ZK93426促进小鼠精子获能相关蛋白酪氨酸磷酸化的机制研究

Mechanisms of ZK93426 on promoting mouse sperm capacitation-associated increase in protein tyrosine phosphorylation

目的探究苯二氮卓受体拮抗剂ZK93426促进小鼠精子获能相关蛋白酪氨酸化升高的作用机制。方法ZK93426与小鼠精子共孵育后,通过钙离子荧光探针Fluo4-AM检测小鼠精子胞内钙离子浓度(Intracellular calcium concentration,[Ca])变化,通过pH荧光探针BCECF-AM检测胞内pH(Intracellular pH,pH)变化,通过氯离子荧光探针MQAE检测胞内氯离子浓度(Intracellular chloride concentration,[Cl~-])变化,通过细胞膜电位荧光探针Di SC(5)检测细胞膜电位Em的变化。结果相比于DMSO溶剂对照组,ZK93426促进小鼠精子获能相关蛋白酪氨酸磷酸化升高的同时,伴随着精子细胞[Ca]、pH升高,[Cl~-]降低。ZK93426还抑制了精子获能过程中细胞膜的超极化。结论ZK93426可能通过调控[Ca],[Cl~-],pH及Em变化来共同调节小鼠精子获能相关蛋白酪氨酸磷酸化的升高。

Objective To investigate the mechanism of benzodiazepine receptor antagonist ZK93426 on promoting capacitation-associated increase in protein tyrosine phosphorylation.Methods After the mouse sperm incubated with ZK93426,changes of intracellular calcium ion concentration([Ca])was measured using calcium fluorescent probe Fluo4-AM;Changes of intracellular pH(pH)was measured using p H fluorescent probe BCECF-AM;Changes of intracellular chloride ion concentration([Cl])was measured using chloride fluorescent probe MQAE;And cell membrane potential(Em)were detected using fluorescence probe Di SC(5).Results Compared to DMSO control group,ZK93426 promoted the capacitationassociated increase in protein tyrosine phosphorylation in mice,accompanied by the increase of[Ca]and p Hand the decrease of[Cl].Membrane hyperpolarization was inhibited by ZK93426 as well.Conclusion ZK93426 may regulate the capacitation-associated increase in tyrosine phosphorylation of mouse sperm by modulating[Ca],[Cl],pHand Em.