and K796 of PARP1 are crosstalk hotspots. <b><i>Conclusion</i></b> : The loss of acetylated hotspot sites results in the loss of ubiquitination and SUMOylation function on nearby sites, which is relatively higher when compared to the gain of function.

The number of microbiome-related studies has notably increased the availability of data on human microbiome composition and function. These studies provide the essential material to deeply explore host-microbiome associations and their relation to the development and progression of various complex diseases. Improved data-analytical tools are needed to exploit all information from these biological datasets, taking into account the peculiarities of microbiome data, i.e., compositional, heterogeneous and sparse nature of these datasets. The possibility of predicting host-phenotypes based on taxonomy-informed feature selection to establish an association between microbiome and predict disease states is beneficial for personalized medicine. In this regard, machine learning (ML) provides new insights into the development of models that can be used to predict outputs, such as classification and prediction in microbiology, infer host phenotypes to predict diseases and use microbial communities to stratify patients by their characterization of state-specific microbial signatures. Here we review the state-of-the-art ML methods and respective software applied in human microbiome studies, performed as part of the COST Action ML4Microbiome activities. This scoping review focuses on the application of ML in microbiome studies related to association and clinical use for diagnostics, prognostics, and therapeutics. Although the data presented here is more related to the bacterial community, many algorithms could be applied in general, regardless of the feature type. This literature and software review covering this broad topic is aligned with the scoping review methodology. The manual identification of data sources has been complemented with: (1) automated publication search through digital libraries of the three major publishers using natural language processing (NLP) Toolkit, and (2) an automated identification of relevant software repositories on GitHub and ranking of the related research papers relying on learning to rank approach.

Recently, it was found that proteomes from poliovirus, measles virus, dengue virus, and severe acute respiratory syndrome-related Coronavirus 2 (SARS-CoV-2) have high molecular mimicry at the heptapeptide level with the human proteome, while heptapeptide commonality is minimal or absent with proteomes from nonhuman primates, that is, gorilla, chimpanzee, and rhesus macaque. To acquire more data on the issue, analyses here have been expanded to Ebola virus, <i>Francisella tularensis</i> , human immunodeficiency virus-1 (HIV-1), <i>Toxoplasma gondii</i> , Variola virus, and <i>Yersinia pestis</i> . Results confirm that heptapeptide overlap is high between pathogens and <i>Homo sapiens</i> , but not between pathogens and primates. Data are discussed in light of the possible genetic bases that differently model primate phenomes, thus possibly underlying the zero/low level of molecular mimicry between infectious agents and primates. Notably, this study might help address preclinical vaccine tests that currently utilize primates as animal models, since autoimmune cross-reactions and the consequent adverse events cannot occur <i>in absentia</i> of shared sequences.

Recent discoveries of distinct molecular subtypes have led to remarkable advances in treatment for a variety of diseases. While subtyping via unsupervised clustering has received a great deal of interest, most methods rely on basic statistical or machine learning methods. At the same time, techniques based on graph clustering, particularly clique-based strategies, have been successfully used to identify disease biomarkers and gene networks. A graph theoretical approach based on the paraclique algorithm is described that can easily be employed to identify putative disease subtypes and serve as an aid in outlier detection as well. The feasibility and potential effectiveness of this method is demonstrated on publicly available gene co-expression data derived from patient samples covering twelve different disease families.

<h4>ABSTRACT</h4> <h4>Background</h4> Oncogene-induced replication stress characterizes many aggressive cancers, including triple-negative breast cancer (TNBC). Several drugs are being developed that target replication stress, although it is unclear how tumors with high levels of replication stress can be identified. We aimed to develop a gene expression signature of oncogene-induced replication stress. <h4>Methods</h4> TNBC and non-transformed RPE1- TP53 wt and RPE1- TP53 mut cell lines were engineered to overexpress the oncogenes CDC25A , CCNE1 or MYC . DNA fiber analysis was used to measure replication kinetics. Analysis of RNA sequencing data of cell lines and patient-derived tumor samples (TCGA n=10,592) was used to identify differential gene expression. Immunohistochemical validation was conducted on breast cancer samples (n=330). <h4>Results</h4> RNA sequencing revealed 52 commonly upregulated genes after induction of CDC25A , CCNE1 or MYC in our cell line panel. Integration with gene expression data of TGCA samples with amplification of replication stress-inducing oncogenes ( CDC25A , CCNE1 , MYC , CCND1 , MYB , MOS , KRAS , ERBB2 , and E2F1 ), yielded a six-gene signature of oncogene-induced replication stress ( NAT10 , DDX27 , ZNF48 , C8ORF33 , MOCS3, and MPP6) . Expression of NAT10 in breast cancer samples was correlated with phospho-RPA (R=0.451, p=1.82×10 −20 ) and γH2AX (R=0.304, p=2.95×10 −9 ). Applying the oncogene-induced replication stress signature to patient samples (TCGA n=8,862 and GEO n=13,912) defined the replication stress landscape across 27 tumor subtypes, and identified diffuse large B cell lymphoma, ovarian cancer, TNBC and colorectal carcinoma as cancer subtypes with high levels of oncogene-induced replication stress. <h4>Conclusion</h4> We developed a gene expression signature of oncogene-induced replication stress, which may facilitate patient selection for agents that target replication stress.

As a self-degradative mechanism, macroautophagy/autophagy has a role in the maintenance of energy homeostasis during critical periods in the development of cells. It also controls cellular damage through the eradication of damaged proteins and organelles. This process is accomplished by tens of ATG (autophagy-related) proteins. Recent studies have shown the involvement of non-coding RNAs in the regulation of autophagy. These transcripts mostly modulate the expression of <i>ATG</i> genes. Both long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) have been shown to modulate the autophagy mechanism. Levels of several lncRNAs and miRNAs are altered in this process. In the present review, we discuss the role of lncRNAs and miRNAs in the regulation of autophagy in diverse contexts such as cancer, deep vein thrombosis, spinal cord injury, diabetes and its complications, acute myocardial infarction, osteoarthritis, pre-eclampsia and epilepsy.<b>Abbreviations</b>: AMI: acute myocardial infarction; ATG: autophagy-related; lncRNA: long non-coding RNA; miRNA: microRNA.

This study aimed to investigate the effects of eugenol on growth, viability, antrum formation and mRNA expression of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase 1 (GPX1) and peroxiredoxin 6 (PRDX6) in bovine secondary follicles cultured in vitro. To this end, bovine ovaries were collected from a local slaughterhouse and in the laboratory the follicles were isolated from the ovarian cortex. The follicles were then cultured in TCM-199+ alone or supplemented with different concentrations of eugenol (0.5, 5.0 and 50.0 μM). Follicular diameters and antrum formation were evaluated on days 0, 6, 12 and 18. Viability analysis was performed using calcein and ethidium homodimer. Real-time PCR was used to quantify mRNA levels for SOD, CAT, GPX1 and PRDX6 in cultured follicles. Follicular diameters and mRNA levels in follicles cultured in vitro were compared using analysis of variance and Kruskal-Wallis tests, while follicular survival and antrum formation were compared using the chi-squared test (P < 0.05). The results showed that secondary follicles cultured with eugenol maintained similar morphology and viability to follicles cultured in the control group. A progressive increase in follicular diameter was observed between days 0 and 12 for all treatments, except for follicles cultured with 50 µM eugenol. Eugenol (5.0 and 50.0 μM) increased mRNA levels for GPX1 in cultured follicles, but 0.5 μM eugenol reduced mRNA levels for SOD. The addition of eugenol did not influence mRNA expression for CAT and PRDX6. In conclusion, eugenol supplementation reduces mRNA levels for SOD and increases mRNA levels of GPX1 in bovine secondary follicles cultured in vitro.

Cells metabolize nutrients for biosynthetic and bioenergetic needs to fuel growth and proliferation. The uptake of nutrients from the environment and their intracellular metabolism is a highly controlled process that involves cross talk between growth signaling and metabolic pathways. Despite constant fluctuations in nutrient availability and environmental signals, normal cells restore metabolic homeostasis to maintain cellular functions and prevent disease. A central signaling molecule that integrates growth with metabolism is the mechanistic target of rapamycin (mTOR). mTOR is a protein kinase that responds to levels of nutrients and growth signals. mTOR forms two protein complexes, mTORC1, which is sensitive to rapamycin, and mTORC2, which is not directly inhibited by this drug. Rapamycin has facilitated the discovery of the various functions of mTORC1 in metabolism. Genetic models that disrupt either mTORC1 or mTORC2 have expanded our knowledge of their cellular, tissue, as well as systemic functions in metabolism. Nevertheless, our knowledge of the regulation and functions of mTORC2, particularly in metabolism, has lagged behind. Since mTOR is an important target for cancer, aging, and other metabolism-related pathologies, understanding the distinct and overlapping regulation and functions of the two mTOR complexes is vital for the development of more effective therapeutic strategies. This review discusses the key discoveries and recent findings on the regulation and metabolic functions of the mTOR complexes. We highlight findings from cancer models but also discuss other examples of the mTOR-mediated metabolic reprogramming occurring in stem and immune cells, type 2 diabetes/obesity, neurodegenerative disorders, and aging.

Obstructive sleep apnea (OSA) results from episodes of airway collapse and intermittent hypoxia (IH) and is associated with a host of health complications. Although the lung is the first organ to sense changes in oxygen levels, little is known about the consequences of IH to the lung hypoxia-inducible factor-responsive pathways. We hypothesized that exposure to IH would lead to cell-specific up- and downregulation of diverse expression pathways. We identified changes in circadian and immune pathways in lungs from mice exposed to IH. Among all cell types, endothelial cells showed the most prominent transcriptional changes. Upregulated genes in myofibroblast cells were enriched for genes associated with pulmonary hypertension and included targets of several drugs currently used to treat chronic pulmonary diseases. A better understanding of the pathophysiologic mechanisms underlying diseases associated with OSA could improve our therapeutic approaches, directing therapies to the most relevant cells and molecular pathways.

Treatment for many viral infections of the central nervous system (CNS) remains only supportive. Here we address a remaining gap in our knowledge regarding how the CNS and immune systems interact during viral infection. By examining the regulation of the immune and nervous system processes in a nonhuman primate model of West Nile virus neurological disease, we show that virus infection disrupts the homeostasis of the immune-neural-synaptic axis via induction of pleiotropic genes with distinct functions in each component of the axis. This pleiotropic gene regulation suggests an unintended off-target negative impact of virus-induced host immune responses on the neurotransmission, which may be a common feature of various viral infections of the CNS.

The synthesis of purine conjugates with natural amino acids is one of the promising directions in search for novel therapeutic agents, including antimycobacterial agents. The purpose of this study was to synthesize N-(purin-6-yl)dipeptides containing the terminal fragment of (S)-glutamic acid. To obtain the target compounds, two synthetic routes were tested. The first of them is based on coupling of N-(purin-6-yl)-(S)-amino acids to dimethyl (S)-glutamate in the presence of carbodiimide coupling agent followed by the removal of ester groups. However, it turned out that this coupling process was accompanied by racemization of the chiral center of N-(purin-6-yl)-α-amino acids and in all cases led to mixtures of (S,S)- and (R,S)-diastereomers (6:4). Individual (S,S)-diastereomers were obtained using an alternative approach based on the nucleophilic substitution of chlorine in 6-chloropurine or 2-amino-6-chloropurine with corresponding dipeptides as nucleophiles. The enantiomeric purity of the target compounds was confirmed by chiral HPLC. To test the assumption that racemization of the chiral center of N-(purin-6-yl)-α-amino acids occurs with the participation of nitrogen atoms of the imidazole ring via the stage of formation of a chirally labile intermediate, we obtained such structural analogs of N-(purin-6-yl)-(S)-alanine as N-(9-benzylpurin-6-yl)-(S)-alanine and N-(7-deazapurin-6-yl)-(S)-alanine. It was found that coupling of these compounds to dimethyl (S)-glutamate was also accompanied by racemization. This indicates that the imidazole fragment does not play a crucial role in this process. When testing the antimycobacterial activity of some of the obtained compounds, conjugates with moderate activity against the laboratory Mycobacterium tuberculosis H37Rv strain (MIC 3.1-6.25 μg/mL) were identified.

In the fruit fly Drosophila melanogaster, male-specific splicing and translation of the Fruitless transcription factor (FruM) alters the presence, anatomy, and/or connectivity of >60 types of central brain neurons that interconnect to generate male-typical behaviors. While the indispensable function of FruM in sex-specific behavior has been understood for decades, the molecular mechanisms underlying its activity remain unknown. Here, we take a genome-wide, brain-wide approach to identifying regulatory elements whose activity depends on the presence of FruM. We identify 436 high-confidence genomic regions differentially accessible in male fruitless neurons, validate candidate regions as bona fide, differentially regulated enhancers, and describe the particular cell types in which these enhancers are active. We find that individual enhancers are not activated universally but are dedicated to specific fru+ cell types. Aside from fru itself, genes are not dedicated to or common across the fru circuit; rather, FruM appears to masculinize each cell type differently, by tweaking expression of the same effector genes used in other circuits. Finally, we find FruM motifs enriched among regulatory elements that are open in the female but closed in the male. Together, these results suggest that FruM acts cell-type-specifically to decommission regulatory elements in male fruitless neurons.

<h4>Background</h4>Human induced pluripotent stem cells (iPSC) have opened new avenues for regenerative medicine. Consequently, iPSC-derived motor neurons have emerged as potentially viable therapies for spinal cord injuries and neurodegenerative disorders including Amyotrophic Lateral Sclerosis. However, direct clinical application of iPSC bears in itself the risk of tumorigenesis and other unforeseeable genetic or epigenetic abnormalities.<h4>Results</h4>Employing RNA-seq technology, we identified and characterized gene regulatory networks triggered by in vitro chemical reprogramming of iPSC into cells with the molecular features of motor neurons (MNs) whose function in vivo is to innervate effector organs. We present meta-transcriptome signatures of 5 cell types: iPSCs, neural stem cells, motor neuron progenitors, early motor neurons, and mature motor neurons. In strict response to the chemical stimuli, along the MN differentiation axis we observed temporal downregulation of tumor growth factor-β signaling pathway and consistent activation of sonic hedgehog, Wnt/β-catenin, and Notch signaling. Together with gene networks defining neuronal differentiation (neurogenin 2, microtubule-associated protein 2, Pax6, and neuropilin-1), we observed steady accumulation of motor neuron-specific regulatory genes, including Islet-1 and homeobox protein HB9. Interestingly, transcriptome profiling of the differentiation process showed that Ca²⁺ signaling through cAMP and LPC was downregulated during the conversion of the iPSC to neural stem cells and key regulatory gene activity of the pathway remained inhibited until later stages of motor neuron formation. Pathways shaping the neuronal development and function were well-represented in the early motor neuron cells including, neuroactive ligand-receptor interactions, axon guidance, and the cholinergic synapse formation. A notable hallmark of our in vitro motor neuron maturation in monoculture was the activation of genes encoding G-coupled muscarinic acetylcholine receptors and downregulation of the ionotropic nicotinic acetylcholine receptors expression. We observed the formation of functional neuronal networks as spontaneous oscillations in the extracellular action potentials recorded on multi-electrode array chip after 20 days of differentiation.<h4>Conclusions</h4>Detailed transcriptome profile of each developmental step from iPSC to motor neuron driven by chemical induction provides the guidelines to novel therapeutic approaches in the re-construction efforts of muscle innervation.

<h4>Background</h4>The quality and yield of wool determine the economic value of the fine-wool sheep. Therefore, discovering markers or genes relevant to wool traits is the cornerstone for the breeding of fine-wool sheep. In this study, we used the Illumina HiSeq X Ten platform to re-sequence 460 sheep belonging to four different fine-wool sheep breeds, namely, Alpine Merino sheep (AMS), Chinese Merino sheep (CMS), Aohan fine-wool sheep (AHS) and Qinghai fine-wool sheep (QHS). Eight wool traits, including fiber diameter (FD), fiber diameter coefficient of variance (FDCV), fiber diameter standard deviation (FDSD), staple length (SL), greasy fleece weight (GFW), clean wool rate (CWR), staple strength (SS) and staple elongation (SE) were examined. A genome-wide association study (GWAS) was performed to detect the candidate genes for the eight wool traits.<h4>Results</h4>A total of 8.222 Tb of raw data was generated, with an average of approximately 8.59X sequencing depth. After quality control, 12,561,225 SNPs were available for analysis. And a total of 57 genome-wide significant SNPs and 30 candidate genes were detected for the desired wool traits. Among them, 7 SNPs and 6 genes are related to wool fineness indicators (FD, FDCV and FDSD), 10 SNPs and 7 genes are related to staple length, 13 SNPs and 7 genes are related to wool production indicators (GFW and CWR), 27 SNPs and 10 genes associated with staple elongation. Among these candidate genes, UBE2E3 and RHPN2 associated with fiber diameter, were found to play an important role in keratinocyte differentiation and cell proliferation. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment results, revealed that multitude significant pathways are related to keratin and cell proliferation and differentiation, such as positive regulation of canonical Wnt signaling pathway (GO:0090263).<h4>Conclusion</h4>This is the first GWAS on the wool traits by using re-sequencing data in Chinese fine-wool sheep. The newly detected significant SNPs in this study can be used in genome-selective breeding for the fine-wool sheep. And the new candidate genes would provide a good theoretical basis for the fine-wool sheep breeding.

<h4>Background</h4>Population health and well-being in Latin America, the current epicenter of the COVID-19 pandemic, has been severely affected during the past semester. Despite the growing evidence about the link between the pandemic, its control measures, and mental health worldwide, there is still no regional evidence of the potential mental health impact. We describe the prevalence and distribution of depressive symptoms across demographic and socioeconomic risk factors in the Peruvian population amidst a national lockdown during the COVID-19 pandemic.<h4>Methods</h4>Cross-sectional study conducted during the community transmission phase and national lockdown in Peru (May 4th-16th, 2020). We recorded 64,493 responses from adult Peruvian residents through an opt-in online questionnaire. All analyses were weighted using raking based on proportions of sociodemographic variables from the last Peruvian census in 2017. The prevalence of depressive symptoms was calculated using the Patient Health Questionnaire (PHQ-9) score of 10 or more. We identified associated demographic and socioeconomic factors by prior mental health diagnosis. Sensitivity analysis considered an alternative cut-off point for depressive symptoms of PHQ-9 ≥ 14.<h4>Results</h4>A total of 57,446 participants were included in the analytical sample. A third of the participants (n = 23,526, unweighted) showed depressive symptoms in the 2 weeks prior to the study. Participants who reported a previous mental health diagnosis doubled the sample prevalence of depressive symptoms (59, 95%CI 56.7, 61.4%) of those without a prior diagnosis. Psychosocial and functioning reactions were largely more prevalent among females and young adults. A dose-response relationship was found between household income and depressive symptoms across previous mental health diagnosis strata, being as low as 32% less in the wealthiest than the most impoverished group (PR: 0.68, 95%CI 0.58,0.79). Other critical factors associated with a higher burden of depressive symptoms were lower education level, single, unemployed, and chronic comorbidity.<h4>Conclusions</h4>An increased burden of depressive symptoms and psychosocial reactions has emerged during the COVID-19 pandemic in Peru compared to previous years. The mental health burden disproportionately affects women, the younger population, and those with low income and education. As the country eases the social distancing measures, it is crucial to use local evidence to adjust public health policies and mental health services to the renewed population needs.

<h4>Background</h4>Apoptosis-related genes(Args)play an essential role in the occurrence and progression of hepatocellular carcinoma(HCC). However, few studies have focused on the prognostic significance of Args in HCC. In the study, we aim to explore an efficient prognostic model of Asian HCC patients based on the Args.<h4>Methods</h4>We downloaded mRNA expression profiles and corresponding clinical data of Asian HCC patients from The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) databases. The Args were collected from Deathbase, a database related to cell death, combined with the research results of GeneCards、National Center for Biotechnology Information (NCBI) databases and a lot of literature. We used Wilcoxon-test and univariate Cox analysis to screen the differential expressed genes (DEGs) and the prognostic related genes (PRGs) of HCC. The intersection genes of DEGs and PGGs were seen as crucial Args of HCC. The prognostic model of Asian HCC patients was constructed by least absolute shrinkage and selection operator (lasso)- proportional hazards model (Cox) regression analysis. Kaplan-Meier curve, Principal Component Analysis (PCA) analysis, t-distributed Stochastic Neighbor Embedding (t-SNE) analysis, risk score curve, receiver operating characteristic (ROC) curve, and the HCC data of ICGC database and the data of Asian HCC patients of Kaplan-Meier plotter database were used to verify the model.<h4>Results</h4>A total of 20 of 56 Args were differentially expressed between HCC and adjacent normal tissues (p < 0.05). Univariate Cox regression analysis showed that 10 of 56 Args were associated with survival time and survival status of HCC patients (p < 0.05). There are seven overlapping genes of these 20 and 10 genes, including BAK1, BAX, BNIP3, CRADD, CSE1L, FAS, and SH3GLB1. Through Lasso-Cox analysis, an HCC prognostic model composed of BAK1, BNIP3, CSE1L, and FAS was constructed. Kaplan-Meier curve, PCA, t-SNE analysis, risk score curve, ROC curve, and secondary verification of ICGC database and Kaplan-Meier plotter database all support the reliability of the model.<h4>Conclusions</h4>Lasso-Cox regression analysis identified a 4-gene prognostic model, which integrates clinical and gene expression and has a good effect. The expression of Args is related to the prognosis of HCC patients, but the specific mechanism remains to be further verified.

<h4>Background</h4>Recent advances in therapeutic options may prevent deterioration related to Huntington's disease (HD), even at the pre-symptomatic stage. Be that as it may, a well-characterized patient population is essential for screening and monitoring outcome. Accordingly, the aim of this study was to describe the characteristics of a Hungarian subpopulation of HD patients and mutation carriers diagnosed at the University of Szeged.<h4>Methods</h4>We conducted a search for International Classification of Diseases (ICD) code G10H0 in the local medical database for the period of 1 January 1998 to 31 December 2018.<h4>Results</h4>We identified 90 HD cases (male: 45, female: 45) and 34 asymptomatic carriers (male: 15, female: 19). The median age of onset was 45 years (range: 16-79). There were 3 cases of juvenile onset (3.3%), and 7 of late disease onset (7.8%). The median repeat length was 43 (range: 36-70) for the pathological and 19 for the non-pathological alleles (range: 9-35). 17.5% of the pathological alleles were in the decreased penetrance range, while 7% of non-pathological alleles were intermediate.<h4>Conclusions</h4>The genetic and clinical features of the population examined in the present study were in line with the previous Hungarian study, as well as with international literature. The exceptions were the higher ratio of reduced penetrance and intermediate alleles.

<h4>Background</h4>To validate markers for cervical carcinoma (CC) and precancerous lesions related with HPV infections.<h4>Methods</h4>Three different cervical cancer cell lines C-33A, SiHa and Caski were used for secretome profiling by label-free quantitative proteomics. Cervical exfoliated cells and matching serum samples were collected from 284 patients with normal control (n = 75, 26.41 %), precancerous lesions (n = 88, 30.99 %) and early stage cervical squamous carcinoma (n = 121, 42.61 %). HPV subtyping and quantification was performed by PCR and hybridization. 20 candidate proteins identified in previous screening studies (tissue, plasma, cells) were quantified by ELISA. Finally, highly quantitative parallel reaction monitoring mass spectrometry was used to assess the specificities and sensitivities of candidate serum markers.<h4>Results</h4>While CC was found to be associated with high-risk HPV subtypes, serum antibodies for high risk HPV were not significantly related to the progression of cervical cancer. Significant differences between patient groups were detected for the four proteins CLU, APOA4, APOE and MLH3, but none would allow clinical application due to insufficient sensitivity and specificity and large variability. Subsequent proteomic secretome analysis of cervical cancer cell lines identified a set of 729 common proteins. Cross referencing this dataset with ELISA measurements revealed six candidate proteins of which two, FBLN1 and ANT3, showed co-occurrence with HPV infection (75.7 % and 85 %, respectively) and had promising diagnostic ability in terms of sensitivity and specificity. After the loss of E6/E7 by using CRISPR/Cas9 gene editing, the content of ANT3 and FBLN1 in KoE6/E7 SiHa were downregulated, which indicated the expression of ANT3 and FBLN1 in cervical cancer may be affected by HPV infection.<h4>Conclusions</h4>FBLN1 and ANT3 might be potential tumor- and HPV-associated serum markers.