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Background Exposure to endocrine-disrupting chemicals, such as Bisphenol A (BPA) and Bisphenol S (BPS), is widespread and has negative implications on embryonic development. Preliminary evidence revealed that in women undergoing IVF treatment, urinary BPA levels were associated with low serum anti-Mullerian hormone, however a definitive relationship between the two has not yet been characterized. Methods This study aimed to evaluate BPA and BPS effects on in vitro oocyte maturation and early preimplantation embryo development through i) analysis of anti-Mullerian hormone (AMH) and anti-Mullerian hormone receptor II (AMHRII), ii) investigation of developmental parameters, such as cleavage, blastocyst rates and developmental arrest, iii) detection of apoptosis and iv) assessment of possible sex ratio skew. An in vitro bovine model was used as a translational model for human early embryonic development. We first assessed AMH and AMHRII levels after bisphenol exposure during oocyte maturation. Zygotes were also analyzed during cleavage and blastocysts stages. Techniques used include in vitro fertilization, quantitative polymerase chain reaction (qPCR), western blotting, TUNEL and immunofluorescence. Results Our findings show that BPA significantly decreased cleavage (p < 0.001), blastocyst (p < 0.005) and overall developmental rates as well as significantly increased embryonic arrest at the 2–4 cell stage (p < 0.05). Additionally, both BPA and BPS significantly increased DNA fragmentation in 2–4 cells, 8–16 cells and blastocyst embryos (p < 0.05). Furthermore, BPA and BPS alter AMH and AMHRII at the mRNA and protein level in both oocytes and blastocysts. BPA, but not BPS, also significantly skews sex ratios towards female blastocysts (p < 0.05). Conclusion This study shows that BPA affects AMH and AMHRII expression during oocyte maturation and that BPS exerts its effects to a greater extent after fertilization and therefore may not be a safer alternative to BPA. Our data lay the foundation for future functional studies, such as receptor kinetics, downstream effectors, and promoter activation/inhibition to prove a functional relationship between bisphenols and the AMH signalling system.

Bisphenol S (BPS) is used as an alternative plasticizer to Bisphenol A (BPA), despite limited knowledge of potential adverse effects. BPA exhibits endocrine disrupting effects during development. This article focuses on the impact of bisphenols during oocyte maturation. Connexins (Cx) are gap junctional proteins that may be affected by bisphenols, providing insight into their mechanism during development. Cxs 37 and 43 are crucial in facilitating cell communication between cumulus cells and oocytes. Cumulus-oocyte complexes (COCs), denuded oocytes, and cumulus cells were exposed to 0.05 mg/mL BPA or BPS for 24 hours. Both compounds had no effect on Cx43. Cumulus cells exhibited a significant increase in Cx37 expression following BPA (p = 0.001) and BPS (p = 0.017) exposure. COCs treated with BPA had increased Cx37 protein expression, whilst BPS showed no effects, suggesting BPA and BPS act through different mechanisms. Experiments conducted in in vitro cultured cumulus cells, obtained by stripping germinal vesicle oocytes, showed significantly increased expression of Cx37 in BPA, but not the BPS, treated group. BPA significantly increased Cx37 protein expression, while BPS did not. Disrupted Cx37 following BPA exposure provides an indication of possible effects of bisphenols on connexins during the early stages of development.

Exposure to chemicals with known endocrine-disrupting effects, such as bisphenol A (BPA) and bisphenol S (BPS), leads to repercussions on oocyte development and, ultimately, on fertility. Bisphenol A is a plasticizer used worldwide that has been detected in blood, urine, tissue samples and follicular fluid. Due to its widely reported detrimental effects, BPA has been substituted with its analogue BPS. Previous experiments in our laboratory have shown that exposure of bovine oocytes to physiologically relevant doses of BPA resulted in spindle abnormalities, reduced meiosis progression, decreased blastocyst rate and gene expression changes. However, the effects of BPS have not yet been investigated. Anti-Müllerian hormone (AMH) has been reported to be a good marker of ovarian reserve and oocyte developmental capability and is commonly used in assisted reproduction for diagnostic measurements. There is evidence that women undergoing IVF with higher BPA levels have lower AMH levels and pregnancy success. The aim of this study was to assess the effect of BPA and BPS on AMH and its receptor as measures of oocyte developmental capability. Abattoir-derived bovine cumulus-oocyte complexes (COC) were matured in vitro in 4 groups: (1) control, (2) vehicle (0.1% ethanol), (3) BPA (0.05mg mL−1 in 0.1% ethanol), and (4) BPS (0.05mg mL−1 in 0.1% ethanol). Pools of 30 COC, 30 denuded oocytes, and cumulus cells corresponding to denuded oocytes were collected for each of the 4 experimental groups, and a minimum of 4 biological replicates were used for each analysis. Anti-Müllerian hormone and AMH receptor mRNA expression was measured in COC, denuded oocytes, and their corresponding cumulus cells using quantitative real-time PCR. Statistical analyses were performed using 1-way ANOVA. Results showed a decrease (P&lt;0.05) in AMH mRNA expression in BPA-treated oocytes (without cumulus cells). In addition, there was an increase (P&lt;0.05) in mRNA AMH receptor levels in COC when treated with BPS. Finally, analyses on cumulus cells alone showed an increase (P&lt;0.05) in the AMH receptor mRNA levels in BPA-treated cells. These results suggest that BPA has an effect on AMH mRNA transcript levels in oocytes, while affecting the receptor expression in cumulus cells. Conversely, BPS affects AMH indirectly, increasing the mRNA levels of its receptor only. Further investigation of the effects of BPA and BPS on AMH expression in in vitro-produced blastocysts derived from treated oocytes will aid in understanding the potential consequences of exposure to BPA and its analogues on early embryonic development.

Background Exposure to endocrine-disrupting chemicals, such as Bisphenol A (BPA) and Bisphenol S (BPS), is widespread and has negative implications on embryonic development. Preliminary evidence revealed that in women undergoing IVF treatment, urinary BPA levels were associated with low serum anti-Mullerian hormone, however a definitive relationship between the two has not yet been characterized. Methods This study aimed to evaluate BPA and BPS effects on in vitro oocyte maturation and early preimplantation embryo development through i) analysis of anti-Mullerian hormone (AMH) and anti-Mullerian hormone receptor II (AMHRII), ii) investigation of developmental parameters, such as cleavage, blastocyst rates and developmental arrest, iii) detection of apoptosis and iv) assessment of possible sex ratio skew. An in vitro bovine model was used as a translational model for human early embryonic development. We first assessed AMH and AMHRII levels after bisphenol exposure during oocyte maturation. Zygotes were also analyzed during cleavage and blastocysts stages. Techniques used include in vitro fertilization, quantitative polymerase chain reaction (qPCR), western blotting, TUNEL and immunofluorescence. Results Our findings show that BPA significantly decreased cleavage (p < 0.001), blastocyst (p < 0.005) and overall developmental rates as well as significantly increased embryonic arrest at the 2–4 cell stage (p < 0.05). Additionally, both BPA and BPS significantly increased DNA fragmentation in 2–4 cells, 8–16 cells and blastocyst embryos (p < 0.05). Furthermore, BPA and BPS alter AMH and AMHRII at the mRNA and protein level in both oocytes and blastocysts. BPA, but not BPS, also significantly skews sex ratios towards female blastocysts (p < 0.05) Conclusion This study shows that BPA affects AMH and AMHRII expression during oocyte maturation and that BPS exerts its effects to a greater extent after fertilization and therefore may not be a safer alternative to BPA. Our data lay the foundation for future functional studies, such as receptor kinetics, downstream effectors, and promoter activation/inhibition to prove a functional relationship between bisphenols and the AMH signalling system.

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