Piasecka Małgorzata, Kawiak Jerzy
Department of Histology and Embryology, Pomeranian Medical University, Szczecin, Poland.
Folia Histochem Cytobiol. 2003;41(3):125-39.
Studies were performed on ejaculated human spermatozoa (32 subjects with normal sperm motility and 25 subjects with low sperm motility). Morphology of sperm midpiece was evaluated in light, fluorescent and transmission or scanning electron microscope. Changes in mitochondrial membrane potential (delta(psi)m) and mass of mitochondria were analysed by flow cytometry using mitochondrial specific probes JC-1 and Mito Tracker Green FM. Moreover, oxidoreductive capability of sperm mitochondria was assessed using cytochemical reaction for NADH-dependent dehydrogenases. In flow cytometry analysis of JC-1-stained spermatozoa, two asthenozoospermic subpopulations were distinguished: patients with a high percentage (76 +/- 11%, 13 subjects) and patients with a low percentage (29 +/- 14%,12 subjects) of spermatozoa with functional-polarized mitochondria with high delta(psi)m. Our microscopic investigations of spermatozoa of seven asthenozoospermic patients reveal that the deformed and unusually thickened sperm midpieces (50-70% of cells), occasionally with persistent cytoplasmic droplet, contain supernumerary mitochondria with normal substructure, full oxidoreductive capability and high delta(psi)m. The midpiece deformations cause nonprogressive movement or immotility. They can also appear in smaller number of spermatozoa (5-35% of cells) in patients with normal sperm motility. Moreover, in three cases of asthenozoospermia midpiece malformations were accompanied by abnormal morphology of outer dense fibers and axoneme. The cytochemical, fluorescence and SEM studies showed the absence of midpieces in many (60-80%) spermatozoa in some other cases of asthenozoospermia. The morphological observations corresponded with flow cytometry analysis of Mito Tracker Green FM-stained spermatozoa. Our results suggest that in some cases of asthenozoospermia the sperm mitochondria can be functionally active and display high delta(psi)m in large number of cells. The results may suggest that asthenozoospermia does not necessarily result from energetic disturbances of sperm mitochondria. The low sperm motility may be associated with deformations of the mitochondrial sheath containing functional mitochondria. The combination of fluorescence microscopy and flow cytometry with electron microscopic investigations is a sensitive, precise and comprehensive examination which helps discover sperm abnormalities responsible for asthenozoospermia.
对射出的人类精子进行了研究（32名精子活力正常的受试者和25名精子活力低的受试者）。在光学显微镜、荧光显微镜以及透射或扫描电子显微镜下评估精子中段的形态。使用线粒体特异性探针JC-1和线粒体示踪绿荧光染料（Mito Tracker Green FM）通过流式细胞术分析线粒体膜电位（Δψm）和线粒体质量的变化。此外，利用针对依赖NADH的脱氢酶的细胞化学反应评估精子线粒体的氧化还原能力。在对JC-1染色精子的流式细胞术分析中，区分出两个弱精子症亚群：线粒体功能极化且Δψm高的精子比例高的患者（76±11%，13名受试者）和该比例低的患者（2９±14%，12名受试者）。我们对7名弱精子症患者精子的显微镜检查显示，变形且异常增厚的精子中段（占细胞的50 - 70%），偶尔伴有持续存在的细胞质滴，含有超数目的线粒体，其亚结构正常，具有完全的氧化还原能力且Δψm高。中段变形导致精子非进行性运动或不动。它们也可能出现在精子活力正常的患者数量较少的精子中（占细胞的5 - 35%）。此外，在3例弱精子症中，中段畸形伴有外致密纤维和轴丝的形态异常。细胞化学、荧光和扫描电镜研究显示，在其他一些弱精子症病例中，许多（60 - 80%）精子没有中段。形态学观察结果与线粒体示踪绿荧光染料染色精子的流式细胞术分析结果一致。我们的结果表明，在某些弱精子症病例中，精子线粒体在大量细胞中可能功能活跃并显示出高Δψm。这些结果可能表明，弱精子症不一定是由精子线粒体的能量紊乱引起的。精子活力低可能与含有功能正常线粒体的线粒体鞘变形有关。荧光显微镜、流式细胞术与电子显微镜检查相结合是一种灵敏、精确且全面的检查方法，有助于发现导致弱精子症的精子异常情况。
