An alkaline follicular fluid fraction induces capacitation and limited release of oviduct epithelium-bound stallion sperm
An alkaline follicular fluid fraction induces capacitation and limited release of oviduct epithelium-bound stallion sperm
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DOI：
10.1530/rep-15-0178
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发表时间：
2015-09-01
期刊：
影响因子：
3.8
通讯作者：
Van Soom, Ann
中科院分区：
文献类型：
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作者：
Leemans, Bart;Gadella, Bart M.;Van Soom, Ann
超活化运动的诱导被认为是触发释放输卵管结合的哺乳动物精子准备受精所必需的。在本研究中，将输卵管结合的种马精子暴露于：i）排卵前和ii）排卵后输卵管液; iii）100%和iv）10%卵泡液（FF）; v）卵丘细胞，vi）成熟马卵母细胞，vii）获能和viii）非获能培养基2 h。这些都没有触发精子释放或过度激活的运动。有趣的是，天然FF对精子活力有害，热灭活、活性炭处理和30 kDa过滤单独或组合可消除这种影响。此外，精子悬浮液暴露于处理FF在pH 7.9，但不是pH 7.4显示钙依赖性运动亢进。精子的Fluo-4 AM染色显示，与在pH 7.9的限定获能条件下的适度响应和在pH 7.4的FF处理中无响应相比，在pH 7.9的FF处理中暴露于FF处理的超活化种马精子中的细胞质Ca 2+升高。此外，1小时的孵育在碱性，处理FF诱导蛋白酪氨酸磷酸化的精子中的20%。测试的条件都没有诱导预先结合到输卵管上皮的精子的广泛释放。然而，超活化条件确实诱导每个输卵管外植体释放70-120个精子，其中48%显示蛋白酪氨酸磷酸化，并且所有精子都是顶体完整的，但能够响应钙离子载体而进行顶体胞吐。我们的结论是，在升高的pH值和细胞外Ca 2+的存在下，
Induction of hyperactivated motility is considered essential for triggering the release of oviduct-bound mammalian spermatozoa in preparation for fertilization. In this study, oviduct-bound stallion spermatozoa were exposed for 2 h to: i) pre-ovulatory and ii) post-ovulatory oviductal fluid; iii) 100% and iv) 10% follicular fluid (FF); v) cumulus cells, vi) mature equine oocytes, vii) capacitating and viii) non-capacitating medium. None of these triggered sperm release or hyperactivated motility. Interestingly, native FF was detrimental to sperm viability, an effect that was negated by heat inactivation, charcoal treatment and 30 kDa filtration alone or in combination. Moreover, sperm suspensions exposed to treated FF at pH 7.9 but not pH 7.4 showed Ca2+-dependent hypermotility. Fluo-4 AM staining of sperm showed elevated cytoplasmic Ca2+ in hyperactivated stallion spermatozoa exposed to treated FF at pH 7.9 compared to a modest response in defined capacitating conditions at pH 7.9 and no response in treated FF at pH 7.4. Moreover, 1 h incubation in alkaline, treated FF induced protein tyrosine phosphorylation in 20% of spermatozoa. None of the conditions tested induced widespread release of sperm pre-bound to oviduct epithelium. However, the hyperactivating conditions did induce release of 70-120 spermatozoa per oviduct explant, of which 48% showed protein tyrosine phosphorylation and all were acrosome-intact, but capable of acrosomal exocytosis in response to calcium ionophore. We conclude that, in the presence of elevated pH and extracellular Ca2+, a heat-resistant, hydrophilic,