不同质量精子线粒体DNA拷贝数及完整性的比较研究

目的探讨精子线粒体DNA（mtDNA）拷贝数与完整性与男性不育的相关性。方法选取正常精液样本18例,异常精液样本34例,采用Percoll液密度梯度离心将样本分为30%层和80%层精子,并采用实时荧光定量PCR技术测定精子mtDNA拷贝数及长链PCR技术测定mtDNA完整性。结果在80%层精子中,少精子症组mtDNA拷贝数（1.52±0.68）个]和少弱精子症组（1.93±0.65）个]与正常组（0.80±0.45）个]比较,差异有统计学意义（P〈0.01）;少弱精子症组mtDNA完整性（0.93±0.27）与正常组（2.70±1.74）、弱精子症组（3.24±2.18）及少精子症组（1.76±0.76）比较,差异有统计学意义（P〈0.01）。在30%层精子中,少精子症组mtDNA拷贝数（13.33±9.96）和弱精子症组（1.92±1.29）与正常组（4.85±3.00）比较,差异有统计学意义（P〈0.05）。结论质量差的精子中存在mtDNA拷贝数增加及完整性减弱,mtDNA的检测可能成为预测精子质量的一个指标。

A comparative study on mitochondrial DNA copy number and integrity in different quality sperm

Objective To detect the copy number and integrity of sperm mitochondrial DNA （mtDNA）, and to investigate the relationship between sperm mtDNA and mate infertility. Methods Using density gradient separation, we selected motile sperm from 18 normal and 34 abnormal sperm samples. Quantitative long polymerase chain reaction （PCR） and realtime PCR were used to measure the mtDNA integrity and copy number in spermatozoa. Results The mtDNA copy numbers were increased in the 30% density gradient layers than the 80% layers in 52 sperm samples, the differences was statistically significant （P〈0.05）. In the 80% layers, a significant increasing in mtDNA copy number was observed in oligozoospermia （1.52±0. 68） and oligoasthenozoospermia （1.93 ±0.65 ） compared to the patients with normal semen parameters （0.80±0. 45, P〈0. 01）. and lower mtDNA integrity was observed in oligoasthenozoospermia （0.93±0.27） compared to the patients with normal semen （2. 70±1.74）, asthenozoospermia （3.24±2.18） and oligozoospermia （1.76±0. 76, P〈0.01）. In the 30% layers, the mtDNA copy number from oligozoospermia （13.33±9.96） and asthenozoospermia （1.92±1.29） were significantly higher than normal semen samples （4.85±3.00, P〈0.05）. Conclusions A decreasing mtDNA integrity and increasing copy number were observed in poor sperm samples. We suggest that mtDNA quantity and quality may serve as a useful diagnostic markers of sperm quality.