A Study On The Effects Of Bisphenol A And Sialic Acids On Human Sperm Function

Various environmental factors can cause reproductive health problems such as infertility. An increasing number of studies suggest that long-term exposure to harmful substances in the environment will cause a decline in semen quality and damage male fertility. This study aims to investigate the effects of typical environmental factors?bisphenol A and sialic acids? on human sperm together with the underlying mechanisms, wishing to provide insights for understanding male infertility.Bisphenol A?BPA?, a plastic monomer and plasticizer, is widely used in polycarbonate plastics and epoxy resins production. Owing to its extensive application, BPA becomes one of the chemicals produced worldwide with highest volume. Well-known as one of the endocrine disrupting chemicals?EDCs?, BPA has estrogen-like effect, and has been shown to impair mammalian male fertility. However, the studies of BPA on male fertility were mainly concentrated on animal models, and the BPA concentrations used usually were not physiological relevant. On the other hand, the studies regarding the direct effect of BPA on sperm were limited. BPA is detectable in blood, urine and semen of a healthy male, with the concentration of 2.0 ng/ml? 8.8 nM?, 3.25 ng/ml?¹4.3 nM? and ⁰.07 ng/ml?⁰.31 nM?, respectively. Our previous work has indicated that exposure of in vitro BPA at low concentrations, even as low as 1 pM, significantly inhibited motility and acrosome reaction of mouse sperm. However, it is unclear whether BPA at those physiologically relevant concentrations affects human sperm or not. This paper is the first study aiming to investigate the direct effects of physiologically relevant BPA on human sperm, hoping to gain new insight into the cause of male infertility. Objective: The objective of the present study is to investigate the effects of different concentrations of BPA?0, 10-3, 10-2, 10-1, 10, 103 nM? on human sperm functions in vitro, and further explore the underlying mechanisms. Methods:?1? An eosin-nigrosin staining kit was used to examine sperm viability.?2? Sperm motion parameters were measured by CASA.?3? Sperm penetration into methylcellulose was used to assess sperm hyperactivation.?4? Chlorotetracycline?CTC? staining was applied to evaluate human sperm capacitation.?5? Ca²⁺ signaling of human sperm was examined by a fluorescence plate reader.?6? The protein tyrosine phosphorylation related to sperm capacitation was analyzed qualitatively by western blotting. Results:BPA at low concentrations resulted in a remarkable decline in human sperm viability, motility and progressive motility, hyperactivation and capacitation. In addition, BPA suppressed human sperm protein tyrosine phosphorylation level, which is related to sperm capacitation. However, BPA had no effect on intracellular Ca²⁺ concentration in human sperm. Conclusions: BPA impaired sperm functions possibly via suppressing protein tyrosine phosphorylation of human sperm. The inhibition of human sperm functions by BPA at physiologically relevant concentrations suggests that environmental BPA pollution may be a candidate cause leading to impaired male fertility.Sialic acid?Sia? is a family of acid nine-carbon sugar acids. The most ubiquitous Sias are N-acetylneuraminic acid?Neu5Ac? and N-glycolyneuraminic acid?Neu5Gc?. However, human beings fail to express Neu5Ac due to the deletion of CMAH gene. Sias, the main components of glycoproteins and glycolipids, are the major glycan of sperm and zona pellucida, suggesting that they play important roles in modulating fertilization processes. A growing body of evidence has revealed that Sias regulate sperm functions in several ways, such as sperm immunization protection, sperm maturation in epididymal and sperm-egg binding. However, the direct effect of Sias on sperm has not been reported. Sia is not only one of the typical environmental factors, but also a crucial physiological factor in human body, which is closely related to human health. Exogenous Sias are ingested from diet with one part being absorbed into body and excreted into urine and the other part being incorporated into newly synthesized endogenous glycoproteins, leading to the production of antibodies and then affecting reproductive function. During capacitation, free Sia monosaccharides are cleaved by sialidase from bound Sias of sperm. However, whether those free Sias are only metabolites or may affect sperm function remains to be explored. The previous studies regarding the effects of Sias on reproductive tract were performed in animal models. In order to reveal the effects of Sias on human fertility as both physiological and environmental factors, this study is designed to perform on human sperm in vitro. Objective: The present study is designed to investigate the effects of Neu5Ac and Neu5Gc at different concentrations?0, 0.1, 1, 10 ?M? on sperm functions in vitro and the related mechanisms. Methods: The effects of Neu5Ac and Neu5Gc on human sperm were performed as follows:?1? Sperm motion parameters were measured by CASA.?2? Sperm penetration into methylcellulose was used to assess human sperm hyperactivation.?3? Human sperm acrosome reaction was performed by chlorotetracycline?CTC? staining.?5? Ca²⁺ signaling of human sperm was examined by a fluorescence plate reader.?6? The protein tyrosine phosphorylation was performed by western blotting. Results: Both Neu5Ac and Neu5Gc significantly inhibited human sperm functions, including motility, progressive motility, hyperactivation and acrosome reaction. Moreover, both Neu5Ac and Neu5Gc reduced intracellular Ca²⁺ concentration of human sperm and suppressed human sperm protein tyrosine phosphorylation levels, which were associated with sperm capacitation. Conclusions: Both Neu5Ac and Neu5Gc impaired sperm functions possibly via reducing intracellular Ca²⁺ concentration and suppressing protein tyrosine phosphorylation of human sperm. These results sugget that endogenous Neu5Ac may participate in regulating the timing of sperm acrosome reaction, while exogenous Neu5Gc existed in the female may has an adverse effect on reproduction.