Osteopontin (OPN) is a secreted extracellular matrix phosphoprotein identified in various tissues and fluids including those of the male and female reproductive tracts. OPN was previously identified as a 55 kDa high fertility marker in Holstein bull seminal plasma, produced by the ampulla and the vesicular gland. The objectives of this study were to characterize OPN on ejaculated and cauda epididymal sperm using immunofluorescence and western blot analysis, and to assess the role of sperm OPN in fertilization. Solubilized sperm membrane proteins from ejaculated and cauda epididymal sperm were separated by 1D SDS-PAGE, transferred to nitrocellulose, and probed with an antibody to bovine milk OPN. A 35 kDa protein was detected by this antibody in both ejaculated and cauda epididymal sperm membranes. Analyses also recognized OPN at 55 and 25 kDa in cauda epididymal fluid and testicular parenchyma homogenates respectively. Immunofluorescent analysis of ejaculated and cauda epididymal sperm showed OPN localization in a well-defined band in the postacrosomal region of the sperm head and also on the midpiece. Results of in vitro fertilization experiments showed that sperm treated with an antibody to OPN fertilized fewer oocytes than sperm treated with control medium while increasing incidence of polyspermy, suggesting a role of sperm-associated OPN in fertilization and a block to polyspermy. These studies demonstrate that OPN exists at multiple molecular weight forms in the bull reproductive tract and its presence on ejaculated sperm may signal its importance in fertilization by interacting with integrins or other proteins on the oocyte plasma membrane.

译文

:骨桥蛋白(OPN)是一种分泌的细胞外基质磷蛋白,可在各种组织和体液(包括男性和女性生殖道的组织和体液)中鉴定出。在壶腹和水泡腺产生的荷斯坦公牛精浆中,OPN先前被鉴定为55 kDa的高生育力标记。这项研究的目的是使用免疫荧光和蛋白质印迹分析来表征射精和马尾附睾精子上的OPN,并评估精子OPN在受精中的作用。通过1D SDS-PAGE分离射精和附睾附睾精子中的可溶精子膜蛋白,将其转移到硝酸纤维素膜上,并用针对牛乳OPN的抗体进行探测。用此抗体在射精和马尾附睾精子膜中检测到35 kDa的蛋白质。分析还分别在马尾附睾液和睾丸薄壁组织匀浆中分别以55和25 kDa的浓度识别了OPN。射精和马尾附睾精子的免疫荧光分析显示,OPN定位在精子头部顶体后区域以及中段的明确定义的条带中。体外受精实验的结果表明,用抗OPN抗体处理的精子比用对照培养基处理的精子受精的卵子少,同时增加了多精子的发生率,这表明与精子相关的OPN在受精中的作用和对多精子的阻滞。这些研究表明,OPN在牛的生殖道中以多种分子量形式存在,并且其在精子上的存在可能通过与卵母细胞质膜上的整联蛋白或其他蛋白质相互作用来表明其在受精中的重要性。

+1
+2
100研值 100研值 ¥99课程
检索文献一次
下载文献一次

去下载>

成功解锁2个技能,为你点赞

《SCI写作十大必备语法》
解决你的SCI语法难题!

技能熟练度+1

视频课《玩转文献检索》
让你成为检索达人!

恭喜完成新手挑战

手机微信扫一扫,添加好友领取

免费领《Endnote文献管理工具+教程》

微信扫码, 免费领取

手机登录

获取验证码
登录