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首页> 外文期刊>Cryobiology: International Journal of Low Temperature Biology and Medicine >Effect of cooling (4 degrees C) and cryopreservation on cytoskeleton actin and protein tyrosine phosphorylation in buffalo spermatozoa
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Effect of cooling (4 degrees C) and cryopreservation on cytoskeleton actin and protein tyrosine phosphorylation in buffalo spermatozoa

机译：冷却（4℃）和冷冻保存对水牛精子细胞骨架肌动蛋白和蛋白质酪氨酸磷酸化的影响

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摘要

Semen cryopreservation is broadly utilized as a part of the bovine reproducing industry, a large portion of the spermatozoa does not survive and the majority of those that do survive experience various molecular and physiological changes that influence their fertilizing capacity. The main aim of this study is to determine the effect of cooling (4 degrees C) and cryopreservation on cytoskeleton actin, tyrosine phosphorylation and quality of buffalo spermatozoa, and to determine the similarity between in vitro capacitation and cryopreservation induced capacitation like changes. To achieve this, Western blot was used to examine the changes in actin expression and protein tyrosine phosphorylation, whereas changes in actin polymerization, localization of actin and protein tyrosine phosphorylation during capacitation and cryopreservation were evaluated by indirect immunofluorescence technique. Localization studies revealed that the actin localized to flagella and acrosome membrane regions and following, capacitation it migrated towards the acrosome region of sperm. Time dependent increase in actin polymerization and protein tyrosine phosphorylation was observed during in vitro capacitation. The cooling phase (4 degrees C) and cryopreservation processes resulted in the loss/damage of cytoskeleton actin. In addition, we performed the actin polymerization and protein tyrosine phosphorylation in cooled and cryopreserved buffalo spermatozoa. Interestingly, cooling and cryopreservation induces actin polymerization and protein tyrosine phosphorylation, which were similar to in vitro capacitation (cryo-capacitation). These changes showed 1.3 folds reduction in the sperm quality parameters which includes motility, viability and plasma membrane integrity. Furthermore, our findings indicate that cooling and cryopreservation damages the cytoskeleton actin and also induces capacitation like changes such as protein tyrosine phosphorylation and actin polymerization. This could be one of the main reasons for reduced sperm quality and fertility failure of cryopreserved spermatozoa. (C) 2015 Elsevier Inc. All rights reserved.
机译：冷冻精液被广泛用作牛繁殖业的一部分，大部分精子不能存活，并且大多数存活的精子会经历各种影响其受精能力的分子和生理变化。这项研究的主要目的是确定冷却（4°C）和冷冻保存对水牛精子细胞骨架肌动蛋白，酪氨酸磷酸化和质量的影响，并确定体外获能和冷冻保存诱导的获能之类的变化之间的相似性。为此，采用蛋白质印迹法检查肌动蛋白表达和蛋白酪氨酸磷酸化的变化，而通过间接免疫荧光技术评估了获能和冷冻保存过程中肌动蛋白聚合，肌动蛋白的定位和蛋白酪氨酸磷酸化的变化。定位研究表明，肌动蛋白定位于鞭毛和顶体膜区域，继而获能后，肌动蛋白向精子的顶体区域迁移。在体外获能过程中观察到肌动蛋白聚合和蛋白质酪氨酸磷酸化的时间依赖性增加。冷却阶段（4摄氏度）和冷冻保存过程导致细胞骨架肌动蛋白的丢失/损坏。此外，我们在冷却和冷冻保存的水牛精子中进行了肌动蛋白聚合和蛋白质酪氨酸磷酸化。有趣的是，冷却和冷冻保存诱导肌动蛋白聚合和蛋白质酪氨酸磷酸化，这类似于体外捕获（冷冻捕获）。这些变化表明精子质量参数降低了1.3倍，其中包括活力，生存力和质膜完整性。此外，我们的发现表明，冷却和冷冻保存会破坏细胞骨架肌动蛋白，并且还会诱导获能，例如蛋白质酪氨酸磷酸化和肌动蛋白聚合等变化。这可能是精子质量下降和冷冻保存的精子失败的主要原因之一。 （C）2015 Elsevier Inc.保留所有权利。

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