权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Role of μ-Calpain in Human Sperm for Fertilization

μ-钙蛋白酶在人类精子受精中的作用

基本信息

批准号：
14571577
负责人：
OZAKI Yasuhiko
金额：
$ 1.86万
依托单位：
Nagoya City University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14571577/
关键词：
calpain human sparm acrosome reaction calcium ion proteolysis 受精能獲得男性不妊症

项目摘要

We evaluated the role of μ-calpain in human sperm for fertilization.Semen collected manually from healthy donors with informed consent was liquefied and following percoll gradient centrifugation. After exposure to different concentration of progesterone (P) or under hypoxic condition (H), the samples were used for immunostaining, SDS-PAGE and western blot analysis. The increase of calcium ion concentration in the sperm was observed by fluorescent microscope (ARGUS-50CA) system using Fura 2-AM. The role of calpain for fertilization was speculated from the results of Hamster penetration test, Hamilton Motility Analyzer, Triple satin method and Acrobeads test using calpain inhibitors.Immunostaining in human sperm was observed using antibodies against the μ-calpain in the types of whole acrosome, equatorial segment, head segment, neck segment or neck and tail segment. Most of these demonstrated acrosome type staining with anti-pro μ-calpain antibody. Western blot analysis revealed P or H treatment to cause a concentration or time-dependent activation of μ-calpain.In addition, calpain inhibitors significantly reduced the acrosome reaction and penetration.The results suggest that μ-calpain in human sperm plays an important role for the fertilization.

我们评估了μ-Calain在人类精子受精中的作用。在知情同意的情况下，手动收集来自健康捐赠者的精子，液化并经过Percoll梯度离心法。分别在不同浓度孕酮(P)和低氧条件下(H)处理后，进行免疫染色、SDS-PAGE和蛋白质印迹分析。用Fura 2-AM荧光显微镜(ARGUS-50CA)系统观察精子内钙离子浓度的升高。通过金黄地鼠穿透试验、哈密尔顿动力分析仪、三倍缎法和使用Calain抑制剂的Acrobeds试验，推测Calain在受精过程中的作用。用抗μ-Calain的抗体观察人类精子的顶体、赤道段、头段、颈段或颈部和尾段的免疫染色。抗μ原-钙蛋白酶抗体阳性的细胞多为顶体型。Western印迹分析显示，P或H处理均可引起μ-calain的浓度依赖或时间依赖性激活，此外，calain抑制剂显著降低顶体反应和穿透能力。结果表明，人精子中的μ-calain在受精过程中起着重要作用。