Gene Literature Dashboard

<h4>Purpose</h4>The physicochemical properties of a xenograft are very important because they strongly influence the bone regeneration capabilities of the graft material. Even though porcine xenografts have many advantages, only a few porcine xenografts are commercially available, and most of their physicochemical characteristics have yet to be reported. Thus, in this work we aimed to investigate the physicochemical characteristics of a porcine bone grafting material and compare them with those of 2 commercially available bovine xenografts to assess the potential of xenogenic porcine bone graft materials for dental applications.<h4>Methods</h4>We used various characterization techniques, such as scanning electron microscopy, the Brunauer-Emmett-Teller adsorption method, atomic force microscopy, Fourier-transform infrared spectroscopy, X-ray diffraction, and others, to compare the physicochemical properties of xenografts of different origins.<h4>Results</h4>The porcine bone grafting material had relatively high porosity (78.4%) and a large average specific surface area (SSA; 69.9 m²/g), with high surface roughness (10-point average roughness, 4.47 µm) and sub-100-nm hydroxyapatite crystals on the surface. Moreover, this material presented a significant fraction of sub-100-nm pores, with negligible amounts of residual organic substances. Apart from some minor differences, the overall characteristics of the porcine bone grafting material were very similar to those of one of the bovine bone grafting material. However, many of these morphostructural properties were significantly different from the other bovine bone grafting material, which exhibited relatively smooth surface morphology with a porosity of 62.0% and an average SSA of 0.5 m²/g.<h4>Conclusions</h4>Considering that both bovine bone grafting materials have been successfully used in oral surgery applications in the last few decades, this work shows that the porcine-derived grafting material possesses most of the key physiochemical characteristics required for its application as a highly efficient xenograft material for bone replacement.

Myopia is the most common cause of visual impairment worldwide. Genetic and environmental factors contribute to the development of myopia. Studies on the molecular genetics of myopia are well established and have implicated the important role of genetic factors. With linkage analysis, association studies, sequencing analysis, and experimental myopia studies, many of the loci and genes associated with myopia have been identified. Thus far, there has been no systemic review of the loci and genes related to non-syndromic and syndromic myopia based on the different approaches. Such a systemic review of the molecular genetics of myopia will provide clues to identify additional plausible genes for myopia and help us to understand the molecular mechanisms underlying myopia. This paper reviews recent genetic studies on myopia, summarizes all possible reported genes and loci related to myopia, and suggests implications for future studies on the molecular genetics of myopia.

<h4>Aim of the study</h4>was to analyze the outcome of treatment and factors predicting results of sorafenib therapy in inoperable/metastatic CD117-positive GIST patients after failure on imatinib and sunitinib.<h4>Material and methods</h4>We identified 60 consecutive patients (40 men, 20 women) with advanced inoperable/metastatic GIST after failure on at least imatinib and sunitinib treated in one sarcoma center with sorafenib at initial dose 2 × 400 mg daily in 2007-2015 (in 56 cases it was 3^rd line therapy). Median follow-up time was 39 months.<h4>Results</h4>One year progression-free survival (PFS; calculated from the date of the start of sorafenib to disease progression) rate was 23% and median PFS = 7.7 months. The median overall survival (OS) was 13.5 months calculated from sorafenib start (1-year OS rate = 57%) and 7 years from imatinib start. Three patients (5%) had objective partial responses to therapy, 31 patients (52%) had stabilization of disease > 4 months. Primary tumor mutational status was known in 43 cases (73%), but we have not identified the differences in PFS between tumors carrying different KIT/PDGFRA mutations. The most common adverse events were: diarrhoea, hand and foot syndrome, fatigue, loss of weight and skin reactions; grade 3-5 toxicity occurred in 35% of patients. 23 patients required sorafenib dose reductions due to AEs.<h4>Conclusions</h4>We confirmed that many advanced GIST patients benefit from sorafenib therapy after imatinib/sunitinib failure with OS > 1 year.

Butyrophilins belonging to the immunoglobulin superfamily are new immune system regulators because they are present on lymphocytes, dendritic cells, monocytes, macrophages, neutrophils and eosinophils, and they exert a stimulatory and (or) inhibitory effect on them. The role of butyrophilins is associated and results from their similarity to the regulatory B7 protein family involved in the modulation of immune phenomena. Butyrophilins are glycoproteins built of two extracellular immunoglobulin domains, stabilized with disulfide bonds: constant IgC, and variable IgV and a transmembrane region. Most of these proteins contain a conserved domain encoded by a single exon - B30.2, also referred to as PRYSPRY. In humans, the family of butyrophilins includes 7 butyrophilin proteins, 5 butyrophilin-like proteins and the SKINT-like factor. Butyrophilins have been also demonstrated to play a role in various infections, e.g. tuberculosis or diseases that include sarcoidosis, systemic lupus erythematosus, rheumatoid arthritis, genetic metabolic diseases, ulcerative colitis, cancer and kidney disease.

The blood-tumor barrier (BTB) restricts the efficient delivery of anti-glioma drugs to cranial glioma tissues. Increased BTB permeability may allow greater delivery of the therapeutic agents. Increasing evidence has revealed that PIWI proteins and PIWI-interacting RNAs (piRNAs) play an important role in tumor progression. However, whether PIWI proteins and piRNAs regulate BTB permeability remains unclear. In the present study, we demonstrated that the PIWIL1/piRNA-DQ593109 (piR-DQ593109) complex was the predominant regulator of BTB permeability. Briefly, PIWIL1 was upregulated in glioma endothelial cells (GECs). Furthermore, piR-DQ593109 was also overexpressed in GECs, as revealed via a piRNA microarray. Downregulation of PIWIL1 or piR-DQ593109 increased the permeability of the BTB. Moreover, PIWIL1 and piR-DQ593109, which formed a piRNA-induced silencing complex, degraded the long non-coding RNA maternally expressed 3 (MEG3) in a sequenced-dependent manner. Furthermore, restoring MEG3 released post-transcriptional inhibition of Runt related transcription factor 3 (RUNX3) by sponging miR-330-5p. In addition, RUNX3 bounded to the promoter regions and reduced the promoter activities of ZO-1, occludin, and claudin-5, which significantly impaired the expression levels of ZO-1, occludin, and claudin-5. In conclusion, downregulating PIWIL1 and piR-DQ593109 increased BTB permeability through the MEG3/miR-330-5p/RUNX3 axis. These data may provide insight into glioma treatment.

<h4>ABSTRACT</h4> Specific class I members of the TRIM family of E3 ubiquitin ligases have been implicated in neuronal development from invertebrates to mammals. The single invertebrate class I TRIM and mammalian TRIM9 regulate axon branching and guidance in response to the axon guidance cue netrin-1, whereas mammalian TRIM46 establishes the axon initial segment. In humans, mutations in TRIM1 and TRIM18 are implicated in Optiz Syndrome, characterized by midline defects and often mild intellectual disability. TRIM67 is the most evolutionarily conserved vertebrate class I TRIM, yet is the least studied. Here we show that TRIM67 interacts with both its closest paralog TRIM9 and the netrin receptor DCC, and is differentially enriched in specific brain regions at specific developmental points. We describe the anatomical and behavioral consequences of deletion of murine Trim67 . While viable, mice lacking Trim67 display severe impairments in spatial memory, cognitive flexibility, social novelty preference, muscle function and sensorimotor gating. Additionally, they exhibit abnormal anatomy of several brain regions, including the hippocampus, striatum and thalamus, as well as the corpus callosum. This study demonstrates the necessity for TRIM67 in appropriate brain development and function. <h4>SIGNIFICANCE STATEMENT</h4> As a family, class I TRIM E3 ubiquitin ligases play important roles in neuronal development and function, potentially cooperatively. TRIM67 is the most evolutionarily conserved class I TRIM and is developmentally regulated and brain-enriched. Deletion of murine Trim67 results in malformations of a subset subcortical brain regions and of cortical and subcortical myelinated fiber tracts, as well as deficits in spatial memory, motor function, sociability and sensorimotor gating. We conclude that TRIM67 is critical for appropriate brain development and behavior, potentially downstream of the axon guidance cue netrin, and in cooperation with class I TRIM9.

Plasma extracellular vesicles (EVs) are lipid membrane vesicles involved in several biological processes including coagulation. Both coagulation and lipid metabolism are strongly associated with cardiovascular events. Lowering very-low- and low-density lipoprotein ((V)LDL) particles via dextran sulphate LDL apheresis also removes coagulation proteins. It remains unknown, however, how coagulation proteins are removed in apheresis. We hypothesize that plasma EVs that contain high levels of coagulation proteins are concomitantly removed with (V)LDL particles by dextran sulphate apheresis. For this, we precipitated (V)LDL particles from human plasma with dextran sulphate and analyzed the abundance of coagulation proteins and EVs in the precipitate. Coagulation pathway proteins, as demonstrated by proteomics and a bead-based immunoassay, were over-represented in the (V)LDL precipitate. In this precipitate, both bilayer EVs and monolayer (V)LDL particles were observed by electron microscopy. Separation of EVs from (V)LDL particles using density gradient centrifugation revealed that almost all coagulation proteins were present in the EVs and not in the (V)LDL particles. These EVs also showed a strong procoagulant activity. Our study suggests that dextran sulphate used in LDL apheresis may remove procoagulant EVs concomitantly with (V)LDL particles, leading to a loss of coagulation proteins from the blood.

Detecting prostate cancer (PCa) using non-invasive diagnostic markers still remains a challenge. The aim of this study was the identification of urine proteins that are sufficiently sensitive and specific to detect PCa in the early stages. Comparative proteomics profiling of urine from patients with PCa, benign prostate hyperplasia, bladder cancer, and renal cancer, coupled with bioinformatics analysis, were performed. Statistically significant difference in abundance showed 20 and 85 proteins in the 2-D DIGE/MS and label-free LC-MS/MS experiments, respectively. In silico analysis indicated activation, binding, and cell movement of subset of immune cells as the top affected cellular functions in PCa, together with the down-regulation of Acute Phase Response Signaling and Liver X Receptor/ Retinoid X Receptor (LXR/RXR) activation pathways. The most promising biomarkers were 35, altered in PCa when compared to more than one group. Half of these have confirmed localization in normal or PCa tissues. Twenty proteins (CD14, AHSG, ENO1, ANXA1, CLU, COL6A1, C3, FGA, FGG, HPX, PTGDS, S100A9, LMAN2, ITIH4, ACTA2, GRN, HBB, PEBP1, CTSB, SPP1) are oncogenes, tumor suppressors, and multifunctional proteins with highly confirmed involvement in PCa, while 9 (AZU1, IGHG1, RNASE2, PZP, REG1A, AMY1A, AMY2A, ACTG2, COL18A1) have been associated with different cancers, but not with PCa so far, and may represent novel findings. LC-MS/MS data are available via ProteomeXchange with identifier PXD008407.

<h4>Objective</h4>Cartilage homeostasis relies on a balance of catabolism and anabolism of cartilage matrix. Our goal was to evaluate the burden of radiographic osteoarthritis and serum levels of type IIA procollagen amino terminal propeptide (sPIIANP), a biomarker representing type II collagen synthesis, in osteoarthritis.<h4>Methods</h4>OA burden was quantified on the basis of radiographic features as total joint faces with an osteophyte, joint space narrowing, or in the spine, disc space narrowing. sPIIANP was measured in 1,235 participants from the Genetics of Generalized Osteoarthritis study using a competitive enzyme-linked immunosorbent assay. Separate multivariable linear regression models, adjusted for age, sex, and body mass index and additionally for ipsilateral osteophytes or joint/disc space narrowing, were used to assess the independent association of sPIIANP with osteophytes and with joint/disc space narrowing burden in knees, hips, hands and spine, individually and together.<h4>Results</h4>After full adjustment, sPIIANP was significantly associated with a lesser burden of hip joint space narrowing and knee osteophytes. sPIIANP was associated with a lesser burden of hand joint space narrowing but a greater burden of hand osteophytes; these results were only evident upon adjustment for osteoarthritic features in all other joints. There were no associations of sPIIANP and features of spine osteoarthritis.<h4>Conclusions</h4>Higher cartilage collagen synthesis, as reflected in systemic PIIANP concentrations, was associated with lesser burden of osteoarthritic features in lower extremity joints (knees and hips), even accounting for osteoarthritis burden in hands and spine, age, sex and body mass index. These results suggest that pro-anabolic agents may be appropriate for early treatment to prevent severe lower extremity large joint osteoarthritis.

<h4>Background</h4>Breast cancer brain metastases (BCBM) develop in about 20-30% of breast cancer (BC) patients. BCBM are associated with dismal prognosis not at least due to lack of valuable molecular therapeutic targets. The aim of the study was to identify new molecular biomarkers and targets in BCBM by using complementary state-of-the-art techniques.<h4>Methods</h4>We compared array expression profiles of three BCBM with 16 non-brain metastatic BC and 16 primary brain tumors (prBT) using a false discovery rate (FDR) p < 0.05 and fold change (FC) > 2. Biofunctional analysis was conducted on the differentially expressed probe sets. High-density arrays were employed to detect copy number variations (CNVs) and whole exome sequencing (WES) with paired-end reads of 150 bp was utilized to detect gene mutations in the three BCBM.<h4>Results</h4>The top 370 probe sets that were differentially expressed between BCBM and both BC and prBT were in the majority comparably overexpressed in BCBM and included, e.g. the coding genes BCL3, BNIP3, BNIP3P1, BRIP1, CASP14, CDC25A, DMBT1, IDH2, E2F1, MYCN, RAD51, RAD54L, and VDR. A number of small nucleolar RNAs (snoRNAs) were comparably overexpressed in BCBM and included SNORA1, SNORA2A, SNORA9, SNORA10, SNORA22, SNORA24, SNORA30, SNORA37, SNORA38, SNORA52, SNORA71A, SNORA71B, SNORA71C, SNORD13P2, SNORD15A, SNORD34, SNORD35A, SNORD41, SNORD53, and SCARNA22. The top canonical pathway was entitled, role of BRCA1 in DNA damage response. Network analysis revealed key nodes as Akt, ERK1/2, NFkB, and Ras in a predicted activation stage. Downregulated genes in a data set that was shared between BCBM and prBT comprised, e.g. BC cell line invasion markers JUN, MMP3, TFF1, and HAS2. Important cancer genes affected by CNVs included TP53, BRCA1, BRCA2, ERBB2, IDH1, and IDH2. WES detected numerous mutations, some of which affecting BC associated genes as CDH1, HEPACAM, and LOXHD1.<h4>Conclusions</h4>Using complementary molecular genetic techniques, this study identified shared and unshared molecular events in three highly aberrant BCBM emphasizing the challenge to detect new molecular biomarkers and targets with translational implications. Among new findings with the capacity to gain clinical relevance is the detection of overexpressed snoRNAs known to regulate some critical cellular functions as ribosome biogenesis.

<h4>Purpose</h4>The lack of specific symptoms at early tumor stages, together with a high biological aggressiveness of the tumor contribute to the high mortality rate for pancreatic cancer (PC), which has a 5-year survival rate of about 7%. Recent failures of targeted therapies inhibiting kinase activity in clinical trials have highlighted the need for new approaches towards combating this deadly disease.<h4>Methods</h4>In this study, we have identified genes that are significantly downregulated in PC, through a meta-analysis of large number of microarray datasets. We have used qRT-PCR to confirm the downregulation of selected genes in a panel of PC cell lines.<h4>Results</h4>This study has yielded several novel candidate tumor-suppressor genes (TSGs) including GNMT, CEL, PLA2G1B and SERPINI2. We highlight the role of GNMT, a methyl transferase associated with the methylation potential of the cell, and CEL, a lipase, as potential therapeutic targets. We have uncovered genetic links to risk factors associated with PC such as smoking and obesity. Genes important for patient survival and prognosis are also discussed, and we confirm the dysregulation of metabolic pathways previously observed in PC.<h4>Conclusions</h4>While many of the genes downregulated in our dataset are associated with protein products normally produced by the pancreas for excretion, we have uncovered some genes whose downregulation appear to play a more causal role in PC. These genes will assist in providing a better understanding of the disease etiology of PC, and in the search for new therapeutic targets and biomarkers.

During S phase, replicated DNA must be assembled into nucleosomes using both newly synthesized and parental histones in a process that is tightly coupled to DNA replication. This DNA replication-coupled process is regulated by multitude of histone chaperones as well as by histone-modifying enzymes. In recent years novel insights into nucleosome assembly of new H3-H4 tetramers have been gained through studies on the classical histone chaperone CAF-1 and the identification of novel factors involved in this process. Moreover, in vitro reconstitution of chromatin replication has shed light on nucleosome assembly of parental H3-H4, a process that remains elusive. Finally, recent studies have revealed that the replication-coupled nucleosome assembly is important for the determination and maintenance of cell fate in multicellular organisms.

<h4>Introduction</h4>Ensuring reliable administration and reporting of cognitive screening tests are fundamental in establishing good clinical practice and research. This study captured the rate and type of errors in clinical practice, using the Addenbrooke's Cognitive Examination-III (ACE-III), and then the reduction in error rate using a computerized alternative, the ACEmobile app.<h4>Methods</h4>In study 1, we evaluated ACE-III assessments completed in National Health Service (NHS) clinics (<i>n</i> = 87) for administrator error. In study 2, ACEmobile and ACE-III were then evaluated for their ability to capture accurate measurement.<h4>Results</h4>In study 1, 78% of clinically administered ACE-IIIs were either scored incorrectly or had arithmetical errors. In study 2, error rates seen in the ACE-III were reduced by 85%-93% using ACEmobile.<h4>Discussion</h4>Error rates are ubiquitous in routine clinical use of cognitive screening tests and the ACE-III. ACEmobile provides a framework for supporting reduced administration, scoring, and arithmetical error during cognitive screening.

<h4>Objective</h4>To assess the role of differentially expressed proteins as a resource for potential biomarker identification of infertility, as male infertility is of rising concern in reproductive medicine and evidence pertaining to its aetiology at a molecular level particularly proteomic as spermatozoa lack transcription and translation. Proteomics is considered as a major field in molecular biology to validate the target proteins in a pathophysiological state. Differential expression analysis of sperm proteins in infertile men and bioinformatics analysis offer information about their involvement in biological pathways.<h4>Materials and methods</h4>Literature search was performed on PubMed, Medline, and Science Direct databases using the keywords 'sperm proteomics' and 'male infertility'. We also reviewed the relevant cross references of retrieved articles and included them in the review process. Articles written in any language other than English were excluded.<h4>Results</h4>Of 575 articles identified, preliminary screening for relevant studies eliminated 293 articles. At the next level of selection, from 282 studies only 80 articles related to male infertility condition met the selection criteria and were included in this review.<h4>Conclusion</h4>In this molecular era, sperm proteomics has created a platform for enhanced understanding of male reproductive physiology as a potential tool for identification of novel protein biomarkers related to sperm function in infertile men. Therefore, it is believed that proteomic biomarkers can overcome the gaps in information from conventional semen analysis that are of limited clinical utility.

Fluorescently encoded microbeads are in demand for multiplexed applications in different fields. Compared to organic dye-based commercially available Luminex's xMAP technology, upconversion nanoparticles (UCNPs) are better alternatives due to their large anti-Stokes shift, photostability, nil background, and single wavelength excitation. Here, we developed a new multiplexed detection system using UCNPs for encoding poly(ethylene glycol) diacrylate (PEGDA) microbeads as well as for labeling reporter antibody. However, to prepare UCNPs-encoded microbeads, currently used swelling-based encapsulation leads to non-uniformity, which is undesirable for fluorescence-based multiplexing. Hence, we utilized droplet microfluidics to obtain encoded microbeads of uniform size, shape, and UCNPs distribution inside. Additionally, PEGDA microbeads lack functionality for probe antibodies conjugation on their surface. Methods to functionalize the surface of PEGDA microbeads (acrylic acid incorporation, polydopamine coating) reported thus far quench the fluorescence of UCNPs. Here, PEGDA microbeads surface was coated with silica followed by carboxyl modification without compromising the fluorescence intensity of UCNPs. In this study, droplet microfluidics-assisted UCNPs-encoded microbeads of uniform shape, size, and fluorescence were prepared. Multiple color codes were generated by mixing UCNPs emitting red and green colors at different ratios prior to encapsulation. UCNPs emitting blue color were used to label the reporter antibody. Probe antibodies were covalently immobilized on red UCNPs-encoded microbeads for specific capture of human serum albumin (HSA) as a model protein. The system was also demonstrated for multiplexed detection of both human C-reactive protein (hCRP) and HSA protein by immobilizing anti-hCRP antibodies on green UCNPs.

Direct reprogramming of other somatic cells into neurons is an alternative strategy for the recovery of an injured nervous system. In this work, we developed a new non-viral gene carrier based on Porphyra yezoensis polysaccharide (PYP). After modification with ethylenediamine, the cationized PYP (Ed-PYP) was combined with plasmids encoding Ascl1, Brn2 and FoxA1 to form spherical nanoscale particles (Ed-PYP-pABF nanoparticles). Cytotoxicity assays proved that Ed-PYP-pABF nanoparticles had a better safety profile than Lipofectamine 2000 and polyetherimide. Characterization tests illustrated that the Ed-PYP-pABF nanoparticles at an Ed-PYP:pABF weight ratio of 40:1 is a potential candidate for gene delivery, which was further supported by Western blot and plasmid encoding enhanced green fluorescence protein transfection. Based on this transfection strategy, we co-delivered pABF to 3T6 cells using Ed-PYP. ELISA indicated that the levels of brain-derived neurotrophic factor, nerve growth factors and sonic hedgehog reached a maximum at 14 days after the last transfection. Immunofluorescence and Western blot further exhibited positive expression of neurofilament 200, Nestin, glial fibrillary acidic protein, growth associated protein-43, β-3tubulin, and microtubule associated protein 2, proving the successful conversion of 3T6 cells into neurons. Taken together, these results illustrated that a natural polysaccharide-based gene co-delivery system is a promising strategy for neural reprogramming.

<h4>Background</h4>Newcastle disease virus (NDV) is a threat to poultry production worldwide. A better understanding of mechanisms of resistance and susceptibility to this virus will improve measures for NDV prevention and control. Males and females from resistant Fayoumi and susceptible Leghorn lines were either challenged with a lentogenic strain of the virus or given a mock infection at 3 weeks of age. The lung transcriptomes generated by RNA-seq were studied using contrasts across the challenged and nonchallenged birds, the two lines, and three time points post-infection, and by using Weighted Gene Co-expression Network Analysis (WGNCA).<h4>Results</h4>Genetic line and sex had a large impact on the lung transcriptome. When contrasting the challenged and nonchallenged birds, few differentially expressed genes (DEG) were identified within each line at 2, 6, and 10 days post infection (dpi), except for the more resistant Fayoumi line at 10 dpi, for which several pathways were activated and inhibited at this time. The interaction of challenge and line at 10 dpi significantly impacted 131 genes (False Discovery Rate (FDR) <0.05), one of which was PPIB. Many DEG were identified between the Fayoumi and Leghorns. The number of DEG between the two lines in the challenged birds decreased over time, but increased over time in the nonchallenged birds. The nonchallenged Fayoumis at 10 dpi showed enrichment of immune type cells when compared to 2 dpi, suggesting important immune related development at this age. These changes between 10 and 2 dpi were not identified in the challenged Fayoumis. The energy allocated to host defense may have interrupted normal lung development. WGCNA identified important modules and driver genes within those modules that were associated with traits of interest, several of which had no known associated function.<h4>Conclusions</h4>The lines' unique response to NDV offers insights into the potential means of their resistance and susceptibility. The lung transcriptome shows a unique response to lentogenic NDV compared to a previous study on the trachea of the same birds. It is important to analyze multiple tissues in order to best understand the chicken's overall response to NDV challenge and improve strategies to combat this devastating disease.

The expression patterns of endogenous circular RNA (circRNA) molecules during epidermal stem cell (EpSC) differentiation have not previously been explored. Here, we show that circRNAs are abundantly expressed in EpSCs and that their expression change dramatically during differentiation in a coordinated manner. Overall, circRNAs are expressed at higher levels in the differentiated cells, and many upregulated circRNAs are derived from developmental genes, including four different circRNAs from DLG1. The observed changes in circRNA expression were largely independent of host gene expression, and circRNAs independently upregulated upon differentiation are more prone to AGO2 binding and have more predicted miRNA binding sites compared to stably expressed circRNAs. In particular, upregulated circRNAs from the HECTD1 and ZNF91 genes have exceptionally high numbers of AGO2 binding sites and predicted miRNA target sites, and circZNF91 contains 24 target sites for miR-23b-3p, which is known to play important roles in keratinocyte differentiation. We also observed that upregulated circRNAs are less likely to be flanked by homologues inverted Alu repeats compared to stably expressed circRNAs. This coincide with DHX9 being upregulated in the differentiated keratinocytes. Finally, none of the circRNAs upregulated upon differentiation were also upregulated upon DNMT3A or DNMT3B knockdown, making it unlikely that epigenetic mechanisms are governing the observed circRNA expression changes. Together, we provide a map of circRNA expression in EpSCs and their differentiated counterparts and shed light on potential function and regulation of differentially expressed circRNAs.

<i>Staphylococcus aureus</i> is the most common contagious pathogen associated with bovine subclinical mastitis. Current diagnosis of <i>S. aureus</i> mastitis is based on bacteriological culture of milk samples and somatic cell counts, which lack either sensitivity or specificity. Identification of milk proteins that contribute to host defense and their variable responses to pathogenic stimuli would enable the characterization of putative biomarkers of subclinical mastitis. To accomplish this, milk whey samples from healthy and mastitic dairy cows were analyzed using a label-free quantitative proteomics approach. In total, 90 proteins were identified, of which 25 showed significant differential abundance between healthy and mastitic samples. In silico functional analyses indicated the involvement of the differentially abundant proteins in biological mechanisms and signaling pathways related to host defense including pathogen-recognition, direct antimicrobial function, and the acute-phase response. This proteomics and bioinformatics analysis not only facilitates the identification of putative biomarkers of <i>S. aureus</i> subclinical mastitis but also recapitulates previous findings demonstrating the abundance of host defense proteins in intramammary infection. All mass spectrometry data are available via ProteomeXchange with identifier <i>PXD007516</i>.

The diversity of processed transcripts in eukaryotic genomes poses a challenge for the classification of their biological functions. Sparse sequence conservation in non-coding sequences and the unreliable nature of RNA structure predictions further exacerbate this conundrum. Here, we describe a computational method, DotAligner, for the unsupervised discovery and classification of homologous RNA structure motifs from a set of sequences of interest. Our approach outperforms comparable algorithms at clustering known RNA structure families, both in speed and accuracy. It identifies clusters of known and novel structure motifs from ENCODE immunoprecipitation data for 44 RNA-binding proteins.

To develop a catalog of regulatory sites in two major model organisms, <i>Drosophila melanogaster</i> and <i>Caenorhabditis elegans</i>, the modERN (model organism Encyclopedia of Regulatory Networks) consortium has systematically assayed the binding sites of transcription factors (TFs). Combined with data produced by our predecessor, modENCODE (Model Organism ENCyclopedia Of DNA Elements), we now have data for 262 TFs identifying 1.23 M sites in the fly genome and 217 TFs identifying 0.67 M sites in the worm genome. Because sites from different TFs are often overlapping and tightly clustered, they fall into 91,011 and 59,150 regions in the fly and worm, respectively, and these binding sites span as little as 8.7 and 5.8 Mb in the two organisms. Clusters with large numbers of sites (so-called high occupancy target, or HOT regions) predominantly associate with broadly expressed genes, whereas clusters containing sites from just a few factors are associated with genes expressed in tissue-specific patterns. All of the strains expressing GFP-tagged TFs are available at the stock centers, and the chromatin immunoprecipitation sequencing data are available through the ENCODE Data Coordinating Center and also through a simple interface (http://epic.gs.washington.edu/modERN/) that facilitates rapid accessibility of processed data sets. These data will facilitate a vast number of scientific inquiries into the function of individual TFs in key developmental, metabolic, and defense and homeostatic regulatory pathways, as well as provide a broader perspective on how individual TFs work together in local networks and globally across the life spans of these two key model organisms.

Neuronal stimulation leads to immediate early gene (IEG) expression through calcium-dependent mechanisms. In recent years, considerable attention has been devoted to the transcriptional responses after neuronal stimulation, but relatively little is known about the changes in chromatin dynamics that follow neuronal activation. Here, we use fluorescence recovery after photobleaching, biochemical fractionations, and chromatin immunoprecipitation to show that KCl-induced depolarization in primary cultured cortical neurons causes a rapid release of the linker histone H1 from chromatin, concomitant with IEG expression. H1 release is repressed by PARP inhibition, PARP1 deletion, a non-PARylatable H1, as well as phosphorylation inhibitions and a nonphosphorylatable H1, leading to hindered IEG expression. Further, H1 is replaced by PARP1 on IEG promoters after neuronal stimulation, and PARP inhibition blocks this reciprocal binding response. Our results demonstrate the relationship between neuronal excitation and chromatin plasticity by identifying the roles of polyadenosine diphosphate ribosylation and phosphorylation of H1 in regulating H1 chromatin eviction and IEG expression in stimulated neurons.

Acute myeloid leukemia (AML) is the most common and severe form of acute leukemia diagnosed in adults. Owing to its heterogeneity, AML is divided into classes associated with different treatment outcomes and specific gene expression profiles. Based on previous studies on AML, in this study, we designed and generated an AML-array containing 900 oligonucleotide probes complementary to human genes implicated in hematopoietic cell differentiation and maturation, proliferation, apoptosis and leukemic transformation. The AML-array was used to hybridize 118 samples from 33 patients with AML of the M1 and M2 subtypes of the French-American‑British (FAB) classification and 15 healthy volunteers (HV). Rigorous analysis of the microarray data revealed that 83 genes were differentially expressed between the patients with AML and the HV, including genes not yet discussed in the context of AML pathogenesis. The most overexpressed genes in AML were STMN1, KITLG, CDK6, MCM5, KRAS, CEBPA, MYC, ANGPT1, SRGN, RPLP0, ENO1 and SET, whereas the most underexpressed genes were IFITM1, LTB, FCN1, BIRC3, LYZ, ADD3, S100A9, FCER1G, PTRPE, CD74 and TMSB4X. The overexpression of the CPA3 gene was specific for AML with mutated NPM1 and FLT3. Although the microarray-based method was insufficient to differentiate between any other AML subgroups, quantitative PCR approaches enabled us to identify 3 genes (ANXA3, S100A9 and WT1) whose expression can be used to discriminate between the 2 studied AML FAB subtypes. The expression levels of the ANXA3 and S100A9 genes were increased, whereas those of WT1 were decreased in the AML-M2 compared to the AML-M1 group. We also examined the association between the STMN1, CAT and ABL1 genes, and the FLT3 and NPM1 mutation status. FLT3+/NPM1- AML was associated with the highest expression of STMN1, and ABL1 was upregulated in FLT3+ AML and CAT in FLT3- AML, irrespectively of the NPM1 mutation status. Moreover, our results indicated that CAT and WT1 gene expression levels correlated with the response to therapy. CAT expression was highest in patients who remained longer under complete remission, whereas WT1 expression increased with treatment resistance. On the whole, this study demonstrates that the AML-array can potentially serve as a first-line screening tool, and may be helpful for the diagnosis of AML, whereas the differentiation between AML subgroups can be more successfully performed with PCR-based analysis of a few marker genes.

<h4>Aim</h4>Inspiratory muscle (diaphragm) function declines with age, contributing to exercise intolerance and impaired airway clearance. Studies of diaphragm dysfunction in rodents have focused on moderate aging (~24months); thus, the impact of advanced age on the diaphragm and potential mechanisms of dysfunction are less clear. Therefore, we aimed to define the effects of advanced age on the mechanics, morphology, and global and redox proteome of the diaphragm.<h4>Methods</h4>We studied diaphragm from young (6months) and very old male mice (30months). Diaphragm function was evaluated using isolated muscle bundles. Proteome analyses followed LC-MS/MS processing of diaphragm muscle.<h4>Results</h4>Advanced aging decreased diaphragm peak power by ~35% and maximal isometric specific force by ~15%, and prolonged time to peak twitch tension by ~30% (P<0.05). These changes in contractile properties were accompanied, and might be caused by, decreases in abundance of calsequestrin, sarcoplasmic reticulum C^a2+-ATPase, sarcalumenin, and parvalbumin that were revealed by our label-free proteomics data. Advanced aging also increased passive stiffness (P<0.05), which might be a consequence of an upregulation of cytoskeletal and extracellular matrix proteins identified by proteomics. Analyses of cysteine redox state indicated that the main diaphragm abnormalities with advanced aging are in metabolic enzymes and mitochondrial proteins.<h4>Conclusion</h4>Our novel findings are that the most pronounced impact of advanced aging on the diaphragm is loss of peak power and disrupted cysteine redox homeostasis in metabolic enzymes and mitochondrial proteins.

Mechanisms of selective autophagy of the ER, known as ER-phagy, require molecular delineation, particularly in vivo. It is unclear how these events control ER proteostasis and cellular health. Here, we identify cell-cycle progression gene 1 (CCPG1), an ER-resident protein with no known physiological role, as a non-canonical cargo receptor that directly binds to core autophagy proteins via an LIR motif to mammalian ATG8 proteins and, independently and via a discrete motif, to FIP200. These interactions facilitate ER-phagy. The CCPG1 gene is inducible by the unfolded protein response and thus directly links ER stress to ER-phagy. In vivo, CCPG1 protects against ER luminal protein aggregation and consequent unfolded protein response hyperactivation and tissue injury of the exocrine pancreas. Thus, via identification of this autophagy protein, we describe an unexpected molecular mechanism of ER-phagy and provide evidence that this may be physiologically relevant in ER luminal proteostasis.

<h4>Background</h4>Gray whale, Eschrichtius robustus (E. robustus), is a single member of the family Eschrichtiidae, which is considered to be the most primitive in the class Cetacea. Gray whale is often described as a "living fossil". It is adapted to extreme marine conditions and has a high life expectancy (77 years). The assembly of a gray whale genome and transcriptome will allow to carry out further studies of whale evolution, longevity, and resistance to extreme environment.<h4>Results</h4>In this work, we report the first de novo assembly and primary analysis of the E. robustus genome and transcriptome based on kidney and liver samples. The presented draft genome assembly is complete by 55% in terms of a total genome length, but only by 24% in terms of the BUSCO complete gene groups, although 10,895 genes were identified. Transcriptome annotation and comparison with other whale species revealed robust expression of DNA repair and hypoxia-response genes, which is expected for whales.<h4>Conclusions</h4>This preliminary study of the gray whale genome and transcriptome provides new data to better understand the whale evolution and the mechanisms of their adaptation to the hypoxic conditions.

The duplication of 5' segment of <i>KMT2A</i> is a rare molecular event in childhood leukemia, and the influence on prognosis is unknown. Here, we report on a boy who developed acute monocytic leukemia. Fluorescence in situ hybridization revealed the duplication of the 5' segment with 2 normal alleles at <i>KMT2A</i> which was eventually found to be fused with <i>MLLT10</i>. Chemotherapy promptly induced the first complete remission in the patient at our facility, and the patient remained in first complete remission with negative minimal residual disease at 3.5 years from diagnosis. Our case is similar to two previously reported patients who had partial duplication of the 5' segment of <i>KMT2A</i> with a <i>KMT2A</i>-<i>MLLT10</i> rearrangement. Further studies and experience with this cryptic translocation may shed more light on the management of acute myeloid leukemia.

Hereditary antithrombin (AT) deficiency is an autosomal-dominant disorder predisposing to venous and arterial thrombosis. Homozygosity resulting in severe AT deficiency is not compatible with life, apart from homozygous mutations affecting the heparin-binding site representing the most severe thrombophilia.<h4>Patients and methods</h4>A 12-year-old previously healthy boy of Romani origin presented with a swollen, painful left leg and fever. Imaging revealed signs of inferior vena cava (IVC) thrombosis, the presence of interrupted intrahepatic IVC with azygos continuation and bilateral iliofemoral thrombosis with enlargement of the azygous and hemiazygos venous system. In addition, right pleural effusion and signs of bilateral renal pericortical cysts, possibly caused by retroperitoneal lymphangiectasia, were disclosed. Thrombophilia screening involving protein C, Protein S, Antithrombin (chromogenic assays based on the inhibition of FIIa and FXa, antigen concentration), APC resistance, prothrombin mutation and Lupus anticoagulants was performed using standard laboratory methods. Genetic analysis of the SERPINC1 gene was done by direct sequencing.<h4>Results</h4>Thrombophilia screening showed isolated decreased AT activity at 21% (RR 80-120%). AT levels were retested and remained decreased (33-36%) on two consecutive occasions. SERPINC1 gene analysis revealed a previously described homozygous mutation (Budapest III) causing type IIB deficiency (c.391C>T; p.Leu131Phe). A family study confirmed the same mutation in both parents and three siblings. The patient improved significantly following warfarin therapy and over the past 2.5 years did not experience new thromboembolism.<h4>Conclusions</h4>This case represents a rare combination of two risk factors provoking manifestation of spontaneous venous thromboembolism at a young age requiring permanent anticoagulant therapy.

There are various types of receptors in the central nervous system (CNS). G protein-coupled receptors (GPCRs) have the highest expression with a wide range of physiological functions. A newer sub group of these receptors namely orphan GPCRs have been discovered. GPR3, GPR6, GPR17, GPR26, GPR37, GPR39, GPR40, GPR50, GPR52, GPR54, GPR55, GPR85, GPR88, GPR103, and GPR139 are the selected orphan GPCRs for this article. Their roles in the central nervous system have not been understood well so far. However, recent studies show that they may have very important functions in the CNS. Hence, in the present study, we reviewed most recent findings regarding the physiological roles of the selected orphan GPCRs in the CNS. After a brief presentation of each receptor, considering the results from genetic and pharmacological manipulation of the receptors, their roles in the pathophysiology of different diseases and disorders including anxiety, depression, schizophrenia, epilepsy, Alzheimer's disease, Parkinson's disease, and substance abuse will be discussed. At present, our knowledge regarding the role of GPCRs in the brain is very limited. However, previous limited studies show that orphan GPCRs have an important place in psychopharmacology and these receptors are potential new targets for the treatment of major CNS diseases.

Objective Since the majority of direct-acting antivirals (DAAs) that are used in the treatment of hepatitis C virus (HCV) infection are mainly metabolized by CYP3A4, it is hypothesized that inter-individual differences in CYP3A4 activity may be associated with the bioavailability of these agents. Methods The level of serum 4β-hydroxycholesterol (4βHC), a surrogate marker of CYP3A4 activity, was determined by LC-MS/MS in samples obtained from patients with HCV infection (CHCs) as well as healthy control subjects (CTLs). Serum samples obtained from patients treated with either asunaprevir/daclatasvir (ASV/DCV) or ombitasvir/paritaprevir/ritonavir (OTV/PTV/r) were used for additional assays. Results The serum 4βHC level in CHCs was significantly higher than that in CTLs, and a gender difference was seen among CHCs. In patients treated with OTV/PTV/r, the serum 4βHC level was observed to gradually decrease during the treatment period. In the cohort treated with ASV/DCV, 4 of 83 patients showed virological treatment failure. In pretreatment testing, an Invader assay detected a low prevalence of resistance-associated variants in these four patients. The average serum concentration of DCV/ASV in the treatment-failed group tended to be lower than that in the sustained virological response (SVR) group. The pretreatment serum 4βHC level in patients with treatment failure was significantly higher than that in patients with an SVR but in whom the prevalence of resistance-associated variants was low in the pretreatment setting. Conclusion The evaluation of CYP3A4 activity by measuring 4βHC before treatment may provide additional information that can potentially be used to select cost- and efficacy-optimized treatment of HCV.

The fabrication and performance of mechanically responsive multilayer superhydrophobic composites are reported. The application of tensile strain triggers the release of small molecules and proteins from these composites, with different tensile strain magnitudes and coating thickness influencing agent release. These mechanoresponsive composites consist of an absorbent drug core surrounded by an electrosprayed superhydrophobic protective coating that limits drug release in the absence of tensile strain. Coating thickness and applied tensile strain control release of chemotherapeutic cisplatin and enzyme β-galactosidase, as measured by atomic absorption and UV-vis spectrophotometry, respectively, with preserved in vitro activity. Such mechanically responsive drug delivery devices, when coupled to existing dynamic mechanical forces in the body or integrated with mechanical medical devices, such as stents, will provide local controlled dosing.

Dumps of a mining-metallurgical complex of post-Soviet Republics have accumulated a huge amount of technogenic waste products. Out of them, Kazakhstan alone has preserved about 20 billion tons. In the field of technogenic waste treatment, there is still no technical solution that leads it to be a profitable process. Recent global trends prompted scientists to focus on developing energy-saving and a highly efficient melting unit that can significantly reduce specific fuel consumption. This paper reports, the development of a new technological method-smelt layer of inversion phase. The introducing method is characterized by a combination of ideal stirring and ideal displacement regimes. Using the method of affine modelling, recalculation of pilot plant's test results on industrial sample has been obtained. Experiments show that in comparison with bubbling and boiling layers of smelt, the degree of zinc recovery increases in the layer of inversion phase. That indicates the reduction of the possibility of new formation of zinc silicates and ferrites from recombined molecules of ZnO, SiO2, and Fe2O3. Calculations show that in industrial samples of the pilot plant, the consumption of natural gas has reduced approximately by two times in comparison with fuming-furnace. The specific fuel consumption has reduced by approximately four times in comparison with Waelz-kiln.

Aggregation of huntingtin protein arising from expanded polyglutamine (polyQ) sequences in the exon-1 region of mutant huntingtin plays a central role in the pathogenesis of Huntington's disease. The huntingtin aggregation pathways are of therapeutic and diagnostic interest, but obtaining critical information from the physiologically relevant htt exon-1 (Httex1) protein has been challenging. Using biophysical techniques and an expression and purification protocol that generates clean, monomeric Httex1, we identified and mapped three distinct aggregation pathways: 1) unseeded in solution; 2) seeded in solution; and 3) membrane-mediated. In solution, aggregation proceeded in a highly stepwise manner, in which the individual domains (N terminus containing 17 amino acids (N17), polyQ, and proline-rich domain (PRD)) become ordered at very different rates. The aggregation was initiated by an early oligomer requiring a pathogenic, expanded Gln length and N17 α-helix formation. In the second phase, β-sheet forms in the polyQ. The slowest step is the final structural maturation of the PRD. This stepwise mechanism could be bypassed by seeding, which potently accelerated aggregation and was a prerequisite for prion-like spreading <i>in vivo</i> Remarkably, membranes could catalyze aggregation even more potently than seeds, in a process that caused significant membrane damage. The N17 governed membrane-mediated aggregation by anchoring Httex1 to the membrane, enhancing local concentration and promoting collision via two-dimensional diffusion. Considering its central roles in solution and in membrane-mediated aggregation, the N17 represents an attractive target for inhibiting multiple pathways. Our approach should help evaluate such inhibitors and identify diagnostic markers for the misfolded forms identified here.

Infected or transformed cells must present peptides derived from endogenous proteins on MHC class I molecules to be recognized and targeted for elimination by Ag-specific cytotoxic T cells. In the first step of peptide generation, proteins are degraded by the proteasome. In this study, we investigated the role of the ubiquitin-specific protease 14 (Usp14), a proteasome-associated deubiquitinase, in direct Ag presentation using a ligand-stabilized model protein expressed as a self-antigen. Chemical inhibition of Usp14 diminished direct presentation of the model antigenic peptide, and the effect was especially pronounced when presentation was restricted to the defective ribosomal product (DRiP) form of the protein. Additionally, presentation specifically from DRiP Ags was diminished by expression of a catalytically inactive form of Usp14. Usp14 inhibition did not appreciably alter protein synthesis and only partially delayed protein degradation as measured by a slight increase in the half-life of the model protein when its degradation was induced. Taken together, these data indicate that functional Usp14 enhances direct Ag presentation, preferentially of DRiP-derived peptides, suggesting that the processing of DRiPs is in some ways different from other forms of Ag.

<h4>Objective</h4>To investigate the feasibility of microRNA (miRNA) levels in CSF as biomarkers for prodromal Huntington disease (HD).<h4>Methods</h4>miRNA levels were measured in CSF from 60 PREDICT-HD study participants using the HTG protocol. Using a CAG-Age Product score, 30 prodromal HD participants were selected based on estimated probability of imminent clinical diagnosis of HD (i.e., low, medium, high; n = 10/group). For comparison, participants already diagnosed (n = 15) and healthy controls (n = 15) were also selected.<h4>Results</h4>A total of 2,081 miRNAs were detected and 6 were significantly increased in the prodromal HD gene expansion carriers vs controls at false discovery rate q < 0.05 (miR-520f-3p, miR-135b-3p, miR-4317, miR-3928-5p, miR-8082, miR-140-5p). Evaluating the miRNA levels in each of the HD risk categories, all 6 revealed a pattern of increasing abundance from control to low risk, and from low risk to medium risk, which then leveled off from the medium to high risk and HD diagnosed groups.<h4>Conclusions</h4>This study reports miRNAs as CSF biomarkers of prodromal and diagnosed HD. Importantly, miRNAs were detected in the prodromal HD groups furthest from diagnosis where treatments are likely to be most consequential and meaningful. The identification of potential biomarkers in the disease prodrome may prove useful in evaluating treatments that may postpone disease onset.<h4>Clinicaltrialsgov identifier</h4>NCT00051324.

Synapsins serve as flagships among the presynaptic proteins due to their abundance on synaptic vesicles and contribution to synaptic communication. Several studies have emphasized the importance of this multi-gene family of neuron-specific phosphoproteins in maintaining brain physiology. In the recent times, increasing evidence has established the relevance of alterations in synapsins as a major determinant in many neurological disorders. Here, we give a comprehensive description of the diverse roles of the synapsin family and the underlying molecular mechanisms that contribute to several neurological disorders. These physiologically important roles of synapsins associated with neurological disorders are just beginning to be understood. A detailed understanding of the diversified expression of synapsins may serve to strategize novel therapeutic approaches for these debilitating neurological disorders.

Spatiotemporal control over the release or activation of biomacromolecules such as siRNA remains a significant challenge. Light-controlled release has gained popularity in recent years; however, a major limitation is that most photoactivable compounds/systems respond only to UV irradiation, but not near-infrared (NIR) light that offers a deeper tissue penetration depth and better biocompatibility. This paper reports a simple NIR-to-UV upconversion nanoparticle (UCNP)-based siRNA nanocarrier for NIR-controlled gene silencing. siRNA is complexed onto a NaYF₄:Yb/Tm/Er UCNP through an azobenzene (Azo)-cyclodextrin (CD) host-guest interaction. The UV emission generated by the NIR-activated UCNP effectively triggers the trans-to-cis photoisomerization of azobenzene, thus leading to the release of siRNA due to unmatched host-guest pairs. The UCNP-siRNA complexes are also functionalized with PEG (i.e., UCNP-(CD/Azo)-siRNA/PEG NPs), targeting ligands (i.e., EGFR-specific GE11 peptide), acid-activatable cell-penetrating peptides (i.e., TH peptide), and imaging probes (i.e., Cy5 fluorophore). The UCNP-(CD/Azo)-siRNA/PEG NPs with both GE11 and TH peptides display a high level of cellular uptake and an excellent endosomal/lysosomal escape capability. More importantly, NIR-controlled spatiotemporal knockdown of GFP expression is successfully achieved in both a 2D monolayer cell model and a 3D multicellular tumor spheroid model. Thus, this simple and versatile nanoplatform has great potential for the selective activation or release of various biomacromolecules.

The rapid expansion of flow cytometry applications has outpaced the functionality of traditional manual analysis tools used to interpret flow cytometry data. Scientists are faced with the daunting prospect of manually identifying interesting cell populations in 50-dimensional datasets, equalling the complexity previously only reached in mass cytometry. Data can no longer be analyzed or interpreted fully by manual approaches. While automated gating has been the focus of intense efforts, there are many significant additional steps to the analytical pipeline (e.g., cleaning the raw files, event outlier detection, extracting immunophenotypes). We review the components of a customized automated analysis pipeline that can be generally applied to large scale flow cytometry data. We demonstrate these methodologies on data collected by the International Mouse Phenotyping Consortium (IMPC).

Nonalcoholic fatty liver disease (NAFLD) is the most prevalent chronic liver disease in the world. The complete natural history of NAFLD is unknown because few high-quality follow-up studies have been conducted. Our aim was to find variables predicting disease severity through an extended follow-up with serial biopsies. In a prospective cohort study, 129 patients who enrolled between 1988 and 1993 were asked to participate in a follow-up study on two occasions; biochemical, clinical, and histologic data were documented. The mean time between biopsies was 13.7 (±1.7) and 9.3 (±1.0) years, respectively. At the end of the study period, 12 patients (9.3%) had developed end-stage liver disease and 34% had advanced fibrosis. Out of the 113 patients with baseline low fibrosis (<3), 16% developed advanced fibrosis. Fibrosis progression did not differ among the different stages of baseline fibrosis (<i>P</i> = 0.374). Fifty-six patients (43%) had isolated steatosis, of whom 9% developed advanced fibrosis (3 patients with biopsy-proven fibrosis stage F3-F4 and 2 patients with end-stage liver disease). Fibrosis stage, ballooning, and diabetes were more common in patients who developed end-stage liver disease; however, there were no baseline clinical, histologic, or biochemical variables that predicted clinical significant disease progression. <i>Conclusion</i>: NAFLD is a highly heterogeneous disease, and it is surprisingly hard to predict fibrosis progression. Given enough time, NAFLD seems to have a more dismal prognosis then previously reported, with 16% of patients with fibrosis stage <3 developing advanced fibrosis and 9.3% showing signs of end-stage liver disease. (<i>Hepatology Communications</i> 2018;2:199-210).

Sirtuins and their pharmacological activators/inhibitors have been associated with a range of neuroprotective effects or disease modifying influences in neurological disorders. Huntington's disease (HD) is an autosomal-dominant, progressive neurodegenerative disease characterized by movement disorder, psychiatric symptoms and cognitive decline. The monogenic mutation in HD encodes a variant of the protein Huntingtin (HTT). The disease is a consequence of a CAG repeat extension leading to an abnormally long polyglutamine (Q) stretch at HTT's N-terminus, which likely confers a toxic gain of function to the mutant polypeptide. HD has currently no effective disease-modifying therapy or preventive measures. In the past 2decades, a sizable body of work on Sirtuins' modification of HD pathology using HD cell and animal models has accumulated. In this chapter, evidence for Sirtuin activities as potential modifiers of HD pathology is reviewed. The conflicting findings of the impacts of mammalian Sirtuin paralogs on HD pathogenesis and disease progression are highlighted. The possible cellular and molecular mechanisms underlying Sirtuin activities in HD are discussed with reference to pathophysiological mechanisms of transcription perturbation, proteostasis, mitochondrial function, and microtubule dynamics. A brief therapeutic perspective on the use of Sirtuin activators and inhibitors is also presented.

Cancer prevention using natural micronutrition on epigenetic mechanisms primarily revolves around plant extracts. However, the role of macronutrition, including animal peptides, on epigenetic modification in cancer has been elusive. In traditional Chinese medicine, the soft-shelled turtle has a long-history of being a functional food that strengthens immunity through unknown mechanisms. The present study aimed to investigate the impact of soft-shelled turtle peptide on microRNA (miRNA) expression in gastric cancer (GC) cells and to analyze the potential anticancer mechanisms for GC. Affymetrix GeneChip miRNA 3.0 Array and quantitative polymerase chain reaction were used to detect the miRNA expression profile in human GC AGS cells treated with the soft-shelled turtle peptide. The results demonstrated that 101 miRNAs (49 upregulated miRNAs and 52 downregulated miRNAs) were significantly differentially expressed in the AGS cells following soft-shelled turtle peptide treatment. Several tumor suppressor miRNAs were upregulated markedly, including miRNA-375, let-7d, miRNA-429, miRNA-148a/148b and miRNA-34a. Pathway analysis indicated that soft-shelled turtle peptide may function with anticancer properties through the Hippo signaling pathway and the forkhead box O signaling pathway. Therefore, these results demonstrated that soft-shelled turtle peptide has the capacity to influence cancer-related pathways through the regulation of miRNA expression in GC cells.

The discovery of the causative gene for Huntington's disease (HD) has promoted numerous efforts to uncover cellular pathways that lower levels of mutant huntingtin protein (mHtt) and potentially forestall the appearance of HD-related neurological defects. Using a cell-based model of pathogenic huntingtin expression, we identified a class of compounds that protect cells through selective inhibition of a lipid kinase, PIP4Kγ. Pharmacological inhibition or knock-down of PIP4Kγ modulates the equilibrium between phosphatidylinositide (PI) species within the cell and increases basal autophagy, reducing the total amount of mHtt protein in human patient fibroblasts and aggregates in neurons. In two <i>Drosophila</i> models of Huntington's disease, genetic knockdown of PIP4K ameliorated neuronal dysfunction and degeneration as assessed using motor performance and retinal degeneration assays respectively. Together, these results suggest that PIP4Kγ is a druggable target whose inhibition enhances productive autophagy and mHtt proteolysis, revealing a useful pharmacological point of intervention for the treatment of Huntington's disease, and potentially for other neurodegenerative disorders.

Genome amplification in the form of rings or giant rod-shaped marker chromosomes (RGMs) is a common genetic alteration in soft tissue tumors. The mitotic stability of these structures is often rescued by perfectly functioning analphoid neocentromeres, which therefore significantly contribute to cancer progression. Here, we disentangled the genomic architecture of many neocentromeres stabilizing marker chromosomes in well-differentiated liposarcoma and lung sarcomatoid carcinoma samples. In cells carrying heavily rearranged RGMs, these structures were assembled as patchworks of multiple short amplified sequences, disclosing an extremely high level of complexity and definitely ruling out the existence of regions prone to neocentromere seeding. Moreover, by studying two well-differentiated liposarcoma samples derived from the onset and the recurrence of the same tumor, we documented an expansion of the neocentromeric domain that occurred during tumor progression, which reflects a strong selective pressure acting toward the improvement of the neocentromeric functionality in cancer. In lung sarcomatoid carcinoma cells we documented, extensive "centromere sliding" phenomena giving rise to multiple, closely mapping neocentromeric epialleles on separate coexisting markers occur, likely due to the instability of neocentromeres arising in cancer cells. Finally, by investigating the transcriptional activity of neocentromeres, we came across a burst of chimeric transcripts, both by extremely complex genomic rearrangements, and <i>cis</i>/<i>trans</i>-splicing events. Post-transcriptional editing events have been reported to expand and variegate the genetic repertoire of higher eukaryotes, so they might have a determining role in cancer. The increased incidence of fusion transcripts, might act as a driving force for the genomic amplification process, together with the increased transcription of oncogenes.

<h4>Background</h4>Alchemilla mollis (lady's mantle) is a common ingredient in skin care products. However, the protective mechanism of A. mollis against skin problems has not been elucidated.<h4>Purpose</h4>This study was to investigate the effects of A. mollis ethanolic extract (AM) on UVB-irradiated normal human dermal fibroblasts (NHDF) and hairless mice.<h4>Methods</h4>The in vitro anti-photoaging effect of AM was performed in NHDFs. The antioxidant activities were assessed through DPPH, ABTS, and reactive oxygen species (ROS) assays. Matrix metalloproteinase 1 (MMP-1), IL-6, procollagen type I, and transforming growth factor-β1 (TGF-β1) were measured by kits. The protein levels of p-c-Jun, p-c-Fos, Nrf2, NQO-1, HO-1, nuclear NFATc1 and cytosolic p-NFATc1 were evaluated by western blotting. In in vivo, H&E and Masson's trichrome staining were carried out. Skin texture was analyzed using the roughness parameters. The expression of MMP-1, procollagen type I, TGF-β1 and elastin were measured by western blot.<h4>Results</h4>AM included gallic acid as an active constituent. AM exhibited a strong antioxidant effect by inhibiting DPPH and ABTS free radicals, as well as ROS production. It was also found to upregulate transforming growth factor β1, type I procollagen and elastin expression, and to downregulate matrix metalloproteinase-1 and interleukin-6 expression in AM-treated NHDFs under UVB irradiation. These effects were attributed to AP-1 and Nrf2/ARE signaling pathways. Significantly, it was demonstrated that AM regulated the UVB-induced NFATc1 dephosphorylation in nucleus. Based on dietary data, AM was effective for the prevention of wrinkle formation, skin thickening, water loss, and erythema in UVB-exposed mouse skin.<h4>Conclusion</h4>Our data indicate that A. mollis provides protection from UVB exposure in both hairless mice skin in vivo and NHDFs in vitro. AM might therefore be useful as a cosmetic material and functional food for the prevention of UVB-induced human skin photoaging.

Electroacupuncture (EA) can relieve various pains. However, its mechanism in terms of the transcriptome is still not well-known. To explore the full profile of EA-induced molecular modification in the central nerve system, three twins of goats were selected for a match-paired experiment: EA stimulation (60 Hz, 30 min) and none-EA (control). Goats in the EA group showed an increased (<i>p</i> < 0.05) nociceptive threshold compared with the control goats. Experimental goats were sacrificed at 4 h of the experiment, and the periaqueductal grays were harvested for RNA sequencing. As a result, 2651 differentially expressed genes (1803 up-regulated and 848 down-regulated genes) were found and enriched in 30 Kyoto Encyclopedia of Genes and Genomes pathways and 149 gene ontology terms. EA-regulated five neuropeptide genes (<i>proenkephalin</i>, <i>proopiomelanocortin</i>, <i>preprodynorphin</i>, <i>diazepam-binding inhibitor</i> and <i>proprotein convertase 1 inhibitor</i>) were validated with quantitative PCR. Furthermore, up-regulated glutamate receptors, glutamate transporters, γ-aminobutyric acid (<i>GABA</i>) receptors, GABA transporters, synaptotagmins or mitogen-activated protein kinase (<i>MAPK</i>) genes might contribute to EA-induced analgesia through regulating the glutamatergic synapse, GABAergic synapse, MAPKs, ribosome or ubiquitin-proteasome pathways. Our findings reveal a full profile of molecular modification in response to EA and provide a solid experimental framework for exploring the mechanisms underlying EA-induced analgesia.

Mammalian ULK1 (unc-51 like kinase 1) and ULK2, Caenorhabditis elegans UNC-51, and Drosophila melanogaster Atg1 are serine/threonine kinases that regulate flux through the autophagy pathway in response to various types of cellular stress. C. elegans UNC-51 and D. melanogaster Atg1 also promote axonal growth and defasciculation; disruption of these genes results in defective axon guidance in invertebrates. Although disrupting ULK1/2 function impairs normal neurite outgrowth in vitro, the role of ULK1 and ULK2 in the developing brain remains poorly characterized. Here, we show that ULK1 and ULK2 are required for proper projection of axons in the forebrain. Mice lacking Ulk1 and Ulk2 in their central nervous systems showed defects in axonal pathfinding and defasciculation affecting the corpus callosum, anterior commissure, corticothalamic axons and thalamocortical axons. These defects impaired the midline crossing of callosal axons and caused hypoplasia of the anterior commissure and disorganization of the somatosensory cortex. The axon guidance defects observed in ulk1/2 double-knockout mice and central nervous system-specific (Nes-Cre) Ulk1/2-conditional double-knockout mice were not recapitulated in mice lacking other autophagy genes (i.e., Atg7 or Rb1cc1 [RB1-inducible coiled-coil 1]). The brains of Ulk1/2-deficient mice did not show stem cell defects previously attributed to defective autophagy in ambra1 (autophagy/Beclin 1 regulator 1)- and Rb1cc1-deficient mice or accumulation of SQSTM1 (sequestosome 1)⁺ or ubiquitin⁺ deposits. Together, these data demonstrate that ULK1 and ULK2 regulate axon guidance during mammalian brain development via a noncanonical (i.e., autophagy-independent) pathway.

<h4>Introduction</h4>The relationship between blood metabolites and hemoglobin degradation products (BMHDPs) formed in the cerebrospinal fluid and the development of vasospasm and delayed cerebral ischemia (DCI) after aneurysmal subarachnoid hemorrhage (aSAH) has been the focus of several previous studies, but their molecular and cellular targets remain to be elucidated.<h4>Methods</h4>Because BMHDP-induced changes in Ca_v 2.3 channel function are thought to contribute to DCI after aSAH, we studied their modulation by unconjugated bilirubin (UCB) in an organotypical neuronal network from wild-type (WT) and Ca_v 2.3-deficient animals (KO). Murine retinae were isolated from WT and KO and superfused with nutrient solution. Electroretinograms were recorded before, during, and after superfusion with UCB. Transretinal signaling was analyzed as b-wave, implicit time, and area under the curve (AUC).<h4>Results</h4>Superfusion of UCB significantly attenuated the b-wave amplitude in the isolated retina from wild-type mice by 14.9% (P < 0.05), followed by gradual partial recovery (P = 0.09). Correspondingly, AUC decreased significantly with superfusion of UCB (P < 0.05). During washout, the b-wave amplitude returned to baseline (P = 0.2839). The effects of UCB were absent in Ca_v 2.3-deficient mice, lacking the expression of Ca_v 2.3 as proofed on the biochemical level.<h4>Conclusions</h4>Ex vivo neuronal recording in the murine retina is able to detect transient impairment of transretinal signaling by UCB in WT, but not in KO. This new model may be useful to further clarify the role of calcium channels in neuronal signal alteration in the presence of BHMDPs.

A lot of solid tumors are characterized by uncontrolled signal transduction triggered by receptors related to cellular growth. The targeting of these cell receptors with antitumor drugs is essential to improve chemotherapy efficacy. This can be achieved by conjugation of an active targeting agent to the polymer portion of a colloidal drug delivery system loaded with an antitumor drug. The goal of this minireview is to report and discuss some recent results in epidermal growth factor receptor targeting by the GE11 peptide combined with colloidal drug delivery systems as smart carriers for antitumor drugs. The minireview chapters will focus on explaining and discussing: (i) Epidermal growth factor receptor (EGFR) structures and functions; (ii) GE11 structure and biologic activity; (iii) examples of GE11 conjugation and GE11-conjugated drug delivery systems. The rationale is to contribute in gathering information on the topic of active targeting to tumors. A case study is introduced, involving research on tumor cell targeting by the GE11 peptide combined with polymer nanoparticles.

Antisense Oligonucleotides (ASOs) represent very attractive therapeutic compounds for the treatment of numerous diseases. The antisense field has remarkably progressed over the last few years with the approval of the first antisense drugs and with promising developments of more potent and nuclease resistant chemistries. Despite these recent clinical successes and advances in chemistry and design, effective delivery of ASOs to their target tissues remains a major issue. This review will describe the latest advances obtained with the tricyclo-DNA (tcDNA) chemistry which displays unique pharmacological properties and unprecedented uptake in many tissues after systemic administration. We will examine the variety of therapeutic approaches using both fully modified tcDNA-ASOs and gapmers, including splice switching applications, correction of aberrant splicing, steric blocking strategies and targeted gene knock-down mediated by RNase H recruitment. We will then discuss the merits and potential liabilities of the tcDNA chemistry in the context of ASO drug development.

<h4>Background</h4>Huntington's disease (HD) is an autosomal dominant neurodegenerative condition caused by an expanded CAG repeat in the gene encoding huntingtin (HTT). Optimizing peripheral quantification of huntingtin throughout the course of HD is valuable not only to illuminate the natural history and pathogenesis of disease, but also to detect peripheral effects of drugs in clinical trial.<h4>Rationale</h4>We previously demonstrated that mutant HTT (mHTT) was significantly elevated in purified HD patient leukocytes compared with controls and that these levels track disease progression. Our present study investigates whether the same result can be achieved with a simpler and more scalable collection technique that is more suitable for clinical trials.<h4>Methods</h4>We collected whole blood at 133 patient visits in two sample sets and generated peripheral blood mononuclear cells (PBMCs). Levels of mHTT, as well as N-, and C-terminal and mid-region huntingtin were measured in the PBMCs using ELISA-based Meso Scale Discovery (MSD) electrochemiluminescence immunoassay platforms, and we evaluated the relationship between different HTT species, disease stage, and brain atrophy on magnetic resonance imaging.<h4>Conclusions</h4>The assays were sensitive and accurate. We confirm our previous findings that mHTT increases with advancing disease stage in patient PBMCs, this time using a simple collection protocol and scalable assay.

The ability to assign expression patterns to the individual cell types that constitute a tissue is a major challenge. This especially applies to brain, given its plethora of different, functionally interconnected cell types. Here, we derived cell type-specific transcriptome signatures from existing single cell RNA data and integrated these signatures with a newly generated dataset of expression (bulk RNA-Seq) of the postnatal developing mouse hippocampus. This integrated analysis allowed us to provide a comprehensive and unbiased prediction of the differentiation drivers for 11 different hippocampal cell types and describe how the different cell types interact to support crucial developmental stages. Our results provide a reliable resource of predicted differentiation drivers and insights into the multifaceted aspects of the cells in hippocampus during development.

Clouds' efficiency at reflecting solar radiation and trapping the terrestrial radiation is strongly modulated by the diurnal cycle of clouds (DCC). Much attention has been paid to mean cloud properties due to their critical role in climate projections; however, less research has been devoted to the DCC. Here we quantify the mean, amplitude, and phase of the DCC in climate models and compare them with satellite observations and reanalysis data. While the mean appears to be reliable, the amplitude and phase of the DCC show marked inconsistencies, inducing overestimation of radiation in most climate models. In some models, DCC appears slightly shifted over the ocean, likely as a result of tuning and fortuitously compensating the large DCC errors over the land. While this model tuning does not seem to invalidate climate projections because of the limited DCC response to global warming, it may potentially increase the uncertainty of climate predictions.

Canine distemper virus (CDV), a paramyxovirus, causes a severe highly contagious lethal disease in carnivores, such as mink. Mink lung epithelial cells (Mv.1.Lu cells) are sensitive to CDV infection and are homologous to the natural host system of mink. The current study analyzed the response of Mv.1.Lu cells to CDV infection by iTRAQ combined with LC-MS/MS. In total, 151 and 369 differentially expressed proteins (DEPs) were markedly up-regulated or down-regulated, respectively. Thirteen DEPs were validated via real-time RT-PCR or western blot analysis. Network and KEGG pathway analyses revealed several regulated proteins associated with the NF-κB signaling pathway. Further validation was performed by western blot analysis and immunofluorescence assay, which demonstrated that different CDV strains induced NF-κB P65 phosphorylation and nuclear translocation. Moreover, the results provided interesting information that some identified DEPs possibly associated with the pathogenesis and the immune response upon CDV infection. This study is the first overview of the responses to CDV infection in Mv.1.Lu cells, and the findings will help to analyze further aspects of the molecular mechanisms involved in viral pathogenesis and the immune responses upon CDV infection.

No abstract available.

<h4>Background</h4>Besides antiviral activities against hepatitis B virus (HBV), telbivudine has an extrahepatic pharmaceutical effect: to improve renal function assessed by estimated glomerular filtration rate (eGFR). However, the durability of this effect after withdrawal of telbivudine or switching to other antivirals has never been investigated.<h4>Methods</h4>We conducted a postmarketing, real-world observation study for telbivudine treatment. The durability of telbivudine-associated renal function improvement was examined following withdrawal/switching of antivirals.<h4>Results</h4>Of 160 telbivudine-treated, chronic hepatitis B patients, 21, 6, and 2 patients were loss to follow-up, dead, and pregnant during the study, respectively. Of the remaining 131 patients, 26, 47, 28, and 30 patients experienced telbivudine withdrawal, continuous use of telbivudine, switching to entecavir, or switching to tenofovir, respectively. During the first 2 years, eGFR in telbivudine-treated patients significantly improved before withdrawal/switching of antivirals (<i>P</i> = .009). Thereafter, eGFR remained unchanged for >1 year in the withdrawal (<i>P</i> = .100) and continuous use (<i>P</i> = .517) subgroups, but decreased significantly in the switching to entecavir (<i>P</i> = .002) and switching to tenofovir (<i>P</i> < .001) subgroups. Multivariate logistic regression analysis revealed that switching to tenofovir and poor liver functional reserve were predictors for eGFR deterioration.<h4>Conclusions</h4>Telbivudine-associated renal function improvement was durable after withdrawal or continuous use of telbivudine. However, renal function deteriorated if patients were switched to entecavir or tenofovir.

<b>Objective:</b> Overweight (body mass index or BMI ≥ 25 kg/m²) and stress interact with each other in complex ways. Overweight promotes chronic low-inflammation states, while stress is known to mediate caloric intake. Both conditions are linked to several avoidable health problems and to cognitive decline, brain atrophy, and dementia. Since it was proposed as a framework for the onset of mental illness, the allostatic load model has received increasing attention. Although changes in health and cognition related to overweight and stress are well-documented separately, the association between allostatic load and brain integrity has not been addressed in depth, especially among overweight subjects. <b>Method:</b> Thirty-four healthy overweight-to-obese and 29 lean adults underwent blood testing, neuropsychological examination, and magnetic resonance imaging to assess the relationship between cortical thickness and allostatic load, represented as an index of 15 biomarkers (this is, systolic and diastolic arterial tension, glycated hemoglobin, glucose, creatinine, total cholesterol, HDL and LDL cholesterol, triglycerides, c-reactive protein, interleukin-6, insulin, cortisol, fibrinogen, and leptin). <b>Results:</b> Allostatic load indexes showed widespread positive and negative significant correlations (<i>p</i> < 0.01) with cortical thickness values depending on body-weight status. <b>Conclusion:</b> The increase of allostatic load is linked to changes in the gray matter composition of regions monitoring behavior, sensory-reward processing, and general cognitive function.

Alzheimer's disease (AD) is a progressive neurodegenerative disease. Relationships of 5HT2A and 5HTTLPR polymorphisms and AD risk have been widely investigated previously, whereas results derived from these studies were inconclusive and controversial. The aim of this study was to investigate the association of the 5-HT2A and 5HTTLPR polymorphisms and AD using a meta-analysis of existing literatures. Studies were collected using PubMed, Web of Science, the Cochrane Library databases, Chinese National Knowledge Infrastructure (CNKI) and Embase. Pooled odds ratios (ORs) with 95% confidence intervals (CIs) were used to assess associations. As a result, a total of 7 publications for 5-HT2A T102C and 16 publications for 5HTTLPR (L/S) comprised 3255 cases and 3690 controls fulfilled the inclusion criteria. Significant association was covered between allelic and recessive models of 5-HT2A T102C and AD (allelic model: <i>p</i> = 0.003, OR [95% CI] = 1.23 [1.07, 1.40]; recessive model: <i>p</i> = 0.03, OR [95% CI] = 1.28 [1.02, 1.59]). Subsequently, we conducted subgroup analysis for 5-HT2A T102C polymorphism based on ethnicities and APOE ε4, and identified a significantly increased risk for the allelic and dominant models of 5-HT2A T102C and AD in Asian subgroup (allelic model: <i>p</i> = 0.002, OR [95% CI] = 1.42 [1.14, 1.78]; dominant model: <i>p</i> = 0.02, OR [95% CI] = 1.60 [1.09, 2.35]) and subgroup without APOE ε4 (allelic model: <i>p</i> = 0.02, OR [95% CI] = 1.44 [1.05, 1.99]; dominant model: <i>p</i> = 0.0008, OR [95% CI] = 2.49 [1.46, 4.25]). Nevertheless, the pooled analyses suggested no significant association between allelic, dominant, and recessive models of 5HTTLPR (L/S) and AD (<i>p</i> > 0.05). In conclusion, our meta-analysis demonstrates that 5HT2A C10T, but not 5HTTLPR (L/S), might increase risk for AD.

<h4>Background</h4>Nasal gene expression profiling is a promising method to characterize COPD non-invasively. We aimed to identify a nasal gene expression profile to distinguish COPD patients from healthy controls. We investigated whether this COPD-associated gene expression profile in nasal epithelium is comparable with the profile observed in bronchial epithelium.<h4>Methods</h4>Genome wide gene expression analysis was performed on nasal epithelial brushes of 31 severe COPD patients and 22 controls, all current smokers, using Affymetrix Human Gene 1.0 ST Arrays. We repeated the gene expression analysis on bronchial epithelial brushes in 2 independent cohorts of mild-to-moderate COPD patients and controls.<h4>Results</h4>In nasal epithelium, 135 genes were significantly differentially expressed between severe COPD patients and controls, 21 being up- and 114 downregulated in COPD (false discovery rate < 0.01). Gene Set Enrichment Analysis (GSEA) showed significant concordant enrichment of COPD-associated nasal and bronchial gene expression in both independent cohorts (FDR_GSEA < 0.001).<h4>Conclusion</h4>We identified a nasal gene expression profile that differentiates severe COPD patients from controls. Of interest, part of the nasal gene expression changes in COPD mimics differentially expressed genes in the bronchus. These findings indicate that nasal gene expression profiling is potentially useful as a non-invasive biomarker in COPD.<h4>Trial registration</h4>ClinicalTrials.gov registration number NCT01351792 (registration date May 10, 2011), ClinicalTrials.gov registration number NCT00848406 (registration date February 19, 2009), ClinicalTrials.gov registration number NCT00807469 (registration date December 11, 2008).

Although it is known that the susceptibility of mouse spermatozoa to freezing-thawing varies greatly with genetic background, the underlying mechanisms remain to be elucidated. In this study, to map genetic regions responsible for the susceptibility of spermatozoa to freezing-thawing, we performed in vitro fertilization using spermatozoa from recombinant inbred mice derived from the C57BL/6J and DBA/2J strains, whose spermatozoa showed distinct fertilization abilities after freezing. Genome-wide interval mapping identified two suggestive quantitative trait loci (QTL) associated with fertilization on chromosomes 1 and 11. The strongest QTL on chromosome 11 included 70 genes at 59.237260-61.324742 Mb and another QTL on chromosome 1 included 43 genes at 153.969506-158.217850 Mb. These regions included at least 15 genes involved with testicular expression and possibly with capacitation or sperm motility. Specifically, the Abl2 gene on chromosome 1, which may affect subcellular actin distribution, had polymorphisms between C57BL/6J and DBA/2J that caused at least three amino acid substitutions. A correlation analysis using recombinant inbred strains revealed that the fertilization rate was strongly correlated with the capacitation rate of frozen-thawed spermatozoa after preincubation. This result is consistent with the fact that C57BL/6J frozen-thawed spermatozoa recover their fertilization capacity following treatment with methyl-β-cyclodextrin to enhance sperm capacitation. Thus, our data provide important clues to the molecular mechanisms underlying cryodamage to mouse spermatozoa.

Hereditary hyperferritinaemia-cataract syndrome (HHCS) is a rare disorder usually caused by heterozygous mutations in the iron-responsive element (IRE) in the 5' untranslated region (5'UTR) of the L-ferritin gene (FTL), disturbing the binding of iron regulatory proteins (IRPs) and the post-transcriptional regulation of ferritin expression. Here, the proband of a consanguineous family displayed moderate bilateral cataracts and elevated serum ferritin in the absence of iron overload. The parents and siblings showed variable degrees of mild bilateral cataracts combined with elevated levels of circulating ferritin. Sequencing of FTL identified a novel 5'UTR mutation c.-151A > G, also named "Ghent +49A > G". The zygosity of the mutation, occurring in homozygous and heterozygous state in the proband and other affected family members respectively, correlated well with severity of ophthalmological and hematological manifestations. The substitution is expected to impair the secondary structure of the upper IRE stem. Functional characterization of +49A > G by electrophoretic mobility shift assays demonstrated a reduced binding affinity for IRP1 compared to the wild-type IRE of FTL. Overall, we have expanded the repertoire of deleterious biallelic FTL IRE mutations in HHCS with this novel +49A > G mutation, the zygosity of which correlated well with the disease expression.

Humans have a number of nonclassical major histocompatibility complex (MHC) class I molecules that are quite divergent from the classical ones, and that may have separated from the classical lineage in pre-mammalian times. To estimate when in evolution the respective nonclassical lineages separated from the classical lineage, we first identified "phylogenetic marker motifs" within the evolution of classical MHC class I; the selected motifs are rather specific for and rather stably inherited within clades of species. Distribution of these motifs in nonclassical MHC class I molecules indicates that the lineage including the nonclassical MHC class I molecules CD1 and PROCR separated from the classical lineage before the emergence of tetrapod species, and that the human nonclassical MHC class I molecules FCGRT, MIC/ULBP/RAET, HFE, MR1, and ZAG show similarity with classical MHC class I at the avian/reptilian level. An MR1-like α1 exon sequence was identified in turtle. Our system furthermore indicates that the lineage UT, hitherto only found in non-eutherian mammals, predates tetrapod existence, and we identified UT genes in reptiles. If only accepting wide distribution of a lineage among extant species as true evidence for ancientness, the oldest identified nonclassical MHC class I lineage remains the fish-specific lineage Z, which was corroborated in the present study by finding both Z and classical-type MHC class I sequences in a primitive fish, the bichir. In short, we gained important new insights into the evolution of classical MHC class I motifs and the probable time of origin of nonclassical MHC class I lineages.

We evaluated the impact of early embryonic deletion of huntingtin (htt) from pyramidal neurons on cortical development, cortical neuron survival and motor behavior, using a cre-loxP strategy to inactivate the mouse htt gene (Hdh) in emx1-expressing cell lineages. Western blot confirmed substantial htt reduction in cerebral cortex of these Emx-htt^KO mice, with residual cortical htt in all likelihood restricted to cortical interneurons of the subpallial lineage and/or vascular endothelial cells. Despite the loss of htt early in development, cortical lamination was normal, as revealed by layer-specific markers. Cortical volume and neuron abundance were, however, significantly less than normal, and cortical neurons showed reduced brain-derived neurotrophic factor (BDNF) expression and reduced activation of BDNF signaling pathways. Nonetheless, cortical volume and neuron abundance did not show progressive age-related decline in Emx-htt^KO mice out to 24months. Although striatal neurochemistry was normal, reductions in striatal volume and neuron abundance were seen in Emx-htt^KO mice, which were again not progressive. Weight maintenance was normal in Emx-htt^KO mice, but a slight rotarod deficit and persistent hyperactivity were observed throughout the lifespan. Our results show that embryonic deletion of htt from developing pallium does not substantially alter migration of cortical neurons to their correct laminar destinations, but does yield reduced cortical and striatal size and neuron numbers. The Emx-htt^KO mice were persistently hyperactive, possibly due to defects in corticostriatal development. Importantly, deletion of htt from cortical pyramidal neurons did not yield age-related progressive cortical or striatal pathology.

Complex network topology is characteristic of many biological systems, including anatomical and functional brain networks (connectomes). Here, we first constructed a structural covariance network from MRI measures of cortical thickness on 296 healthy volunteers, aged 14-24 years. Next, we designed a new algorithm for matching sample locations from the Allen Brain Atlas to the nodes of the SCN. Subsequently we used this to define, transcriptomic brain networks by estimating gene co-expression between pairs of cortical regions. Finally, we explored the hypothesis that transcriptional networks and structural MRI connectomes are coupled. A transcriptional brain network (TBN) and a structural covariance network (SCN) were correlated across connection weights and showed qualitatively similar complex topological properties: assortativity, small-worldness, modularity, and a rich-club. In both networks, the weight of an edge was inversely related to the anatomical (Euclidean) distance between regions. There were differences between networks in degree and distance distributions: the transcriptional network had a less fat-tailed degree distribution and a less positively skewed distance distribution than the SCN. However, cortical areas connected to each other within modules of the SCN had significantly higher levels of whole genome co-expression than expected by chance. Nodes connected in the SCN had especially high levels of expression and co-expression of a human supragranular enriched (HSE) gene set that has been specifically located to supragranular layers of human cerebral cortex and is known to be important for large-scale, long-distance cortico-cortical connectivity. This coupling of brain transcriptome and connectome topologies was largely but not entirely accounted for by the common constraint of physical distance on both networks.

Many tumors produce platelet-derived growth factor (PDGF)-DD, which promotes cellular proliferation, epithelial-mesenchymal transition, stromal reaction, and angiogenesis through autocrine and paracrine PDGFRβ signaling. By screening a secretome library, we found that the human immunoreceptor NKp44, encoded by NCR2 and expressed on natural killer (NK) cells and innate lymphoid cells, recognizes PDGF-DD. PDGF-DD engagement of NKp44 triggered NK cell secretion of interferon gamma (IFN)-γ and tumor necrosis factor alpha (TNF-α) that induced tumor cell growth arrest. A distinctive transcriptional signature of PDGF-DD-induced cytokines and the downregulation of tumor cell-cycle genes correlated with NCR2 expression and greater survival in glioblastoma. NKp44 expression in mouse NK cells controlled the dissemination of tumors expressing PDGF-DD more effectively than control mice, an effect enhanced by blockade of the inhibitory receptor CD96 or CpG-oligonucleotide treatment. Thus, while cancer cell production of PDGF-DD supports tumor growth and stromal reaction, it concomitantly activates innate immune responses to tumor expansion.

Macroscopic cortical networks are important for cognitive function, but it remains challenging to construct anatomically plausible individual structural connectomes from human neuroimaging. We introduce a new technique for cortical network mapping based on inter-regional similarity of multiple morphometric parameters measured using multimodal MRI. In three cohorts (two human, one macaque), we find that the resulting morphometric similarity networks (MSNs) have a complex topological organization comprising modules and high-degree hubs. Human MSN modules recapitulate known cortical cytoarchitectonic divisions, and greater inter-regional morphometric similarity was associated with stronger inter-regional co-expression of genes enriched for neuronal terms. Comparing macaque MSNs with tract-tracing data confirmed that morphometric similarity was related to axonal connectivity. Finally, variation in the degree of human MSN nodes accounted for about 40% of between-subject variability in IQ. Morphometric similarity mapping provides a novel, robust, and biologically plausible approach to understanding how human cortical networks underpin individual differences in psychological functions.

Bone biomaterials play a vital role in bone repair by providing the necessary substrate for cell adhesion, proliferation, and differentiation and by modulating cell activity and function. In past decades, extensive efforts have been devoted to developing bone biomaterials with a focus on the following issues: (1) developing ideal biomaterials with a combination of suitable biological and mechanical properties; (2) constructing a cell microenvironment with pores ranging in size from nanoscale to submicro- and microscale; and (3) inducing the oriented differentiation of stem cells for artificial-to-biological transformation. Here we present a comprehensive review of the state of the art of bone biomaterials and their interactions with stem cells. Typical bone biomaterials that have been developed, including bioactive ceramics, biodegradable polymers, and biodegradable metals, are reviewed, with an emphasis on their characteristics and applications. The necessary porous structure of bone biomaterials for the cell microenvironment is discussed, along with the corresponding fabrication methods. Additionally, the promising seed stem cells for bone repair are summarized, and their interaction mechanisms with bone biomaterials are discussed in detail. Special attention has been paid to the signaling pathways involved in the focal adhesion and osteogenic differentiation of stem cells on bone biomaterials. Finally, achievements regarding bone biomaterials are summarized, and future research directions are proposed.

The generation and differentiation of an appropriate number of neurons, as well as its distribution in different parts of the brain, is crucial for the proper establishment, maintenance and plasticity of neural circuitries. Newborn neurons travel along the brain in a process known as neuronal migration, to finalize their correct position in the nervous system. Defects in neuronal migration produce abnormalities in the brain that can generate neurodevelopmental pathologies, such as autism, schizophrenia and intellectual disability. In this review, we present an overview of the developmental origin of the different telencephalic subdivisions and a description of migratory pathways taken by distinct neural populations traveling long distances before reaching their target position in the brain. In addition, we discuss some of the molecules implicated in the guidance of these migratory paths and transcription factors that contribute to the correct migration and integration of these neurons.

The phosphorylation of the ribosomal protein S6 (rpS6) is widely used to track neuronal activity. Although it is generally assumed that rpS6 phosphorylation has a stimulatory effect on global protein synthesis in neurons, its exact biological function remains unknown. By using a phospho-deficient rpS6 knockin mouse model, we directly tested the role of phospho-rpS6 in mRNA translation, plasticity and behavior. The analysis of multiple brain areas shows for the first time that, in neurons, phospho-rpS6 is dispensable for overall protein synthesis. Instead, we found that phospho-rpS6 controls the translation of a subset of mRNAs in a specific brain region, the nucleus accumbens (Acb), but not in the dorsal striatum. We further show that rpS6 phospho-mutant mice display altered long-term potentiation (LTP) in the Acb and enhanced novelty-induced locomotion. Collectively, our findings suggest a previously unappreciated role of phospho-rpS6 in the physiology of the Acb, through the translation of a selective subclass of mRNAs, rather than the regulation of general protein synthesis.

The nutrient responsive O-GlcNAcylation is a dynamic post-translational protein modification found on several nucleocytoplasmic proteins. Previous studies have suggested that hyperglycemia induces the levels of total O-GlcNAcylation inside the cells. Hyperglycemia mediated increase in protein O-GlcNAcylation has been shown to be responsible for various pathologies including insulin resistance and Alzheimer's disease. Since maternal hyperglycemia during pregnancy is associated with adverse neurodevelopmental outcomes in the offspring, it is intriguing to identify the effect of increased protein O-GlcNAcylation on embryonic neurogenesis. Herein using human embryonic stem cells (hESCs) as model, we show that increased levels of total O-GlcNAc is associated with decreased neural progenitor proliferation and premature differentiation of cortical neurons, reduced AKT phosphorylation, increased apoptosis and defects in the expression of various regulators of embryonic corticogenesis. As defects in proliferation and differentiation during neurodevelopment are common features of various neurodevelopmental disorders, increased O-GlcNAcylation could be one mechanism responsible for defective neurodevelopmental outcomes in metabolically compromised pregnancies such as diabetes.

This review summarizes the mechanisms of the initiator protein DnaA in replication initiation and its regulation in <i>Escherichia coli</i>. The chromosomal origin (<i>oriC</i>) DNA is unwound by the replication initiation complex to allow loading of DnaB helicases and replisome formation. The initiation complex consists of the DnaA protein, DnaA-initiator-associating protein DiaA, integration host factor (IHF), and <i>oriC</i>, which contains a duplex-unwinding element (DUE) and a DnaA-oligomerization region (DOR) containing DnaA-binding sites (DnaA boxes) and a single IHF-binding site that induces sharp DNA bending. DiaA binds to DnaA and stimulates DnaA assembly at the DOR. DnaA binds tightly to ATP and ADP. ATP-DnaA constructs functionally different sub-complexes at DOR, and the DUE-proximal DnaA sub-complex contains IHF and promotes DUE unwinding. The first part of this review presents the structures and mechanisms of <i>oriC</i>-DnaA complexes involved in the regulation of replication initiation. During the cell cycle, the level of ATP-DnaA level, the active form for initiation, is strictly regulated by multiple systems, resulting in timely replication initiation. After initiation, regulatory inactivation of DnaA (RIDA) intervenes to reduce ATP-DnaA level by hydrolyzing the DnaA-bound ATP to ADP to yield ADP-DnaA, the inactive form. RIDA involves the binding of the DNA polymerase clamp on newly synthesized DNA to the DnaA-inactivator Hda protein. In <i>datA</i>-dependent DnaA-ATP hydrolysis (DDAH), binding of IHF at the chromosomal locus <i>datA</i>, which contains a cluster of DnaA boxes, results in further hydrolysis of DnaA-bound ATP. SeqA protein inhibits untimely initiation at <i>oriC</i> by binding to newly synthesized <i>oriC</i> DNA and represses <i>dnaA</i> transcription in a cell cycle dependent manner. To reinitiate DNA replication, ADP-DnaA forms oligomers at DnaA-reactivating sequences (<i>DARS1</i> and <i>DARS2</i>), resulting in the dissociation of ADP and the release of nucleotide-free apo-DnaA, which then binds ATP to regenerate ATP-DnaA. <i>In vivo, DARS2</i> plays an important role in this process and its activation is regulated by timely binding of IHF to <i>DARS2</i> in the cell cycle. Chromosomal locations of <i>DARS</i> sites are optimized for the strict regulation for timely replication initiation. The last part of this review describes how DDAH and DARS regulate DnaA activity.

Maintenance of metal homeostasis is crucial for many different enzymatic activities and in turn for cell function and survival. In addition, cells display detoxification and protective mechanisms against toxic accumulation of metals. Perturbation of any of these processes normally leads to cellular dysfunction and finally to cell death. In the last years, loss of metal regulation has been described as a common pathological feature in many human neurodegenerative diseases. However, in most cases, it is still a matter of debate whether such dyshomeostasis is a primary or a secondary downstream defect. In this review, we will summarize and critically evaluate the contribution of <i>Drosophila</i> to model human diseases that involve altered metabolism of metals or in which metal dyshomeostasis influence their pathobiology. As a prerequisite to use <i>Drosophila</i> as a model, we will recapitulate and describe the main features of core genes involved in copper and zinc metabolism that are conserved between mammals and flies. <i>Drosophila</i> presents some unique strengths to be at the forefront of neurobiological studies. The number of genetic tools, the possibility to easily test genetic interactions <i>in vivo</i> and the feasibility to perform unbiased genetic and pharmacological screens are some of the most prominent advantages of the fruitfly. In this work, we will pay special attention to the most important results reported in fly models to unveil the role of copper and zinc in cellular degeneration and their influence in the development and progression of human neurodegenerative pathologies such as Parkinson's disease, Alzheimer's disease, Huntington's disease, Friedreich's Ataxia or Menkes, and Wilson's diseases. Finally, we show how these studies performed in the fly have allowed to give further insight into the influence of copper and zinc in the molecular and cellular causes and consequences underlying these diseases as well as the discovery of new therapeutic strategies, which had not yet been described in other model systems.

<h4>Background</h4>Although post-traumatic stress disorder (PTSD) is primarily a mental disorder, it can cause additional symptoms that do not seem to be directly related to the central nervous system, which PTSD is assumed to directly affect. PTSD-mediated heart diseases are some of such secondary disorders. In spite of the significant correlations between PTSD and heart diseases, spatial separation between the heart and brain (where PTSD is primarily active) prevents researchers from elucidating the mechanisms that bridge the two disorders. Our purpose was to identify genes linking PTSD and heart diseases.<h4>Methods</h4>In this study, gene expression profiles of various murine tissues observed under various types of stress or without stress were analyzed in an integrated manner using tensor decomposition (TD).<h4>Results</h4>Based upon the obtained features, ∼ 400 genes were identified as candidate genes that may mediate heart diseases associated with PTSD. Various gene enrichment analyses supported biological reliability of the identified genes. Ten genes encoding protein-, DNA-, or mRNA-interacting proteins-ILF2, ILF3, ESR1, ESR2, RAD21, HTT, ATF2, NR3C1, TP53, and TP63-were found to be likely to regulate expression of most of these ∼ 400 genes and therefore are candidate primary genes that cause PTSD-mediated heart diseases. Approximately 400 genes in the heart were also found to be strongly affected by various drugs whose known adverse effects are related to heart diseases and/or fear memory conditioning; these data support the reliability of our findings.<h4>Conclusions</h4>TD-based unsupervised feature extraction turned out to be a useful method for gene selection and successfully identified possible genes causing PTSD-mediated heart diseases.

A series of hybrid compounds based on the natural products artemisinin and thymoquinone was synthesized and investigated for their biological activity against the malaria parasite <i>Plasmodium falciparum</i> 3D7 strain, human cytomegalovirus (HCMV), and two leukemia cell lines (drug-sensitive CCRF-CEM and multidrug-resistant subline CEM/ADR5000). An unprecedented one-pot method of selective formation of C-10α-acetate <b>14</b> starting from a 1:1 mixture of C-10α- to C-10β-dihydroartemisinin was developed. The key step of this facile method is a mild decarboxylative activation of malonic acid mediated by DCC/DMAP. Ether-linked thymoquinone-artemisinin hybrids <b>6a/b</b> stood out as the most active compounds in all categories, while showing no toxic side effects toward healthy human foreskin fibroblasts and thus being selective. They exhibited EC₅₀ values of 0.2 μM against the doxorubicin-sensitive as well as the multidrug-resistant leukemia cells and therefore can be regarded as superior to doxorubicin. Moreover, they showed to be five times more active than the standard drug ganciclovir and nearly eight times more active than artesunic acid against HCMV. In addition, hybrids <b>6a/b</b> possessed excellent antimalarial activity (EC₅₀ = 5.9/3.7 nM), which was better than that of artesunic acid (EC₅₀ = 8.2 nM) and chloroquine (EC₅₀ = 9.8 nM). Overall, most of the presented thymoquinone-artemisinin-based hybrids exhibit an excellent and broad variety of biological activities (anticancer, antimalarial, and antiviral) combined with a low toxicity/high selectivity profile.

ZipA is essential for cell division in <i>Escherichia coli</i>, acting early in the process to anchor polymers of FtsZ to the cytoplasmic membrane. Along with FtsA, FtsZ and ZipA form a proto-ring at midcell that recruits additional proteins to eventually build the division septum. Cells carrying the thermosensitive <i>zipA1</i> allele divide fairly normally at 30°C in rich medium but cease dividing at temperatures above 34°C, forming long filaments. In a search for suppressors of the <i>zipA1</i> allele, we found that deletions of specific genes involved in amino acid biosynthesis could partially rescue cell growth and division at 34°C or 37°C but not at 42°C. Notably, although a diverse group of amino acid biosynthesis gene deletions could partially rescue the growth of <i>zipA1</i> cells at 34°C, only deletions of genes related to the biosynthesis of threonine, glycine, serine, and methionine could rescue growth at 37°C. Adding exogenous pyridoxal 5-phosphate (PLP), a cofactor for many of the enzymes affected by this study, partially suppressed <i>zipA1</i> mutant thermosensitivity. For many of the deletions, PLP had an additive rescuing effect on the <i>zipA1</i> mutant. Moreover, added PLP partially suppressed the thermosensitivity of <i>ftsQ</i> and <i>ftsK</i> mutants and weakly suppressed an <i>ftsI</i> mutant, but it failed to suppress <i>ftsA</i> or <i>ftsZ</i> thermosensitive mutants. Along with the ability of a deletion of <i>metC</i> to partially suppress the <i>ftsK</i> mutant, our results suggest that perturbations of amino acid metabolic pathways, particularly those that redirect the flow of carbon away from the synthesis of threonine, glycine, or methionine, are able to partially rescue some cell division defects.<b>IMPORTANCE</b> Cell division of bacteria, such as <i>Escherichia coli</i>, is essential for their successful colonization. It is becoming increasingly clear that nutritional status and central metabolism can affect bacterial size and shape; for example, a metabolic enzyme (OpgH) can moonlight as a regulator of FtsZ, an essential cell division protein. Here, we demonstrate a link between amino acid metabolism and ZipA, another essential cell division protein that binds directly to FtsZ and tethers it to the cytoplasmic membrane. Our evidence suggests that altering flux through the methionine-threonine-glycine-serine pathways and supplementing with the enzyme cofactor pyridoxal-5-phosphate can partially compensate for an otherwise lethal defect in ZipA, as well as several other cell division proteins.

Malignant glioma cells invade the surrounding brain parenchyma, by migrating along the blood vessels, thus promoting cancer growth. The biological bases of these activities are grounded in profound alterations of the metabolism and the structural organization of the cells, which consequently acquire the ability to modify the surrounding microenvironment, by altering the extracellular matrix and affecting the properties of the other cells present in the brain, such as normal glial-, endothelial- and immune-cells. Most of the effects on the surrounding environment are probably exerted through the release of a variety of extracellular vesicles (EVs), which contain many different classes of molecules, from genetic material to defined species of lipids and enzymes. EV-associated molecules can be either released into the extracellular matrix (ECM) and/or transferred to neighboring cells: as a consequence, both deep modifications of the recipient cell phenotype and digestion of ECM components are obtained, thus causing cancer propagation, as well as a general brain dysfunction. In this review, we first analyze the main intracellular and extracellular transformations required for glioma cell invasion into the brain parenchyma; then we discuss how these events may be attributed, at least in part, to EVs that, like the pawns of a dramatic chess game with cancer, open the way to the tumor cells themselves.

<h4>Background</h4>Chondrogenesis is the earliest stage of skeletal development and is a highly dynamic process, integrating the activities and functions of transcription factors, cell signaling molecules and extracellular matrix proteins. The molecular mechanisms underlying chondrogenesis have been extensively studied and multiple key regulators of this process have been identified. However, a genome-wide overview of the gene regulatory network in chondrogenesis has not been achieved.<h4>Results</h4>In this study, employing RNA sequencing, we identified 332 protein coding genes and 34 long non-coding RNA (lncRNA) genes that are highly selectively expressed in human fetal growth plate chondrocytes. Among the protein coding genes, 32 genes were associated with 62 distinct human skeletal disorders and 153 genes were associated with skeletal defects in knockout mice, confirming their essential roles in skeletal formation. These gene products formed a comprehensive physical interaction network and participated in multiple cellular processes regulating skeletal development. The data also revealed 34 transcription factors and 11,334 distal enhancers that were uniquely active in chondrocytes, functioning as transcriptional regulators for the cartilage-selective genes.<h4>Conclusions</h4>Our findings revealed a complex gene regulatory network controlling skeletal development whereby transcription factors, enhancers and lncRNAs participate in chondrogenesis by transcriptional regulation of key genes. Additionally, the cartilage-selective genes represent candidate genes for unsolved human skeletal disorders.

A chromosome is a single long DNA molecule assembled along its length with nucleosomes and proteins. During interphase, a mammalian chromosome exists as a highly organized supramolecular globule in the nucleus. Here, we discuss new insights into how genomic DNA is packaged and organized within interphase chromosomes. Our emphasis is on the structural principles that underlie chromosome organization, with a particular focus on the intrinsic contributions of the 10-nm chromatin fiber, but not the regular 30-nm fiber. We hypothesize that the hierarchical globular organization of an interphase chromosome is fundamentally established by the self-interacting properties of a 10-nm zig-zag array of nucleosomes, while histone post-translational modifications, histone variants, and chromatin-associated proteins serve to mold generic chromatin domains into specific structural and functional entities.

Neurological disorders are a major threat to public health. Stem cell-based regenerative medicine is now a promising experimental paradigm for its treatment, as shown in pre-clinical animal studies. Initial attempts have been on the replacement of neuronal cells only, but glial progenitors (GPs) are now becoming strong alternative cellular therapeutic candidates to replace oligodendrocytes and astrocytes as knowledge accumulates about their important emerging role in various disease processes. There are many examples of successful therapeutic outcomes for transplanted GPs in small animal models, but clinical translation has proved to be challenging due to the 1,000-fold larger volume of the human brain compared to mice. Human GPs transplanted into the mouse brain migrate extensively and can induce global cell replacement, but a similar extent of migration in the human brain would only allow for local rather than global cell replacement. We review here the mechanisms that govern cell migration, which could potentially be exploited to enhance the migratory properties of GPs through cell engineering pre-transplantation. We furthermore discuss the (dis)advantages of the various cell delivery routes that are available, with particular emphasis on intra-arterial injection as the most suitable route for achieving global cell distribution in the larger brain. Now that therapeutic success has proven to be feasible in small animal models, future efforts will need to be directed to enhance global cell delivery and migration to make bench-to-bedside translation a reality.

<h4>Background</h4>This study was aimed at understanding whether bronchial biopsy specimen can be used as a surrogate for DNA methylation analysis in surgically resected lung cancer.<h4>Methods</h4>A genome-wide methylation was analyzed in 42 surgically resected tumor tissues, 136 bronchial washing, 12 sputum, and 8 bronchial biopsy specimens using the Infinium HumanMethylation450 BeadChip, and models for prediction of lung cancer were evaluated using TCGA lung cancer data.<h4>Results</h4>Four thousand seven hundred and twenty-six CpGs (<i>P</i> < 1.0E-07) that were highly methylated in tumor tissues were identified from 42 lung cancer patients. Ten CpGs were selected for prediction of lung cancer. Genes including the 10 CpGs were classified into three categories: (i) transcription (<i>HOXA9</i>, <i>SOX17</i>, <i>ZNF154</i>, <i>HOXD13</i>); (ii) cell signaling (<i>HBP1</i>, <i>SFRP1</i>, <i>VIPR2</i>); and (iii) adhesion (<i>PCDH17</i>, <i>ITGA5</i>, <i>CD34</i>). Three logistic regression models based on the 10 CpGs classified 897 TCGA primary lung tissues with a sensitivity of 95.0~97.8% and a specificity of 97.4~98.7%. However, the classification performance of the models was very poor in bronchial washing samples: the area under the curve (AUC) was equal to 0.72~0.78. The methylation levels of the 10 CpGs in bronchial biopsy were not significantly different from those in surgically resected tumor tissues (<i>P</i> > 0.05, Wilcoxon rank-sum test). However, their methylation levels were significantly different between paired bronchial biopsy and washing (<i>P</i> < 0.05, Wilcoxon signed-rank test).<h4>Conclusions</h4>The present study suggests that bronchial biopsy specimen may be used as a surrogate for DNA methylation analysis in patient with inoperable lung cancer.

The PITX1 transcription factor is expressed during hindlimb development, where it plays a critical role in directing hindlimb growth and the specification of hindlimb morphology. While it is known that PITX1 regulates hindlimb formation, in part, through activation of the Tbx4 gene, other transcriptional targets remain to be elucidated. We have used a combination of ChIP-seq and RNA-seq to investigate enhancer regions and target genes that are directly regulated by PITX1 in embryonic mouse hindlimbs. In addition, we have analyzed PITX1 binding sites in hindlimbs of Anolis lizards to identify ancient PITX1 regulatory targets. We find that PITX1-bound regions in both mouse and Anolis hindlimbs are strongly associated with genes implicated in limb and skeletal system development. Gene expression analyses reveal a large number of misexpressed genes in the hindlimbs of Pitx1^-/- mouse embryos. By intersecting misexpressed genes with genes that have neighboring mouse PITX1 binding sites, we identified 440 candidate targets of PITX1. Of these candidates, 68 exhibit ultra-conserved PITX1 binding events that are shared between mouse and Anolis hindlimbs. Among the ancient targets of PITX1 are important regulators of cartilage and skeletal muscle development, including Sox9 and Six1. Our data suggest that PITX1 promotes chondrogenesis and myogenesis in the hindlimb by direct regulation of several key members of the cartilage and muscle transcriptional networks.

Inactivation of the androgen receptor (AR) pathway by androgen deprivation therapy (ADT) is the mainstay of (metastatic) prostate cancer therapy. Ultimately, the AR pathway will be re-activated despite castrate levels of circulating androgens. Thereby, maintaining its role even in castration resistant prostate cancer (CRPC). The recent STAMPEDE and CHAARTED trials showed that docetaxel in combination with ADT increased survival in hormone sensitive prostate cancer patients, suggesting cross-talk between AR signaling and chemotherapy efficacy. We hypothesized that a similar interaction may also apply for CRPC that is treated with cabazitaxel. We studied the impact of androgen status on the efficacy, pharmacodynamics and -kinetics of cabazitaxel in a unique and clinically relevant patient derived xenograft model of castration resistant disease. We found that cabazitaxel is highly effective in a castrate setting with strongly reduced AR activation, while tumor growth inhibition by cabazitaxel was completely abolished in the presence of high AR pathway activity. Moreover, additional experiments showed that intratumoral cabazitaxel levels were 3.5 times higher in tumors from castrated mice as compared to tumors from androgen-supplemented animals. We confirmed that cabazitaxel pharmacokinetics were not affected by testosterone, suggesting that androgen status might influence cabazitaxel tumor uptake directly. This study reveals the impact of androgen status on cabazitaxel efficacy and supports the potential of combination of taxane chemotherapeutics with AR axis targeting agents.

The hepatic repair complex in the setting of cirrhosis has received increasing attention, as it implies the regression of cirrhosis, which was traditionally taken to be an irreversible state. In this brief review, the patterns of fibrosis, the existing staging systems for chronic liver disease and the histopathological features of cirrhosis regression are discussed.

The present study aimed to classify gastric cancer (GC) into subtypes and to screen the subtype‑specific genes, their targeted microRNAs (miRNAs) and enriched pathways to explore the putative mechanism of each GC subtypes. The GSE13861 data set was downloaded from the Gene Expression Omnibus and used to screen differential expression genes (DEGs) in GC samples based on the detection of imbalanced differential signal algorithm. The specific genes in each subtype were identified with the cut‑off criterion of U>0.04, pathway enrichment analysis was performed and the subtype‑specific subpaths of miRNA‑target pathway were determined. A total of 1,263 DEGs were identified in the primary gastric adenocarcinoma (PGD) samples, which were subsequently divided into four subtypes, according to the hierarchy cluster analysis. Identification of the subpaths of each subtype indicated that the subpath related to subtype 1 was miRNA (miR)‑202/calcium voltage‑gated channel subunit α1 (CACNA1E)/type II diabetes mellitus. The nuclear factor‑κB signaling pathway was the most significantly specific pathway and subpath identified for subtype 2, which was regulated by miR‑338‑targeted suppression of C‑C motif chemokine ligand 21 (CCL21). For subtype 3, significant related pathways included ubiquitin‑mediated proteolysis and proteasome, and the important subpath was miR‑146B/proteasome 26S subunit, non‑ATPase 3 (PSMD3)/proteasome; focal adhesion was the significant pathway indicated for subtype 4, and the subpaths were miR‑34A/vinculin (VCL)/focal adhesion and miR‑34C/VCL/focal adhesion. In addition, Helicobacter pylori infection was higher in GC subtype 1 than in other subtypes. Specific genes, such as CACNA1E, CCL21, PSMD3 and VCL, may be used as potential feature genes to identify different subtypes of GC, and their associated subpaths may partially explain the pathogenetic mechanism of each GC subtype.

This study aimed to analyze epigenetically and genetically altered genes in melanoma to get a better understanding of the molecular circuitry of melanoma and identify potential gene targets for the treatment of melanoma. The microarray data of GSE31879, including mRNA expression profiles (seven melanoma and four melanocyte samples) and DNA methylation profiles (seven melanoma and five melanocyte samples), were downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) and differentially methylated positions (DMPs) were screened using the linear models for microarray data (limma) package in melanoma compared with melanocyte samples. Gene ontology (GO) and pathway enrichment analysis of the DEGs were carried out using the Database for Annotation, Visualization, and Integrated Discovery. Moreover, differentially methylated genes (DMGs) were identified, and a transcriptional regulatory network was constructed using the University of California Santa Cruz genome browser database. A total of 1,215 DEGs (199 upregulated and 1,016 downregulated) and 14,094 DMPs (10,450 upregulated and 3,644 downregulated) were identified in melanoma compared with melanocyte samples. Additionally, the upregulated and downregulated DEGs were significantly associated with different GO terms and pathways, such as pigment cell differentiation, biosynthesis, and metabolism. Furthermore, the transcriptional regulatory network showed that DMGs such as Aristaless-related homeobox (<i>ARX</i>), damage-specific DNA binding protein 2 (<i>DDB2</i>), and myelin basic protein (<i>MBP</i>) had higher node degrees. Our results showed that several methylated genes (<i>ARX</i>, <i>DDB2</i>, and <i>MBP</i>) may be involved in melanoma progression.

Synthesis of hydroxyapatite (HA) through sol-gel process in different solvent systems is reported. Calcium nitrate tetrahydrate (CNTH) and diammonium hydrogen phosphate (DAHP) were used as calcium and phosphorus precursors, respectively. Three different synthesis reactions were carried out by changing the solvent media, while keeping all other process parameters constant. A measure of 0.5 M aqueous DAHP solution was used in all reactions while CNTH was dissolved in distilled water, tetrahydrofuran (THF) and <i>N</i>,<i>N</i>-dimethylformamide (DMF) at a concentration of 0.5 M. Ammonia solution (28-30%) was used to maintain the pH of the reaction mixtures in the 10-12 range. All reactions were carried out at 40 ± 2°C for 4 h. Upon completion of the reactions, products were filtered, washed and calcined at 500°C for 2 h. It was clearly demonstrated through various techniques that the dielectric constant and polarity of the solvent mixture strongly influence the chemical structure and morphological properties of calcium phosphate synthesized. Water-based reaction medium, with highest dielectric constant, mainly produced β-calcium pyrophosphate (β-CPF) with a minor amount of HA. DMF/water system yielded HA as the major phase with a very minor amount of β-CPF. THF/water solvent system with the lowest dielectric constant resulted in the formation of pure HA.

There is accumulating evidence that miRNA might serve as potential diagnostic and prognostic markers for various types of cancer. Hepatocellular carcinoma (HCC) is the most common type of malignant lesion but the significance of miRNAs in HCC remains largely unknown. The present study aimed to establish the diagnostic value of miR-101-3p/5p in HCC and then further investigate the prospective molecular mechanism via a bioinformatic analysis. First, the miR-101 expression profiles and parallel clinical parameters from 362 HCC patients and 50 adjacent non-HCC tissue samples were downloaded from The Cancer Genome Atlas (TCGA). Second, we aggregated all miR-101-3p/5p expression profiles collected from published literature and the Gene Expression Omnibus and TCGA databases. Subsequently, target genes of miR-101-3p and miR-101-5p were predicted by using the miRWalk database and then overlapped with the differentially expressed genes of HCC identified by natural language processing. Finally, bioinformatic analyses were conducted with the overlapping genes. The level of miR-101 was significantly lower in HCC tissues compared with adjacent non-HCC tissues (<i>P</i> < 0.001), and the area under the curve of the low miR-101 level for HCC diagnosis was 0.925 (<i>P</i> < 0.001). The pooled summary receiver operator characteristic (SROC) of miR-101-3p was 0.86, and the combined SROC curve of miR-101-5p was 0.80. Bioinformatic analysis showed that the target genes of both miR-101-3p and miR-101-5p are involved in several pathways that are associated with HCC. The hub genes for miR-101-3p and miR-101-5p were also found. Our results suggested that both miR-101-3p and miR-101-5p might be potential diagnostic markers in HCC, and that they exert their functions via targeting various prospective genes in the same pathways.

Adeno-associated virus (AAV) has shown promising therapeutic efficacy with a good safety profile in a wide range of animal models and human clinical trials. With the advent of clustered regulatory interspaced short palindromic repeat (CRISPR)-based genome-editing technologies, AAV provides one of the most suitable viral vectors to package, deliver, and express CRISPR components for targeted gene editing. Recent discoveries of smaller Cas9 orthologues have enabled the packaging of Cas9 nuclease and its chimeric guide RNA into a single AAV delivery vehicle for robust <i>in vivo</i> genome editing. Here, we discuss how the combined use of small Cas9 orthologues, tissue-specific minimal promoters, AAV serotypes, and different routes of administration has advanced the development of efficient and precise <i>in vivo</i> genome editing and comprehensively review the various AAV-CRISPR systems that have been effectively used in animals. We then discuss the clinical implications and potential strategies to overcome off-target effects, immunogenicity, and toxicity associated with CRISPR components and AAV delivery vehicles. Finally, we discuss ongoing non-viral-based <i>ex vivo</i> gene therapy clinical trials to underscore the current challenges and future prospects of CRISPR/Cas9 delivery for human therapeutics.

The aim of the present study was to identify the common molecular mechanisms of multiple glioma subtypes, including astrocytoma, glioblastoma and oligodendroglioma, in addition to the specific mechanisms of different types. The gene expression profile set GSE4290 was downloaded from the Gene Expression Omnibus database. Differentially expressed genes (DEGs) from three types of glioma, relative to non-tumor tissue, were calculated by the t-test method with a linear regression model. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of the DEGs was performed. GeneVenn online analysis software was used for the comparison of the DEGs between subtypes. A total of 795 DEGs, including 619 up and 176 downregulated DEGs were screened from the astrocytoma expression profiles; these were enriched in the KEGG pathways of 'neuroactive ligand-receptor interaction' (upregulated) and 'Wnt signaling pathway' (downregulated). Protein-protein interaction networks for astrocytoma, glioblastoma and oligodendroglioma were constructed with 1,617, 7,027 and 1,172 pairs, respectively. A total of 595 common DEGs were obtained between the three subtypes, which were enriched in pathways associated with neural signaling. Glioblastoma is a subtype of astrocytoma; there were 195 DEGs common between these subtypes that were not also associated with oligodendroglioma. DEGs unique to astrocytoma, glioblastoma and oligodendroglioma were associated with the development of the nervous system, the cell cycle and cell matrix components, respectively. The screened DEG p53 gene is likely to be critical for glioma development, including via the Wnt and p53 signaling pathways. Brain-derived neurotrophic factor and cyclin-dependent kinase 1 genes were also likely to be important in the mechanism of glioma development, and were associated with the cell cycle and p53 signaling pathways. Immune system-associated and cell matrix component pathways may be unique signaling pathways associated with astrocytoma and oligodendroglioma, respectively.

The scaffold-free tissue engineering using spheroids is pointed out as an approach for optimizing the delivery system of cartilage construct. In this study, we aimed to evaluate the micromolded nonadhesive hydrogel (MicroTissues®) for spheroid compaction (2-day culture) and spontaneous chondrogenesis (21-day culture) using cartilage progenitors cells (CPCs) from human nasal septum without chondrogenic stimulus. CPC spheroids showed diameter stability (486 <i>μ</i>m ± 65), high percentage of viable cells (88.1 ± 2.1), and low percentage of apoptotic cells (2.3%). After spheroid compaction, the synthesis of TGF-<i>β</i>1, TGF-<i>β</i>2, and TGF-<i>β</i>3 was significantly higher compared to monolayer (<i>p</i> < 0.005). Biomechanical assay revealed that the maximum forces applied to spheroids after chondrogenesis were 2.6 times higher than for those cultured for 2 days. After spontaneous chondrogenesis, CPC spheroids were entirely positive for N-cadherin, collagen type II and type VI, and aggrecan and chondroitin sulfate. Comparing to monolayer, the expression of SOX5 and SOX6 genes analyzed by qPCR was significantly upregulated (<i>p</i> < 0.01). Finally, we observed the capacity of CPC spheroids starting to fuse. To the best of our knowledge, this is the first time in the scientific literature that human CPC spheroids were formed by micromolded nonadhesive hydrogel, achieving a successful scaffold-free cartilage engineering without chondrogenic stimulus (low cost).

<h4>Purpose of review</h4>This review presents an overview of the current knowledge of TRAF molecules in inflammation with an emphasis on available human evidence and direct <i>in vivo</i> evidence of mouse models that demonstrate the contribution of TRAF molecules in the pathogenesis of inflammatory diseases.<h4>Recent findings</h4>The tumor necrosis factor receptor (TNF-R)-associated factor (TRAF) family of cytoplasmic proteins was initially identified as signaling adaptors that bind directly to the intracellular domains of receptors of the TNF-R superfamily. It is now appreciated that TRAF molecules are widely employed in signaling by a variety of adaptive and innate immune receptors as well as cytokine receptors. TRAF-dependent signaling pathways typically lead to the activation of nuclear factor-κBs (NF-κBs), mitogen-activated protein kinases (MAPKs), or interferon-regulatory factors (IRFs). Most of these signaling pathways have been linked to inflammation, and therefore TRAF molecules were expected to regulate inflammation and inflammatory responses since their discovery in 1990s. However, direct <i>in vivo</i> evidence of TRAFs in inflammation and especially in inflammatory diseases had been lacking for many years, partly due to the difficulty imposed by early lethality of TRAF2^-/-, TRAF3^-/-, and TRAF6^-/- mice. With the creation of conditional knockout and lineage-specific transgenic mice of different TRAF molecules, our understanding about TRAFs in inflammation and inflammatory responses has rapidly advanced during the past decade.<h4>Summary</h4>Increasing evidence indicates that TRAF molecules are versatile and indispensable regulators of inflammation and inflammatory responses and that aberrant expression or function of TRAFs contributes to the pathogenesis of inflammatory diseases.

The scaffolds made of polycaprolactone (PCL) are actively employed in different areas of biology and medicine, especially in tissue engineering. However, the usage of unmodified PCL is significantly restricted by the hydrophobicity of its surface, due to the fact that its inert surface hinders the adhesion of cells and the cell interactions on PCL surface. In this work, the surface of PCL nanofibers is modified by Ar/CO₂/C₂H₄ plasma depositing active COOH groups in the amount of 0.57 at % that were later used for the immobilization of platelet-rich plasma (PRP). The modification of PCL nanofibers significantly enhances the viability and proliferation (by hundred times) of human mesenchymal stem cells, and decreases apoptotic cell death to a normal level. According to X-ray photoelectron spectroscopy (XPS), after immobilization of PRP, up to 10.7 at % of nitrogen was incorporated into the nanofibers surface confirming the grafting of proteins. Active proliferation and sustaining the cell viability on nanofibers with immobilized PRP led to an average number of cells of 258 ± 12.9 and 364 ± 34.5 for nanofibers with ionic and covalent bonding of PRP, respectively. Hence, our new method for the modification of PCL nanofibers with PRP opens new possibilities for its application in tissue engineering.

Sensitivity is the key in optical detection of low-abundant analytes, such as circulating RNA or DNA. The enzyme Exonuclease III (Exo III) is a useful tool in this regard; its ability to recycle target DNA molecules results in markedly improved detection sensitivity. Lower limits of detection may be further achieved if the detection background of autofluorescence can be removed. Here we report an ultrasensitive and specific method to quantify trace amounts of DNA analytes in a wash-free suspension assay. In the presence of target DNA, the Exo III recycles the target DNA by selectively digesting the dye-tagged sequence-matched probe DNA strand only, so that the amount of free dye removed from the probe DNA is proportional to the number of target DNAs. Remaining intact probe DNAs are then bound onto upconversion nanoparticles (energy donor), which allows for upconversion luminescence resonance energy transfer (LRET) that can be used to quantify the difference between the free dye and tagged dye (energy acceptor). This scheme simply avoids both autofluorescence under infrared excitation and many tedious washing steps, as the free dye molecules are physically located away from the nanoparticle surface, and as such they remain "dark" in suspension. Compared to alternative approaches requiring enzyme-assisted amplification on the nanoparticle surface, introduction of probe DNAs onto nanoparticles only after DNA hybridization and signal amplification steps effectively avoids steric hindrance. Via this approach, we have achieved a detection limit of 15 pM in LRET assays of human immunodeficiency viral DNA.

The excitement surrounding the potential of gene therapy has been tempered due to the challenges that have thus far limited its successful implementation in the clinic such as issues regarding stability, transfection efficiency, and toxicity. In this study, low molecular weight linear polyethyleneimine (2.5 kDa) was modified by conjugation to a lipid, lithocholic acid, and complexed with a natural polysaccharide, dermatan sulfate (DS), to mask extra cationic charges of the modified polymer. In vitro examination revealed that these modifications improved complex stability with plasmid DNA (pDNA) and transfection efficiency. This novel ternary polyplex (pDNA/3E/DS) was used to investigate if tumor-targeted radiotherapy led to enhanced accumulation and retention of gene therapy vectors in vivo in tumor-bearing mice. Imaging of biodistribution revealed that tumor irradiation led to increased accumulation and retention as well as decreased off-target tissue buildup of pDNA in not only pDNA/3E/DS, but also in associated PEI-based polyplexes and commercial DNA delivery vehicles. The DS-containing complexes developed in this study displayed the greatest increase in tumor-specific pDNA delivery. These findings demonstrate a step forward in nucleic acid vehicle design as well as a promising approach to overall cancer gene therapy through utilization of radiotherapy as a tool for enhanced delivery.

Friedreich's ataxia (FRDA), the most common inherited ataxia, is caused by recessive mutations that reduce the levels of frataxin (FXN), a mitochondrial iron binding protein. We developed an inducible mouse model of <i>Fxn</i> deficiency that enabled us to control the onset and progression of disease phenotypes by the modulation of <i>Fxn</i> levels. Systemic knockdown of <i>Fxn</i> in adult mice led to multiple phenotypes paralleling those observed in human patients across multiple organ systems. By reversing knockdown after clinical features appear, we were able to determine to what extent observed phenotypes represent reversible cellular dysfunction. Remarkably, upon restoration of near wild-type FXN levels, we observed significant recovery of function, associated pathology and transcriptomic dysregulation even after substantial motor dysfunction and pathology were observed. This model will be of broad utility in therapeutic development and in refining our understanding of the relative contribution of reversible cellular dysfunction at different stages in disease.

<h4>Background</h4>Accelerated loss of adipose tissue in cancer is associated with shorter survival, and reduced quality of life. Evidence is emerging suggesting tumour association with alterations in adipose tissue, but much less is known about drug-related mechanisms contributing to adipose atrophy. Identification of mechanisms by which tumour and cancer treatments, such as chemotherapy, affect adipose tissue are required to develop appropriate therapeutic interventions to prevent fat depletion in cancer. This pre-clinical study aimed to assess alterations in adipose tissue during the clinical course of cancer.<h4>Methods</h4>Fischer 344 rats bearing the Ward colorectal tumour were euthanized before chemotherapy, after 1- cycle, or 2-cycles of a combination chemotherapy consisting of Irinotecan (CPT-11) combined with 5-fluorouracil (5-FU), which recapitulates first line treatment for human colorectal cancer. Periuterine adipose tissue was isolated. Healthy rats served as a reference group. Histological analysis (hematoxylin and eosin), Real-time PCR (TaqMan) and proteomic analysis (LC-MS/MS) were performed.<h4>Results</h4>Larger adipocytes (3993.7 ± 52.6 μm²) in tumour-bearing animals compared to the reference group (3227.7 ± 36.7 μm²; p < 0.001) was associated with reduced expression of proteins involved in mitochondrial fatty acid oxidation. The presence of a tumour has a significant effect on phospholipid but not triglyceride fatty acid composition. There were greater proportions of saturated fatty acids concurrent with lower monounsaturated fatty acids within the PL fraction of adipocytes in tumour-bearing animals. Chemotherapy treatment decreased the size of adipocytes (2243.9 ± 30.4 μm²; p < 0.001) and led to depletion of n-3 polyunsaturated fatty acids in adipose tissue triglyceride. Evaluation of the proteome profile revealed decreased expression of proteins involved in ATP generation, β-oxidation, and lipogenesis. Overall, adipose tissue may not be able to efficiently oxidize fatty acids to provide energy to maintain energy demanding pathways like lipogenesis inside the tissue.<h4>Conclusions</h4>In conclusion, metabolic adaptations to mitochondrial impairment may contribute to diminished lipid storage capacity of adipose tissue following chemotherapy delivery.

<h4>Background</h4>Huntington's disease (HD), caused by expansion of a polyglutamine tract within HUNTINGTIN (HTT) protein, is an autosomal dominant neurodegenerative disease associated with a progressive neurodegeneration of striatum and cerebral cortex. Although a few studies have identified substantial microRNA (miRNA) alterations in central nervous tissues from HD patients, it will be more accessible to employ these molecular changes in peripheral tissues as biomarkers for HD.<h4>Methods</h4>We examined the expression levels of 13 miRNAs (miR-1, mirR-9, miR-9*, miR-10b, miR-29a, miR-29b, miR-124a, miR-132, miR-155, miR-196a, miR-196b, miR-330 and miR-615), 10 of which previously demonstrated alterations and 3 of which are potential regulators of differentially-expressed genes in brains of HD patients, in the peripheral leukocytes of 36 HD patients, 8 pre-symptomatic HD carriers and 28 healthy controls.<h4>Results</h4>We found expression levels of miR-9* was significantly lower in HD patients compared with those in healthy controls, while other miRNAs did not show significant difference between these two groups. However, there was no significant correlation between Unified Huntington's Disease Rating Scales (UHDRS) and levels of miR-9* in peripheral leukocytes of HD patients.<h4>Conclusion</h4>Our findings indicate the potential of miR-9* in peripheral leukocyte as a signature of neurodegeneration in HD patients.

<h4>Background</h4>Mitochondrial dysfunction may represent a pathogenic factor in Huntington disease (HD). Physical exercise leads to enhanced mitochondrial function in healthy participants. However, data on effects of physical exercise on HD skeletal muscle remains scarce. We aimed at investigating adaptations of the skeletal muscle mitochondria to endurance training in HD patients.<h4>Methods</h4>Thirteen HD patients and 11 healthy controls completed 26 weeks of endurance training. Before and after the training phase muscle biopsies were obtained from M. vastus lateralis. Mitochondrial respiratory chain complex activities, mitochondrial respiratory capacity, capillarization, and muscle fiber type distribution were determined from muscle samples.<h4>Results</h4>Citrate synthase activity increased during the training intervention in the whole cohort (P = 0.006). There was no group x time interaction for citrate synthase activity during the training intervention (P = 0.522). Complex III (P = 0.008), Complex V (P = 0.043), and succinate cytochrome c reductase (P = 0.008) activities increased in HD patients and controls by endurance training. An increase in mass-specific mitochondrial respiratory capacity was present in HD patients during the endurance training intervention. Overall capillary-to-fiber ratio increased in HD patients by 8.4% and in healthy controls by 6.4% during the endurance training intervention.<h4>Conclusions</h4>Skeletal muscle mitochondria of HD patients are equally responsive to an endurance-training stimulus as in healthy controls. Endurance training is a safe and feasible option to enhance indices of energy metabolism in skeletal muscle of HD patients and may represent a potential therapeutic approach to delay the onset and/or progression of muscular dysfunction.<h4>Trial registration</h4>ClinicalTrials.gov NCT01879267 . Registered May 24, 2012.

<h4>Background</h4>Since the anticancer drugs have diverse inhibited mechanisms to the cancer cells, the use of two or more kinds of anticancer agents may achieve excellent therapeutic effects, especially to the drug-resistant tumors.<h4>Results</h4>In this study, we developed a kind of dual drug [methotrexate (MTX) and 10-hydroxycamptothecine (HCPT)] loaded nanoneedles (DDNDs) with pronounced targeting property, high drug loading and prolonged drug release. The anti-solvent precipitation of the HCPT and MTX modified PEG-b-PLGA (PEG-b-PLGA-MTX, PPMTX) leads to nucleation of nanoneedles with nanocrystalline HCPT as the core wrapped with PPMTX as steric stabilizers. In vitro cell uptake studies showed that the DDNDs revealed an obviously targeting property and entered the HeLa cells easier than the nanoneedles without MTX modification. The cytotoxicity tests illustrated that the DDNDs possessed better killing ability to HeLa cells than the individual drugs or their mixture in the same dose, indicating its good synergistic effect and targeting property. The in vivo studies further confirmed these conclusions.<h4>Conclusions</h4>This approach led to a promising sustained drug delivery system for cancer diagnosis and treatment.

Huntington's disease (HD) is a neurodegenerative disease caused by an expansion in the huntingtin protein (also called Htt) that induces neuronal cell death with age. We found that the treatment of 12-month-old symptomatic heterozygous and homozygous <i>zQ175</i> huntingtin knockin mice for 12 weeks with CTEP, a negative allosteric modulator of metabotropic glutamate receptor 5 (mGluR5), reduced the size and number of huntingtin aggregates, attenuated caspase-3 activity, and reduced both neuronal apoptosis and neuronal loss in brain tissue. Both motor and cognitive impairments were improved in CTEP-treated <i>zQ175</i> mice. The reduction in huntingtin protein aggregate burden by CTEP correlated with the activation of an autophagy pathway mediated by the kinase GSK3β, the transcription factor ZBTB16, and the autophagy factor ATG14. Inhibition of mGluR5 with CTEP also reduced the inhibitory phosphorylation of the autophagosome biogenesis-related kinase ULK1, increased the phosphorylation of the autophagy factor ATG13, and increased the abundance of the autophagy-related protein Beclin1 in homozygous <i>zQ175</i> mice. The findings suggest that mGluR5 antagonism may activate autophagy through convergent mechanisms to promote the clearance of mutant huntingtin aggregates and might be therapeutic in HD patients.

Quantitative Systems Pharmacology (QSP) is a drug discovery approach that integrates computational and experimental methods in an iterative way to gain a comprehensive, unbiased understanding of disease processes to inform effective therapeutic strategies. We report the implementation of QSP to Huntington's Disease, with the application of a chemogenomics platform to identify strategies to protect neuronal cells from mutant huntingtin induced death. Using the STHdh ^Q111 cell model, we investigated the protective effects of small molecule probes having diverse canonical modes-of-action to infer pathways of neuronal cell protection connected to drug mechanism. Several mechanistically diverse protective probes were identified, most of which showed less than 50% efficacy. Specific combinations of these probes were synergistic in enhancing efficacy. Computational analysis of these probes revealed a convergence of pathways indicating activation of PKA. Analysis of phospho-PKA levels showed lower cytoplasmic levels in STHdh ^Q111 cells compared to wild type STHdh ^Q7 cells, and these levels were increased by several of the protective compounds. Pharmacological inhibition of PKA activity reduced protection supporting the hypothesis that protection may be working, in part, through activation of the PKA network. The systems-level studies described here can be broadly applied to any discovery strategy involving small molecule modulation of disease phenotype.

Alzheimer's disease (AD) is the most common form of dementia, characterized by accumulation of amyloid β (Aβ) and neurofibrillary tangles. Oxidative stress and inflammation are considered to play an important role in the development and progression of AD. However, the extent to which these events contribute to the Aβ pathologies remains unclear. We performed inter-species comparative gene expression profiling between AD patient brains and the App ^{NL-G-F/NL-G-F} and 3xTg-AD-H mouse models. Genes commonly altered in App ^{NL-G-F/NL-G-F} and human AD cortices correlated with the inflammatory response or immunological disease. Among them, expression of AD-related genes (C4a/C4b, Cd74, Ctss, Gfap, Nfe2l2, Phyhd1, S100b, Tf, Tgfbr2, and Vim) was increased in the App ^{NL-G-F/NL-G-F} cortex as Aβ amyloidosis progressed with exacerbated gliosis, while genes commonly altered in the 3xTg-AD-H and human AD cortices correlated with neurological disease. The App ^{NL-G-F/NL-G-F} cortex also had altered expression of genes (Abi3, Apoe, Bin2, Cd33, Ctsc, Dock2, Fcer1g, Frmd6, Hck, Inpp5D, Ly86, Plcg2, Trem2, Tyrobp) defined as risk factors for AD by genome-wide association study or identified as genetic nodes in late-onset AD. These results suggest a strong correlation between cortical Aβ amyloidosis and the neuroinflammatory response and provide a better understanding of the involvement of gender effects in the development of AD.

Friedreich's ataxia (FRDA) is a genetic neurodegenerative disorder caused by transcriptional silencing of the frataxin gene (FXN) due to expansions of GAA repeats in intron 1. FRDA manifests with multiple symptoms, which may include ataxia, cardiomyopathy and diabetes mellitus. Expanded GAA tracts are genetically unstable, exhibiting both expansions and contractions. GAA length correlates with severity of FRDA symptoms and inversely with age of onset. Thus, tissue-specific somatic instability of long GAA repeats may be implicated in the development of symptoms and disease progression. Herein, we determined the extent of somatic instability of the GAA repeats in heart, cerebral cortex, spinal cord, cerebellar cortex, and pancreatic tissues from 15 FRDA patients. Results demonstrate differences in the lengths of the expanded GAAs among different tissues, with significantly longer GAA tracts detected in heart and pancreas than in other tissues. The expansion bias detected in heart and pancreas may contribute to disease onset and progression, making the mechanism of somatic instability an important target for therapy. Additionally, we detected significant differences in GAA tract lengths between lymphocytes and fibroblast pairs derived from 16 FRDA patients, with longer GAA tracts present in the lymphocytes. This result urges caution in direct comparisons of data obtained in these frequently used FRDA models. Furthermore, we conducted a longitudinal analysis of the GAA repeat length in lymphocytes collected over a span of 7-9 years and demonstrated progressive expansions of the GAAs with maximum gain of approximately 9 repeats per year. Continuous GAA expansions throughout the patient's lifespan, as observed in FRDA lymphocytes, should be considered in clinical trial designs and data interpretation.

<h4>Objective</h4>This study was intended to identify genes positively selected in Thoroughbred horses (THBs) that potentially contribute to their running performances.<h4>Methods</h4>The genomes of THB and Jeju horses (JH, Korean native horse) were compared to identify genes positively selected in THB. We performed cross-population extended haplotype homozygosity (XP-EHH) and cross-population composite likelihood ratio test (XP-CLR) statistical methods for our analysis using whole genome resequencing data of 14 THB and 6 JH.<h4>Results</h4>We identified 98 (XP-EHH) and 200 (XP-CLR) genes that are under positive selection in THB. Gene enrichment analysis identified 72 gene ontology biological process (GO BP) terms. The genes and GO BP terms explained some of THB's characteristics such as immunity, energy metabolism and eye size and function related to running performances. GO BP terms that play key roles in several cell signaling mechanisms, which affected ocular size and visual functions were identified. GO BP term Eye photoreceptor cell differentiation is among the terms annotated presumed to affect eye size.<h4>Conclusion</h4>Our analysis revealed some positively selected candidate genes in THB related to their racing performances. The genes detected are related to the immunity, ocular size and function, and energy metabolism.

<h4>Background</h4>Genealogy and molecular genetic studies of a Swedish river valley population resulted in a large pedigree, showing that the hereditary hemochromatosis (HH) <i>HFE/p.</i>C282Y mutation is inherited with other recessive disorders such as Wilson´s disease (WND), a rare recessive disorder of copper overload. The population also contain individuals with the Swedish long QT syndrome (LQTS1) founder mutation (<i>KCNQ1</i>/p.Y111C) which in homozygotes causes the Jervell & Lange Nielsen syndrome (JLNS) and hearing loss (HL).Aims of the study were to test whether the Swedish long QT founder mutation originated in an ancestral HFE family and if carriers had an increased risk for hemochromatosis (HH), a treatable disorder. We also aimed to identify the pathogenic mutation causing the hearing loss disorder segregating in the pedigree.<h4>Methods</h4>LQTS patients were asked about their ancestry and possible origin in a HH family. They were also offered a predictive testing for the HFE genotype. Church books were screened for families with hearing loss. One HH family had two members with hearing loss, who underwent molecular genetic analysis of the LQTS founder mutation, connexin 26 and thereafter exome sequencing. Another family with hearing loss in repeat generations was also analyzed for connexin 26 and underwent exome sequencing.<h4>Results</h4>Of nine LQTS patients studied, four carried a HFE mutation (two p.C282Y, two p.H63D), none was homozygous. Three LQTS patients confirmed origin in a female founder ( b 1694, identical to AJ b 1694, a HFE pedigree member from the Fax river. Her descent of 44 HH families, included also 29 families with hearing loss (HL) suggesting JLNS. Eleven LQTS probands confirmed origin in a second founder couple (b 1614/1605) in which the woman b 1605 was identical to a HFE pedigree member from the Fjällsjö river. In her descent there were not only 64 HH, six WND families, one JLNS, but also 48 hearing loss families. Most hearing loss was non syndromic and caused by founder effects of the late 16^th century. One was of Swedish origin carrying the <i>WHRN,</i> c.1977delC, (p.S660Afs*30) mutation, the other was a <i>TMC1</i>(NM_138691),c.1814T>C,(p.L605P) mutation, possibly of Finnish origin.<h4>Conclusions</h4>Deep human HFE genealogies show HFE to be associated with other genetic disorders like Wilson´s disease, LQTS, JLNS, and autosomal recessive hearing loss. Two new homozygous HL mutations in <i>WHRN</i>/p.S660Afs*30 and <i>TMC1/</i>p.L605P were identified,none of them previously reported from Scandinavia. The rarity of JLNS was possibly caused by miscarriage or intrauterine death. Most hearing loss (81.7%) was seen after 1844 when first cousin marriages were permitted. However, only 10 (10.3%) came from 1^st cousin unions and only 2 (2.0 %) was born out of wedlock.

Background/aim: Huntington disease (HD) is a progressive adult-onset neurodegenerative disorder presenting an autosomal dominant inheritance. Since there is no information on the prevalence of HD in northwestern Iran, the aim of the present study was to determine the prevalence of HD and the number of CAG trinucleotide repeats in the population of northwestern Iran.Materials and methods: Genomic DNA was extracted from whole blood by the salting-out method. DNA samples were analyzed to determine the number of CAG trinucleotide repeats of the HD gene. An abnormally large number of CAG repeats, which is a diagnostic factor for the disease, was detected by polymerase chain reaction and agarose gel electrophoresis.Results: Out of 40 cases, we identified 14 nonkindred individuals with one expanded CAG allele at the IT15 gene. The frequency of the HD mutation in our group of patients was 35%. Expanded alleles varied from 36 to 70 CAG repeats, and normal alleles in HD patients varied from 20 to 26 CAG units. Conclusion: We found a significant correlation between age at onset of the disease and length of the expanded CAG tract: the lower the age, the longer the trinucleotide repeats length.

Alterations of lipid metabolism have been frequently associated with Huntington's disease (HD) over the past years. HD is the most common neurodegenerative disorder, with a complex pathogenic profile, typically characterized by progressive striatal and cortical degeneration and associated motor, cognitive and behavioral disturbances. Previous findings from our group support the idea that disturbed sphingolipid metabolism could represent an additional hallmark of the disease. Although such a defect represents a common biological denominator among multiple disease models ranging from cells to humans through mouse models, more efforts are needed to clearly define its clinical significance and the role it may play in the progression of the disease. In this study, we provided the first evidence of a defective <i>de novo</i> biosynthetic pathway of sphingolipids in multiple HD pre-clinical models. qPCR analysis revealed perturbed gene expression of sphingolipid-metabolizing enzymes in both early and late stage of the disease. In particular, reduction in the levels of sptlc1 and cerS1 mRNA in the brain tissues from manifest HD mice resulted in a significant decrease in the content of dihydroSphingosine, dihydroSphingosine-1-phospahte and dihydroCeramide [C18:0] as assessed by mass spectrometry. Moreover, <i>in vitro</i> studies highlighted the relevant role that aberrant sphingolipid metabolism may have in the HD cellular homeostasis. With this study, we consolidate the evidence of disturbed sphingolipid metabolism in HD and demonstrate for the first time that the <i>de novo</i> biosynthesis pathway is also significantly affected in the disease. This finding further supports the hypothesis that perturbed sphingolipid metabolism may represent a crucial factor accounting for the high susceptibility to disease in HD.

Although the function of the cerebellum in neurocognition has been well-documented, the similar role of the brainstem has yet to be fully elucidated. This clinical research study aimed to combine data relating to neuropsychological assessments and P300 to explore cognitive dysfunction and affective impairment following brainstem stroke. Thirty-four patients with isolated brainstem stroke and twenty-six healthy controls were recruited; for each patient, we collated data pertaining to the P300, Addenbrooke's Cognitive Examination III (ACE-III), Montreal Cognitive Assessment Chinese version (MoCA), trail-making test (TMT), Symbol Digit Modalities Test (SDMT), Wechsler Adult Intelligence Scale-Digit Spans (DS), Stroop test, Self Rating Depression Scale (SDS), and Self Rating Anxiety Scale (SAS). Significance was analyzed using an independent <i>T</i>-test or the Mann-Whitney U-test. Correlation was analyzed using Pearson's correlation analysis or Spearman's correlation analysis. Collectively, data revealed that brainstem stroke caused mild cognitive impairment (MCI), and that visuospatial, attention, linguistic, and emotional disturbances may occur after isolated brainstem stroke. Cognitive decline was linked to P300 latency, ACE-III, and MoCA; P300 latency was correlated with ACE-III. Patients with right brainstem lesions were more likely to suffer memory decline. The present study provides initial data relating to the role of the brainstem in neurocognition, and will be useful for further understanding of vascular cognitive and affective impairment.

Over the past decade, a surge of evidence has documented various pathological processes in the retina of patients suffering from mild cognitive impairment, Alzheimer's disease (AD), Parkinson's disease (PD), and other neurodegenerative diseases. Numerous studies have shown that the retina, a central nervous system tissue formed as a developmental outgrowth of the brain, is profoundly affected by AD. Harboring the earliest detectable disease-specific signs, amyloid β-protein (Aβ) plaques, the retina of AD patients undergoes substantial ganglion cell degeneration, thinning of the retinal nerve fiber layer, and loss of axonal projections in the optic nerve, among other abnormalities. More recent investigations described Aβ plaques in the retina located within sites of neuronal degeneration and occurring in clusters in the mid- and far-periphery of the superior and inferior quadrants, regions that had been previously overlooked. Diverse structural and/or disease-specific changes were also identified in the retina of PD, Huntington's disease, and multiple sclerosis patients. The pathological relationship between the retina and brain prompted the development of imaging tools designed to noninvasively detect and monitor these signs in living patients. One such tool is optical coherence tomography (OCT), uniquely providing high-resolution two-dimensional cross-sectional imaging and three-dimensional volumetric measurements. As such, OCT emerged as a prominent approach for assessing retinal abnormalities <i>in vivo</i>, and indeed provided multiple parameters that allowed for the distinction between normal aged individuals and patients with neurodegenerative diseases. Beyond the use of retinal optical fundus imaging, which recently allowed for the detection and quantification of amyloid plaques in living AD patients <i>via</i> a wide-field view of the peripheral retina, a major advantage of OCT has been the ability to measure the volumetric changes in specified retinal layers. OCT has proven to be particularly useful in analyzing retinal structural abnormalities consistent with disease pathogenesis. In this review, we provide a summary of OCT findings in the retina of patients with AD and other neurodegenerative diseases. Future studies should explore the combination of imaging early hallmark signs together with structural-functional biomarkers in the accessible retina as a practical means of assessing risk, disease progression, and therapeutic efficacy in these patients.

Current applications of human induced pluripotent stem cell (hiPSC) technologies in patient-specific models of neurodegenerative and neuropsychiatric disorders tend to focus on neuronal phenotypes. Here, we review recent efforts toward advancing hiPSCs toward non-neuronal cell types of the central nervous system (CNS) and highlight their potential use for the development of more complex <i>in vitro</i> models of neurodevelopment and disease. We present evidence from previous works in both rodents and humans of the importance of these cell types (oligodendrocytes, microglia, astrocytes) in neurological disease and highlight new hiPSC-based models that have sought to explore these relationships <i>in vitro</i>. Lastly, we summarize efforts toward conducting high-throughput screening experiments with hiPSCs and propose methods by which new screening platforms could be designed to better capture complex relationships between neural cell populations in health and disease.

Pediatric adrenocortical carcinoma and adrenocortical adenoma are two rare diseases affecting children. Molecular analyses were performed to identify commonalities in gene expression between the diseases. Differentially expressed genes were identified for the pediatric adrenocortical adenoma and carcinoma tissues, as compared with normal tissues, using the expression dataset. Protein-protein interaction (PPI) networks were constructed for adenoma and carcinoma disease models, and common modules among the diseases were identified. A total of two common modules with 14 nodes and 20 nodes were revealed among the adenoma and carcinoma networks, respectively. Genes of the common modules were also identified to be the common hub genes of the disease models. Enrichment of the genes of the common modules suggested associations with steroid biosynthesis, the proteasome, cell cycle and metabolic pathways. Modularity, topological and functional analysis of the PPI networks revealed common modules among pediatric adenoma and carcinoma disease models, which provided insight into the underlying disease mechanisms and suggesting prospective targets for future study.

Vertigo is a common complaint in patients who present to the emergency department. It can be a manifestation originating from several different disease processes. Although most patients with vertigo, especially younger patients, will have a benign disorder, up to 3% of such patients will have a cerebellar infarct. Although ruling out these types of fatal diagnoses is essential for emergency medicine physicians, this task can be especially complicated. Classic signs of a cerebellar infarct include symptoms suggestive of central vertigo with focal neurologic deficits on physical exam. Up to 10% of patients with cerebellar infarctions, however, present to the emergency department with vertigo and no focal neurologic deficits. The following case report discusses a male in his late twenties with the chief complaint of vertigo. On initial exam, he had no focal neurologic deficits but did have other concerning symptoms including severe ataxia. Imaging subsequently revealed the patient to have sustained a cerebellar infarct. When differentiating benign forms of vertigo from cerebellar infarcts or other central causes, the clinician should take into account risk factors such as central symptoms including neurologic deficits and severe ataxia. Implementing this strategy may decrease morbidity and mortality associated with cerebellar infarctions.

Huntington's disease (HD) is a neurodegenerative disorder that is caused by abnormal expansion of CAG repeats in the HTT gene. The transcribed mutant RNA contains expanded CAG repeats that translate into a mutant huntingtin protein. This expanded CAG repeat also causes mis-splicing of pre-mRNA due to sequestration of muscle blind like-1 splicing factor (MBNL1), and thus both of these elicit the pathogenesis of HD. Targeting the onset as well as progression of HD by small molecules could be a potent therapeutic approach. We have screened a set of small molecules to target this transcript and found Myricetin, a flavonoid, as a lead molecule that interacts with the CAG motif and thus prevents the translation of mutant huntingtin protein as well as sequestration of MBNL1. Here, we report the first solution structure of the complex formed between Myricetin and RNA containing the 5'CAG/3'GAC motif. Myricetin interacts with this RNA via base stacking at the AA mismatch. Moreover, Myricetin was also found reducing the proteo-toxicity generated due to the aggregation of polyglutamine, and further, its supplementation also improves neurobehavioral deficits in the HD mouse model. Our study provides the structural and mechanistic basis of Myricetin as an effective therapeutic candidate for HD and other polyQ related disorders.

We investigated the role of HFE C282Y, H63D, and TMPRSS6 A736V variants in the pathogenesis of iron deficiency anemia (IDA) in celiac disease (CD) patients, at diagnosis and after 1 year of gluten-free diet (GFD). Demographic and clinical features were prospectively recorded for all CD patients between 2013 and 2017. C282Y, H63D, and A736V variants were evaluated for CD patients and controls. Finally, 505 consecutive CD patients and 539 age-matched control subjects were enrolled. At diagnosis, 229 CD subjects had IDA (45.3%), with a subgroup of anemic patients (45.4%) presented persistent IDA at follow-up. C282Y allele frequency was significantly increased in CD compared with controls (1.1% vs 0.2%, P = .001), whereas H63D and A736V allele frequencies were similar among patients and controls (P = .92 and .84, respectively). At diagnosis, C282Y variant in anemic CD patients was significantly increased compared to nonanemic group (2% and 0.5%, P = .04). At follow-up, A736V was significantly increased in IDA persistent than in IDA not persistent (57.7% vs 35.2%, P < .0001). CD patients with H63D mutation showed higher Hb, MCV, serum iron, and ferritin levels than subjects without HFE mutations. Decreased hepcidin values were observed in anemic compared to nonanemic subjects at follow-up (1.22 ± 1.14 vs 2.08 ± 2.15, P < .001). This study suggests a protective role of HFE in IDA CD patients and confirms the role of TMPRSS6 in predicting oral iron response modulating hepcidin action on iron absorption. Iron supplementation therapeutic management in CD could depend on TMPRSS6 genotype that could predict persistent IDA despite iron supplementation and GFD.

<h4>Background</h4>Barrett's esophagus metaplasia is the key precursor lesion of esophageal adenocarcinoma. The aim of this study was to find a subset of markers that may allow the identification of patients at risk for esophageal adenocarcinoma, and to determine genes differentially expressed in esophageal squamous cell carcinoma.<h4>Methods</h4>Laser capture microdissection technique was applied to procure cells from defined regions. Genome-wide RNA profiling was performed on esophageal adenocarcinoma (n = 21), Barrett's esophagus (n = 20), esophageal squamous carcinoma (n = 9) and healthy esophageal biopsies (n = 18) using the Affymetrix Human Genome U133plus 2.0 array. Microarray results were validated by quantitative real-time polymerase chain reaction in a second and independent cohort and by immunohistochemistry of two putative markers in a third independent cohort.<h4>Results</h4>Through unsupervised hierarchical clustering and principal component analysis, samples were separated into four distinct groups that match perfectly with histology. Many genes down-regulated in esophageal cancers belong to the epidermal differentiation complex or the related GO-group "cornified envelope" of terminally differentiated keratinocytes. Similarly, retinol metabolism was strongly down-regulated. Genes showing strong overexpression in esophageal carcinomas belong to the GO groups extracellular region /matrix such as MMP1, CTHRC1, and INHBA. According to an analysis of genes strongly up-regulated in both esophageal adenocarcinoma and Barrett's esophagus, REG4 might be of particular interest as an early marker for esophageal adenocarcinoma.<h4>Conclusions</h4>Our study provides high quality data, which could serve for identification of potential biomarkers of Barrett's esophagus at risk of esophageal adenocarcinoma progression.

Neuropsychiatric disorders, represented by schizophrenia, affect not only neurons but also myelinating oligodendroglia (OL), both contribute to the complex etiology. Although numerous susceptibility genes for schizophrenia have been identified, their function has been primarily studied in neurons. Whether malfunction of risk genes underlies OL defects in schizophrenia pathogenesis remains poorly understood. In this study, we investigated the function and regulation of the well-recognized schizophrenia risk factor, Fasciculation and Elongation Protein Zeta-1 (FEZ1), in OL. We found that FEZ1 is expressed in oligodendroglia progenitor cells (OPCs) derived from rodent brains and human induced pluripotent stem cells (iPSCs) in culture and in myelinating oligodendrocytes in the brain. In addition, a vigorous upregulation of FEZ1 occurs during OPC differentiation and myelinogenesis, whereas knockdown of FEZ1 significantly attenuates the development of OL process arbors. We further showed that transcription of the Fez1 gene in OL cells is governed by a sophisticated functional interplay between histone acetylation-mediated chromatin modification and transcription factors that are dysregulated in schizophrenia. At the post-transcriptional level, the selective RNA-binding protein QKI, a glia-specific risk factor of schizophrenia, binds FEZ1 mRNA. Moreover, QKI deficiency results in a marked reduction of FEZ1 specifically in OL cells of the quakingviable (qk^v) hypomyelination mutant mice. These observations have uncovered novel pathways that involve multifaceted genetic lesions and/or epigenetic dysregulations in schizophrenia, which converge on FEZ1 regulation and cause OL impairment in neuropsychiatric disorders.

In the present study, we tested whether there were proteomic differences in blood between schizophrenia patients after the initial onset of the disorder and controls; and whether those differences were also present at birth among neonates who later developed schizophrenia compared to those without a psychiatric admission. We used multiple reaction monitoring mass spectrometry to quantify 77 proteins (147 peptides) in serum samples from 60 first-onset drug-naive schizophrenia patients and 77 controls, and 96 proteins (152 peptides) in 892 newborn blood-spot (NBS) samples collected between 1975 and 1985. Both serum and NBS studies showed significant alterations in protein levels. Serum results revealed that Haptoglobin and Plasma protease C1 inhibitor were significantly upregulated in first-onset schizophrenia patients (corrected P < 0.05). Alpha-2-antiplasmin, Complement C4-A and Antithrombin-III were increased in first-onset schizophrenia patients (uncorrected P-values 0.041, 0.036 and 0.013, respectively) and also increased in newborn babies who later develop schizophrenia (P-values 0.0058, 0.013 and 0.044, respectively). We also tested whether protein abundance at birth was associated with exposure to an urban environment during pregnancy and found highly significant proteomic differences at birth between urban and rural environments. The prediction model for urbanicity had excellent predictive performance in both discovery (area under the receiver operating characteristic curve (AUC) = 0.90) and validation (AUC = 0.89) sample sets. We hope that future biomarker studies based on stored NBS samples will identify prognostic disease indicators and targets for preventive measures for neurodevelopmental conditions, particularly those with onset during early childhood, such as autism spectrum disorder.

Low-density lipoprotein receptor-related protein-1 (LRP1) is a large endocytic and signaling molecule broadly expressed by neurons and glia. In adult mice, global inducible (<i>Lrp1^flox/flox;CAG-CreER</i>) or oligodendrocyte (OL)-lineage specific ablation (<i>Lrp1^flox/flox;Pdgfra-CreER</i>) of <i>Lrp1</i> attenuates repair of damaged white matter. In oligodendrocyte progenitor cells (OPCs), <i>Lrp1</i> is required for cholesterol homeostasis and differentiation into mature OLs. <i>Lrp1-</i>deficient OPC/OLs show a strong increase in the sterol-regulatory element-binding protein-2 yet are unable to maintain normal cholesterol levels, suggesting more global metabolic deficits. Mechanistic studies revealed a decrease in peroxisomal biogenesis factor-2 and fewer peroxisomes in OL processes. Treatment of <i>Lrp1^-/-</i> OPCs with cholesterol or activation of peroxisome proliferator-activated receptor-γ with pioglitazone alone is not sufficient to promote differentiation; however, when combined, cholesterol and pioglitazone enhance OPC differentiation into mature OLs. Collectively, our studies reveal a novel role for <i>Lrp1</i> in peroxisome biogenesis, lipid homeostasis, and OPC differentiation during white matter development and repair.

Insufficient folding capacity of the endoplasmic reticulum (ER) activates the unfolded protein response (UPR) to restore homeostasis. Yet, how the UPR achieves ER homeostatic readjustment is poorly investigated, as in most studies the ER stress that is elicited cannot be overcome. Here we show that a proteostatic insult, provoked by persistent expression of the secretory heavy chain of immunoglobulin M (µ_s), is well-tolerated in HeLa cells. Upon µ_s expression, its levels temporarily eclipse those of the ER chaperone BiP, leading to acute, full-geared UPR activation. Once BiP is in excess again, the UPR transitions to chronic, submaximal activation, indicating that the UPR senses ER stress in a ratiometric fashion. In this process, the ER expands about three-fold and becomes dominated by BiP. As the UPR is essential for successful ER homeostatic readjustment in the HeLa-µ_s model, it provides an ideal system for dissecting the intricacies of how the UPR evaluates and alleviates ER stress.

Whereas individual steps of protein-coding gene expression in eukaryotes can be studied in isolation in vitro, it has become clear that these steps are intimately connected within cells. Connections not only ensure quality control but also fine-tune the gene expression process, which must adapt to environmental changes while remaining robust. In this review, we systematically present proven and potential mechanisms by which sequence-specific DNA-binding transcription factors can alter gene expression beyond transcription initiation and regulate pre-mRNA splicing, and thereby mRNA isoform production, by (i) influencing transcription elongation rates, (ii) binding to pre-mRNA to recruit splicing factors, and/or (iii) blocking the association of splicing factors with pre-mRNA. We propose various mechanistic models throughout the review, in some cases without explicit supportive evidence, in hopes of providing fertile ground for future studies.

The host innate immune response mediated by type I interferon (IFN) and the resulting up-regulation of hundreds of interferon-stimulated genes (ISGs) provide an immediate barrier to virus infection. Studies of the type I 'interferome' have mainly been carried out at a single species level, often lacking the power necessary to understand key evolutionary features of this pathway. Here, using a single experimental platform, we determined the properties of the interferomes of multiple vertebrate species and developed a webserver to mine the dataset. This approach revealed a conserved 'core' of 62 ISGs, including genes not previously associated with IFN, underscoring the ancestral functions associated with this antiviral host response. We show that gene expansion contributes to the evolution of the IFN system and that interferomes are shaped by lineage-specific pressures. Consequently, each mammal possesses a unique repertoire of ISGs, including genes common to all mammals and others unique to their specific species or phylogenetic lineages. An analysis of genes commonly down-regulated by IFN suggests that epigenetic regulation of transcription is a fundamental aspect of the IFN response. Our study provides a resource for the scientific community highlighting key paradigms of the type I IFN response.

Nuclear transport receptors (NTRs) recognize localization signals of cargos to facilitate their passage across the central channel of nuclear pore complexes (NPCs). About 30 different NTRs constitute different transport pathways in humans and bind to a multitude of different cargos. The exact cargo spectrum of the majority of NTRs, their specificity and even the extent to which active nucleocytoplasmic transport contributes to protein localization remains understudied because of the transient nature of these interactions and the wide dynamic range of cargo concentrations. To systematically map cargo-NTR relationships <i>in situ</i>, we used proximity ligation coupled to mass spectrometry (BioID). We systematically fused the engineered biotin ligase BirA* to 16 NTRs. We estimate that a considerable fraction of the human proteome is subject to active nuclear transport. We quantified the specificity and redundancy in NTR interactions and identified transport pathways for cargos. We extended the BioID method by the direct identification of biotinylation sites. This approach enabled us to identify interaction interfaces and to discriminate direct versus piggyback transport mechanisms. Data are available via ProteomeXchange with identifier PXD007976.

The intercalated cells (ITCs) of the amygdala have been shown to be critical regulatory components of amygdalar circuits, which control appropriate fear responses. Despite this, the molecular processes guiding ITC development remain poorly understood. Here we establish the zinc finger transcription factor Tshz1 as a marker of ITCs during their migration from the dorsal lateral ganglionic eminence through maturity. Using germline and conditional knock-out (cKO) mouse models, we show that <i>Tshz1</i> is required for the proper migration and differentiation of ITCs. In the absence of <i>Tshz1</i>, migrating ITC precursors fail to settle in their stereotypical locations encapsulating the lateral amygdala and BLA. Furthermore, they display reductions in the ITC marker Foxp2 and ectopic persistence of the dorsal lateral ganglionic eminence marker Sp8. <i>Tshz1</i> mutant ITCs show increased cell death at postnatal time points, leading to a dramatic reduction by 3 weeks of age. In line with this, <i>Foxp2</i>-null mutants also show a loss of ITCs at postnatal time points, suggesting that <i>Foxp2</i> may function downstream of <i>Tshz1</i> in the maintenance of ITCs. Behavioral analysis of male <i>Tshz1</i> cKOs revealed defects in fear extinction as well as an increase in floating during the forced swim test, indicative of a depression-like phenotype. Moreover, <i>Tshz1</i> cKOs display significantly impaired social interaction (i.e., increased passivity) regardless of partner genetics. Together, these results suggest that <i>Tshz1</i> plays a critical role in the development of ITCs and that fear, depression-like and social behavioral deficits arise in their absence.<b>SIGNIFICANCE STATEMENT</b> We show here that the zinc finger transcription factor Tshz1 is expressed during development of the intercalated cells (ITCs) within the mouse amygdala. These neurons have previously been shown to play a crucial role in fear extinction. <i>Tshz1</i> mouse mutants exhibit severely reduced numbers of ITCs as a result of abnormal migration, differentiation, and survival of these neurons. Furthermore, the loss of ITCs in mouse <i>Tshz1</i> mutants correlates well with defects in fear extinction as well as the appearance of depression-like and abnormal social interaction behaviors reminiscent of depressive disorders observed in human patients with distal 18q deletions, including the <i>Tshz1</i> locus.

Neuroticism is a relatively stable personality trait characterized by negative emotionality (for example, worry and guilt) ¹ ; heritability estimated from twin studies ranges from 30 to 50% ² , and SNP-based heritability ranges from 6 to 15% ^3-6 . Increased neuroticism is associated with poorer mental and physical health ^7,8 , translating to high economic burden ⁹ . Genome-wide association studies (GWAS) of neuroticism have identified up to 11 associated genetic loci ^3,4 . Here we report 116 significant independent loci from a GWAS of neuroticism in 329,821 UK Biobank participants; 15 of these loci replicated at P < 0.00045 in an unrelated cohort (N = 122,867). Genetic signals were enriched in neuronal genesis and differentiation pathways, and substantial genetic correlations were found between neuroticism and depressive symptoms (r _g = 0.82, standard error (s.e.) = 0.03), major depressive disorder (MDD; r _g = 0.69, s.e. = 0.07) and subjective well-being (r _g = -0.68, s.e. = 0.03) alongside other mental health traits. These discoveries significantly advance understanding of neuroticism and its association with MDD.

The present study aimed to examine potential crucial genes associated with Parkinson's disease (PD) in addition to the interactions and regulators of these genes. The chip data (GSE7621) were obtained from the Gene Expression Omnibus and standardized using the robust multi‑array average in the Affy package of R software. The differentially expressed genes (DEGs) were then screened using the Samr package with a false discovery rate (FDR) <0.05 and |log2 fold change (FC)|>1. Crucial PD‑associated genes were predicted using the Genetic Association Database in the Database for Annotation, Visualization and Integrated Discovery and sequence alignment. Furthermore, transcription factors (TFs) of the crucial PD‑associated genes were predicted, and protein‑protein interactions (PPIs) between the crucial PD‑associated genes were analyzed using the Search Tool for the Retrieval of Interacting Genes/Proteins. Additionally, another dataset of PD was used to validate the expression of crucial PD‑associated genes. A total of 670 DEGs (398 upregulated and 272 downregulated genes) were identified in the PD samples. Of these, 10 DEGs enriched in pathways associated with the nervous system were predicted to be crucial in PD, including C‑X‑C chemokine receptor type 4 (CXCR4), deleted in colorectal cancer (DCC) and NCL adaptor protein 2 (NCK2). All 10 genes were associated with neuron development and differentiation. They were simultaneously modulated by multiple TFs, including GATA, E2F and E4 promoter‑binding protein 4. The PPI networks showed that DCC and CXCR4 were hub proteins. The DCC‑netrin 1‑roundabout guidance receptor 2‑slit guidance ligand 1 interaction pathway, and several genes, including TOX high mobility group box family member 4, kinase insert domain receptor and zymogen granule protein 16B, which interacted with CXCR4, were novel findings. Additionally, CXCR4 and NCK2 were upregulated in another dataset (GSE8397) of PD. These genes, interactions of proteins and TFs may be important in the progression of PD.

The number of obesity cases is rapidly increasing in developed and developing countries, thereby causing significant health problems worldwide. The pathologic factors of obesity at the molecular level are not fully characterized, although the imbalance between energy intake and consumption is widely recognized as the main reason for fat accumulation. Previous studies reported that obesity can be caused by the dysfunction of genes associated with other diseases, such as myocardial infarction, hence providing new insights into dissecting the pathogenesis of obesity by investigating its associations with other diseases. In this study, we investigated the relationship between obesity and diseases from Online Mendelian Inheritance in Man (OMIM) databases on the protein-protein interaction (PPI) network. The obesity genes and genes of one OMIM disease were mapped onto the network, and the interaction scores between the two gene sets were investigated on the basis of the PPI of individual gene pairs, thereby inferring the relationship between obesity and this disease. Results suggested that diseases related to nutrition and endocrine are the top two diseases that are closely associated with obesity. This finding is consistent with our general knowledge and indicates the reliability of our obtained results. Moreover, we inferred that diseases related to psychiatric factors and bone may also be highly related to obesity because the two diseases followed the diseases related to nutrition and endocrine according to our results. Numerous obesity-disease associations were identified in the literature to confirm the relationships between obesity and the aforementioned four diseases. These new results may help understand the underlying molecular mechanisms of obesity-disease co-occurrence and provide useful insights for disease prevention and intervention.

Inter-individual variability in blood pressure (BP) is influenced by both genetic and non-genetic factors including socioeconomic and psychosocial stressors. A deeper understanding of the gene-by-socioeconomic/psychosocial factor interactions on BP may help to identify individuals that are genetically susceptible to high BP in specific social contexts. In this study, we used a genomic region-based method for longitudinal analysis, Longitudinal Gene-Environment-Wide Interaction Studies (LGEWIS), to evaluate the effects of interactions between known socioeconomic/psychosocial and genetic risk factors on systolic and diastolic BP in four large epidemiologic cohorts of European and/or African ancestry. After correction for multiple testing, two interactions were significantly associated with diastolic BP. In European ancestry participants, outward/trait anger score had a significant interaction with the <i>C10orf107</i> genomic region (<i>p</i> = 0.0019). In African ancestry participants, depressive symptom score had a significant interaction with the <i>HFE</i> genomic region (<i>p</i> = 0.0048). This study provides a foundation for using genomic region-based longitudinal analysis to identify subgroups of the population that may be at greater risk of elevated BP due to the combined influence of genetic and socioeconomic/psychosocial risk factors.

Increasing evidence suggests that undernutrition during the fetal period may lead to glucose intolerance, impair the insulin response and induce insulin resistance (IR). Considering the importance of chromium (Cr) in maintaining carbohydrate metabolism, the present study aimed to determine the effects of maternal low Cr (LC) on glucose metabolism in C57BL mice offspring, and the involved mechanisms. Weaned C57BL mice were born from mothers fed a control diet or LC diet, and were then fed a control or LC diet for 13 weeks. Subsequently, the liver microRNA (miRNA/miR) expression profile was analyzed by miRNA array analysis. A maternal LC diet increased fasting serum glucose (P<0.05) and insulin levels (P<0.05), homeostasis model assessment of IR index (P<0.01), and the area under curve for glucose concentration during oral glucose tolerance test (P<0.01). In addition, 8 upregulated and 6 downregulated miRNAs were identified in the maternal LC group (fold change ≥2, P<0.05). miRNA‑gene networks, Kyoto Encyclopedia of Genes and Genomes pathway analysis of differentially expressed miRNAs, and miRNA overexpression in HepG2 cells revealed the critical role of insulin signaling, via miR‑327, miR‑466f‑3p and miR‑223‑3p, in the effects of early life Cr restriction on glucose metabolism. In conclusion, maternal Cr restriction may irreversibly increase IR, which may involve a specific miRNA affecting the insulin signaling pathway.

Thousands of long noncoding RNAs (lncRNAs) have been identified in eukaryotic genomes, many of which are expressed in spatially and temporally restricted patterns. Nonetheless, the roles of the majority of these transcripts are still unknown. One of the mechanisms by which lncRNAs function is through the modulation of chromatin states. To assess the functions of lncRNAs, we developed RNA-DamID, a novel approach that detects lncRNA-genome interactions in a cell-type-specific manner in vivo with high sensitivity and accuracy. Identifying the cell-type-specific genome occupancy of lncRNAs is vital to understanding their mechanisms of action in development and disease. We used RNA-DamID to investigate targeting of the lncRNAs in the Drosophila dosage-compensation complex (DCC) and show that initial targeting is cell-type specific.

Trace elements are chemical elements needed in minute amounts for normal physiology. Some of the physiologically relevant trace elements include iodine, copper, iron, manganese, zinc, selenium, cobalt and molybdenum. Of these, some are metals, and in particular, transition metals. The different electron shells of an atom carry different energy levels, with those closest to the nucleus being lowest in energy. The number of electrons in the outermost shell determines the reactivity of such an atom. The electron shells are divided in sub-shells, and in particular the third shell has s, p and d sub-shells. Transition metals are strictly defined as elements whose atom has an incomplete d sub-shell. This incomplete d sub-shell makes them prone to chemical reactions, particularly redox reactions. Transition metals of biologic importance include copper, iron, manganese, cobalt and molybdenum. Zinc is not a transition metal, since it has a complete d sub-shell. Selenium, on the other hand, is strictly speaking a nonmetal, although given its chemical properties between those of metals and nonmetals, it is sometimes considered a metalloid. In this review, we summarize the current knowledge on the inborn errors of metal and metalloid metabolism.

<h4>Background</h4>According to the Compendium of Materia Medica, Gastrodia elata (GE) Blume is a top-grade herbal medicine frequently used to treat dizziness, headaches, tetanus, and epilepsy, suggesting that it affects neurological functions. Although studies have supported its effects in preventing diverse neurodegenerations such as Huntington's disease (HD), its mechanisms require further investigation.<h4>Purpose</h4>To investigate the ability of the molecular mechanism of GE to prevent mutant huntingtin (mHTT) protein aggregation by focusing on mitochondrial function and biogenesis, which have been proposed as the therapeutic targets of HD.<h4>Study design/methods</h4>mHtt overexpression in pheochromocytoma (PC12) cells was used as an in vitro cell model of HD. A retardation assay was applied to measure protein aggregation during Htt expression. Cotransfection with transcriptional genes was used to test their relationships with HTT aggregates by monitoring with a confocal laser scanning microscope. Western blot analysis was used to estimate protein expression under different drug treatments or when cotransfected with other related genes.<h4>Results</h4>Mutant, abnormal Htt overexpression resulted in significant protein aggregation in PC12 cells. GE dose-dependently attenuated mHTT aggregates and increased cyclic-AMP response element-binding protein (CREB) phosphorylation. Adenosine A_2A-R receptor (A_2A-R) antagonist counteracted these phenomena. CREB overexpression significantly attenuated mHTT aggregation. GE increased the promoter activity and expression of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). Furthermore, wild-type PGC-1α but not mutant PGC-1α overexpression attenuated mHTT aggregates.<h4>Conclusion</h4>GE attenuated mHtt aggregation by mediating mitochondrial function and biogenesis through the A_2A-R/PKA/CREB/PGC-1α-dependent pathway.

Genome-wide association studies have uncovered common variants at many loci influencing human complex traits and diseases, such as high-density lipoprotein cholesterol (HDL-C). However, the contribution of the identified genes is difficult to ascertain from current efforts interrogating common variants with small effects. Thus, there is a pressing need for scalable, cost-effective strategies for uncovering causal variants, many of which may be rare and noncoding. Here, we used a multiplexed inversion probe (MIP) target capture approach to resequence both coding and regulatory regions at seven HDL-C associated loci in 797 individuals with extremely high HDL-C vs. 735 low-to-normal HDL-C controls. Our targets included protein-coding regions of GALNT2, APOA5, APOC3, SCARB1, CCDC92, ZNF664, CETP , and LIPG (>9 kb), and proximate noncoding regulatory features (>42 kb). Exome-wide genotyping in 1,114 of the 1,532 participants yielded a >90% genotyping concordance rate with MIP-identified variants in ~90% of participants. This approach rediscovered nearly all established GWAS associations in GALNT2, CETP , and LIPG loci with significant and concordant associations with HDL-C from our phenotypic-extremes design at 0.1% of the sample size of lipid GWAS studies. In addition, we identified a novel, rare, CETP noncoding variant enriched in the extreme high HDL-C group (P<0.01, Score Test). Our targeted resequencing of individuals at the HDL-C phenotypic extremes offers a novel, efficient, and cost-effective approach for identifying rare coding and noncoding variation differences in extreme phenotypes and supports the rationale for applying this methodology to uncover rare variation—particularly non-coding variation--underlying myriad complex traits.

Macroautophagy/autophagy is the main intracellular catabolic pathway in neurons that eliminates misfolded proteins, aggregates and damaged organelles associated with ageing and neurodegeneration. Autophagy is regulated by both MTOR-dependent and -independent pathways. There is increasing evidence that autophagy is compromised in neurodegenerative disorders, which may contribute to cytoplasmic sequestration of aggregation-prone and toxic proteins in neurons. Genetic or pharmacological modulation of autophagy to promote clearance of misfolded proteins may be a promising therapeutic avenue for these disorders. Here, we demonstrate robust autophagy induction in motor neuronal cells expressing SOD1 or TARDBP/TDP-43 mutants linked to amyotrophic lateral sclerosis (ALS). Treatment of these cells with rilmenidine, an anti-hypertensive agent and imidazoline-1 receptor agonist that induces autophagy, promoted autophagic clearance of mutant SOD1 and efficient mitophagy. Rilmenidine administration to mutant SOD1^G93A mice upregulated autophagy and mitophagy in spinal cord, leading to reduced soluble mutant SOD1 levels. Importantly, rilmenidine increased autophagosome abundance in motor neurons of SOD1^G93A mice, suggesting a direct action on target cells. Despite robust induction of autophagy in vivo, rilmenidine worsened motor neuron degeneration and symptom progression in SOD1^G93A mice. These effects were associated with increased accumulation and aggregation of insoluble and misfolded SOD1 species outside the autophagy pathway, and severe mitochondrial depletion in motor neurons of rilmenidine-treated mice. These findings suggest that rilmenidine treatment may drive disease progression and neurodegeneration in this mouse model due to excessive mitophagy, implying that alternative strategies to beneficially stimulate autophagy are warranted in ALS.

Pathological developments leading to amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) are associated with misbehavior of several key proteins, such as SOD1 (superoxide dismutase 1), TARDBP/TDP-43, FUS, C9orf72, and dipeptide repeat proteins generated as a result of the translation of the intronic hexanucleotide expansions in the C9orf72 gene, PFN1 (profilin 1), GLE1 (GLE1, RNA export mediator), PURA (purine rich element binding protein A), FLCN (folliculin), RBM45 (RNA binding motif protein 45), SS18L1/CREST, HNRNPA1 (heterogeneous nuclear ribonucleoprotein A1), HNRNPA2B1 (heterogeneous nuclear ribonucleoprotein A2/B1), ATXN2 (ataxin 2), MAPT (microtubule associated protein tau), and TIA1 (TIA1 cytotoxic granule associated RNA binding protein). Although these proteins are structurally and functionally different and have rather different pathological functions, they all possess some levels of intrinsic disorder and are either directly engaged in or are at least related to the physiological liquid-liquid phase transitions (LLPTs) leading to the formation of various proteinaceous membrane-less organelles (PMLOs), both normal and pathological. This review describes the normal and pathological functions of these ALS- and FTLD-related proteins, describes their major structural properties, glances at their intrinsic disorder status, and analyzes the involvement of these proteins in the formation of normal and pathological PMLOs, with the ultimate goal of better understanding the roles of LLPTs and intrinsic disorder in the "Dr. Jekyll-Mr. Hyde" behavior of those proteins.

The E3 ubiquitin ligase NEDD4 has been intensively studied in processes involved in viral infections, such as virus budding. However, little is known about its functions in bacterial infections. Our investigations into the role of NEDD4 in intracellular bacterial infections demonstrate that Mycobacterium tuberculosis and Listeria monocytogenes, but not Mycobacterium bovis BCG, replicate more efficiently in NEDD4 knockdown macrophages. In parallel, NEDD4 knockdown or knockout impaired basal macroautophagy/autophagy, as well as infection-induced autophagy. Conversely, NEDD4 expression promoted autophagy in an E3 catalytic activity-dependent manner, thereby restricting intracellular Listeria replication. Mechanistic studies uncovered that endogenous NEDD4 interacted with BECN1/Beclin 1 and this interaction increased during Listeria infection. Deficiency of NEDD4 resulted in elevated K48-linkage ubiquitination of endogenous BECN1. Further, NEDD4 mediated K6- and K27- linkage ubiquitination of BECN1, leading to elevated stability of BECN1 and increased autophagy. Thus, NEDD4 participates in killing of intracellular bacterial pathogens via autophagy by sustaining the stability of BECN1.

Pathogen-host protein-protein interaction systems examine the interactions between the protein repertoires of 2 distinct organisms. Some of these pathogen proteins interact with the host protein system and may manipulate it for their own advantages. In this work, we designed an R script by concatenating 2 functions called rowDM and rowCVmed to infer pathogen-host interaction using previously reported microarray data, including host gene enrichment analysis and the crossing of interspecific domain-domain interactions. We applied this script to the <i>Toxoplasma</i>-host system to describe pathogen survival mechanisms from human, mouse, and <i>Toxoplasma</i> Gene Expression Omnibus series. Our outcomes exhibited similar results with previously reported microarray analyses, but we found other important proteins that could contribute to toxoplasma pathogenesis. We observed that <i>Toxoplasma</i> ROP38 is the most differentially expressed protein among toxoplasma strains. Enrichment analysis and KEGG mapping indicated that the human retinal genes most affected by <i>Toxoplasma</i> infections are those related to antiapoptotic mechanisms. We suggest that proteins PIK3R1, PRKCA, PRKCG, PRKCB, HRAS, and c-JUN could be the possible substrates for differentially expressed <i>Toxoplasma</i> kinase ROP38. Likewise, we propose that <i>Toxoplasma</i> causes overexpression of apoptotic suppression human genes.

The microRNA (miRNA) expression profiles and their biological functions in calcium oxalate nephrolithiasis remain unclear. In this study, we investigate the miRNA and mRNA expression profiles of kidney tissues in calcium oxalate stone rats. 16 Sprague Dawley rats were divided into control group and stone-forming group. 24-hour urine samples and kidney tissues were collected for biochemical and histological determination after 4 weeks. MiRNA and mRNA microarray were applied to evaluate the miRNA and mRNA expression profiles. To validate the microarray results, the quantitative real-time PCR (qRT-PCR) was performed. A total of 38 miRNAs and 2728 mRNAs were significantly and differentially expressed in kidney tissues of stone-forming group versus control group. Gene Ontology (GO) analysis revealed that most of the target genes were enriched in terms of oxidation reduction, ion transport, inflammatory response, and response to wounding. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis of these targets highlights their critical role in cytokine-cytokine receptor interaction, gap junction, and chemokine signaling pathway. Furthermore, the reliability of the microarray-based results was confirmed by using qRT-PCR determination. The miRNA and mRNA expressions in calcium oxalate stone rat kidneys might provide a basis for further research on urolithiasis mechanism.

<h4>Background</h4>Pseudomonas aeruginosa is an opportunistic pathogen that causes serious infections in immunocompromised hosts including severely burned patients. In burn patients, P. aeruginosa infection often leads to septic shock and death. Despite numerous studies, the influence of severe thermal injuries on the pathogenesis of P. aeruginosa during systemic infection is not known. Through RNA-seq analysis, we recently showed that the growth of P. aeruginosa strain UCBPP-PA14 (PA14) in whole blood obtained from severely burned patients significantly altered the expression of the PA14 transcriptome when compared with its growth in blood from healthy volunteers. The expression of PA14_23430 and the adjacent gene, PA14_23420, was enhanced by seven- to eightfold under these conditions.<h4>Results</h4>Quantitative real-time PCR analysis confirmed the enhancement of expression of both PA14_23420 and PA14_23430 by growth of PA14 in blood from severely burned patients. Computer analysis revealed that PA14_23430 (hepP) encodes a potential heparinase while PA14_23420 (zbdP) codes for a putative zinc-binding dehydrogenase. This analysis further suggested that the two genes form an operon with zbdP first. Presence of the operon was confirmed by RT-PCR experiments. We characterized hepP and its protein product HepP. hepP was cloned from PA14 by PCR and overexpressed in E. coli. The recombinant protein (rHepP) was purified using nickel column chromatography. Heparinase assays using commercially available heparinase as a positive control, revealed that rHepP exhibits heparinase activity. Mutation of hepP resulted in delay of pellicle formation at the air-liquid interface by PA14 under static growth conditions. Biofilm formation by PA14ΔhepP was also significantly reduced. In the Caenorhabditis elegans model of slow killing, mutation of hepP resulted in a significantly lower rate of killing than that of the parent strain PA14.<h4>Conclusions</h4>Changes within the blood of severely burned patients significantly induced expression of hepP in PA14. The heparinase encoded by hepP is a potential virulence factor for PA14 as HepP influences pellicle formation as well as biofilm development by PA14 and the protein is required for full virulence in the C. elegans model of slow killing.

<h4>Purpose</h4>Metronomic capecitabine (MC) is a well-tolerated systemic treatment showing promising results in one retrospective study, as second-line therapy after sorafenib failure, in patients with hepatocellular carcinoma (HCC).<h4>Methods</h4>117 patients undergoing MC were compared to 112 patients, eligible for this treatment, but undergoing best supportive care (BSC) after sorafenib discontinuation for toxicity or HCC progression. The two groups were compared for demographic and clinical features. A multivariate regression analysis was conducted to detect independent prognostic factors. To balance confounding factors between the two groups, a propensity score model based on independent prognosticators (performance status, neoplastic thrombosis, causes of sorafenib discontinuation and pre-sorafenib treatment) was performed.<h4>Results</h4>Patients undergoing MC showed better performance status, lower tumor burden, lower prevalence of portal vein thrombosis, and better cancer stage. Median (95% CI) post-sorafenib survival (PSS) was longer in MC than in BSC patients [9.5 (7.5-11.6) vs 5.0 (4.2-5.7) months (p < 0.001)]. Neoplastic thrombosis, cause of sorafenib discontinuation, pre-sorafenib treatment and MC were independent prognosticators. The benefit of capecitabine was confirmed in patients after matching with propensity score [PSS: 9.9 (6.8-12.9) vs. 5.8 (4.8-6.8) months, (p = 0.001)]. MC lowered the mortality risk by about 40%. MC achieved better results in patients who stopped sorafenib for adverse events than in those who progressed during it [PSS: 17.3 (10.5-24.1) vs. 7.8 (5.2-10.1) months, (p = 0.035)]. Treatment toxicity was low and easily manageable with dose modulation.<h4>Conclusions</h4>MC may be an efficient and safe second-line systemic therapy for HCC patients who discontinued sorafenib for toxicity or tumor progression.

Over the past few years, multiple immune checkpoint blockers (ICBs) have achieved unprecedented clinical success and have been approved by regulatory agencies for the treatment of an increasing number of malignancies. However, only a limited fraction of patients responds to ICBs employed as a standalone intervention, calling for the development of combinatorial regimens. Radiation therapy (RT) stands out as a very promising candidate for this purpose. Indeed, RT mediates antineoplastic effects not only by cytotoxic and cytostatic mechanisms, but also by modulating immunological functions, both locally (within the irradiated field) and systemically. As combinatorial regimens involving RT and ICBs are being developed and clinically tested at an accelerating pace, it is paramount to identify biomarkers that reliably predict the likelihood of individual patients to respond. Here, we discuss emerging biomarkers that may potentially predict the response of cancer patients to RT plus ICBs.

<h4>Objective</h4>To assess the impact of osteoarthritis (OA) on the meniscus by comparing transcripts and biological processes in the meniscus between patients with and without OA.<h4>Design</h4>RNA microarrays were used to identify transcripts differentially expressed (DE) in meniscus obtained from 12 OA and 12 non-OA patients. The non-OA specimens were obtained at the time of arthroscopic partial meniscectomy. Real-time PCR was performed on selected transcripts. Biological processes and gene-networking was examined computationally. Transcriptome signatures were mapped with 37 OA-related transcripts to evaluate how meniscus gene expression relates to that of OA cartilage.<h4>Results</h4>We identified 168 transcripts significantly DE between OA (75 elevated, 93 repressed) and non-OA samples (≥1.5-fold). Among these, CSN1S1, COL10A1, WIF1, and SPARCL1 were the most prominent transcripts elevated in OA meniscus, POSTN and VEGFA were most highly repressed in OA meniscus. Transcripts elevated in OA meniscus represented response to external stimuli, cell migration and cell localization while those repressed in OA meniscus represented histone deacetylase activity (related to epigenetics) and skeletal development. Numerous long non-coding RNAs (lncRNAs) were DE between the two groups. When segregated by OA-related transcripts, two distinct clustering patterns appeared: OA meniscus appeared to be more inflammatory while non-OA meniscus exhibited a "repair" phenotype.<h4>Conclusions</h4>Numerous transcripts with potential relevance to the pathogenesis of OA are DE in OA and non-OA meniscus. These data suggest an involvement of epigenetically regulated histone deacetylation in meniscus tears as well as expression of lncRNAs. Patient clustering based on transcripts related to OA in articular cartilage confirmed distinct phenotypes between injured (non-OA) and OA meniscus.

Although genome-wide association studies (GWAS) have identified several variants linked to depression, few GWAS of non-European populations have been performed. We conducted a genome-wide analysis of depression in a large, population-based sample of Hispanics/Latinos. Data came from 12,310 adults in the Hispanic Community Health Study/Study of Latinos (HCHS/SOL). Past-week depressive symptoms were assessed using the 10-item Center for Epidemiological Studies of Depression Scale. Three phenotypes were examined: a total depression score, a total score modified to account for psychiatric medication use, and a score excluding anti-depressant medication users. We estimated heritability due to common variants (h²_SNP), and performed a GWAS of the three phenotypes. Replication was attempted in three independent Hispanic/Latino cohorts. We also performed sex-stratified analyses, analyzed a binary trait indicating probable depression, and conducted three trans-ethnic analyses. The three phenotypes exhibited significant heritability (h²_SNP = 6.3-6.9%; p = .002) in the total sample. No SNPs were genome-wide significant in analyses of the three phenotypes or the binary indicator of probable depression. In sex-stratified analyses, seven genome-wide significant SNPs (one in females; six in males) were identified, though none were supported through replication. Four out of 24 loci identified in prior GWAS were nominally associated in HCHS/SOL. There was no evidence of overlap in genetic risk factors across ancestry groups, though this may have been due to low power. We conducted the largest GWAS of depression-related phenotypes in Hispanic/Latino adults. Results underscore the genetic complexity of depressive symptoms as a phenotype in this population and suggest the need for much larger samples.

<h4>Objective</h4>To determine whether indeterminate (Likert-score 3/5) peripheral zone (PZ) multiparametric MRI (mpMRI) studies are classifiable by prostate-specific antigen (PSA), PSA density (PSAD), Prostate Imaging Reporting And Data System version 2 (PI-RADS_v2) rescoring and morphological MRI features.<h4>Methods</h4>Men with maximum Likert-score 3/5 within their PZ were retrospectively selected from 330 patients who prospectively underwent prostate mpMRI (3 T) without an endorectal coil, followed by 20-zone transperineal template prostate mapping biopsies ＋/- focal lesion-targeted biopsy. PSAD was calculated using pre-biopsy PSA and MRI-derived volume. Two readers A and B independently assessed included men with both Likert-assessment and PI-RADS_v2. Both readers then classified mpMRI morphological features in consensus. Men were divided into two groups: significant cancer (≥ Gleason 3 ＋ 4) or insignificant cancer (≤ Gleason 3 ＋ 3)/no cancer. Comparisons between groups were made separately for PSA & PSAD using Mann-Whitney test and morphological descriptors with Fisher's exact test. PI-RADS_v2 and Likert-assessment were descriptively compared and percentage inter-reader agreement calculated.<h4>Results</h4>76 males were eligible for PSA & PSAD analyses, 71 for PI-RADS scoring, and 67 for morphological assessment (excluding significant image artefacts). Unlike PSA (p = 0.915), PSAD was statistically different (p = 0.004) between the significant [median: 0.19 ng ml^-² (interquartile range: 0.13-0.29)] and non-significant/no cancer [median: 0.13 ng ml^-² (interquartile range: 0.10-0.17)] groups. Presence of mpMRI morphological features was not significantly different between groups. Subjective Likert-assessment discriminated patients with significant cancer better than PI-RADS_v2. Inter-reader percentage agreement was 83% for subjective Likert-assessment and 56% for PI-RADS_v2.<h4>Conclusion</h4>PSAD may categorize presence of significant cancer in patients with Likert-scored 3/5 PZ mpMRI findings. Advances in knowledge: PSAD may be used in indeterminate PZ mpMRI to guide decisions between biopsy vs monitoring.

Black rice (<i>Oryza sativa</i> L.) is considered to be a healthy food due to its high content of anthocyanins in the pericarp. The synthetic pathway of anthocyanins in black rice grains has been identified, however, the proteomic profile of leaves during grain development is still unclear. Here, isobaric Tags Relative and Absolute Quantification (iTRAQ) MS/MS was carried out to identify statistically significant changes of leaf proteome in the black rice during grain development. Throughout three sequential developmental stages, a total of 3562 proteins were detected and 24 functional proteins were differentially expressed 3-10 days after flowering (DAF). The detected proteins are known to be involved in various biological processes and most of these proteins were related to gene expression regulatory (33.3%), signal transduction (16.7%) and developmental regulation and hormone-like proteins (12.5%). The coordinated changes were consistent with changes in regulatory proteins playing a leading role in leaves during black rice grain development. This indicated that signal transduction between leaves and grains may have an important role in anthocyanin biosynthesis and accumulation during grain development of black rice. In addition, four identified up-regulated proteins associated with starch metabolism suggested that the remobilization of nutrients for starch synthesis plays a potential role in anthocyanin biosynthesis of grain. The mRNA transcription for eight selected proteins was validated with quantitative real-time PCR. Our results explored the proteomics of the coordination between leaf and grain in anthocyanins biosynthesis of grain, which might be regulated by signal transduction and sugar metabolism in black rice leaf.

<h4>Objectives</h4>The collagen VI related muscular dystrophies (COL6-RD), Ullrich congenital muscular dystrophy (UCMD) and Bethlem myopathy (BM) are among the most common congenital muscular dystrophies and are characterized by distal joint laxity and a combination of distal and proximal joint contractures. Inheritance can be dominant negative (DN) or recessive depending on the type and location of the mutation. DN mutations allow incorporation of abnormal chains into secreted tetramers and are the most commonly identified mutation type in COL6-RD. Null alleles (nonsense, frameshift, and large deletions) do not allow incorporation of abnormal chains and act recessively. To better define the pathways disrupted by mutations in collagen VI, we have used a transcriptional profiling approach with RNA-Seq to identify differentially expressed genes in COL6-RD individuals from controls.<h4>Methods</h4>RNA-Seq allows precise detection of all expressed transcripts in a sample and provides a tool for quantification of expression data on a genomic scale. We have used RNA-Seq to identify differentially expressed genes in cultured dermal fibroblasts from 13 COL6-RD individuals (8 dominant negative and 5 null) and 6 controls. To better assess the transcriptional changes induced by abnormal collagen VI in the extracellular matrix (ECM); we compared transcriptional profiles from subjects with DN mutations and subjects with null mutations to transcriptional profiles from controls.<h4>Results</h4>Differentially expressed transcripts between COL6-RD and control fibroblasts include upregulation of ECM components and downregulation of factors controlling matrix remodeling and repair. DN and null samples are differentiated by downregulation of genes involved with DNA replication and repair in null samples.<h4>Conclusions</h4>Differentially expressed genes identified here may help identify new targets for development of therapies and biomarkers to assess the efficacy of treatments.

Many Preterm-born children suffer from neurobehavioral disorders. Premature birth terminates the hypoxic <i>in utero</i> environment and supply of maternal hormones. As the production of interneurons continues until the end of pregnancy, we hypothesized that premature birth would disrupt interneuron production and that restoration of the hypoxic milieu or estrogen treatment might reverse interneuron generation. To test these hypotheses, we compared interneuronal progenitors in the medial ganglionic eminences (MGEs), lateral ganglionic eminences (LGEs), and caudal ganglionic eminences (CGEs) between preterm-born [born on embryonic day (E) 29; examined on postnatal day (D) 3 and D7] and term-born (born on E32; examined on D0 and D4) rabbits at equivalent postconceptional ages. We found that both total and cycling Nkx2.1⁺, Dlx2⁺, and Sox2⁺ cells were more abundant in the MGEs of preterm rabbits at D3 compared with term rabbits at D0, but not in D7 preterm relative to D4 term pups. Total Nkx2.1⁺ progenitors were also more numerous in the LGEs of preterm pups at D3 compared with term rabbits at D0. Dlx2⁺ cells in CGEs were comparable between preterm and term pups. Simulation of hypoxia by dimethyloxalylglycine treatment did not affect the number of interneuronal progenitors. However, estrogen treatment reduced the density of total and proliferating Nkx2.1⁺ and Dlx2⁺ cells in the MGEs and enhanced Ascl1 transcription factor. Estrogen treatment also reduced Ki67, c-Myc, and phosphorylation of retinoblastoma protein, suggesting inhibition of the G1-to-S phase transition. Hence, preterm birth disrupts interneuron neurogenesis in the MGE and estrogen treatment reverses interneuron neurogenesis in preterm newborns by cell-cycle inhibition and elevation of Ascl1. We speculate that estrogen replacement might partially restore neurogenesis in human premature infants.<b>SIGNIFICANCE STATEMENT</b> Prematurity results in developmental delays and neurobehavioral disorders, which might be ascribed to disturbances in the development of cortical interneurons. Here, we show that preterm birth disrupts interneuron neurogenesis in the medial ganglionic eminence (MGE) and, more importantly, that estrogen treatment reverses this perturbation in the population of interneuron progenitors in the MGE. The estrogen seems to restore neurogenesis by inhibiting the cell cycle and elevating Ascl1 expression. As preterm birth causes plasma estrogen level to drop 100-fold, the estrogen replacement in preterm infants is physiological. We speculate that estrogen replacement might ameliorate disruption in production of interneurons in human premature infants.

Parasitic B chromosomes invade and persist in natural populations through several mechanisms for transmission advantage (drive). They may contain gene-derived sequences which, in some cases, are actively transcribed. A further interesting question is whether B-derived transcripts become functional products. In the grasshopper Eyprepocnemis plorans, one of the gene-derived sequences located on the B chromosome shows homology with the gene coding for the CAP-G subunit of condensin I. We show here, by means of fluorescent in situ hybridization coupled with tyramide signal amplification (FISH-TSA), that this gene is located in the distal region of the B24 chromosome variant. The DNA sequence located in the B chromosome is a pseudogenic version of the CAP-G gene (B-CAP-G). In two Spanish populations, we found active transcription of B-CAP-G, but it did not influence the expression of CAP-D2 and CAP-D3 genes coding for corresponding condensin I and II subunits, respectively. Our results indicate that the transcriptional regulation of the B-CAP-G pseudogene is uncoupled from the standard regulation of the genes that constitute the condensin complex, and suggest that some of the B chromosome known effects may be related with its gene content and transcriptional activity, thus opening new exciting avenues for research.

<h4>Purpose</h4>Retinoblastoma (Rb) is the most common intraocular tumor in children. MicroRNAs (miRNAs) play a crucial role in gene regulation and cell growth/apoptosis/differentiation. The current study aimed to investigate the role of miR-498 in Rb.<h4>Methods</h4>Quantitative real-time polymerase chain reaction (QRT-PCR) was used to test mRNA level of miR-498. http://www.targetscan.org and http://www.microrna.org were applied to predict target of miR-498. Dual-luciferase reporter assay was applied to investigate if miR-498 targeted cell cycle progression 1 (CCPG1). Western blot (WB) was carried out to assess CCPG1 protein levels. 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) assay was used to evaluate cell proliferation. Annexin-V Fluorescein (FITC) was adopted to explore cell apoptosis.<h4>Results</h4>In Y79 cells, miR-498 was higher than in normal ARPE-19 cells. MiR-498 could recognize CCPG1-3' untranslated region (UTR). CCPG1 protein level was remarkably decreased when overexpressed miR-498, nevertheless, significantly increased when inhibiting miR-498. Y79 cells that were transfected with miR-498 mimics manifested notable cell apoptosis down-regulation and cell proliferation promotion; whereas, those transfected with miR-498 inhibitor displayed significant cell apoptosis up-regulation and cell proliferation inhibition compared with control group.<h4>Conclusion</h4>Taken together, miR-498 promotes cell proliferation and inhibits cell apoptosis in Rb by directly targeting CCPG1.

The brain has long been known to display the most complex pattern of alternative splicing, thereby producing diverse protein isoforms compared to other tissues. Recent evidence indicates that many alternative exons are neuron-specific, evolutionarily conserved, and found in regulators of transcription including DNA-binding protein and histone modifying enzymes. This raises a possibility that neurons adopt unique mechanisms of transcription. Given that transcriptional machineries are frequently mutated in neurodevelopmental disorders with cognitive dysfunction, it is important to understand how neuron-specific alternative splicing contributes to proper transcriptional regulation in the brain. In this review, we summarize current knowledge regarding how neuron-specific splicing events alter the function of transcriptional regulators and shape unique gene expression patterns in the brain and the implications of neuronal splicing to the pathophysiology of neurodevelopmental disorders.

Oligodendrocytes are essential for the development, function, and health of the vertebrate central nervous system. These cells maintain axon myelination to ensure saltatory propagation of action potentials. Oligodendrocyte develops from neural progenitor cells, in a step-wise process that involves oligodendrocyte precursor specification, proliferation, and differentiation. The lineage progression requires coordination of transcriptional and epigenetic circuits to mediate the stage-specific intricacies of oligodendrocyte development. Epigenetic mechanisms involve DNA methylation, histone modifications, ATP-dependent chromatin remodeling, and non-coding RNA modulation that regulate the chromatin state over regulatory genes, which must be expressed or repressed to establish oligodendrocyte identity and lineage progression. In this review, we will focus on epigenetic programming associated with histone modification enzymes, chromatin remodeling, and non-coding RNAs that regulate oligodendrocyte lineage progression, and discuss how these mechanisms might be harnessed to induce myelin repair for treatment of demyelinating diseases such as multiple sclerosis.

Cellular glycosylation processes are vital to cell functioning. In malignant cells, they are profoundly altered. We used time-course gene expression data from the NCI-60 cancer cell lines treated with 11 antitumor agents to analyze expression changes of genes involved in glycosylation pathways, genes encoding glycosylation targets or regulators, and members of cancer pathways affected by glycosylation. We also identified glycosylation genes for which pretreatment expression levels or changes after treatment were correlated with drug sensitivity. Their products are involved in <i>N-</i>glycosylation and <i>O-</i>glycosylation, fucosylation, biosynthesis of poly-<i>N</i>-acetyllactosamine, removal of misfolded proteins, binding to hyaluronic acid and other glycans, and cell adhesion. Tumor cell sensitivity to multiple agents was correlated with transcriptional response of <i>C1GALT1C1, FUCA1, SDC1, MUC1</i>; members of the <i>MGAT, GALNT, B4GALT, B3GNT, MAN</i>, and <i>EDEM</i> families; and other genes. These genes may be considered as potential candidates for drug targeting in combination therapy to enhance treatment response.

The gastric ganglion is the largest visceral ganglion in cephalopods. It is connected to the brain and is implicated in regulation of digestive tract functions. Here we have investigated the neurochemical complexity (through <i>in silico</i> gene expression analysis and immunohistochemistry) of the gastric ganglion in <i>Octopus vulgaris</i> and tested whether the expression of a selected number of genes was influenced by the magnitude of digestive tract parasitic infection by <i>Aggregata octopiana</i>. Novel evidence was obtained for putative peptide and non-peptide neurotransmitters in the gastric ganglion: cephalotocin, corticotrophin releasing factor, FMRFamide, gamma amino butyric acid, 5-hydroxytryptamine, molluscan insulin-related peptide 3, peptide PRQFV-amide, and tachykinin-related peptide. Receptors for cholecystokinin_A and cholecystokinin_B, and orexin₂ were also identified in this context for the first time. We report evidence for acetylcholine, dopamine, noradrenaline, octopamine, small cardioactive peptide related peptide, and receptors for cephalotocin and octopressin, confirming previous publications. The effects of <i>Aggregata</i> observed here extend those previously described by showing effects on the gastric ganglion; in animals with a higher level of infection, genes implicated in inflammation (NFκB, fascin, serpinB10 and the toll-like 3 receptor) increased their relative expression, but TNF-α gene expression was lower as was expression of other genes implicated in oxidative stress (i.e., superoxide dismutase, peroxiredoxin 6, and glutathione peroxidase). Elevated <i>Aggregata</i> levels in the octopuses corresponded to an increase in the expression of the cholecystokinin_A receptor and the small cardioactive peptide-related peptide. In contrast, we observed decreased relative expression of cephalotocin, dopamine β-hydroxylase, peptide PRQFV-amide, and tachykinin-related peptide genes. A discussion is provided on (i) potential roles of the various molecules in food intake regulation and digestive tract motility control and (ii) the difference in relative gene expression in the gastric ganglion in octopus with relatively high and low parasitic loads and the similarities to changes in the enteric innervation of mammals with digestive tract parasites. Our results provide additional data to the described neurochemical complexity of <i>O. vulgaris</i> gastric ganglion.

Twenty-five genome-wide association studies (GWAS) of asthma were published between 2007 and 2016, the largest with a sample size of 157242 individuals. Across these studies, 39 genetic variants in low linkage disequilibrium (LD) with each other were reported to associate with disease risk at a significance threshold of <i>P</i><5 × 10^-8, including 31 in populations of European ancestry. Results from analyses of the UK Biobank data (<i>n</i>=380 503) indicate that at least 28 of the 31 associations reported in Europeans represent true-positive findings, collectively explaining 2.5% of the variation in disease liability (median of 0.06% per variant). We identified 49 transcripts as likely target genes of the published asthma risk variants, mostly based on LD with expression quantitative trait loci (eQTL). Of these genes, 16 were previously implicated in disease pathophysiology by functional studies, including <i>TSLP</i>, <i>TNFSF4</i>, <i>ADORA1</i>, <i>CHIT1</i> and <i>USF1</i>. In contrast, at present, there is limited or no functional evidence directly implicating the remaining 33 likely target genes in asthma pathophysiology. Some of these genes have a known function that is relevant to allergic disease, including <i>F11R, CD247, PGAP3, AAGAB</i>, <i>CAMK4</i> and <i>PEX14</i>, and so could be prioritized for functional follow-up. We conclude by highlighting three areas of research that are essential to help translate GWAS findings into clinical research or practice, namely validation of target gene predictions, understanding target gene function and their role in disease pathophysiology and genomics-guided prioritization of targets for drug development.

Melanoma is an aggressive skin cancer; an early detection of the primary tumor may improve its prognosis. Despite many genes have been shown to be involved in melanoma, the full framework of melanoma transformation has not been completely explored. The characterization of pathways involved in tumor restraint in <i>in vitro</i> models may help to identify oncotarget genes. We therefore aimed to probe novel oncotargets through an integrated approach involving proteomic, gene expression and bioinformatic analysis We investigated molecular modulations in melanoma cells treated with ascorbic acid, which is known to inhibit cancer growth at high concentrations. For this purpose a proteomic approach was applied. A deeper insight into ascorbic acid anticancer activity was achieved; the discovery of deregulated processes suggested further biomarkers. In addition, we evaluated the expression of identified genes as well as the migration ability in several melanoma cell lines. Data obtained by a multidisciplinary approach demonstrated the involvement of Enolase 1 (ENO1), Parkinsonism-associated deglycase (PARK7), Prostaglansin E synthase 3 (PTGES3), Nucleophosmin (NPM1), Stathmin 1 (STMN1) genes in cell transformation and identified Single stranded DNA binding protein 1 (SSBP1) as a possible onco-suppressor in melanoma cancer.

One of the aims of synthetic biology is bottom-up construction of reconstituted human cells for medical uses. To that end, we generated giant unilamellar vesicles (GUVs) that contained a HeLa cell extract, which comprises a cell-free protein synthesis (CFPS) system. Then we expressed Huntingtin protein fragments that contained polyglutamine (polyQ) sequences (Htt-polyQ), a hallmark of Huntington's disease. That system produced polyQ-dependent protein aggregates, as previously demonstrated in living cells. We next simplified the system by generating GUVs that contained purified human factors, which reconstituted a CFPS system. Htt-polyQ fragments expressed in these GUVs also formed protein aggregates. Moreover, an N-terminal deletion mutant, which had failed to form protein aggregates in living cells, also failed to form protein aggregates in the reconstituted GUVs. Thus, the GUV systems that encapsulated a human CFPS system could serve as reconstituted cells for studying neurological diseases.

This article reports the degradation and biological properties of as-drawn Mg-4Zn-1Sr (designated as ZSr41) and pure Mg (P-Mg) wires as bioresorbable intramedullary pins for bone repair. Specifically, their cytocompatibility with bone marrow derived mesenchymal stem cells (BMSCs) and degradation in vitro, and their biological effects on peri-implant tissues and in vivo degradation in rat tibiae were studied. The as-drawn ZSr41 pins showed a significantly faster degradation than P-Mg in vitro and in vivo. The in vivo average daily degradation rates of both ZSr41 and P-Mg intramedullary pins were significantly greater than their respective in vitro degradation rates, likely because the intramedullary site of implantation is highly vascularized for removal of degradation products. Importantly, the concentrations of Mg²⁺, Zn²⁺, and Sr²⁺ ions in the BMSC culture in vitro and their concentrations in rat blood in vivo were all lower than their respective therapeutic dosages, i.e., in a safe range. Despite of rapid degradation with a complete resorption time of 8 weeks in vivo, the ZSr41 intramedullary pins showed a significant net bone growth because of stimulatory effects of the metallic ions released. However, proportionally released OH^- ions and hydrogen gas caused adverse effects on bone marrow cells and resulted in cavities in surrounding bone. Thus, properly engineering the degradation properties of Mg-based implants is critical for harvesting the bioactivities of beneficial metallic ions, while controlling adverse reactions associated with the release of OH^- ions and hydrogen gas. It is necessary to further optimize the alloy processing conditions and/or modify the surfaces, for example, applying coatings onto the surface, to reduce the degradation rate of ZSr41 wires for skeletal implant applications.

<h4>Background</h4>Little is currently known regarding the immunologic mechanism(s) that initiate peanut allergy. Notably, peanut proteins have been detected in house dust, and their levels correlate with peanut allergy prevalence.<h4>Objective</h4>This study aimed to develop a new mouse model for peanut allergy and to investigate the immunologic mechanisms involved in peanut allergen sensitization.<h4>Methods</h4>To mimic environmental exposure, naive mice were exposed to peanut flour by inhalation for up to 4 weeks. We then analyzed serum levels of IgE antibody and challenged mice with peanut proteins. Immunological mechanisms involved in sensitization were analyzed using cytokine reporter mice, an adoptive cell transfer model, and gene knockout mice.<h4>Results</h4>When exposed to peanut flour by inhalation, both BALB/c and C57BL/6 mice developed peanut allergy, as demonstrated by the presence of peanut-specific IgE antibodies and manifestation of acute anaphylaxis on challenge. A large number of follicular helper T (Tfh) cells were also detected in draining lymph nodes of allergic mice. These cells produced IL-4 and IL-21, and they more robustly promoted peanut-specific IgE production than T_h2 cells did. Genetic depletion of Tfh cells decreased IgE antibody levels and protected mice from anaphylaxis, without affecting T_h2 cells. Furthermore, peanut flour exposure increased lung levels of IL-1α and IL-1β, and mice deficient in the receptor for these cytokines showed a significant decrease in Tfh cells compared with in wild-type mice.<h4>Conclusions</h4>Tfh cells play a key role in peanut allergy, and the IL-1 pathway is involved in the Tfh response to peanut allergen exposure.

Tissue regeneration requires dynamic cellular adaptation to the wound environment. It is currently unclear how this is orchestrated at the cellular level and how cell fate is affected by severe tissue damage. Here we dissect cell fate transitions during colonic regeneration in a mouse dextran sulfate sodium (DSS) colitis model, and we demonstrate that the epithelium is transiently reprogrammed into a primitive state. This is characterized by de novo expression of fetal markers as well as suppression of markers for adult stem and differentiated cells. The fate change is orchestrated by remodeling the extracellular matrix (ECM), increased FAK/Src signaling, and ultimately YAP/TAZ activation. In a defined cell culture system recapitulating the extracellular matrix remodeling observed in vivo, we show that a collagen 3D matrix supplemented with Wnt ligands is sufficient to sustain endogenous YAP/TAZ and induce conversion of cell fate. This provides a simple model for tissue regeneration, implicating cellular reprogramming as an essential element.

Specificity of the ubiquitin proteasome system is controlled by ubiquitin E3 ligases, including their major representatives, the multisubunit cullin-RING ubiquitin (Ub) ligases (CRLs). More than 200 different CRLs are divided into seven families according to their cullin scaffolding proteins (CUL1-7) around which they are assembled. Research over two decades has revealed that different CRL families are specialized to fulfill specific cellular functions. Whereas many CUL1-based CRLs (CRL1s) ubiquitylate cell cycle regulators, CRL4 complexes often associate with chromatin to control DNA metabolism. Based on studies about differentiation programs of mesenchymal stem cells (MSCs), including myogenesis, neurogenesis, chondrogenesis, osteogenesis and adipogenesis, we propose here that CRL3 complexes evolved to fulfill a pivotal role in mammalian cell differentiation.

<h4>Purpose</h4>To identify the characteristics and initial disease severity of patients with nonalcoholic fatty liver disease (NAFLD) and assess incidence and risk factors for disease progression in a retrospective study.<h4>Methods</h4>Patients ≥18 years of age without alcoholism or other liver diseases (eg, hepatitis B/C) were selected from Geisinger Health System electronic medical record data from 2004 to 2015. Initial disease stage was stratified into uncomplicated NAFLD, advanced fibrosis, cirrhosis, hepatocellular carcinoma (HCC), and liver transplant using clinical biomarkers, diagnosis, and procedure codes. Disease progression was defined as stage progression or death and analyzed via Kaplan-Meier plots and multistate models.<h4>Results</h4>In the NAFLD cohort (N=18,754), 61.5% were women, 39.0% had type 2 diabetes mellitus (T2DM), and the mean body mass index was 38.2±10.2 kg/m². At index, 69.9% had uncomplicated NAFLD, 11.7% had advanced fibrosis, and 17.8% had cirrhosis. Of 18,718 patients assessed for progression, 17.3% progressed (11.0% had stage progression, 6.3% died without evidence of stage progression) during follow-up (median=842 days). Among subgroups, 12.3% of those without diabetes mellitus progressed vs 24.7% of those with T2DM. One-year mortality increased from 0.5% in uncomplicated NAFLD to 22.7% in HCC. After liver transplant, mortality decreased to 5.6% per year.<h4>Conclusions</h4>In 2.3 years of follow-up, approximately 17% of patients progressed or died without evidence of stage progression. T2DM was associated with approximately twice the risk of disease progression, and mortality risk increased with disease stage. Early diagnosis and monitoring of disease progression, especially in patients with T2DM, is warranted.

[This corrects the article on p. 1401 in vol. 7, PMID: 27703463.].

Pediatric spinal ependymomas (SEPN) are important albeit uncommon malignant central nervous system tumors with limited treatment options. Our current knowledge about the underlying biology of these tumors is limited due to their rarity. To begin to elucidate molecular mechanisms that give rise to pediatric SEPN, we compared the transcriptomic landscape of SEPNs to that of intracranial ependymomas using genome-wide mRNA and microRNA (miRNA) expression profiling in primary tumour samples. We found that pediatric SEPNs are characterized by increased expression of genes involved in developmental processes, oxidative phosphorylation, cellular respiration, electron transport chain, and cofactor metabolic process. Next, we compared pediatric spinal and intracranial ependymomas with the same tumours in adults and found a relatively low number of genes in pediatric tumours that were shared with adult tumours (12.5%). In contrast to adult SEPN, down-regulated genes in pediatric SEPN were not enriched for position on chromosome 22. At the miRNA level, we found ten miRNAs that were perturbed in pediatric SEPN and we identified regulatory relationships between these miRNAs and their putative targets mRNAs using the integrative miRNA-mRNA network and predicted miRNA target analysis. These miRNAs include the oncomiR <i>hsa-miR-10b</i> and its family member <i>hsa-miR-10a</i>, both of which are upregulated and target chromatin modification genes that are down regulated in pediatric SEPN. The tumor suppressor, <i>hsa-miR-124</i>, was down regulated in pediatric SEPN and it normally represses genes involved in cell-cell communication and metabolic processes. Together, our findings suggest that pediatric SEPN is characterized by a distinct transcriptional landscape from that of pediatric intracranial EPNs or adult tumors (both SEPNs and intracranial EPNs). Although confirmatory studies are needed, our study reveals novel molecular pathways that may drive tumorigenesis and could serve as biomarkers or rational therapeutic targets.

We previously demonstrated that treatment of the H295R adrenocortical cancer cell line with the non-steroidal, high-affinity GPER (G protein-coupled estrogen receptor 1) agonist G-1 reduced tumor growth <i>in vitro</i> and <i>in vivo</i> through a GPER independent action. Moreover, we observed that G-1 treatment induces cell-cycle arrest and apoptosis following a sustained ERK1/2 activation. However, the precise mechanisms causing these effects were not clarified. Starting from our preliminary published results, we performed a microarray study that clearly evidenced a strong and significative up-regulation of EGR-1 gene in H295R cells treated for 24h with micromolar concentration of G-1. The microarray findings were confirmed by RT-PCR and Western-blot analysis as well as by immunofluorescence that revealed a strong nuclear staining for EGR-1 after G-1 treatment. EGR-1 is a point of convergence of many intracellular signaling cascades that control tumor cell growth and proliferation as well as others that relate to cell death machinery. Here we found that the increased Egr-1 expression was a consequence of G-1-mediated ROS-dependent ERK activation that were promptly reversed by the presence of the antioxidant n-acetyl-cysteine. Finally, we observed that silencing EGR-1 gene expression reversed the main effects induced by G-1 in ACC cells, including upregulation of the negative regulator of cell cycle, p21^Waf1/Cip1 and the positive regulator of mitochondrial apoptotic pathway, BAX, as well as the cell growth inhibition. The identified ROS/MAPK/Egr-1/BAX pathway as a potential off-target effect of the G-1 could be useful in implementing the pharmacological approach for ACC therapy.

Two or more anions constrained in close proximity within a single pocket are found in a number of natural systems but a less common motif in artificial systems. This review summarizes work on anion receptors capable of stabilizing anionic dimers, trimers, tetramers and clusters in a well-defined fashion. These systems may provide insights into the fundamental chemistry of anion-anion interactions and provide a guide for understanding in greater detail a number of biological and environmental processes, as well as key tenants of relevance to supramolecular chemistry, extraction, transport, crystal engineering, and the like. The primary goal of this review is to provide a general introduction into multi-anion recognition chemistry for the benefit of supramolecular and non-supramolecular chemists alike.

<h4>Background/objective</h4>This is a multicentre, randomised, double-blind, placebo-controlled clinical trial to investigate the safety and efficacy of Chinese herbal Fufang Xian Ling Gu Bao (XLGB) with antiadipogenic compounds for the prevention of corticosteroid (CS)-induced osteonecrosis of femoral head (ONFH).<h4>Methods</h4>Patients of both genders, aged between 18 and 65 years, with diseases such as systemic lupus erythematosus, nephrosis, dermatosis and rheumatoid arthritis indicated for CS treatment and who did not show magnetic resonance imaging of ONFH at baseline were recruited into the study and then randomised into either XLGB group (n = 129) with daily oral administration of XLGB or placebo group (n = 146).<h4>Results</h4>Magnetic resonance imaging revealed a total of 30 ONFH cases at 6 months after CS treatment, with 6.98% (9 of 129 cases) and 14.4% (21 of 146 cases) in the XLGB group and placebo group, respectively, (<i>p</i> < 0.05), i.e., a 2-fold significantly less ONFH identified in the XLGB treatment group. Blood tests suggested that XLGB significantly inhibited the elevation of activated protein C resistance induced by CS treatment.<h4>Conclusion</h4>This is the first multicentre clinical study to demonstrate that the antiadipogenic compounds-rich herbal Fufang (formula) XLGB is effective in preventing CS-associated ONFH in patients with immune-inflammatory diseases under CS treatment.<h4>The translational potential of this article</h4>The translation potential of this clinical trial is that the initially officially approved clinical indication for XLGB for treatment of osteoporosis has been now also proven to be effective for a new clinical application.

<h4>Aims</h4>Iron-overload disorders are common and could lead to significant morbidity and mortality worldwide. Due to limited treatment options, there is a great need to develop novel strategies to remove the excess body iron. To discover potential epigenetic modulator in hepcidin upregulation and subsequently decreasing iron burden, we performed an epigenetic screen. The in vivo effects of the identified compounds were further tested in iron-overload mouse models, including Hfe^-/-, Hjv^-/-, and hepatocyte-specific Smad4 knockout (Smad4^fl/fl;Alb-Cre⁺) mice.<h4>Results</h4>Entinostat (MS-275), the clinical used histone deacetylase 1 (HDAC1) inhibitor, was identified the most potent hepcidin agonist. Consistently, Hdac1-deficient mice also presented higher hepcidin levels than wild-type controls. Notably, the long-term treatment with entinostat in Hfe^-/- mice significantly alleviated iron overload through upregulating hepcidin transcription. In contrast, entinostat showed no effect on hepcidin expression and iron levels in Smad4^fl/fl;Alb-Cre⁺ mice. Further mechanistic studies revealed that HDAC1 suppressed expression of hepcidin through interacting with SMAD4 rather than deacetylation of SMAD4 or histone-H3 on the hepcidin promoter.<h4>Innovation</h4>The findings uncovered HDAC1 as a novel hepcidin suppressor through complexing with SMAD4 but not deacetylation of either histone 3 or SMAD4. In addition, our study suggested a novel implication of entinostat in treating iron-overload disorders.<h4>Conclusions</h4>Based on our results, we conclude that entinostat strongly activated hepcidin in vivo and in vitro. HDAC1 could serve as a novel hepcidin suppressor by binding to SMAD4, effect of which is independent of BMP/SMAD1/5/8 signaling. Antioxid. Redox Signal. 28, 1224-1237.

Lipid droplets (LDs) are the main energy resource for porcine preimplantation embryonic development. PLIN3 has been implicated in LD formation and regulation. Therefore, this study aimed to detect the dynamic pattern of PLIN3 in pig oocytes and cumulus cells (CC) during in vitro maturation (IVM), and to determine the relationship between PLIN3 and LD content. IVM with cumulus-enclosed oocytes (CEO), cumulus-denuded oocytes (DO) and the CCs denuded from the corresponding oocytes (DCC) was performed in porcine follicular fluid (PFF) or PFF-free optimized medium. DO and the DCC were cultured together under the same conditions as described above, while the DO was named DTO and the DCC was named DTCC in this group. Firstly, our results revealed LDs distributed widely in oocytes and CC, while the PLIN3 protein coated these LDs and spread out ubiquitously in the cytoplasm. Secondly, not only the mRNA level but also at protein level of PLIN3 in immature naked oocytes (IO) was higher than that in matured CEO, DO and DTO. Although PLIN3 was expressed at lower levels in CC from immature oocytes (ICC), the protein level of PLIN3 was comparably higher in the ECC and DCC groups. The triglyceride (TG) content in CEO and DO was significantly less abundant compared with that in IO. Therefore, our results indicated that co-culturing of oocytes and CC might affect PLIN3 expression levels in CC but not in oocytes. Lipid accumulation in pig oocytes during maturation might be affected by PLIN3 cross-talk between oocytes and CC.

The aim of this retrospective study was to establish a simple self-assessed scale for individual risk of cirrhosis in patients with chronic hepatitis B. A total of 1808 consecutive patients were enrolled and analyzed. According to the results of multivariate logistic regression analysis, a simple nomogram was calculated for cirrhosis. The area under receiver operating characteristic curves (AUROCs) were calculated to compare the diagnostic accuracy of nomogram with aspartate aminotransferase to platelet ratio index (APRI), fibrosis index based on the four factors (FIB-4), and S index. The AUROCs of nomogram for cirrhosis were 0.807 (adjusted AUROC 0.876) in model group and 0.794 (adjusted AUROC0.866) in validation group. DeLong's test and Brier Score further demonstrated that nomogram was superior to APRI, FIB-4 and S index in both model group and validation group. The patients with nomogram <0.07 could be defined as low risk group with cirrhosis prevalence lower than 4.3% (17/397). The patients with nomogram >0.52 could be defined as high risk group with cirrhosis prevalence higher than 73.0% (119/163). In conclusion, as a self-assessed style, simple, non-invasive, economical, convenient, and repeatable scale, nomogram is suitable to serve as a massive health screening tool for cirrhosis in CHB patients and further external validation is needed.

Zinc (Zn), which is an essential trace element, is involved in numerous mammalian physiological events; therefore, either a deficiency or excess of Zn impairs cellular machineries and influences physiological events, such as systemic growth, bone homeostasis, skin formation, immune responses, endocrine function, and neuronal function. Zn transporters are thought to mainly contribute to Zn homeostasis within cells and in the whole body. Recent genetic, cellular, and molecular studies of Zn transporters highlight the dynamic role of Zn as a signaling mediator linking several cellular events and signaling pathways. Dysfunction in Zn transporters causes various diseases. This review aims to provide an update of Zn transporters and Zn signaling studies and discusses the remaining questions and future directions by focusing on recent progress in determining the roles of SLC39A/ZIP family members in vivo.

The RNA-binding protein, UPF1, is best known for its central role in the nonsense-mediated RNA decay (NMD) pathway. Feng et al. now report a new function for UPF1-it is an E3 ubiquitin ligase that specifically promotes the decay of a key pro-muscle transcription factor: MYOD. UPF1 achieves this through its RING-like domain, which confers ubiquitin E3 ligase activity. Feng et al. provide evidence that the ability of UPF1 to destabilize MYOD represses myogenesis. In the future, it will be important to define other protein substrates of UPF1-driven ubiquitination and to determine whether this biochemical activity is responsible for some of UPF1's previously defined biological functions, including in development and stress responses. The exciting findings presented by Feng et al. open up the possibility that protein turnover and RNA turnover are coupled processes.

No abstract available.

Condensin is a protein complex with diverse functions in chromatin packaging and chromosome condensation and segregation. We studied condensin in the evolutionarily distant protist model <i>Tetrahymena</i>, which features noncanonical nuclear organization and divisions. In <i>Tetrahymena</i>, the germline and soma are partitioned into two different nuclei within a single cell. Consistent with their functional specializations in sexual reproduction and gene expression, condensins of the germline nucleus and the polyploid somatic nucleus are composed of different subunits. Mitosis and meiosis of the germline nucleus and amitotic division of the somatic nucleus are all dependent on condensins. In condensin-depleted cells, a chromosome condensation defect was most striking at meiotic metaphase, when <i>Tetrahymena</i> chromosomes are normally most densely packaged. Live imaging of meiotic divisions in condensin-depleted cells showed repeated nuclear stretching and contraction as the chromosomes failed to separate. Condensin depletion also fundamentally altered chromosome arrangement in the polyploid somatic nucleus: multiple copies of homologous chromosomes tended to cluster, consistent with a previous model of condensin suppressing default somatic pairing. We propose that failure to form discrete chromosome territories is the common cause of the defects observed in the absence of condensins.

SMG-1,a member of the phosphoinositide kinase-like kinase family, functioned as a tumor suppressor gene. However, the role of SMG-1 in GC remain uncharacterized. In this study, regulation of SMG-1 by miR-192 and-215, along with the biological effects of this modulation, were studied in GC. We used gene microarrays to screening and luciferase reporter assays were to verify the potential targets of miR-192 and-215. Tissue microarrays analyses were applied to measure the levels of SMG-1 in GC tissues. Western blot assays were used to assess the signaling pathway of SMG-1 regulated by miR-192 and-215 in GC. SMG-1 was significantly downregulated in GC tissues.The proliferative and invasive properties of GC cells were decreased by inhibition of miR-192 and-215, whereas an SMG-1siRNA rescued the inhibitory effects. Finally, SMG-1 inhibition by miR-192 and-215 primed Wnt signaling and induced EMT. Wnt signaling pathway proteins were decreased markedly by inhibitors of miR-192 and-215, while SMG-1 siRNA reversed the inhibition apparently. Meanwhile, miR-192 and-215 inhitibtors increased E-cadherin expression and decreased N-cadherin and cotransfection of SMG-1 siRNA reversed these effects. In summary, these findings illustrate that SMG-1 is suppressed by miR-192 and-215 and functions as a tumor suppressor in GC by inactivating Wnt signaling and suppressing EMT.

In the nervous system, Netrin-1 serves as a neural guide, mediating the neuronal development. However, it remains blurred whether Netrin-1 can protect neurons from apoptosis induced by cerebral stroke. In the current study, the cultured rat primary cortical neurons were transfected with Netrin-1-encoding lentivirus before the oxygen-glucose-deprivation (OGD) treatment. Cell death and apoptosis were evaluated by lactate dehydrogenase (LDH) release and flow cytometry. We found that Netrin-1 attenuated OGD-induced cell death and neuronal apoptosis at 24 h after OGD treatment, and that the overexpression of Netrin-1 activated the ERK signaling pathway. These effects were partly abolished by blocking its receptor deleted in colorectal cancer (DCC) or U0126, an inhibitor of the ERK signaling pathway. Netrin-1 overexpression in neurons elevated the expression of DCC, on mRNA level and protein level. Netrin-1 also reduced DNA damage. Taken together, our findings suggest that Netrin-1 attenuates cell death and neuronal apoptosis via the DCC/ERK signaling pathway in the cultured primary cortical neurons after OGD injury, which may involve the mediation of DNA damage in the neurons.

The CINdex Bioconductor package addresses an important area of high-throughput genomic analysis. It calculates the chromosome instability (CIN) index, a novel measurement that quantitatively characterizes genome-wide copy number alterations (CNAs) as a measure of CIN. The advantage of this package is an ability to compare CIN index values between several groups for patients (case and control groups), which is a typical use case in translational research. The differentially changed cytobands or chromosomes can then be linked to genes located in the affected genomic regions, as well as pathways. This enables in-depth systems biology-based network analysis and assessment of the impact of CNA on various biological processes or clinical outcomes. This package was successfully applied to analysis of DNA copy number data in colorectal cancer as a part of multi-omics integrative study as well as for analysis of several other cancer types. The source code, along with an end-to-end tutorial, and example data are freely available in Bioconductor at http://bioconductor.org/packages/CINdex/.

Zn²⁺ is an essential element for cell survival/growth, and its deficiency is linked to many disorders. Extracellular Zn²⁺ concentration changes participate in modulating fundamental cellular processes such as proliferation, secretion, ion transport, and cell signal transduction in a mechanism that is not well understood. Here, we hypothesize that the Zn²⁺-sensing receptor ZnR/G protein-coupled receptor 39 (GPR39), found in tissues where dynamic Zn²⁺ homeostasis takes place, enables extracellular Zn²⁺ to trigger intracellular signaling pathways regulating key cell functions in vascular cells. Thus, we investigated how extracellular Zn²⁺ regulates cell viability, proliferation, motility, angiogenesis, vascular tone, and inflammation through ZnR/GPR39 in endothelial cells. Knockdown of GPR39 through siRNA largely abolished Zn²⁺-triggered cellular activity changes, Ca²⁺ responses, as well as the downstream activation of Gαq-PLC pathways. Extracellular Zn²⁺ promoted vascular cell survival/growth through activation of cAMP and Akt as well as overexpressing of platelet-derived growth factor-α receptor and vascular endothelial growth factor A. It also enhanced cell adhesion and mobility, endothelial tubule formation, and cytoskeletal reorganization. Such effects from extracellular Zn²⁺ were not observed in GPR39^-/- endothelial cells. Zn²⁺ also regulated inflammation-related key molecules such as heme oxygenase-1, selectin L, IL-10, and platelet endothelial cell adhesion molecule 1, as well as vascular tone-related prostaglandin I2 synthase and nitric oxide synthase-3. In sum, extracellular Zn²⁺ regulates endothelial cell activity in a ZnR/GPR39-dependent manner and through the downstream G_αq-PLC pathways. Thus, ZnR/GPR39 may be a therapeutic target for regulating endothelial activity.

<h4>Background & aims</h4>Nonalcoholic steatohepatitis (NASH) is associated with oxidative stress. We surmised that pharmacologic activation of NF-E2 p45-related factor 2 (Nrf2) using the acetylenic tricyclic bis(cyano enone) TBE-31 would suppress NASH because Nrf2 is a transcriptional master regulator of intracellular redox homeostasis.<h4>Methods</h4><i>Nrf2</i>^<i>+/+</i> and <i>Nrf2</i>^<i>-/-</i> C57BL/6 mice were fed a high-fat plus fructose (HFFr) or regular chow diet for 16 weeks or 30 weeks, and then treated for the final 6 weeks, while still being fed the same HFFr or regular chow diets, with either TBE-31 or dimethyl sulfoxide vehicle control. Measures of whole-body glucose homeostasis, histologic assessment of liver, and biochemical and molecular measurements of steatosis, endoplasmic reticulum (ER) stress, inflammation, apoptosis, fibrosis, and oxidative stress were performed in livers from these animals.<h4>Results</h4>TBE-31 treatment reversed insulin resistance in HFFr-fed wild-type mice, but not in HFFr-fed Nrf2-null mice. TBE-31 treatment of HFFr-fed wild-type mice substantially decreased liver steatosis and expression of lipid synthesis genes, while increasing hepatic expression of fatty acid oxidation and lipoprotein assembly genes. Also, TBE-31 treatment decreased ER stress, expression of inflammation genes, and markers of apoptosis, fibrosis, and oxidative stress in the livers of HFFr-fed wild-type mice. By comparison, TBE-31 did not decrease steatosis, ER stress, lipogenesis, inflammation, fibrosis, or oxidative stress in livers of HFFr-fed Nrf2-null mice.<h4>Conclusions</h4>Pharmacologic activation of Nrf2 in mice that had already been rendered obese and insulin resistant reversed insulin resistance, suppressed hepatic steatosis, and mitigated against NASH and liver fibrosis, effects that we principally attribute to inhibition of ER, inflammatory, and oxidative stress.

<h4>ABSTRACT</h4> Huntington disease (HD) is a progressive neurodegenerative disease that initially affects the striatum leading to changes in behavior and loss of motor coordination. It is caused by an expansion in the polyglutamine repeat at the N-terminus of huntingtin (HTT) that leads to aggregation of mutant HTT. The loss of wildtype function, in combination with the toxic gain of function mutation, initiates various cell death pathways. Wildtype and mutant HTT are regulated by different post-translational modifications that can positively or negatively regulate their function or toxicity. In particular, we have previously shown that caspase cleavage of mutant HTT at amino acid position aspartate 586 (D586) by caspase-6 is critical for the pathogenesis of the disease in an HD mouse model. Herein, we describe the identification of a new caspase cleavage site at position D572 that is mediated by caspase-1. Inhibition of caspase-1 also inhibits cleavage at D586 through inhibition of caspase-6. Inhibition of caspase cleavage at D572 significantly decreases mutant HTT aggregation and significantly increased the turnover of soluble mutant HTT. This suggests that caspase-1 may be a viable target to inhibit caspase cleavage of mutant HTT at both D572 and D586 to promote mutant HTT clearance.

<h4>Aims</h4>Lack of Cu,Zn-superoxide dismutase (CuZnSOD) in homozygous knockout mice (Sod1^-/-) leads to accelerated age-related muscle loss and weakness, but specific deletion of CuZnSOD in skeletal muscle (mSod1KO mice) or neurons (nSod1KO mice) resulted in only mild muscle functional deficits and failed to recapitulate the loss of mass and function observed in Sod1^-/- mice. To dissect any underlying cross-talk between motor neurons and skeletal muscle in the degeneration in Sod1^-/- mice, we characterized neuromuscular changes in the Sod1^-/- model compared with mSod1KO mice and examined degenerative molecular mechanisms and pathways in peripheral nerve and skeletal muscle.<h4>Results</h4>In contrast to mSod1KO mice, myofiber atrophy in Sod1^-/- mice was associated with increased muscle oxidative damage, neuromuscular junction degeneration, denervation, nerve demyelination, and upregulation of proteins involved in maintenance of myelin sheaths. Proteomic analyses confirmed increased proteasomal activity and adaptive stress responses in muscle of Sod1^-/- mice that were absent in mSod1KO mice. Peripheral nerve from neither Sod1^-/- nor mSod1KO mice showed increased oxidative damage or molecular responses to increased oxidation compared with wild type mice. Differential cysteine (Cys) labeling revealed a specific redox shift in the catalytic Cys residue of peroxiredoxin 6 (Cys47) in the peripheral nerve from Sod1^-/- mice. Innovation and Conclusion: These findings demonstrate that neuromuscular integrity, redox mechanisms, and pathways are differentially altered in nerve and muscle of Sod1^-/- and mSod1KO mice. Results support the concept that impaired redox signaling, rather than oxidative damage, in peripheral nerve plays a key role in muscle loss in Sod1^-/- mice and potentially sarcopenia during aging. Antioxid. Redox Signal. 28, 275-295.

<h4>Significance</h4>Particulate matter (PM) air pollution is a leading cause of global cardiovascular morbidity and mortality. Understanding the biological action of PM is of particular importance in improvement of public health. Recent Advances: Both fine (PM <2.5 μM) and ultrafine particles (<0.1 μM) are widely believed to mediate their effects through redox regulated pathways. A rather simplistic graded ramp model of redox stress has been replaced by a more sophisticated understanding of the role of oxidative stress in signaling, and the realization that many of the observed effects may involve disruption and/or enhancement of normal endogenous redox signaling and induction of a potent immune-mediated response, through entrainment of multiple reactive oxygen species (ROS).<h4>Critical issues</h4>The molecular events by which pulmonary oxidative stress in response to inhalational exposure to air pollution triggers inflammation, major ROS (e.g., superoxide, hydroxyl radical, nitric oxide, and peroxynitrite) generated in air pollution exposure, types of oxidative tissue damage in target organs, contributions of nonimmune and immune cells in inflammation, and the role of protective proteins (e.g., surfactant, proteins, and antioxidants) are highly complex and may differ depending on models and concomitant disease states.<h4>Future directions</h4>While the role of oxidative stress in the lung has been well demonstrated, the role of oxidative stress in mediating systemic effects especially in inflammation and injury processes needs further work. The role of antioxidant defenses with chronic exposure will also need further exploration. Antioxid. Redox Signal. 28, 797-818.

<h4>Objective</h4>The aim of the study was to assess the distribution, frequency, and specific location of mutant huntingtin protein (mHTT) aggregates-the pathological hallmark of Huntington disease (HD)-within the various compartments of the spinal cord and their potential impact on the local vasculature and blood-spinal cord barrier (BSCB).<h4>Methods</h4>We performed a series of postmortem immunohistochemical and immunofluorescent stainings, as well as Western blot analyses, on cervical and lumbar sections of the spinal cord in patients diagnosed with HD (n = 11 of all grades of disease severity) along with sex- and age-matched healthy controls (n = 9).<h4>Results</h4>We observed that mHTT was preferably expressed within the anterior horn of the gray matter, in both cervical and lumbar sections. At the cellular level, mHTT aggregates were more often encountered in the extracellular matrix but could also be observed within cell bodies and neurites as well as within the endothelium of blood vessels with an increase in the density of small blood vessels in cervical sections of HD cases. These vasculature changes were accompanied with features of BSCB leakage, as assessed by the presence of increased levels of fibrinogen in the surrounding parenchyma and enhanced leukocyte infiltration.<h4>Interpretation</h4>This alteration in BSCB integrity may be explained, in part, by the dysregulation we found in some of the main proteins associated with it such as junctional adhesion molecule-1 and vascular endothelial cadherin. These observations have important implications for our understanding of HD pathology and may also have significant therapeutic implications. Ann Neurol 2017;82:981-994.

<h4>Background and objectives</h4>Despite the obligate iron loss from blood donation, some donors present with hyperferritinaemia that can result from a wide range of acute and chronic conditions including hereditary haemochromatosis (HH). The objective of our study was to investigate the causes of hyperferritinaemia in the blood donor population and explore the value of extensive HH mutational analyses.<h4>Materials and methods</h4>Forty-nine consecutive donors (f = 6, m = 43) were included prospectively from the Capital Regional Blood Center. Inclusion criteria were a single ferritin value >1000 μg/l or repeated hyperferritinaemia with at least one value >500 μg/l. All donors were questioned about their medical history and underwent a physical examination, biochemical investigations and next-generation sequencing of HH-related genes, including the HFE gene, the haemojuvelin gene (HFE2/HJV), the hepcidin gene (HAMP), the ferroportin 1 gene (SLC40A1) and the transferrin receptor 2 gene (TFR2).<h4>Results</h4>Forty of 49 donors were mutation positive with a combined 69 mutations, 54 of which were located in the HFE gene. There were 11 mutations in the TFR2 gene, two mutations in the HFE2 gene and two mutations in the HAMP gene. Only four donors had apparent alternative causes of hyperferritinaemia.<h4>Conclusion</h4>HH-related mutations were the most frequent cause of hyperferritinaemia in a Danish blood donor population, and it appears that several different HH-genotypes can contribute to hyperferritinaemia. HH screening in blood donors with high ferritin levels could be warranted. HH-related iron overload should not in itself result in donor ineligibility.

<h4>Background</h4>Annexin-1 (ANXA1) plays pivotal roles in regulating various physiological processes including inflammation, proliferation and apoptosis, and deregulation of ANXA1 functions has been associated with tumorigenesis and metastasis events in several types of cancer. Though ANXA1 levels correlate with breast cancer disease status and outcome, its distinct functional involvement in breast cancer initiation and progression remains unclear. We hypothesized that ANXA1-responsive kinase signaling alteration and associated phosphorylation signaling underlie early events in breast cancer initiation events and hence profiled ANXA1-dependent phosphorylation changes in mammary gland epithelial cells.<h4>Methods</h4>Quantitative phosphoproteomics analysis of mammary gland epithelial cells derived from ANXA1-heterozygous and ANXA1-deficient mice was carried out using stable isotope labeling with amino acids in cell culture (SILAC)-based mass spectrometry. Kinase and signaling changes underlying ANXA1 perturbations were derived by upstream kinase prediction and integrated network analysis of altered proteins and phosphoproteins.<h4>Results</h4>We identified a total of 8110 unique phosphorylation sites, of which 582 phosphorylation sites on 372 proteins had ANXA1-responsive changes. A majority of these phosphorylation changes occurred on proteins associated with cytoskeletal reorganization spanning the focal adhesion, stress fibers, and also the microtubule network proposing new roles for ANXA1 in regulating microtubule dynamics. Comparative analysis of regulated global proteome and phosphoproteome highlighted key differences in translational and post-translational effects of ANXA1, and suggested closely coordinated rewiring of the cell adhesion network. Kinase prediction analysis suggested activity modulation of calmodulin-dependent protein kinase II (CAMK2), P21-activated kinase (PAK), extracellular signal-regulated kinase (ERK), and IκB kinase (IKK) upon loss of ANXA1. Integrative analysis revealed regulation of the WNT and Hippo signaling pathways in ANXA1-deficient mammary epithelial cells, wherein there is downregulation of transcriptional effects of TEA domain family (TEAD) suggestive of ANXA1-responsive transcriptional rewiring.<h4>Conclusions</h4>The phosphoproteome landscape uncovered several novel perspectives for ANXA1 in mammary gland biology and highlighted its involvement in key signaling pathways modulating cell adhesion and migration that could contribute to breast cancer initiation.

Autophagy is a vital lysosomal degradation pathway that serves as a quality control mechanism. It rids the cell of damaged, toxic or excess cellular components, which if left to persist could be detrimental to the cell. It also serves as a recycling pathway to maintain protein synthesis under starvation conditions. A key initial event in autophagy is formation of the autophagosome, a unique double-membrane organelle that engulfs the cytosolic cargo destined for degradation. This step is mediated by the serine/threonine protein kinase ULK1 (unc-51-like kinase 1), which functions in a complex with at least three protein partners: FIP200 (focal adhesion kinase family interacting protein of 200 kDa), ATG (autophagy-related protein) 13 (ATG13), and ATG101. In this artcile, we focus on the regulation of the ULK1 complex during autophagy initiation. The complex pattern of upstream pathways that converge on ULK1 suggests that this complex acts as a node, converting multiple signals into autophagosome formation. Here, we review our current understanding of this regulation and in turn discuss what happens downstream, once the ULK1 complex becomes activated.

The endoplasmic reticulum (ER) is a key site for lipid biosynthesis and folding of nascent transmembrane and secretory proteins. These processes are maintained by careful homeostatic control of the environment within the ER lumen. Signalling sensors within the ER detect perturbations within the lumen (ER stress) and employ downstream signalling cascades that engage effector mechanisms to restore homeostasis. The most studied signalling mechanism that the ER employs is the unfolded protein response (UPR), which is known to increase a number of effector mechanisms, including autophagy. In this chapter, we will discuss the emerging role of autophagy as a UPR effector pathway. We will focus on the recently discovered selective autophagy pathway for ER, ER-phagy, with particular emphasis on the structure and function of known mammalian ER-phagy receptors, namely FAM134B, SEC62, RTN3 and CCPG1. Finally, we conclude with our view of where the future of this field can lead our understanding of the involvement of ER-phagy in ER homeostasis.

Cellular protein homeostasis is maintained by two major degradation pathways, namely the ubiquitin-proteasome system (UPS) and autophagy. Until recently, the UPS and autophagy were considered to be largely independent systems targeting proteins for degradation in the proteasome and lysosome, respectively. However, the identification of crucial roles of molecular players such as ubiquitin and p62 in both of these pathways as well as the observation that blocking the UPS affects autophagy flux and vice versa has generated interest in studying crosstalk between these pathways. Here, we critically review the current understanding of how the UPS and autophagy execute coordinated protein degradation at the molecular level, and shed light on our recent findings indicating an important role of an autophagy-associated transmembrane protein EI24 as a bridging molecule between the UPS and autophagy that functions by regulating the degradation of several E3 ligases with Really Interesting New Gene (RING)-domains.

Roquin proteins preclude spontaneous T cell activation and aberrant differentiation of T follicular helper (Tfh) or T helper 17 (Th17) cells. Here we showed that deletion of Roquin-encoding alleles specifically in regulatory T (Treg) cells also caused the activation of conventional T cells. Roquin-deficient Treg cells downregulated CD25, acquired a follicular Treg (Tfr) cell phenotype, and suppressed germinal center reactions but could not protect from colitis. Roquin inhibited the PI3K-mTOR signaling pathway by upregulation of Pten through interfering with miR-17∼92 binding to an overlapping cis-element in the Pten 3' UTR, and downregulated the Foxo1-specific E3 ubiquitin ligase Itch. Loss of Roquin enhanced Akt-mTOR signaling and protein synthesis, whereas inhibition of PI3K or mTOR in Roquin-deficient T cells corrected enhanced Tfh and Th17 or reduced iTreg cell differentiation. Thereby, Roquin-mediated control of PI3K-mTOR signaling prevents autoimmunity by restraining activation and differentiation of conventional T cells and specialization of Treg cells.

Proteotoxic stress plays an important role in the pathogenesis of Huntington's disease (HD). Autophagy is proposed as a compensatory mechanism to remove protein aggregates under proteotoxic stress by up-regulating p62 expression. In the present study, we investigated the molecular action of p62 to proteotoxic stress in HD cells. Using two different HD cellular models, STHdhQ7 and STHdhQ111 cells derived from wild type and HD knock-in mice and human fibroblasts from healthy and HD patients, we found that HD cells are more vulnerable to cell death under proteotoxic stress and during stress recovery. We further showed that P62 was up-regulated in both STHdhQ7 and STHdhQ111 cells in response to the stress with distinct subcellular localization patterns. While dispersed p62 puncti were found in STHdhQ7 cells, p62 bodies were initially present in the lysosomes and accumulated to the juxtanuclear regions of STHdhQ111 cells as MG132 incubation continued. Unlike in STHdhQ7 cells, p62 puncti were not associated with K48-linked polyubiquitinated protein aggregates or proteasomal components in STHdhQ111. Interestingly, addition of cysteine during MG132 incubation rescued cell death in STHdhQ111 cells caused by stress recovery and altered the subcellular distribution of p62. Our data suggest that aberrant positioning of p62 affects the proteasomal clearance of protein aggregates and may contribute to the increased vulnerability to proteotoxic stress-induced cell death in HD cells.

<h4>Background</h4>Forty years ago the actin cytoskeleton was determined to be disrupted in fibroblasts from persons with DNA repair-defective, hereditary colon cancer, with no clear connection between the cytoskeleton and DNA repair defects at that time. Recently, the large number of sequenced genomes has indicated that mammalian mutagenesis has a large stochastic component. As a result, large coding regions are large mutagen targets. Cytoskeletal protein-related coding regions (CPCRs), including extra-cellular matrix proteins, are among the largest coding regions in the genome and are indeed very commonly mutated in cancer.<h4>Methods</h4>To determine whether mutagen sensitivity of the actin cytoskeleton could be assessed experimentally, we treated tissue culture cells with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone and quantified overall cytoskeleton integrity with rhodamine-phalloidin stains for F-actin.<h4>Results</h4>The above approach indicated cytoskeletal degradation with increasing mutagen exposure, consistent with increased mutagenesis of CPCRs in TCGA, smoker samples, where overall mutation rates correlate with CPCR mutation rates (R² = 0.8694; p < 0.00001). In addition, mutagen exposure correlated with a decreasing cell perimeter to area ratio, raising questions about potential decreasing, intracellular diffusion and concentrations of chemotherapy drugs, with increasing mutagenesis and decreasing cytoskeleton integrity.<h4>Conclusion</h4>Determination of cytoskeletal integrity may provide the opportunity to assess mutation burdens in nonclonal cell populations, such as in intact tissues, where DNA sequencing for heterogeneous mutation burdens can be challenging.

Ipomoeassin F, a plant-derived macrolide, exhibited single-digit nanomolar growth inhibition activity against many cancer cell lines. In this report, a series of 5-oxa/aza analogues was prepared and screened for cytotoxicity. Replacement of 5-CH₂ with O/NH simplified the synthesis and led to only a small activity loss. N-methylation almost completely restored the potency. Further studies with additional 5-oxa analogues suggested, for the first time, that size and flexibility of the ring also significantly influence the bioactivity of ipomoeassin F.

Secondary impairment of blood-brain barrier (BBB) occurs in the remote thalamus after ischemic stroke. Netrin-1, an axonal guidance molecule, presents bifunctional effects on blood vessels through receptor-dependent pathways. This study investigates whether netrin-1 protects BBB against secondary injury. Netrin-1 (600 ng/d for 7 days) was intracerebroventricularly infused 24 h after middle cerebral artery occlusion (MCAO) in hypertensive rats. Neurological function was assessed 8 and 14 days after MCAO, and the permeability of BBB in the ipsilateral thalamus was detected. The viability of brain microvascular endothelial cells was determined after being disposed with netrin-1 (50 ng/mL) before oxygen-glucose deprivation (OGD). The role of netrin-1 was further explored by examining its receptors and their function. We found that netrin-1 infusion improved neurological function, attenuated secondary impairment of BBB by up-regulating the levels of tight junction proteins and diminishing extravasation of albumin, with autophagy activation 14 days after MCAO. Netrin-1 also enhanced cell survival and autophagy activity in OGD-treated cells, inhibited by UNC5H2 siRNA transfection. Furthermore, the beneficial effects of netrin-1 were suppressed by PI3K inhibitors 3-Methyladenine and LY294002. Our results showed that netrin-1 ameliorated BBB impairment secondary to ischemic stroke by promoting tight junction function and endothelial survival. PI3K-mediated autophagy activation depending on UNC5H2 receptor could be an underlying mechanism.

<i>Lactobacilli</i> are bacteria that are beneficial to host health, but information on communication between <i>Lactobacilli</i> and host cells in the intestine is lacking. In this study, we examined the proteomes of the <i>Lactobacillus mucosae</i> strain LM1, as a model of beneficial bacteria, and the intestinal porcine epithelial cell line (IPEC-J2) after co-culture. Label-free proteomics demonstrated the high-throughput capability of the technique, and robust characterization of the functional profiles and changes in the bacteria and intestinal cells was achieved in pure and mixed cultures. After co-culture, we identified totals of 376 and 653 differentially expressed proteins in the LM1 and IPEC-J2 proteomes, respectively. The major proteomic changes in the LM1 strain occurred in the functional categories of transcription, general function, and translation, whereas those in IPEC-J2 cells involved metabolic and cellular processes, and cellular component organization/biogenesis. Among them, elongation factor Tu, glyceraldehyde 3-phosphate dehydrogenase, and phosphocarrier protein HPr, which are known to be involved in bacterial adhesion, were upregulated in LM1. In contrast, proteins involved in tight junction assembly, actin organization, and genetic information processing (i.e., histones and signaling pathways) were significantly upregulated in IPEC-J2 cells. Furthermore, we identified functional pathways that are possibly involved in host-microbe crosstalk and response. These findings will provide novel insights into host-bacteria communication and the molecular mechanism of probiotic establishment in the intestine.

<i>Aspergillus fumigatus</i> is the most common mold species to cause disease in immunocompromised patients. Infection usually begins when its spores (conidia) are inhaled into the airways, where they germinate, forming hyphae that penetrate and destroy the lungs and disseminate to other organs, leading to high mortality. The ability of hyphae to penetrate the pulmonary epithelium is a key step in the infectious process. <i>A. fumigatus</i> produces extracellular proteases that are thought to enhance penetration by degrading host structural barriers. This study explores the role of the <i>A. fumigatus</i> transcription factor XprG in controlling secreted proteolytic activity and fungal virulence. We deleted <i>xprG</i>, alone and in combination with <i>prtT</i>, a transcription factor previously shown to regulate extracellular proteolysis. <i>xprG</i> deletion resulted in abnormal conidiogenesis and formation of lighter colored, more fragile conidia and a moderate reduction in the ability of culture filtrates (CFs) to degrade substrate proteins. Deletion of both <i>xprG</i> and <i>prtT</i> resulted in an additive reduction, generating a mutant strain producing CF with almost no ability to degrade substrate proteins. Detailed proteomic analysis identified numerous secreted proteases regulated by XprG and PrtT, alone and in combination. Interestingly, proteomics also identified reduced levels of secreted cell wall modifying enzymes (glucanases, chitinases) and allergens following deletion of these genes, suggesting they target additional cellular processes. Surprisingly, despite the major alteration in the secretome of the <i>xprG/prtT</i> null mutant, including two to fivefold reductions in the level of 24 proteases, 18 glucanases, 6 chitinases, and 19 allergens, it retained wild-type virulence in murine systemic and pulmonary models of infection. This study highlights the extreme adaptability of <i>A. fumigatus</i> during infection based on extensive gene redundancy.

<h4>Background</h4>The advent of comprehensive genomic profiling has markedly advanced the understanding of the biology of pediatric hematological malignancies, however, its application to clinical care is still unclear. We present our experience integrating genomic data into the clinical management of children with high-risk hematologic malignancies and blood disorders and describe the broad impact that genomic profiling has in multiple aspects of patient care.<h4>Methods</h4>The Precision in Pediatric Sequencing Program at Columbia University Medical Center instituted prospective clinical next-generation sequencing (NGS) for high-risk malignancies and blood disorders. Testing included cancer whole exome sequencing (WES) of matched tumor-normal samples or targeted sequencing of 467 cancer-associated genes, when sample adequacy was a concern, and tumor transcriptome (RNA-seq). A multidisciplinary molecular tumor board conducted interpretation of results and final tiered reports were transmitted to the electronic medical record according to patient preferences.<h4>Results</h4>Sixty-nine samples from 56 patients with high-risk hematologic malignancies and blood disorders were sequenced. Patients carried diagnoses of myeloid malignancy (<i>n</i> = 25), lymphoid malignancy (<i>n</i> = 25), or histiocytic disorder (<i>n</i> = 6). Six patients had only constitutional WES, performed for a suspicion of an inherited predisposition for their disease. For the remaining 50 patients, tumor was sequenced with matched normal tissue when available. The mean number of somatic variants per sample was low across the different disease categories (2.85 variants/sample). Interestingly, a gene fusion was identified by RNA-seq in 58% of samples who had adequate RNA available for testing. Molecular profiling of tumor tissue led to clinically impactful findings in 90% of patients. Forty patients (80%) had at least one targetable gene variant or fusion identified in their tumor tissue; however, only seven received targeted therapy. Importantly, NGS findings contributed to the refinement of diagnosis and prognosis for 34% of patients. Known or likely pathogenic germline alterations were discovered in 24% of patients involving cancer predisposition genes in 12% of cases.<h4>Conclusion</h4>Incorporating whole exome and transcriptome profiling of tumor and normal tissue into clinical practice is feasible, and the value that comprehensive testing provides extends beyond the ability to target-specific mutations.

Iron transport across the intestinal epithelium is facilitated by the divalent metal transporter 1 (DMT1) on the brush border membrane (BBM). The fluorescent metal sensor calcein, which is hydrophilic, membrane-impermeable and quenched by chelation with iron, was used to test our hypothesis that intestinal iron absorption is through the endocytic processes and is involved in a pathway where BBM-derived vesicles fuse with basolateral membrane (BLM)-derived vesicles. To monitor the flux of iron via transcytosis, Caco-2 cells were employed as a polarized cell layer in Transwell chambers. When calcein was added to the basal chamber along with apo-transferrin (apo-Tf), calcein rapidly underwent endocytosis and co-localized with apo-Tf. Calcein was quenched by adding an iron-ascorbate complex and then restored by adding 2,2'-dipyridyl into the apical chamber. These results were confirmed by live-cell imaging. When hemin from the apical surface and calcein from the basal chamber were added to the Caco-2 cells, internalization of DMT1 and quenching of calcein were not observed until 2 h later. These results indicated that absorbed hemin required processing before hemin-derived iron was available to BLM-derived vesicles. These studies suggest that iron is transported in Caco-2 cells by transcytosis with apical-derived vesicles that are fused to BLM-derived vesicles.

<h4>Aims</h4>Ferroportin (FPN) is an iron exporter that plays an important role in cellular and systemic iron metabolism. Our previous work has demonstrated that FPN is decreased in prostate tumors. We sought to identify the molecular pathways regulated by FPN in prostate cancer cells.<h4>Results</h4>We show that overexpression of FPN induces profound effects in cells representative of multiple histological subtypes of prostate cancer by activating different but converging pathways. Induction of FPN induces autophagy and activates the transcription factors tumor protein 53 (p53) and Kruppel-like factor 6 (KLF6) and their common downstream target, cyclin-dependent kinase inhibitor 1A (p21). FPN also induces cell cycle arrest and stress-induced DNA-damage genes. Effects of FPN are attributable to its effects on intracellular iron and can be reproduced with iron chelators. Importantly, expression of FPN not only inhibits proliferation of all prostate cancer cells studied but also reduces growth of tumors derived from castrate-resistant adenocarcinoma C4-2 cells in vivo.<h4>Innovation</h4>We use a novel model of FPN expression to interrogate molecular pathways triggered by iron depletion in prostate cancer cells. Since prostate cancer encompasses different subtypes with a highly variable clinical course, we further explore how histopathological subtype influences the response to iron depletion. We demonstrate that prostate cancer cells that derive from different histopathological subtypes activate converging pathways in response to FPN-mediated iron depletion. Activation of these pathways is sufficient to significantly reduce the growth of treatment-refractory C4-2 prostate tumors in vivo.<h4>Conclusions</h4>Our results may explain why FPN is dramatically suppressed in cancer cells, and they suggest that FPN agonists may be beneficial in the treatment of prostate cancer.

Huntington disease (HD) is a severe neuropsychiatric disorder caused by a cytosine-adenine-guanine (CAG) repeat expansion in the HTT gene. Although HD is frequently complicated by depression, it is still unknown to what extent common HTT CAG repeat size variations in the normal range could affect depression risk in the general population. Using binary logistic regression, we assessed the association between HTT CAG repeat size and depression risk in two well-characterized Dutch cohorts─the Netherlands Study of Depression and Anxiety and the Netherlands Study of Depression in Older Persons─including 2165 depressed and 1058 non-depressed persons. In both cohorts, separately as well as combined, there was a significant non-linear association between the risk of lifetime depression and HTT CAG repeat size in which both relatively short and relatively large alleles were associated with an increased risk of depression (β = -0.292 and β = 0.006 for the linear and the quadratic term, respectively; both P < 0.01 after adjustment for the effects of sex, age, and education level). The odds of lifetime depression were lowest in persons with a HTT CAG repeat size of 21 (odds ratio: 0.71, 95% confidence interval: 0.52 to 0.98) compared to the average odds in the total cohort. In conclusion, lifetime depression risk was higher with both relatively short and relatively large HTT CAG repeat sizes in the normal range. Our study provides important proof-of-principle that repeat polymorphisms can act as hitherto unappreciated but complex genetic modifiers of depression.

Common brain abnormalities in cortical morphology and functional organization are observed in psychiatric disorders and aging, reflecting shared genetic influences. This preliminary study aimed to examine the contribution of a polygenetic risk for psychiatric disorders (PRS_cross) to aging brain and to identify molecular mechanisms through the use of multimodal brain images, genotypes, and transcriptome data. We showed age-related cortical thinning in bilateral inferior frontal cortex (IFC) and superior temporal gyrus and alterations in the functional connectivity between bilateral IFC and between right IFC and right inferior parietal lobe as a function of PRS_cross. Interestingly, the genes in PRS_cross, that contributed most to aging neurodegeneration, were expressed in the functioanlly connected cortical regions. Especially, genes identified through the genotype-functional connectivity association analysis were commonly expressed in both cortical regions and formed strong gene networks with biological processes related to neural plasticity and synaptogenesis, regulated by glutamatergic and GABAergic transmission, neurotrophin signaling, and metabolism. This study suggested integrating genotype and transcriptome with neuroimage data sheds new light on the mechanisms of aging brain.

<h4>Objective</h4>This systematic review aims to review the evidence for the diagnostic accuracy of the non-English updated versions of Addenbrooke's Cognitive Examination (ACE) - the ACE-Revised (ACE-R) and the ACE-III - in the diagnosis of dementia.<h4>Methods</h4>A systematic search resulted in 16 eligible studies evaluating the diagnostic accuracy of ACE-R and ACE-III in ten different languages. Most studies were assessed as of medium to low quality using Standards for Reporting of Diagnostic Accuracy (STARD) guidance.<h4>Results</h4>The findings of excellent diagnostic accuracy are compromised by the methodological limitations of studies. While studies generally reported excellent diagnostic accuracy across and within different languages, optimal cut-offs even within particular language versions, varied.<h4>Conclusion</h4>There is a need for future research to address these limitations through adherence to STARD guidelines. The ACE-III is particularly under-evaluated and should be a focus of future research. The variance in obtained optimal cut-offs within language versions is an issue compromising clinical utility and could be addressed in future work through use of a-priori defined thresholds.

In cycling human endometrium, menstruation is followed by rapid estrogen-dependent growth. Upon ovulation, progesterone and rising cellular cAMP levels activate the transcription factor Forkhead box O1 (FOXO1) in endometrial stromal cells (EnSCs), leading to cell cycle exit and differentiation into decidual cells that control embryo implantation. Here we show that FOXO1 also causes acute senescence of a subpopulation of decidualizing EnSCs in an IL-8 dependent manner. Selective depletion or enrichment of this subpopulation revealed that decidual senescence drives the transient inflammatory response associated with endometrial receptivity. Further, senescent cells prevent differentiation of endometrial mesenchymal stem cells in decidualizing cultures. As the cycle progresses, IL-15 activated uterine natural killer (uNK) cells selectively target and clear senescent decidual cells through granule exocytosis. Our findings reveal that acute decidual senescence governs endometrial rejuvenation and remodeling at embryo implantation, and suggest a critical role for uNK cells in maintaining homeostasis in cycling endometrium.

Cdk5 is a post-mitotic kinase with complex roles in maintaining neuronal health. The various mechanisms by which Cdk5 inhibits and promotes neurodegeneration are still poorly understood. Here, we show that in <i>Drosophila melanogaster</i> Cdk5 regulates basal autophagy, a key mechanism suppressing neurodegeneration. In a targeted screen, Cdk5 genetically interacted with Acinus (Acn), a primarily nuclear protein, which promotes starvation-independent, basal autophagy. Loss of Cdk5, or its required cofactor p35, reduces S437-Acn phosphorylation, whereas Cdk5 gain-of-function increases pS437-Acn levels. The phospho-mimetic S437D mutation stabilizes Acn and promotes basal autophagy. In <i>p35</i> mutants, basal autophagy and lifespan are reduced, but restored to near wild-type levels in the presence of stabilized Acn^S437D. Expression of aggregation-prone polyQ-containing proteins or the Amyloid-β42 peptide, but not alpha-Synuclein, enhances Cdk5-dependent phosphorylation of S437-Acn. Our data indicate that Cdk5 is required to maintain the protective role of basal autophagy in the initial responses to a subset of neurodegenerative challenges.

Detailed characterization of the cell types in the human brain requires scalable experimental approaches to examine multiple aspects of the molecular state of individual cells, as well as computational integration of the data to produce unified cell-state annotations. Here we report improved high-throughput methods for single-nucleus droplet-based sequencing (snDrop-seq) and single-cell transposome hypersensitive site sequencing (scTHS-seq). We used each method to acquire nuclear transcriptomic and DNA accessibility maps for >60,000 single cells from human adult visual cortex, frontal cortex, and cerebellum. Integration of these data revealed regulatory elements and transcription factors that underlie cell-type distinctions, providing a basis for the study of complex processes in the brain, such as genetic programs that coordinate adult remyelination. We also mapped disease-associated risk variants to specific cellular populations, which provided insights into normal and pathogenic cellular processes in the human brain. This integrative multi-omics approach permits more detailed single-cell interrogation of complex organs and tissues.

We conducted a critical review of human papillomavirus (HPV) integration into the host genome in oral/oropharyngeal cancer, reviewed the literature for HPV-induced cancers, and obtained current data for HPV-related oral and oropharyngeal cancers. In addition, we performed studies to identify HPV integration sites and the relationship of integration to viral-host fusion transcripts and whether integration is required for HPV-associated oncogenesis. Viral integration of HPV into the host genome is not required for the viral life cycle and might not be necessary for cellular transformation, yet HPV integration is frequently reported in cervical and head and neck cancer specimens. Studies of large numbers of early cervical lesions revealed frequent viral integration into gene-poor regions of the host genome with comparatively rare integration into cellular genes, suggesting that integration is a stochastic event and that site of integration may be largely a function of chance. However, more recent studies of head and neck squamous cell carcinomas (HNSCCs) suggest that integration may represent an additional oncogenic mechanism through direct effects on cancer-related gene expression and generation of hybrid viral-host fusion transcripts. In HNSCC cell lines as well as primary tumors, integration into cancer-related genes leading to gene disruption has been reported. The studies have shown that integration-induced altered gene expression may be associated with tumor recurrence. Evidence from several studies indicates that viral integration into genic regions is accompanied by local amplification, increased expression in some cases, interruption of gene expression, and likely additional oncogenic effects. Similarly, reported examples of viral integration near microRNAs suggest that altered expression of these regulatory molecules may also contribute to oncogenesis. Future work is indicated to identify the mechanisms of these events on cancer cell behavior.

The cell cycle coordinates core functions such as replication and cell division. However, cell-cycle-regulated transcription in the control of non-core functions, such as cell identity maintenance through specific transcription factors (TFs) and signalling pathways remains unclear. Here, we provide a resource consisting of mapped transcriptomes in unsynchronized HeLa and U2OS cancer cells sorted for cell cycle phase by Fucci reporter expression. We developed a novel algorithm for data analysis that enables efficient visualization and data comparisons and identified cell cycle synchronization of Notch signalling and TFs associated with development. Furthermore, the cell cycle synchronizes with the circadian clock, providing a possible link between developmental transcriptional networks and the cell cycle. In conclusion we find that cell cycle synchronized transcriptional patterns are temporally compartmentalized and more complex than previously anticipated, involving genes, which control cell identity and development.

The aim of this study was to evaluate the potential effects of methyl donor supplementation of pregnant animals in the presence or absence of a concomitant lactation on the methylome of the offspring. Twenty Holstein cows, 10 nulliparous (non-lactating while pregnant) and 10 multiparous (lactating while pregnant) were blocked by parity and randomly assigned to an i.m. weekly injections of a placebo (CTRL) or a solution containing methyl donors (MET). After calving, 5 calves randomly selected from each treatment (two born to non-lactating and three to lactating dams) were blood-sampled to determine their full methylome. There were more than 2,000 CpG differentially methylated between calves born to CTRL and those born to MET, and also between calves born to lactating and non-lactating dams. Most of the differences affected genes involved in immune function, cell growth regulation and differentiation, kinase activity, and ion channeling. We conclude that the coexistence of pregnancy and lactation affects the methylome of the offspring, and that supplementation of methyl donors early in gestation has also consequences on the methylome.

<h4>Background</h4>Systemic morphine has evidence to support its use for reducing breathlessness in patients with severe chronic obstructive pulmonary disease (COPD). The effectiveness of the nebulized route, however, has not yet been confirmed. Recent studies have shown that opioid receptors are localized within epithelium of human trachea and large bronchi, a target site for a dosimetric nebulizer. The aim of this study was to compare any clinical or statistical differences in breathlessness intensity between nebulized 2.0% morphine and 0,9% NaCl in patients with very severe COPD.<h4>Methods</h4>The study was a double-blind, controlled, cross-over trial. Participants received morphine or NaCl during two 4-day periods. Sequence of periods was randomized. The primary outcome measure was reduction of breathlessness intensity now by ≥20 mm using a 100 mm visual analogue scale (VAS) at baseline, 15, 30, 60, 120, 180 and 240 min after daily administration, during normal activities.<h4>Results</h4>Ten of 11 patients included completed the study protocol. All patients experienced clinically and statistically significant (p < 0.0001) breathlessness reduction during morphine nebulization. Mean VAS changes for morphine and 0.9% NaCl periods were 25.4 mm (standard deviation (SD): 9.0; median: 23,0; range: 14.0 to 41,5; confidence interval (CI): 95%) and 6.3 mm (SD: 7.8; median: 6.8; range: -11,5 to 19,5; CI: 95%), respectively. No treatment emergent adverse effects were noted.<h4>Discussion</h4>Our study showed superiority of dosimetrically administered nebulized morphine compared to NaCl in reducing breathlessness. This may have been achieved through morphine's direct action on receptors in large airways, although a systemic effect from absorption through the lungs cannot be excluded.<h4>Trial registration</h4>Retrospectively registered (07.03.2017), ISRCTN14865597.

<h4>Background</h4>Acupuncture has been used as a common therapeutic tool in many disorders including anxiety and depression. Serotonin transporter (SERT) plays an important role in the pathology of anxiety and other mood disorders. The aim of this study was to evaluate the effects of acupuncture on lipopolysaccharide (LPS)-induced anxiety-like behaviors and SERT in the dorsal raphe nuclei (DRN).<h4>Methods</h4>Rats were given acupuncture at ST41 (Jiexi), LI11 (Quchi) or SI3 (Houxi) acupoint in LPS-treated rats. Anxiety-like behaviors of elevated plus maze (EPM) and open field test (OFT) were measured and expressions of SERT and/or c-Fos were also examined in the DRN using immunohistochemistry.<h4>Results</h4>The results showed that 1) acupuncture at ST41 acupoint, but neither LI11 nor SI3, significantly attenuated LPS-induced anxiety-like behaviors in EPM and OFT, 2) acupuncture at ST41 decreased SERT expression increased by LPS in the DRN.<h4>Conclusions</h4>Our results suggest that acupuncture can ameliorate anxiety-like behaviors, possibly through regulation of SERT in the DRN.

Melanoma treatment with the BRAF V600E inhibitor vemurafenib provides therapeutic benefits but the common emergence of drug resistance remains a challenge. We generated A375 melanoma cells resistant to vemurafenib with the goal of investigating changes in miRNA expression patterns that might contribute to resistance. Increased expression of miR-204-5p and miR-211-5p occurring in vemurafenib-resistant cells was determined to impact vemurafenib response. Their expression was rapidly affected by vemurafenib treatment through RNA stabilization. Similar effects were elicited by MEK and ERK inhibitors but not AKT or Rac inhibitors. Ectopic expression of both miRNA in drug-naïve human melanoma cells was sufficient to confer vemurafenib resistance and more robust tumor growth <i>in vivo</i> Conversely, silencing their expression in resistant cells inhibited cell growth. Joint overexpression of miR-204-5p and miR-211-5p durably stimulated Ras and MAPK upregulation after vemurafenib exposure. Overall, our findings show how upregulation of miR-204-5p and miR-211-5p following vemurafenib treatment enables the emergence of resistance, with potential implications for mechanism-based strategies to improve vemurafenib responses.<b>Significance:</b> Identification of miRNAs that enable resistance to BRAF inhibitors in melanoma suggests a mechanism-based strategy to limit resistance and improve clinical outcomes. <i>Cancer Res; 78(4); 1017-30. ©2017 AACR</i>.

Triple-negative breast cancer (TNBC) is a breast cancer subtype characterized by marked differences between White and Black/African-American women. We performed a systems-level analysis on datasets from The Cancer Genome Atlas to elucidate how the expression patterns of mRNAs are shaped by regulatory noncoding RNAs (ncRNA). Specifically, we studied isomiRs, that is, isoforms of miRNAs, and tRNA-derived fragments (tRF). In normal breast tissue, we observed a marked cohesiveness in both the ncRNA and mRNA layers and the associations between them. This cohesiveness was widely disrupted in TNBC. Many mRNAs become either differentially expressed or differentially wired between normal breast and TNBC in tandem with isomiR or tRF dysregulation. The affected pathways included energy metabolism, cell signaling, and immune responses. Within TNBC, the wiring of the affected pathways with isomiRs and tRFs differed in each race. Multiple isomiRs and tRFs arising from specific miRNA loci (e.g., miR-200c, miR-21, the miR-17/92 cluster, the miR-183/96/182 cluster) and from specific tRNA loci (e.g., the nuclear tRNA^Gly and tRNA^Leu, the mitochondrial tRNA^Val and tRNA^Pro) were strongly associated with the observed race disparities in TNBC. We highlight the race-specific aspects of transcriptome wiring by discussing in detail the metastasis-related <i>MAPK</i> and the <i>Wnt/β-catenin</i> signaling pathways, two of the many key pathways that were found differentially wired. In conclusion, by employing a data- and knowledge-driven approach, we comprehensively analyzed the normal and cancer transcriptomes to uncover novel key contributors to the race-based disparities of TNBC.<b>Significance:</b> This big data-driven study comparing normal and cancer transcriptomes uncovers RNA expression differences between Caucasian and African-American patients with triple-negative breast cancer that might help explain disparities in incidence and aggressive character. <i>Cancer Res; 78(5); 1140-54. ©2017 AACR</i>.

The group IVA calcium-dependent cytosolic phospholipase A2 (cPLA2α) enzyme directs a complex "eicosanoid storm" that accompanies the tissue response to injury. cPLA2α and its downstream eicosanoid mediators are also implicated in the pathogenesis of fibrosis in many organs, including the kidney. We aimed to determine the role of cPLA2α in bone marrow-derived cells in a murine model of renal fibrosis, unilateral ureteral obstruction (UUO). WT C57BL/6J mice were irradiated and engrafted with donor bone marrow from either WT mice [WT-bone marrow transplant (BMT)] or mice deficient in cPLA2α (KO-BMT). After full engraftment, mice underwent UUO and kidneys were collected 3, 7, and 14 days after injury. Using picrosirius red, collagen-3, and smooth muscle α actin staining, we determined that renal fibrosis was significantly attenuated in KO-BMT animals as compared with WT-BMT animals. Lipidomic analysis of homogenized kidneys demonstrated a time-dependent upregulation of pro-inflammatory eicosanoids after UUO; KO-BMT animals had lower levels of many of these eicosanoids. KO-BMT animals also had fewer infiltrating pro-inflammatory CD45+CD11b+Ly6C^hi macrophages and reduced message levels of pro-inflammatory cytokines. Our results indicate that cPLA2α and/or its downstream mediators, produced by bone marrow-derived cells, play a major role in eicosanoid production after renal injury and in renal fibrinogenesis.

Cell-surface carbohydrates play important roles in numerous biological processes through their interactions with various protein-binding partners. These interactions are made possible by the vast structural diversity of carbohydrates and the diverse array of carbohydrate presentations on the cell surface. Among the most complex and important carbohydrates are glycosaminoglycans (GAGs), which display varied stereochemistry, chain lengths, and patterns of sulfation. GAG-protein interactions participate in neuronal development, angiogenesis, spinal cord injury, viral invasion, and immune response. Unfortunately, little structural information is available for these complexes; indeed, for the highly sulfated chondroitin sulfate motifs, CS-E and CS-D, there are no structural data. We describe here the development and validation of the GAG-Dock computational method to predict accurately the binding poses of protein-bound GAGs. We validate that GAG-Dock reproduces accurately (<1-Å rmsd) the crystal structure poses for four known heparin-protein structures. Further, we predict the pose of heparin and chondroitin sulfate derivatives bound to the axon guidance proteins, protein tyrosine phosphatase σ (RPTPσ), and Nogo receptors 1-3 (NgR1-3). Such predictions should be useful in understanding and interpreting the role of GAGs in neural development and axonal regeneration after CNS injury.

The neurodegenerative disorder Huntington's disease (HD) is typically characterized by extensive loss of striatal neurons and the midlife onset of debilitating and progressive chorea, dementia, and psychological disturbance. HD is caused by a CAG repeat expansion in the <i>Huntingtin</i> (<i>HTT</i>) gene, translating to an elongated glutamine tract in the huntingtin protein. The pathogenic mechanism resulting in cell dysfunction and death beyond the causative mutation is not well defined. To further delineate the early molecular events in HD, we performed RNA-sequencing (RNA-seq) on striatal tissue from a cohort of 5-y-old <i>OVT73</i>-line sheep expressing a human CAG-expansion <i>HTT</i> cDNA transgene. Our HD <i>OVT73</i> sheep are a prodromal model and exhibit minimal pathology and no detectable neuronal loss. We identified significantly increased levels of the urea transporter <i>SLC14A1</i> in the <i>OVT73</i> striatum, along with other important osmotic regulators. Further investigation revealed elevated levels of the metabolite urea in the <i>OVT73</i> striatum and cerebellum, consistent with our recently published observation of increased urea in postmortem human brain from HD cases. Extending that finding, we demonstrate that postmortem human brain urea levels are elevated in a larger cohort of HD cases, including those with low-level neuropathology (Vonsattel grade 0/1). This elevation indicates increased protein catabolism, possibly as an alternate energy source given the generalized metabolic defect in HD. Increased urea and ammonia levels due to dysregulation of the urea cycle are known to cause neurologic impairment. Taken together, our findings indicate that aberrant urea metabolism could be the primary biochemical disruption initiating neuropathogenesis in HD.