Glutathione peroxidases (GPxs) are regarded as important protectors against oxidative stress. Some members of this protein family were reported to play key roles in protecting sperm against oxidative stress. Whether GPx6 a member of the GPx family also plays a role in protection against oxidative stress is not known to date. The objective of the present study was to evaluate the localization and function of glutathione peroxidase 6 (GPx6) in boar accessory sex glands, seminal plasma, and sperm, as well as the effect of GPx6 on vitality and capacitation in boar sperm. qPCR and Western blot analysis demonstrated the presence of GPx6 in testis, epididymis, bulbourethral glands, prostate, seminal vesicle, sperm and seminal plasma. Incubation of sperm with an GPx6 antibody had no significant effect on the viability of boar sperm prior to capacitation. Surprisingly, when capacitated sperm was incubated with the GPx6 antibody for 240 min, sperm vitality was significantly improved. Western blotting showed that in capacitated sperm without prior pretreatment, GPx6 protein content was reduced compared to sperm before capacitation. To further confirm a role for GPx6 in sperm capacitation, we tested sperm acrosome reaction by ACR.2 and FITC-PSA. The results showed that treatment of sperm with the GPx6 antibody significantly increased sperm capacitation and acrosome reaction. Furthermore, we examined the concentration of cAMP in sperm after capacitation. ELISA demonstrated that the cAMP concentration in the sperm exposed to the GPx6 antibody was significantly higher than that of the control group. In addition, the exposure of sperm to the GPx6 antibody significantly increased the concentration of H2O2, while the expression of SOD3 and CAT were decreased. Based on these observations we would like to postulate that in the boar reproductive tract the GPx6 protein becomes attached to the sperm head preventing the sperm to undergo premature capacitation by affecting components of the antioxidant pathway. How GPx6 expression following ejaculation becomes suppressed to allow sperm capacitation to take place needs further investigation.

中文翻译：

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GPx6 参与体外诱导的猪精子获能和顶体反应

谷胱甘肽过氧化物酶 (GPx) 被认为是抵御氧化应激的重要保护剂。据报道，该蛋白质家族的一些成员在保护精子免受氧化应激方面发挥着关键作用。GPx6 作为 GPx 家族的一员，是否也在抗氧化应激方面发挥作用，目前尚不清楚。本研究的目的是评估谷胱甘肽过氧化物酶 6 (GPx6) 在公猪附属性腺、精浆和精子中的定位和功能，以及 GPx6 对公猪精子活力和获能的影响。qPCR 和蛋白质印迹分析表明 GPx6 在睾丸、附睾、尿道球腺、前列腺、精囊、精子和精浆中存在。在获能前，用 GPx6 抗体孵育精子对公猪精子的活力没有显着影响。令人惊讶的是，当获能精子与 GPx6 抗体孵育 240 分钟时，精子活力显着提高。蛋白质印迹表明，在未经事先预处理的获能精子中，与获能前的精子相比，GPx6 蛋白含量降低。为了进一步确认 GPx6 在精子获能中的作用，我们通过 ACR.2 和 FITC-PSA 测试了精子顶体反应。结果表明，用 GPx6 抗体处理精子显着增加了精子获能和顶体反应。此外，我们检查了获能后精子中 cAMP 的浓度。ELISA 证明暴露于 GPx6 抗体的精子中的 cAMP 浓度显着高于对照组。此外，精子暴露于 GPx6 抗体显着增加了 H2O2 的浓度，而SOD3和CAT的表达降低。基于这些观察，我们想假设在公猪生殖道中 GPx6 蛋白附着在精子头部，通过影响抗氧化途径的成分来防止精子过早获能。射精后 GPx6 表达如何被抑制以允许精子获能需要进一步研究。