老年大鼠睾丸间质细胞自噬的变化  

Study on autophagy in aged rat Leydig cells

作　　者：李维仁[1,2] 侯垒[2] 崔功静 常智杰[3] 贾金铭[1] 辛钟成[4] 

机构地区：[1]中国中医科学院广安门医院男科,北京100053 [2]北京航天总医院泌尿外科 [3]清华大学医学院生物化学系 [4]北京大学第一医院男科中心

出　　处：《中国男科学杂志》2013年第7期3-7,12,共6页Chinese Journal of Andrology

摘　　要：目的探讨老年大鼠睾丸间质细胞自噬的变化。方法取青年组（3月龄）及老年组（24月龄）清洁级雄性SD大鼠各10只，麻醉后取血清检测总睾酮浓度，取睾丸组织用HE染色观察睾丸组织形态学变化，电镜观察睾丸间质细胞的超微结构改变。通过密度梯度离心分离培养原代睾丸间质细胞，一部分细胞转染GFP-LC3质粒48h后共聚焦显微镜观察GFP颗粒分布情况，另一部分细胞用黄体生成素（LH）刺激后，提取细胞总蛋白，用Westernblot比较青年组和老年组睾丸间质细胞中自噬相关蛋白LC3蛋白表达水平的差异。结果HE染色显示老年大鼠睾丸呈老年退行性改变，电镜下观察到老年大鼠睾丸间质细胞内质网肿胀，自噬结构显著减少（P〈0．01）。老年大鼠血清睾酮水平显著低于青年组（P〈0．05）。原代培养的睾丸间质细胞无论LH刺激与否，老年组大鼠睾丸间质细胞中GFP．LC3荧光颗粒所占面积比均低于青年组（LH刺激与否P〈〈0．01），老年大鼠睾丸间质细胞中自噬相关蛋白LC3．II表达均显著低于青年组（LH刺激与否P〈0．01）。结论睾丸问质细胞的衰老与其自噬水平降低密切相关。Objective To investigate the changes of autophagy in aged rat Leydig cells. Methods Ten young （3 months） and ten aged （24 months old） male SD rats of clean grade were utilized as the research model. After anesthesia, blood was collected by vena cava puncture, and the serum total testosterone concentrations were measured by ELISA. Then the histological changes in testis and ultrastructure of Leydig cells were observed by immunohistochemistry staining and electron microscopy （EM） respectively. The density gradient centrifugation was used for isolation of primary Leydig cells from rat testes. Some cells were transfected with GFP-LC3 plasmid and cultured for 48 hours, and the fluorescence particles were observed by confocal microscopy to determine their distribution. The other cells were stimulated by luteinizing hormone （LH）, then their LC3 protein expressions were detected by western blot. Results HE staining showed that some degenerative changes occurred in aged rat testes, and EM showed swollen endoplasmic reticulum and decreased autophagic structures （P〈0.01） and more senescent organelles in Leydig cells of aged rats. The serum testosterone level inaged rats was significantly lower than that of young group （P〈0.05）. In primary Leydig cells, with or without LH stimulation, the area ratio of GFP-LC3 fluorescence particles and LC3- II protein expression in aged group were all significantly lower than those of young group （with or without LH stimulation, P〈0.01）. Conclusion Aging is closely related with autophagy depression in aged Leydig cells.

关 键 词：衰老 自噬 莱迪希细胞 

分 类 号：Q492.4[生物学—生理学]