人精子过氧化氧化还原蛋白2表达与精子DNA损伤的相关性

Correlation between expression of Prdx2 protein in human mature spermatozoa and sperm DNA damage

目的精子DNA损伤的分子机制尚不明确。文中旨在通过检测精子DNA正常者和精子DNA损伤患者过氧化氧化还原蛋白2(Prdx2)表达差异,分析Prdx2表达与精子DNA损伤指数(DFI)的相关性。方法收集2017年7月至2018年7月南京军区南京总医院生殖医学中心门诊67份男性不孕患者的精液标本。按DFI值分为3组:DFI正常组(DFI≤15%,n=38)、轻度损伤组(15%<DFI<30%,n=20)和重度损伤组(DFI≥30%,n=9)。通过计算机辅助精子分析系统(CASA)进行精液检测,获得精液常规参数:前向运动精子百分率(PR)、精子密度及精子总活率。提取精子中的总蛋白,Western blot检测Prdx2的表达,免疫荧光检测精子中Prdx2的定位。并分析Prdx2表达与DFI、精液常规参数的相关性。结果与DFI正常组Prdx2相对表达水平(9.06±1.80)比较,轻度损伤组(6.32±1.89)和重度损伤组(2.87±0.83)分别下降了30.2%、68.2%,差异有统计学意义(P<0.01)。免疫荧光结果显示,Prdx2主要定位于人精子中段线粒体以及精子头部核区域;重度损伤组和轻度损伤组Prdx2表达较DFI正常组均降低,其中重度损伤组的Prdx2阳性反应明显减弱。Prdx2蛋白表达与DFI呈负相关(r=-0.478,P<0.01),与PR、精子总活率呈正相关(r=0.135、0.128,P<0.01)。结论 Prdx2对精子DNA损伤患者中DNA完整性的影响可能通过维持细胞内氧化还原平衡,参与保护精子DNA免受损伤,同时影响精子运动能力来实现。

Objective The molecular mechanisms underlying sperm DNA damage remain uncertain.This study aimed to determine the expression difference of Prdx2 protein between normal and damaged sperm DNA,and analyze the correlation between expression of Prdx2 protein in human mature spermatozoa and sperm DNA damage.Methods Semen specimens of sixty-seven male infertility patients were collected.They were separated in three groups according to the sperm DNA damage levels:normal group,slight damage group,and severe damage group.Total protein of human of mature spermatozoa was extracted.The expression of Prdx2 protein was detected by western blotting.The Localization of Prdx2 in spermatozoa was detected by immunofluorescence.Results Comparing the expression of Prdx2 protein with group normal(9.06±1.80),group slight damage(6.32±1.89)and group severe damage(2.87±0.83)decreased by 30.2%and 68.2%respectively(P<0.01).Immunofluorescence showed that Prdx2 was mainly distributed in the mitochondria and nuclear of spermatozoa.Immunofluorescence results showed that Prdx2 was mainly located in the mitochondria of human sperm midsection and the nucleus region of the sperm head.The expression of Prdx2 in both group slight damage and group severe damage was lower than that in the group normal.The positive Prdx2 response was significantly reduced in group severe damage.The protein expression of Prdx2 was negatively correlated with DFI(r=-0.478,P<0.01),and was positively correlated with progressive motility and total motility(r=0.135、0.128,P<0.01).Conclusion Human sperm Prdx2 protein expression may be involved in protecting sperm DNA and affecting sperm motility by maintaining intracellular redox balance.