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目的探讨糖酵解途径在人精子顶体反应的作用及其分子机制。方法取健康志愿者捐献的精液,采用体外实验的方法,以糖酵解抑制剂S-3-氯-1,2-丙二醇(S-3-MCPD)处理人精子,处理组浓度分别为0.1 mmol/L、0.5 mmol/L和2.5 mmol/L,对照组加入相同体积溶剂。用考染法检测顶体反应发生率,免疫印迹法检测蛋白酪氨酸磷酸化,虫荧光素酶法检测ATP水平。结果 0.5 mmol/L和2.5 mmol/L S-3-MCPD处理组顶体反应发生率分别为34.5%和25.9%,与对照组(57.2%)比较,差异有统计学意义(P<0.01)。S-3-MCPD蛋白酪氨酸磷酸化存在显著剂量-效应关系。ATP含量与S-3-MCPD作用浓度呈显著负相关(r=0.769,P<0.01)。结论糖酵解途径在人精子顶体反应发生过程具有重要作用,其机制可能涉及糖酵解产生的ATP为蛋白酪氨酸磷酸化所需,而后者则是顶体反应的关键调节机制之一。
Objective To investigate the role of glycolytic pathway in acrosome reaction of human sperm and its molecular mechanism. Methods Semen donated by healthy volunteers was used to treat human sperm with the S-3-chloro-1,2-propanediol (S-3-MCPD), a glycolysis inhibitor. /L0.5 mmol / L and 2.5 mmol / L, the same volume of solvent control group. The incidence of acrosome reaction was detected by the test of infection, the protein tyrosine phosphorylation was detected by immunoblotting, and the ATP level by luciferase assay. Results The incidence of acrosome reaction was 34.5% and 25.9% in the 0.5 mmol / L and 2.5 mmol / L S-3-MCPD groups, respectively, which was significantly different from that in the control group (57.2%, P <0.01). S-3-MCPD protein tyrosine phosphorylation there is a significant dose-effect relationship. There was a significant negative correlation between ATP content and the concentration of S-3-MCPD (r = 0.769, P <0.01). Conclusions The glycolytic pathway plays an important role in the acrosome reaction of human sperm. The mechanism may involve that the ATP produced by glycolysis is required for protein tyrosine phosphorylation while the latter is one of the key regulatory mechanisms of acrosome reaction .