Hashemitabar Mahmoud, Sabbagh Susan, Orazizadeh Mahmoud, Ghadiri Atta, Bahmanzadeh Maryam
Cellular and Molecular Research Center, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.
J Assist Reprod Genet. 2015 Jun;32(6):853-63. doi: 10.1007/s10815-015-0465-7. Epub 2015 Apr 1.
Asthenozoospermia is a common cause of human male infertility characterized by reduced sperm motility. The molecular mechanism that impairs sperm motility is not fully understood. This study proposed to identify novel biomarkers by focusing on sperm tail proteomic analysis of asthenozoospermic patients.
Sperm were isolated from normozoospermic and asthenozoospermic semen samples. Tail fractions were obtained by sonication followed by Percoll gradient. The proteins were extracted by solubilization and subjected to two-dimensional gel electrophoresis (2-DE); then, the spots were analyzed using Image Master 2D Platinum software. The significantly increased/decreased amounts of proteins in the two groups were exploited by matrix-assisted laser desorption-ionization time-of-flight/time-of-flight (MALDI-TOF-TOF) mass spectrometry.
Three hundred ninety protein spots were detected in both groups. Twenty-one protein spots that had significantly altered amounts (p < 0.05) were excised and exploited using MALDI-TOF-TOF mass spectrometry. They led to the identification of the following 14 unique proteins: Tubulin beta 2B; glutathione S-transferase Mu 3; keratin, type II cytoskeletal 1; outer dense fiber protein 2; voltage-dependent anion-selective channel protein 2; A-kinase anchor protein 4; cytochrome c oxidase subunit 6B; sperm protein associated with the nucleus on the X chromosome B; phospholipid hydroperoxide glutathione peroxidase-mitochondrial; isoaspartyl peptidase/L-asparaginase; heat shock-related 70 kDa protein 2; stress-70 protein, mitochondrial; glyceraldehyde-3-phosphate dehydrogenase, testis-specific and clusterin.
Fourteen proteins present in different amounts in asthenozoospermic sperm tail samples were identified, four of which are reported here for the first time. These proteins might be used as markers for the better diagnosis of sperm dysfunctions, targets for male contraceptive development, and to predict embryo quality.
弱精子症是男性不育的常见原因，其特征为精子活力降低。目前，损害精子活力的分子机制尚未完全明确。本研究旨在通过聚焦弱精子症患者精子尾部蛋白质组分析来鉴定新的生物标志物。
从正常精子症和弱精子症精液样本中分离精子。通过超声处理和Percoll梯度离心获得尾部片段。蛋白质经溶解提取后进行二维凝胶电泳（2-DE）；然后，使用Image Master 2D Platinum软件分析斑点。通过基质辅助激光解吸电离飞行时间/飞行时间（MALDI-TOF-TOF）质谱法分析两组中蛋白质含量显著增加/减少的情况。
两组均检测到390个蛋白质斑点。切除21个含量有显著变化（p < 0.05）的蛋白质斑点，并使用MALDI-TOF-TOF质谱法进行分析。由此鉴定出以下14种独特蛋白质：微管蛋白β2B；谷胱甘肽S-转移酶Mu 3；细胞角蛋白II型细胞骨架1；外致密纤维蛋白2；电压依赖性阴离子选择性通道蛋白2；A激酶锚定蛋白4；细胞色素c氧化酶亚基6B；与X染色体B上细胞核相关的精子蛋白；磷脂氢过氧化物谷胱甘肽过氧化物酶-线粒体；异天冬氨酰肽酶/L-天冬酰胺酶；热休克相关70 kDa蛋白2；应激70蛋白，线粒体；睾丸特异性甘油醛-3-磷酸脱氢酶和簇集蛋白。
鉴定出14种在弱精子症精子尾部样本中含量不同的蛋白质，其中4种为本研究首次报道。这些蛋白质可作为更好地诊断精子功能障碍的标志物、男性避孕药物开发的靶点以及预测胚胎质量的指标。
