Interaction and dissociation by ligands of estrogen receptor and Hsp90: the antiestrogen RU 58668 induces a protein synthesis-dependent clustering of the receptor in the cytoplasm

The in vivo interaction of estrogen receptor (ER) and Hsp90, demonstrated in the absence of hormone by a nuclear cotranslocation assay of the cytoplasmic Hsp90 with the karyophilic receptor, was disrupted by agonist and antagonist ligands, which, after dissociating the Hsp90, allowed the chaperone protein to be relocalized in the cytoplasm. The pure antiestrogen RU 58668 (RU), which was unable to stimulate an estrogen-dependent reporter gene and completely inhibited its estradiol-induced activity, also profoundly modified the subcellular distribution of ER in a specific time- and dose-dependent manner; ER appeared as speckled fluorescent clusters mainly located in the perinuclear region of the cytoplasm. The kinetics of appearance and reversal of the RU-dependent ER mislocalization in the presence or absence of cycloheximide demonstrated 1) that this effect was reversed by RU withdrawal or estradiol (E2) treatment, and 2) that cycloheximide with RU inhibited and reversed the ER cytoplasmic mislocalization induced by RU alone. These results point to a protein synthesis-dependent step in the mechanism of action of this antiestrogen. After RU treatment, a large portion of ER was found in the particulate fraction of the cytoplasm. However, confocal and electron microscopic analysis showed that ER clusters were not associated with specific cytoplasmic organelles or compartments. Using ER mutants, it was found that the ligand binding domain was sufficient for RU to produce receptor mislocalization, while the constitutive nuclear localization signals were dispensable. We propose that the antiestrogenic properties of RU are primarily due to the induction of an aggregation-prone receptor conformation that cannot undertake the constitutive and the ligand-induced nuclear localization function of the receptor because it is sequestered in the cytoplasm by fast turning over protein(s). We predict that antiestrogens able to block ER nuclear localization will behave as pure antihormones and will inhibit all the nuclear action of ER elicited by agonistic ligands or by ligand-independent mechanisms such as growth factor stimulation.     

HLA class II genes in chronic hepatitis C virus-infection and associated immunological disorders

To investigate the factors that may confer susceptibility or protection to hepatitis C virus (HCV) infection and to HCV-associated immunological disorders, we designed two studies on 420 Sardinian transfusion-dependent thalassemia patients followed in our department in Cagliari since 1974. The first one was an epidemiological survey aimed to evaluate the prevalence of HCV infection and HCV-associated immunological disorders. In the second study, the distribution of different HLA class II genes was examined by DNA analysis in 116 HCV positive patients, 30 HCV negative patients, and 606 healthy controls. Three hundred fourteen patients became infected with HCV (74.7%) after 5.6 +/- 2.8 years of regular transfusion program. Mixed cryoglobulinemia, purpura, arthritis, proteinuria, decreased complement levels, rheumatoid factor and anti-GOR, smooth muscle antibody (SMA), anti-nuclear antibody (ANA), and liver, kidney microsome (LKM) autoantibodies were significantly more represented in HCV positive patients than in negative ones (P < .05). A significant increase of HLA class II DR2 subtype (DRB1*1601,DQB1*0502) was observed in a group of 30 HCV negative patients who despite 10.3 +/- 2.2 years in a regular blood transfusion program did not show any evidence of HCV infection (Pc < .0092). Our results represent clear evidence for a relationship between HCV infection and immune extrahepatic abnormalities. A gene(s) located in the human major histocompatibility complex (MHC) region may play an important role in conferring protection against HCV infection.     

Time of day variations in driving performance

A 25-year-old woman complained of anasarca and was admitted to Sakura National hospital on the presumptive diagnosis of nephrotic syndrome with 10.7 g of 24-hour urinary protein. At first, lupus nephritis with antiphospholipid antibody syndrome was suspected because of prolongation of APTT, existence of lupus anticoagulant and elevation of serum anticardiolipin antibody titer (IgM) in addition to positive ANA, lymphocytopenia and the biologically false positive test for syphilis (BFPTS). On day 28 of hospitalization, renal biopsy findings revealed severe endocapillary cell damage, such as swelling and proliferation of endothelial cells, fragmentation and double contour of the basement membrane walls, which were located only in the capillary lumens with a few thrombi. Immunofluorescent micrography revealed the absence of specific immunoglobulin or complement deposit. Therefore, the diagnosis of lupus nephritis was negated as these findings were suggestive of characteristic glomerulopathy due to primary antiphospholipid antibody syndrome. She was treated initially with oral prednisolone 60 mg and intravenous infusion of heparin 20,000 units daily. Moreover, cyclophosphamide 750 mg was administered intravenously as pulse therapy on day 13 as her serum level of CH50 had fallen suddenly, and hemodialysis was necessary because her renal function had deteriorated and she was suffering from cough and orthopnea with overhydratin. After the combined therapy, BFPTS disappeared and APTT returned to the normal range: dialysis treatment was not required further after the 4th hemodialysis. Thereafter, renal function improved and complete remission of nephrotic syndrome was obtained. This patient was a case of primary antiphospholipid antibody syndrome in which endothelial cell damage was located exclusively in the capillary lumens and pulse cyclophosphamide therapy in addition to prednisolone and anticoagulant was effective. We present this instructive case to promote understanding of the pathogenesis of primary antiphospholipid antibody syndrome.     

Clinical evaluation and microbiology of oropharyngeal infection due to fluconazole-resistant Candida in human immunodeficiency virus-infected patients

The aim of the present study was to investigate pregnancy rates ensuing from transfer of embryos with multinucleated blastomeres. In our in-vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) programme, 1735 embryo transfers were performed from January 1, 1995 to August 31, 1996. In 136 of these transfers at least one embryo with one or more multinucleated blastomeres was present per transfer (study group). For each of these 136 transfers, two matched controls with transfer of exclusively mononucleated embryos were selected (control group). Matching was carried out according to age, method of fertilization (IVF or ICSI), number of transferred embryos and quality score of transferred embryos. In the study group, there were eight transfers of exclusively multinucleated embryos from which one pregnancy ensued and 128 transfers in which multinucleated and mononucleated embryos were transferred together leading to 23 pregnancies. The overall clinical pregnancy rate per transfer was 16.9% in the study group versus 28.7% in the control group (P = 0.01). The ongoing pregnancy rate per transfer was 13.2% in the study group versus 23.2% in the control group (P = 0.03). The implantation rate per transferred embryo was 6.0% in the study group versus 11.3% in the control group (P = 0.003). This study shows that embryos with one or more multinucleated blastomeres have a poorer implantation potential than embryos with mononucleated blastomeres. Transfer of embryos with multinucleated blastomeres should hence only be considered when insufficient numbers of embryos with only mononucleated blastomeres are present.     

A new challenge in clinical research in childhood ALL: the prospective meta-analysis strategy for intergroup collaboration

We consider the problems arising in clinical research on childhood acute lymphoblastic leukemia (ALL). Given the therapeutic progress achieved over the last few decades, any improvement in the outcome for the majority of children with ALL is difficult to assess with the usual size trials. Furthermore, the progress in genetics and molecular biology has now led to the identification of subgroups of children, typically with rare characteristics, for whom new treatments still await evaluation. For both these aspects of clinical research, there is an increasing need for international intergroup cooperation. After a discussion on the role of retrospective meta-analysis and randomized controlled trials in ALL research, we suggest that intergroup studies could be made more feasible, but still scientifically rigorous, by adopting a strategy of prospective meta-analysis. This strategy can be described as follows: i) different groups prospectively plan to ask the same randomized question within their protocols which may differ in other aspects, and to pool their data in order to evaluate treatment effect; ii) the management of the study can be de-centralized, by allowing each group to be responsible for conducting its own protocol. We would like to stimulate the debate on the methodological and practical aspects of research perspectives in ALL (and in pediatric oncology).     

Improving the detection rate of early gastric cancer requires more than open access gastroscopy: a five year study

BACKGROUND/AIMS: To explore the reasons why patients with gastric cancer continue to present with advanced disease despite open access gastroscopy. PATIENTS: All patients diagnosed with gastric cancer between 1 August 1989 and 31 July 1994. METHODS: A retrospective study of the presentation of gastric cancer in South Tees; patients were diagnosed at open access gastroscopy or referred through conventional channels. Primary care records of 81 patients dying between 1991 and 1995 were analysed for previous symptoms, investigations, and antisecretory drug therapy. Findings were compared with 200 age and sex matched controls. RESULTS: The overall incidence of earlier stage gastric cancer remains low at 13%. Diagnostic delay occurs in both primary and secondary care due to a high incidence of previous dyspepsia and investigation. One in six patients had been previously investigated in the three years prior to diagnosis, the majority of whom were on antisecretory drugs. CONCLUSIONS: Early gastric cancer remains rare in South Tees health district. Advantages of open access gastroscopy appear to be compromised by delayed referral to hospital and failure of endoscopists to recognise the early disease; either they are unaware of its appearance or prior treatment with an H2 receptor antagonist masks the disease by allowing mucosal healing.     

Study of acid lability of DNA in patients with leptospirosis]

Measurements of DNA and study of chromatin status in blood cell nuclei help solve numerous basic problems of biology and medicine. Acid lability of DNA was assessed in 5 patients with grave icterohemorrhagic leptospirosis. The distribution of fluorescence peaks on the curve of neutrophil DNA hydrolysis in patients with grave leptospirosis indicates that functionally low-active acid-resistant DNA fraction predominates in their chromatin. Assessment of acid lability of DNA is proposed to be used for assessing the severity of the pathological process.     

Morbidity study of extruder personnel with potential exposure to brominated dioxins and furans. II. Results of clinical laboratory studies

OBJECTIVE: To test whether dioxins affect liver and thyroid function, lipid metabolism and glucose or immunological variables, in workers exposed to brominated dioxins and furans. METHODS: 34 male production employees (29 were extruder operators) and eight technical support personnel were studied, all of whom were potentially exposed to polybrominated dibenzo-p-dioxins (PBDDs) and furans (PBDFs) during production of resins containing polybrominated diphenyl ethers (PBDEs). Controls were from a similar resin producing plant that did not use PBDEs. Blood samples were analysed for tetra, penta, and hexabrominated congeners, but 2,3,7,8-TBDD was the only exposure measure used in the regression analyses. Seven liver function indicators, five measures of blood lipids and glucose, four haematology and blood coagulation measures, and three measures of thyroid function were examined. RESULTS: None of the variables was statistically related to concentration of 2,3,7,8-TBDD in the regression analyses. Cigarette smoking was related to several outcomes at the 0.05 level: aspartate aminotransferase, alanine aminotransferase, glutamate dehydrogenase (GLDH), erythrocyte sedimentation rate, and white blood cell count. Body mass index was also related to alanine aminotransferase, gamma-glutamyltranspeptidase, cholinesterase, GLDH, cholesterol, triglycerides, high density lipoprotein, low density lipoprotein, and glucose concentrations. No definitive associations between liver, blood lipid, thyroid, or immunological variables and exposure to brominated dioxins or blood lipid concentration of 2,3,7,8-TBDD were found. CONCLUSIONS: The study population was small and hence the findings must be interpreted with caution. Nevertheless, these results provide a base for interpreting the results of clinical studies in similarly exposed populations.