Comparison of Short-Term Estrogenicity Tests for Identification of Hormone-Disrupting Chemicals

The aim of this study was to compare results obtained by eight different short-term assays of estrogenlike actions of chemicals conducted in 10 different laboratories in five countries. Twenty chemicals were selected to represent direct-acting estrogens, compounds with estrogenic metabolites, estrogenic antagonists, and a known cytotoxic agent. Also included in the test panel were 17β-estradiol as a positive control and ethanol as solvent control. The test compounds were coded before distribution. Test methods included direct binding to the estrogen receptor (ER), proliferation of MCF-7 cells, transient reporter gene expression in MCF-7 cells, reporter gene expression in yeast strains stably transfected with the human ER and an estrogen-responsive reporter gene, and vitellogenin production in juvenile rainbow trout. 17β-Estradiol, 17α-ethynyl estradiol, and diethylstilbestrol induced a strong estrogenic response in all test systems. Colchicine caused cytotoxicity only. Bisphenol A induced an estrogenic response in all assays. The results obtained for the remaining test compounds—tamoxifen, ICI 182.780, testosterone, bisphenol A dimethacrylate, 4-n-octylphenol, 4-n-nonylphenol, nonylphenol dodecylethoxylate, butylbenzylphthalate, dibutylphthalate, methoxychlor, o,p′-DDT, p,p′-DDE, endosulfan, chlomequat chloride, and ethanol—varied among the assays. The results demonstrate that careful standardization is necessary to obtain a reasonable degree of reproducibility. Also, similar methods vary in their sensitivity to estrogenic compounds. Thus, short-term tests are useful for screening purposes, but the methods must be further validated by additional interlaboratory and interassay comparisons to document the reliability of the methods

Pre-Operative Risk Factors Predict Post-Operative Respiratory Failure after Liver Transplantation

OBJECTIVE: Post-operative pulmonary complications significantly affect patient survival rates, but there is still no conclusive evidence regarding the effect of post-operative respiratory failure after liver transplantation on patient prognosis. This study aimed to predict the risk factors for post-operative respiratory failure (PRF) after liver transplantation and the impact on short-term survival rates. DESIGN: The retrospective observational cohort study was conducted in a twelve-bed adult surgical intensive care unit in northern Taiwan. The medical records of 147 liver transplant patients were reviewed from September 2002 to July 2007. Sixty-two experienced post-operative respiratory failure while the remaining 85 patients did not. MEASUREMENTS AND MAIN RESULTS: Gender, age, etiology, disease history, pre-operative ventilator use, molecular adsorbent re-circulating system (MARS) use, source of organ transplantation, model for end-stage liver disease score (MELD) and Child-Turcotte-Pugh score calculated immediately before surgery were assessed for the two groups. The length of the intensive care unit stay, admission duration, and mortality within 30 days, 3 months, and 1 year were also evaluated. Using a logistic regression model, post-operative respiratory failure correlated with diabetes mellitus prior to liver transplantation, pre-operative impaired renal function, pre-operative ventilator use, pre-operative MARS use and deceased donor source of organ transplantation (p<0.05). Once liver transplant patients developed PRF, their length of ICU stay and admission duration were prolonged, significantly increasing their mortality and morbidity (p<0.001). CONCLUSIONS: The predictive pre-operative risk factors significantly influenced the occurrence of post-operative respiratory failure after liver transplantation

Regulation of human endometrial function: mechanisms relevant to uterine bleeding

This review focuses on the complex events that occur in the endometrium after progesterone is withdrawn (or blocked) and menstrual bleeding ensues. A detailed understanding of these local mechanisms will enhance our knowledge of disturbed endometrial/uterine function – including problems with excessively heavy menstrual bleeding, endometriosis and breakthrough bleeding with progestin only contraception. The development of novel strategies to manage these clinically significant problems depends on such new understanding as does the development of new contraceptives which avoid the endometrial side effect of breakthrough bleeding

Autoimmune and autoinflammatory mechanisms in uveitis

The eye, as currently viewed, is neither immunologically ignorant nor sequestered from the systemic environment. The eye utilises distinct immunoregulatory mechanisms to preserve tissue and cellular function in the face of immune-mediated insult; clinically, inflammation following such an insult is termed uveitis. The intra-ocular inflammation in uveitis may be clinically obvious as a result of infection (e.g. toxoplasma, herpes), but in the main infection, if any, remains covert. We now recognise that healthy tissues including the retina have regulatory mechanisms imparted by control of myeloid cells through receptors (e.g. CD200R) and soluble inhibitory factors (e.g. alpha-MSH), regulation of the blood retinal barrier, and active immune surveillance. Once homoeostasis has been disrupted and inflammation ensues, the mechanisms to regulate inflammation, including T cell apoptosis, generation of Treg cells, and myeloid cell suppression in situ, are less successful. Why inflammation becomes persistent remains unknown, but extrapolating from animal models, possibilities include differential trafficking of T cells from the retina, residency of CD8(+) T cells, and alterations of myeloid cell phenotype and function. Translating lessons learned from animal models to humans has been helped by system biology approaches and informatics, which suggest that diseased animals and people share similar changes in T cell phenotypes and monocyte function to date. Together the data infer a possible cryptic infectious drive in uveitis that unlocks and drives persistent autoimmune responses, or promotes further innate immune responses. Thus there may be many mechanisms in common with those observed in autoinflammatory disorders

DAX-1 expression in human breast cancer: comparison with estrogen receptors ER-α, ER-β and androgen receptor status

BACKGROUND: So far there have been no reports on the expression pattern of DAX-1 (dosage-sensitive sex reversal, adrenal hypoplasia critical region, on chromosome X, gene 1) in human breast cells and its relationship to the estrogen receptors, ER-α and ER-β, and the androgen receptor (AR). METHODS: In this study we evaluated, by immunohistochemistry and Western blot analysis, the presence and distribution of DAX-1 in benign breast disease (BBD), in situ carcinoma (CIS), and ductal and lobular breast carcinomas. RESULTS: In BBD and breast carcinomas, DAX-1 was present in both the nuclei and the cytoplasm of epithelial cells, although in infiltrative carcinomas the percentage of nuclear immunoreaction was higher than in CIS. An important relation was observed between DAX-1 and AR expression and between this orphan receptor and nodal status. CONCLUSION: DAX-1 might modify the AR and ER-β intracellular location, and because a direct positive relation between the expression of these three receptors was found it could be assumed that the presence of DAX-1 in neoplastic cells might indicate a possible failure of endocrine therapies

The utility of pathway selective estrogen receptor ligands that inhibit nuclear factor-κB transcriptional activity in models of rheumatoid arthritis

Rheumatoid arthritis (RA) is a chronic inflammatory disease that produces synovial proliferation and joint erosions. The pathologic lesions of RA are driven through the production of inflammatory mediators in the synovium mediated, in part, by the transcription factor NF-κB. We have identified a non-steroidal estrogen receptor ligand, WAY-169916, that selectively inhibits NF-κB transcriptional activity but is devoid of conventional estrogenic activity. The activity of WAY-169916 was monitored in two models of arthritis, the HLA-B27 transgenic rat and the Lewis rat adjuvant-induced model, after daily oral administration. In both models, a near complete reversal in hindpaw scores was observed as well as marked improvements in the histological scores. In the Lewis rat adjuvant model, WAY-169916 markedly suppresses the adjuvant induction of three serum acute phase proteins: haptoglobin, α1-acid glycoprotein (α1-AGP), and C-reactive protein (CRP). Gene expression experiments also demonstrate a global suppression of adjuvant-induced gene expression in the spleen, liver, and popliteal lymph nodes. Finally, WAY-169916 was effective in suppressing tumor necrosis factor-α-mediated inflammatory gene expression in fibroblast-like synoviocytes isolated from patients with RA. Together, these data suggest the utility of WAY-169916, and other compounds in its class, in treating RA through global suppression of inflammation via selective blockade of NF-κB transcriptional activity

Rapid Insulinotropic Action of Low Doses of Bisphenol-A on Mouse and Human Islets of Langerhans: Role of Estrogen Receptor β

Bisphenol-A (BPA) is a widespread endocrine-disrupting chemical (EDC) used as the base compound in the manufacture of polycarbonate plastics. It alters pancreatic β-cell function and can be considered a risk factor for type 2 diabetes in rodents. Here we used ERβ−/− mice to study whether ERβ is involved in the rapid regulation of KATP channel activity, calcium signals and insulin release elicited by environmentally relevant doses of BPA (1 nM). We also investigated these effects of BPA in β-cells and whole islets of Langerhans from humans. 1 nM BPA rapidly decreased KATP channel activity, increased glucose-induced [Ca2+]i signals and insulin release in β-cells from WT mice but not in cells from ERβ−/− mice. The rapid reduction in the KATP channel activity and the insulinotropic effect was seen in human cells and islets. BPA actions were stronger in human islets compared to mouse islets when the same BPA concentration was used. Our findings suggest that BPA behaves as a strong estrogen via nuclear ERβ and indicate that results obtained with BPA in mouse β-cells may be extrapolated to humans. This supports that BPA should be considered as a risk factor for metabolic disorders in humans

Breast cancer oestrogen independence mediated by BCAR1 or BCAR3 genes is transmitted through mechanisms distinct from the oestrogen receptor signalling pathway or the epidermal growth factor receptor signalling pathway

INTRODUCTION: Tamoxifen is effective for endocrine treatment of oestrogen receptor-positive breast cancers but ultimately fails due to the development of resistance. A functional screen in human breast cancer cells identified two BCAR genes causing oestrogen-independent proliferation. The BCAR1 and BCAR3 genes both encode components of intracellular signal transduction, but their direct effect on breast cancer cell proliferation is not known. The aim of this study was to investigate the growth control mediated by these BCAR genes by gene expression profiling. METHODS: We have measured the expression changes induced by overexpression of the BCAR1 or BCAR3 gene in ZR-75-1 cells and have made direct comparisons with the expression changes after cell stimulation with oestrogen or epidermal growth factor (EGF). A comparison with published gene expression data of cell models and breast tumours is made. RESULTS: Relatively few changes in gene expression were detected in the BCAR-transfected cells, in comparison with the extensive and distinct differences in gene expression induced by oestrogen or EGF. Both BCAR1 and BCAR3 regulate discrete sets of genes in these ZR-75-1-derived cells, indicating that the proliferation signalling proceeds along distinct pathways. Oestrogen-regulated genes in our cell model showed general concordance with reported data of cell models and gene expression association with oestrogen receptor status of breast tumours. CONCLUSIONS: The direct comparison of the expression profiles of BCAR transfectants and oestrogen or EGF-stimulated cells strongly suggests that anti-oestrogen-resistant cell proliferation is not caused by alternative activation of the oestrogen receptor or by the epidermal growth factor receptor signalling pathway

In vitro studies on the modification of low-dose hyper-radiosensitivity in prostate cancer cells by incubation with genistein and estradiol

<p>Abstract</p> <p>Background</p> <p>As the majority of prostate cancers (PC) express estrogen receptors, we evaluated the combination of radiation and estrogenic stimulation (estrogen and genistein) on the radiosensitivity of PC cells in vitro.</p> <p>Methods</p> <p>PC cells LNCaP (androgen-sensitive) and PC-3 (androgen-independent) were evaluated. Estrogen receptor (ER) expression was analyzed by means of immunostaining. Cells were incubated in FCS-free media with genistein 10 μM and estradiol 10 μM 24 h before irradiation and up to 24 h after irradiation. Clonogenic survival, cell cycle changes, and expression of p21 were assessed.</p> <p>Results</p> <p>LNCaP expressed both ER-α and ER-β, PC-3 did not. Incubation of LNCaP and PC-3 with genistein resulted in a significant reduction of clonogenic survival. Incubation with estradiol exhibited in low concentrations (0.01 μM) stimulatory effects, while higher concentrations did not influence survival. Both genistein 10 μM and estradiol 10 μM increased low-dose hyper-radiosensitivity [HRS] in LNCaP, while hormonal incubation abolished HRS in PC-3. In LNCaP cells hormonal stimulation inhibited p21 induction after irradiation with 4 Gy. In PC-3 cells, the proportion of cells in G2/M was increased after irradiation with 4 Gy.</p> <p>Conclusion</p> <p>We found an increased HRS to low irradiation doses after incubation with estradiol or genistein in ER-α and ER-β positive LNCaP cells. This is of high clinical interest, as this tumor model reflects a locally advanced, androgen dependent PC. In contrast, in ER-α and ER-β negative PC-3 cells we observed an abolishing of the HRS to low irradiation doses by hormonal stimulation. The effects of both tested compounds on survival were ER and p53 independent. Since genistein and estradiol effects in both cell lines were comparable, neither ER- nor p53-expression seemed to play a role in the linked signalling. Nevertheless both compounds targeted the same molecular switch. To identify the underlying molecular mechanisms, further studies are needed.</p

Fluoro-deoxi-glucose uptake and angiogenesis are independent biological features in lung metastases

Neoangiogenesis and enhanced glucose metabolism in neoplasms are likely to be activated by the same biochemical stimulus; hypoxia. A correlation between these two parameters has been postulated. The objective of this study was to evaluate the relationship between Fluoro-desoxi-glucose uptake at positron emission tomography scan and angiogenesis in lung metastasis. Fluoro-desoxi-glucose activity, expressed as a standard uptake value, and microvessel intratumoural density, were retrospectively calculated in a series of 43 lung metastasis resected in 19 patients. Primary sites were colorectal cancer in 16 metastases, sarcoma in eight, gynaecological in four and other sites in 15. The correlation between the two parameters was tested by logistic regression and multivariate analysis. Positron emission tomography scan was positive in 17 patients (sensitivity 89%). No correlation was observed between standard uptake value and microvessel intratumoural density in this series of lung metastasis. Positron emission tomography negative and positive nodules presented comparable value of microvessel intratumoural density (12.9 vs 11.3). Standard uptake value was significantly correlated with nodules size and was higher in colon cancer metastasis than in sarcoma ones. Microvessel intratumoural density was independent from nodule size but significantly higher in sarcoma than in colon cancer metastasis. The lack of correlation was confirmed by multivariate analysis after adjustment for tumour type and nodules size. The present study demonstrated that positron emission tomography scan is positive in a high proportion of patients regardless of microvessel density. Glucose uptake and angiogenesis appear to be independent biological features in lung metastasis. This observation may have implications for future antiangiogenic therapies