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Immunocytochemical progesterone receptor (PR) assays employing two different monoclonal antireceptor antibodies (mPRi, JZB39) were compared with each other and with the dextran-coated charcoal assay (DCC) in human breast carcinoma. The immunocytochemical analyses were semiquantitatively scored (histoscore) based on relative staining intensity and the percentage of positively stained carcinoma cell nuclei. The immunocytochemically determined PR status agreed with that obtained by the DCC assay in 76% of the cases when the monoclonal antibody mPRi was used. The agreement was slightly better (82%) with the immunocytochemical assay using the JZB39 antibody. The semiquantitative histoscores correlated significantly with log-transformed cytosol PR concentrations in the DCC assay (mPRi, R = 0.543; JZB39, R = 0.618; P less than 0.0001 for both). The histoscores obtained with the mPRi and JZB39 antibodies were highly correlated with each other in adjacent frozen sections (R = 0.84, P less than 0.0001) the latter antibody giving somewhat higher histoscores. These results further validate the use of immunocytochemical PR assays for routine diagnostic purposes, especially when sufficient material is not available for DCC assays.

To clarify the effects on blood coagulation-fibrinolytic system after transcatheter hepatic arterial therapy for cases of hepatocellular carcinoma (HCC), plasma levels of Plasmin-alpha 2PI complex (PIC), Ddimer and Thrombin-ATIII (TAT) before and after therapy were measured by EIA, in addition to other conventional coagulofibrinolytic parameters. In the group (9 cases) treated with intra-arterial injection of adriamycin, there were no significant changes of coagulofibrinolytic parameters except for Ddimer (P less than 0.05) which was elevated 1-2 days after therapy. However, only two cases in whom plasma PIC, Ddimer and TAT levels were clearly elevated before therapy, showed further marked elevation of those parameters after therapy. In the group (29 cases) treated with intra-arterial injection of adriamycin-lipiodol suspension, whether or not embolized with gelfoam, plasma PIC, Ddimer and TAT levels were significantly elevated (P less than 0.01) after therapy, as well as other conventional coagulofibrinolytic parameters. These results indicate that hypercoagulable and hyperfibrinolytic states were induced by treatment. Moreover, the secondary hyperfibrinolytic state tended to persist longer than the hypercoagulable state. The 14 cases embolized with gelfoam seemed to have more apparent effects on blood coagulation-fibrinolytic system than cases not treated with gelfoam. Therefore, we conclude that caution and prophylaxis for the occurrence of disseminated intravascular coagulation are necessary for transcatheter arterial therapy for cases of HCC.

A mutant of polyomavirus, F9-5000, capable of growing in F9 cell [M. Vasseur et al., J. Virol., 43: 800-808, 1982 (1)], has a deletion in the enhancer from nucleotide 5119 to nucleotide 5142. The oligonucleotide corresponding to the deleted region (delta F9-5000 element) showed silencer activity on gene expression in F9 cells. Mobility shift assay revealed a nuclear factor, PEBP4, in F9 nuclear extract which bound to the delta F9-5000 element. Mutations introduced into the PEBP4 binding site specifically abolished its binding as well as the inhibitory effect on gene expression. After F9 cells were induced to differentiate, two more factors, PEBP2 and PEBP1, a member of AP1 family, became detectable in addition to PEBP4, and at the same time the delta F9-5000 element lost silencer activity and acquired an enhancer activity. The recognition sequence of PEBP2 as well as that of PEBP1 overlapped with that of a repressor, PEBP4. PEBP4 and PEBP3, a factor related to PEBP2, were shown to compete for binding to delta F9-5000. Interplay of a ubiquitous negative factor and differentiation-induced positive factors may represent one aspect of the gene regulation during embryonic development.

The Met mutant of Drosophila melanogaster is highly resistant to juvenile hormone III (JH III) or its chemical analog, methoprene, an insect growth regulator. Five major mechanisms of insecticide resistance were examined in Met and susceptible Met+ flies. These two strains showed only minor differences when penetration, excretion, tissue sequestration, or metabolism of [3H]JH III was measured. In contrast, when we examined JH III binding by a cytosolic binding protein from a JH target tissue, Met strains had a 10-fold lower binding affinity than did Met+ strains. Studies using deficiency-bearing chromosomes provide strong evidence that the Met locus controls the binding protein characteristics and may encode the protein. These studies indicate that resistance in Met flies results from reduced binding affinity of a cytosolic binding protein for JH III.

Two hundred and fifty patients were entered into a randomized clinical study to compare the effectiveness of goserelin (Zoladex) in depot formulation with diethyl stilboestrol in locally advanced or metastatic prostate cancer. In 22 patients from the two arms of the study regular assessments were made of the effect of these hormone treatments on the haemostatic system. Selection of those patients with no recent surgical intervention and those on no drugs liable to interfere with the haemostatic mechanism was done at entry, in order to remove bias and achieve comparable groups. Baseline comparison of the two treatment groups showed no difference in clinical or biochemical measures of disease extent or activity, including serum prostate specific antigen (PSA) levels. There was a significant fall in plasma antithrombin-III (AT-III) activity in the DES treated group both from baseline and compared with the goserelin group. This effect commenced within 1 month and was maintained until monitoring ceased at 12 months. There was also a significant increase of fibrinolytic activity in the DES treated patients compared with those on goserelin. No divergence between the two treatment groups was seen in any other haematological parameters at baseline or on follow-up. A single AT-III estimation was also performed on a larger group of 74 patients at median follow-up time of 17 months (range 3-24). This confirmed the difference noted in the original study group. In the main study thrombotic episodes were noted in 13/126 patients treated with DES and 0/124 treated with goserelin (P less than 0.001). These findings suggest that lowered AT-III is the major factor through which DES affects the coagulation mechanism, and that no such effect is seen with goserelin treatment despite an equivalent therapeutic efficacy.

A 38-year-old patient with cerebral P. falciparum malaria was admitted 12 days after a short trip to Kenya. The serum level of tumor necrosis factor (TNF-alpha) was elevated (251 pg/ml). In contrast, Protein C (plasma activity 36.1%; antigen concentration 31.7%) and protein C inhibitor 1 (activity 0.55 U/ml) levels were decreased. This suggested a state of functional activation of the clotting system which was confirmed by elevated levels (4.8 ng/ml) of circulating thrombin-antithrombin-III-complexes (TAT). Protein S (total and free) and coagulation factor IX levels were within normal range. Under successful antiparasitic therapy, TNF-alpha as well as protein C and protein C inhibitor 1 levels returned to baseline within one week. In the context of other studies that demonstrate procoagulant effects of TNF-alpha, it is remarkable that in the case of complicated P. falciparum malaria, an elevated concentration of TNF-alpha can be paralleled by a decreased plasma level of protein C and an increase in TAT suggesting a procoagulant state.

Human caliciviruses (HCVs) are little known, recently recognized viruses associated with gastroenteritis. We identified HCV infection in an outbreak of gastroenteritis which occurred in one room of a day care center (DCC) participating in a longitudinal study of diarrhea. Utilizing an enzyme-linked immunoassay and immunosorbent electron microscopy to detect HCV, we tested specimens from all children in attendance during the period of the illness outbreak and during prior and subsequent weeks. HCV infection was documented in 14 children, 11 of whom were asymptomatic. Thirteen of the 14 HCV-infected children were 8 months of age or younger. New cases of HCV infection occurred during a 4-week period. Forty percent of children less than 1 year of age were infected with HCV during the period of investigation. Few documented HCV infections have been reported. This may be related to a high attack rate of predominantly asymptomatic infections in early life, resulting in a high prevalence of antibody to HCV by 4 years of age.

In situ hybridization was used to investigate the presence of mRNAs of three members of the serine protease inhibitor (serpin) superfamily, angiotensinogen (AG), alpha 1 protease inhibitor (alpha 1PI), and antithrombin III (ATIII) in normal human liver. The probes were full length 35S radiolabeled complementary DNAs of human AG, alpha 1PI, and ATIII. The three mRNAs were found to be uniformly distributed in all hepatocytes, with no evidence of any special distribution, but the signal was more intense for alpha 1PI than for AG and ATIII. Kupffer cells, biliary epithelial cells, and vascular cells were all negative. The same tissue was studied by peroxidase-antiperoxidase immunohistochemistry using specific antibodies against AG, alpha 1PI, and ATIII. No significant amounts of any of the proteins, alpha 1PI, AG, or ATIII were detected in frozen or fixed sections of normal liver. This study indicates that these proteins are not stored in the normal human hepatocyte, but that their genes are actively expressed and that in situ hybridization is the only technique presently available to detect their presence.

Alteration of antithrombin III (ATIII) activity, glycemia level dependent, exists in diabetes mellitus. In this study the ability of a low molecular weight heparin (LMWH) (Fluxum, Alfa-Wassermann S.p.A., Bologna, Italy), as well as unfractioned héparin, to preserve ATIII activity from glucose-induced alterations, both in vitro and in vivo, is reported. The subcutaneous and intravenous LMWH and heparin administration increases basal depressed ATIII activity in diabetic patients. Heparin shows an equivalent effect on both anti-IIa and anti-Xa activity of ATIII, while LMWH is more effective in preserving the anti-Xa activity. Similarity, heparin preserves ATIII activity from hyperglycemia-induced alterations, during hyperglycemic clamp, and LMWH infusion is able to preserve a significant amount of anti-Xa activity from glucose-induced alterations. Since diabetic patients show a high incidence of thrombotic accidents, LMWH appears to be a promising innovation for the prevention of diabetic thrombophylia.

The diagnostic efficacy of hepatic computed tomography density (HCTD) in comparison with serum ferritin for the detection of iron overload was investigated in uremic patients on maintenance hemodialysis (HD) and in patients with idiopathic hemochromatosis (IHC). Ten IHC patients, 38 HD patients and 40 healthy subjects underwent the CT scanning of the liver and determination of percent saturation of transferrin, serum ferritin concentration and HLA typing. Liver iron content was determined by histochemical grading and direct measurement of liver iron concentration either in IHC patients or in HD patients. Nineteen HD patients were considered to have iron overload on the basis of liver iron concentration exceeding 3.6 mumol/100 mg dry weight. The mean +/- SD values of HCTD in healthy subjects, IHC patients, HD patients with iron overload and without iron overload were 60.2 +/- 5.6, 79 +/- 5.6, 71.4 +/- 3.6, 58 +/- 3.8 Hounsfield units, respectively. HCTD showed positive correlations with liver iron concentration and serum ferritin either in IHC patients or in HD patients. The analysis of the diagnostic efficacy of HCTD in comparison with serum ferritin for the detection of excessive hepatic iron in HD patients demonstrated that HCTD had higher sensitivity, specificity, positive and negative predictive values. Cut-off points were arbitrarily fixed to 66 Hounsfield units for HCTD, 400 micrograms/liter for serum ferritin and 3.6 mumol/100 mg dry weight for liver iron concentration. Seventeen HD patients who possessed the histocompatibility antigens associated with IHC, namely HLA-A3 and/or HLA-B7 and/or HLA-B14, had liver iron concentration, serum ferritin and HCTD values higher than those of the HD patients without these "hemochromatosis alleles".(ABSTRACT TRUNCATED AT 250 WORDS)

We have recently found that phosphotungstate (PTA) has a heparin-like anticoagulant effect. In the present paper, we studied whether PTA is useful as an eluent of antithrombin III (ATIII) on heparin-Sepharose affinity chromatography. Human ATIII adsorbed on the heparin-Sepharose gel was eluted with NaCl buffer at the NaCl level of 1M. Whereas, PTA could elute ATIII at the level of less than 1mM and thus obtained ATIII fraction contained less impurities than such a fraction eluted with NaCl. Residual PTA in the eluate was easily decomposed by alkalization, being convenient for subsequent studies.

A new biological method for the assay of plasma antithrombin-III (AT-III) activity was developed without influence from heparin cofactor II (HC-II). AT-III deficient plasma is used as a substrate and diluted prothrombin time reagent as a reaction trigger for the specific assay of AT-III. The AT-III deficient plasma is prepared by passage of plasma through a heparin-agarose column. In the presence of heparin, AT-III in the sample shows concentration dependent anticoagulant activity and calibration curve is linear on semi-logarithmic graph paper. The results of reproducibility, recoveries and correlation studies with a chromogenic assay indicate that this biological method is reliable and suitable for routine use in clinical laboratories. Influence of HC-II is minimal. The method provides several advantages over those of chromogenic substrate and fibrinogen.

The data of HLA-haplotyping were used as an allele marker controlling hereditary hemochromatosis in 23 patients with beta-thalassemia. The results obtained have permitted a conclusion that hemosiderosis in patients with beta-thalassemia may be caused by association of beta-thalassemia gene with hereditary hemochromatosis. Early diagnosis of hyperferremia is of great prognostic importance as the adequate treatment timely conducted can prevent the development of irreversible changes in the patients.