JC-1单标法流式细胞术检测精子线粒体膜电位的研究

Evaluation of Sperm Mitochondrial Membrane Potential by JC-1 Fluorescent Staining and Flow Cytometry

目的:探讨应用荧光染料JC-1单色标记法进行流式细胞术检测精子线粒体膜电位(MMP)的可行性及其临床意义。方法:收集63例男性精液标本,分为生育组(n=31)和不育组(n=32)。通过计算机辅助精液分析系统进行精液常规分析。精液标本洗涤处理后用JC-1染色后上流式细胞仪分析,用发橙红色荧光精子百分率(JC-1+%)表示MMP正常精子的比例。结果:生育组精子JC-1+%为(75.89±15.69)%,显著高于不育组[(54.04±22.21)%,P=0.000]。63例标本中,JC-1+%与精子活动率呈显著正相关(r=0.610,P=0.000),与(a+b)级精子百分率呈显著正相关(r=0.614,P=0.000),与d级精子百分率呈显著负相关(r=-0.504,P=0.000)。JC-1+%与已建立的罗丹明/碘化吡啶双染法检测结果(Rh123+/PI-%)呈显著正相关(r=0.938,P=0.000)。结论:应用流式细胞术JC-1单标法检测精子MMP具有可行性,精子JC-1+%可作为男性不育的辅助诊断指标。

Objective： To investigate the feasibility and clinical significance of detecting sperm mitochondrial membrane potential （MMP） by JC-1 fluorescent staining and flow cytometry, and to explore the relationship between the results of JC-1 staining and seminal parameters. Methods： Sixty-three semen samples were divided into a fertile （n=31） and an infertile group （ n = 32 ） andunderwent computer-assisted semen analysis （CASA）. All the samples were washed, followed by JC-1 staining and evaluation of sperm MMP by flow eytometry. The percentage of normal sperm MMP was indicated as the percentage of sperm emitting orange-red fluorescence （JC-1^＋ % ）. Results： The JC-1^＋ % was significantly higher in the fertile group than in the infertile one （ [ 75.89 ± 15.69 ] % vs [54.04 ± 22.21 ] %, P = 0.000）, correlated positively with sperm motility （ r = 0.610, P = 0.00（3 ） and the percentage of grade a ＋ b sperm （r = 0.614, P = 0.000） and negatively with grade d sperm （r = -0.504, P = 0.000）. There was a significant positive correlation between the results of JC-1 staining （ JC-1^＋ % ） and that of Rh123/PI dual fluorescent staining （ Rh123^＋ / PI^- % ） （ r = 0.938, P =0.000）. Conclusion： JC-1 staining and flow cytometry could readily and quicldy detect sperm MMP and the sperm JC-1^＋% could be an auxiliary marker for the diagnosis of male infertility. Nail J Androl, 2008, 14（2） ：135-138