Bisphenol A: Potential Factor of Miscarriage in Women in the Context of the Phenomenon of Neutrophil Extracellular Traps.

Humans are exposed to a number of environmental pollutants every day. Among them, endocrine disruptors are particularly harmful to human health. Bisphenol A (BPA) is a xenoestrogen that has been shown to disrupt the endocrine system and cause reproductive toxicity. In this study, we aimed to verify the potential relationship between BPA and miscarriage involving the formation of neutrophil extracellular traps (NETs). Blood samples were collected from healthy women and women who had miscarriage in the first trimester of pregnancy. The serum levels of cytoplasmic anti-PR3 antibody and perinuclear anti-MPO antibody were determined using an immunoenzymatic method. The concentrations of key proinflammatory proteins TNF-α and MCP-1, as well as NADPH oxidase subunits NOX1 and NCF2, were also measured in the serum samples. The serum concentration of BPA was determined using gas chromatography. The results showed that the concentrations of BPA were significantly elevated in the serum of women who had miscarriage compared to the control group, with the highest concentration found in the "NETs-positive" group. The levels of MCP-1 and TNF-α were significantly higher in the "NETs-positive" group compared to the "NETs-negative" and control group. The levels of NOX1 and NCF2 were also higher in the "NETs-positive" group compared to the "NETs-negative" group. The study showed that BPA could play a role in the course of miscarriage through the formation of NETs. The results indicate the need to limit the exposure of women planning pregnancy to xenoestrogens, including BPA.

Identification of the Bisphenol A (BPA) and the Two Analogues BPS and BPF in Cryptorchidism.

Objective: to explore the association of plasma concentrations of bisphenol A (BPA), bisphenol S (BPS), and bisphenol F (BPF) with unilateral cryptorchidism. In addition, to analyze selected demographic and intraoperative characteristics. Design: Retrospective analysis to determine plasma concentrations of total BPA, BPS and BPF using gas chromatography - mass spectrometry (GC-MS) among prepubertal boys with cryptorchidism and prebupertal male control subjects. During operation, the size, turgor and location of the cryptorchid testes were assessed. Main Outcome Measure: Plasma concentrations of total BPA, BPS and BPF. Results: In children with cryptorchidism, plasma levels of BPA, BPS and BPF were significantly higher compared to the control subjects. For BPA, it was: median value: 9.95 ng/mL vs. 5.54 ng/mL, p<0.05. For BPS, it was: median value: 3.93 ng/mL vs. 1.45 ng/mL, p<0.001. For BPF, it was: median value: 3.56 ng/mL vs. 1.83 ng/mL, p<0.05. In cryptorchid group, BPA was detected in 61.4% samples, BPS in 19.3% and BPF in 19.3%. All the three bisphenols were detected in plasma samples of both the healthy subjects and the study cohort. In the latter group, we found significant higher levels of BPA in boys from urban areas. We found a weak positive correlation between the levels of BPS and BPF and reduced turgor of the testes. Furthermore, results showed weak positive correlations between BPA and BPS levels and the age of the children as well as between BPS and BPF concentrations and the place of residence. Conclusions: Results provide a first characterization of prepubertal boys suffering from cryptorchidism and exposed to different kind of bisphenols. Our study suggests that cryptorchid boys are widely exposed to BPA and, to a lesser extent, also to its alternatives, such as BPS and BPF.

Sex-dependent dysregulation of human neutrophil responses by bisphenol A.

BACKGROUND: In the present study, we aimed to investigate selected functions of human neutrophils exposed to bisphenol A (BPA) under in vitro conditions. As BPA is classified among xenoestrogens, we compared its action and effects with those of 17ß-estradiol (E2). METHODS: Chemotaxis of neutrophils was examined using the Boyden chamber. Their phagocytosis and nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) oxidase activity were assessed via Park's method with latex beads and Park's test with nitroblue tetrazolium. To assess the total concentration of nitric oxide (NO), the Griess reaction was utilized. Flow cytometry was used to assess the expression of cluster of differentiation (CD) antigens. The formation of neutrophil extracellular traps (NETs) was analyzed using a microscope (IN Cell Analyzer 2200 system). Expression of the investigated proteins was determined using Western blot. RESULTS: The analysis of results obtained for both sexes demonstrated that after exposure to BPA, the chemotactic capacity of neutrophils was reduced. In the presence of BPA, the phagocytic activity was found to be elevated in the cells obtained from women and reduced in the cells from men. Following exposure to BPA, the percentage of neutrophils with CD14 and CD284 (TLR4) expression, as well as the percentage of cells forming NETs, was increased in the cells from both sexes. The stimulatory role of BPA and E2 in the activation of NADPH oxidase was observed only in female cells. On the other hand, no influence of E2 on the expression of CD14 and CD284, chemotaxis, phagocytosis, and the amount of NET-positive neutrophils was found for both sexes. The study further showed that BPA intensified NO production and iNOS expression in the cells of both sexes. In addition, intensified expression of all tested PI3K-Akt pathway proteins was observed in male neutrophils. CONCLUSIONS: The study demonstrated the influence of BPA on neutrophil functions associated with locomotion and pathogen elimination, which in turn may disturb the immune response of these cells in both women and men. Analysis of the obtained data showed that the effect of this xenoestrogen on the human neutrophils was more pronounced than E2.

Oxytocin antagonism reverses the effects of high oestrogen levels and oxytocin on decidualization and cyclooxygenase activity in endometrial tissues.

RESEARCH QUESTION: What is the in-vitro effect of oxytocin receptor (OTR) antagonism on parameters of receptivity in human endometrial explants and endometrial stromal cell lines cultured in oestradiol-rich conditions mimicking ovarian stimulation? DESIGN: Experimental in-vitro study on endometrial tissue explants collected by aspiration biopsy from 30 women undergoing fertility treatment and cultured endometrial tHESC cell line. The study examined the effects of high oestradiol, oxytocin and OTR antagonist on parameters of decidualization (cell viability and prolactin secretion) as well as cyclooxygenase-1/2 (COX-1/2) activity and prostaglandin F2α (PGF2α) secretion. Changes in expression of OXTR and COX-2 genes were examined using quantitative polymerase chain reaction (qPCR). RESULTS: In experiments on cultured endometrial cell line, high oestradiol and oxytocin similarly limited the viability of cells. In cultured endometrial explants both also decreased the secretion of prolactin (a marker of decidualization) and augmented endometrial COX-2 activity and formation of PGF2α. Oxytocin antagonist atosiban was confirmed to reverse the above effects, both in the endometrial line and endometrial explants. Addition of atosiban to cultures acted analogously in experiments employing both oxytocin and high oestradiol. CONCLUSIONS: Oxytocin antagonist reversed the effects of high oestradiol and oxytocin on parameters related to endometrial receptivity in conditions mimicking ovarian stimulation. This might point to a novel, endometrium-related mechanism to support embryo implantation achieved by the application of oxytocin antagonist prior to embryo transfer.

Simultaneous analysis of bisphenol A fractions in maternal and fetal compartments in early second trimester of pregnancy.

Background Bisphenol A (BPA) is an estrogenic, endocrine-disrupting compound widely used in the industry. It is also a ubiquitous environmental pollutant. Its presence was confirmed in human fetuses, which results from maternal exposure during pregnancy. The mechanisms behind maternal-fetal transfer, and relationships between pregnant women and fetal exposures remain unclear. The aim of this study was to assess the impact of maternal exposure to BPA on the exposure of the fetus. Methods Maternal plasma and amniotic fluid samples were collected from 52 pregnant women undergoing amniocentesis for prenatal diagnosis of chromosomal abnormalities. BPA was measured by gas chromatography-mass spectrometry (GC-MS). The permeability factor - a ratio of fetal-to-maternal BPA concentration - was used as a measure delineating the transplacental transfer of BPA. Results The median concentration of maternal plasma BPA was 8 times higher than the total BPA concentration in the amniotic fluid (8.69 ng/mL, range: 4.3 ng/mL-55.3 ng/mL vs. median 1.03 ng/mL, range: 0.3 ng/mL-10.1 ng/mL). There was no direct relationship between the levels of BPA in maternal plasma and amniotic fluid levels. The permeability factor, in turn, negatively correlated with fetal development (birth weight) (R = -0.54, P < 0.001). Conclusion Our results suggest that the risk of fetal BPA exposure depends on placental BPA permeability rather than the levels of maternal BPA plasma concentration and support general recommendations to become aware and avoid BPA-containing products.

Expression of serine proteases in neutrophils from women and men: Regulation by endocrine disruptor bisphenol A.

Bisphenol A (BPA) is a well-known endocrine disruptor. However, little information is available about its immunological effects. In the present study, we aimed to evaluate cytotoxic activity of BPA on human polymorphonuclear neutrophils (PMNs) according to gender and examine its effect on the expression of neutrophil serine proteases. Results indicated that exposure to BPA (above 16 µM) leads to a decrease in viability of PMNs and to morphological changes in these cells of both genders. The experiments showed different effects of BPA on the expression of proteinase 3, elastase, and cathepsin G in PMNs of both men and women, depending on the gender and concentration used. Thus, our findings suggest for the first time that through dysregulation of the expression of these enzymes, BPA may lead to disorders of the nonspecific cellular response in people exposed to this xenoestrogen.

Cellular, transcriptomic and methylome effects of individual and combined exposure to BPA, BPF, BPS on mouse spermatocyte GC-2 cell line.

Environmental factors, particularly xenoestrogens adversely affect reproductive health and their main mechanism is based on steroid-signaling pathway alterations. The presence of bisphenol A (BPA) in the environment has been confirmed and it is about to be replaced by analogues such as bisphenol F (BPF) and bisphenol S (BPS). Whether the BPF and BPS exert similar adverse effects to BPA has become the subject of intense scientific scrutiny. The aim of the present study was to evaluate and compare the cellular, transcriptomic and methylome effects of exposure to BPA, BPF, BPS individually and in combination on GC-2 spermatocyte cell line. The results show that all studied compounds affect cell viability, induce apoptosis and cause cellular damage. BPA, BPF and BPS also influence GC-2 cell steroid receptor and steroidogenesis related genes expressions. In addition to specific molecular mechanisms, all studied compounds also increase global DNA methylation. Exposure to a combination of all the studied compounds caused comparable effects on cell culture to each of them examined separately. These data suggest that exposure to BPA and its main substitutes- BPF and BPS induced multitude of effects and hence, BPF and BPS are not safe alternative to BPA in terms of male reproductive health.

Oxythiamine improves antifungal activity of ketoconazole evaluated in canine Malassezia pachydermatis strains.

BACKGROUND: Malassezia pachydermatis is an opportunistic yeast involved in skin and ear canal infections of dogs and cats. Reports suggest that strains of M. pachydermatis resistant to commonly used antifungal agents may be emerging. Therefore, new therapeutic strategies should be explored. OBJECTIVES: The synergistic effect of oxythiamine (OT) and ketoconazole (KTC) was analysed using a reference strain and field isolates (n = 66) of M. pachydermatis. Hydrogel formulations containing these components also were evaluated. METHODS AND MATERIALS: The minimum inhibitory concentrations (MICs) and minimum fungicidal concentrations (MFCs) of OT, KTC and their mixtures were determined by a broth macrodilution method. The antifungal effects of hydrogel formulations were determined by a plate diffusion method. RESULTS: The MIC and MFC values of OT were in the range 0.08 × 103 to 10 × 103  mg/L. All M. pachydermatis strains showed higher susceptibility to KTC (MICs and MFCs in the range 0.04-0.32 mg/L). Formulations that combined OT and KTC showed a synergistic effect for all tested isolates (n = 66). Hydrogels that contained OT at a concentration of 10 × 103 or 20 × 103  mg/L and KTC at the concentration of 0.1 × 103  mg/L showed a stronger effect than a commercially available product with KTC alone (20 × 103  mg/L). CONCLUSIONS AND CLINICAL IMPORTANCE: Synergy of these drugs may allow for successful topical treatment which utilizes lower doses of KTC without changing its therapeutic effectiveness. Hydrogel formulations proved to be attractive drug carriers for potential topical use.

Evaluation of Bisphenol A influence on endocannabinoid system in pregnant women.

Bisphenol A (BPA) is a synthetic chemical widely used in the industry, which may potentially evoke negative effects on human health, especially on reproductive processes and fetal development. BPA has been reported to act on estrogen, estrogen-related, androgen, thyroid hormone, pregnane X, peroxisome proliferation-activated, and aryl hydrocarbon receptors. However, other potential mechanisms of BPA action on pregnancy cannot be excluded. Comprehensive evaluation of BPA effect on pregnant women can be performed by use of metabolomics. In the present study LC-MS-based plasma metabolomics was performed in the group of pregnant women with known concentrations of free, conjugated and total BPA. Significant positive correlations were observed between several endocannabinoids (fatty acid amides) and free (r = 0.307-0.557, p-value = 0.05-0.00002) and total (r = 0.413-0.519, p-value = 0.008-0.00006) BPA concentrations. Palmitoleamide was positively correlated with conjugated (r = 0.348, p-value = 0.05) while lysophosphatidylethanolamine 18:0 with free (r = 0.519, p-value = 0.00006) BPA concentration. The docking calculations of BPA and fatty acid amide hydrolase (enzyme degrading endocannabinoids, FAAH) indicated that it can act as a competitive inhibitor by blocking FAAH catalytic residues. In vitro study showed that BPA moderately inhibits FAAH activity (15% decrease for 200 ng mL-1 and almost 50% for 200 µg mL-1 of BPA). In the present study for the first time inhibitory potential of BPA on FAAH hydrolase is reported. Inhibition of FAAH may lead to a rise of plasma endocannabinoids level. BPA exposure and increased level of endocannabinoids are miscarriage risk factors. Based on obtained results it can be hypothesized that BPA may induce adverse pregnancy outcomes by acting on endocannabinoid system.

Amniotic Fluid Angiogenic and Inflammatory Factor Profiling in Foetal Down Syndrome.

OBJECTIVES: Angiogenic factors are proteins that can potentially be related to certain foetal chromosomal abnormalities. The goal of this study was to determine the concentrations of 60 angiogenic factors in the amniotic fluid of women carrying foetuses with Down syndrome (DS). METHODS: After analysis of the karyotyping results, for the purpose of this study, we chose 12 women with foetal DS. For the control group, we selected 12 healthy patients with uncomplicated pregnancies (15-18 weeks of gestation) who delivered healthy newborns at term. To assess the concentrations of proteins in the amniotic fluid, we used a protein macroarray, which enabled the simultaneous determination of 60 angiogenic factors per sample. RESULTS: In the amniotic fluid of women with foetal DS compared to patients with healthy foetuses, we reported significant decreases in the concentrations of 14 angiogenic factors, including leptin, angiopoietin 1 (ANG-1), angiostatin, epidermal growth factor (EGF), interleukin 1-beta (IL-1b), interleukin 4 (IL-4), interleukin 12p40 (IL-12p40), monocyte chemotactic protein 2 (MCP-2), matrix metalloproteinase-1 (MMP-1), matrix metalloproteinase-9 (MMP-9), platelet endothelial cell adhesion molecule 1 (PECAM-1), transforming growth factor alpha (TGF alpha), vascular endothelial growth factor 2 (VEGFR2), and vascular endothelial growth factor 3 (VEGFR3). CONCLUSIONS: Based on our findings, we hypothesise that angiogenic factors may play roles in the pathogenesis of DS. Defining the factors' potential as biochemical factors of DS requires further investigation in a larger group of patients.