Lewis S E, Donnelly E T, Sterling E S, Kennedy M S, Thompson W, Chakravarthy U
Department of Obstetrics and Gynaecology, Queen's University of Belfast, UK.
Mol Hum Reprod. 1996 Nov;2(11):873-8. doi: 10.1093/molehr/2.11.873.
The aim of this study was to investigate the presence of nitric oxide synthase (NOS) and the production of nitric oxide (NO) by human spermatozoa. Immunoreactivity was examined using a polyclonal antibody raised against porcine cerebellar nitric oxide synthase and monoclonal endothelial (eNOS) and brain (bNOS) antibodies. Using each antibody, NOS was observed localized in the head and midpiece regions of the spermatozoon. Immunofluorescence observed for eNOS and bNOS was more intense in normozoospermic samples. Sperm motility was assessed by computer-assisted semen analysis (CASA) in the presence and absence of NG-nitro-L-arginine methyl ester (L-NAME; 10(-5)M), and NO synthesis inhibitor or tumour necrosis factor (TNF)-alpha (20 IU/ml), a superoxide inducer. In the presence of L-NAME, percentage progressive motility, average path velocity (VAP), straight line velocity (VSL) and curvilinear velocity (VCL) were significantly reduced after 30 min. Sperm viability was not decreased by TNF alpha or L-NAME. The accumulation of nitrite (the stable end-product of the NOS/NO pathway) by spermatozoa was measured using the Griess reaction. After 8 h, nitrite concentrations were lower in asthenozoospermic compared to normozoospermic samples. In the presence of TNF alpha, nitrite accumulation was significantly reduced in normozoospermic samples. We conclude that NOS is present in human spermatozoa and that eNOS and bNOS are abundant in normozoospermic samples. Nitric oxide (at endogenous concentrations) appears to be necessary for adequate sperm motility.
本研究的目的是调查人类精子中一氧化氮合酶（NOS）的存在情况以及一氧化氮（NO）的产生。使用针对猪小脑一氧化氮合酶产生的多克隆抗体以及单克隆内皮型（eNOS）和脑型（bNOS）抗体检测免疫反应性。使用每种抗体，观察到NOS定位于精子的头部和中段区域。在正常精子样本中，观察到的eNOS和bNOS的免疫荧光更强。通过计算机辅助精液分析（CASA）在存在和不存在NG-硝基-L-精氨酸甲酯（L-NAME；10⁻⁵M）（一种NO合成抑制剂）或肿瘤坏死因子（TNF）-α（20 IU/ml）（一种超氧化物诱导剂）的情况下评估精子活力。在存在L-NAME的情况下，30分钟后进行性运动百分比、平均路径速度（VAP）、直线速度（VSL）和曲线速度（VCL）显著降低。TNF-α或L-NAME未降低精子活力。使用格里斯反应测量精子中亚硝酸盐（NOS/NO途径的稳定终产物）的积累。8小时后，与正常精子样本相比，弱精子症样本中的亚硝酸盐浓度较低。在存在TNF-α的情况下，正常精子样本中的亚硝酸盐积累显著减少。我们得出结论，人类精子中存在NOS，并且在正常精子样本中eNOS和bNOS含量丰富。一氧化氮（内源性浓度）似乎是精子充分运动所必需的。
