二甲磺基乙烷处理生精小管体外培养模型的建立  

Establishment of culture model of seminiferous tubules treated by ethane dimethanesulfonate in vitro

作　　者：张磊[1,2] 张齐好[3,2] 苏志坚[3,2] 黄亚东[3,2] 

机构地区：[1]广东省生物制品与药物研究所,广州510440 [2]暨南大学医药生物技术研究开发中心,广州510632 [3]暨南大学细胞生物学系,广州510632

出　　处：《上海交通大学学报（医学版）》2015年第3期327-332,共6页Journal of Shanghai Jiao tong University:Medical Science

基　　金：国家自然科学基金(81070477;31000663;31271607);中央高校基本科研业务费专项资金(21611378);广东省高等学校珠江学者岗位计划资助项目(2012)~~

摘　　要：目的建立二甲磺基乙烷(EDS)处理生精小管的体外培养模型。方法雄性SD大鼠于实验前7 d腹腔注射EDS,分离生精小管进行体外培养,分为基础处理组(加入胰岛素-转铁蛋白-亚硒酸钠培养基添加剂ITS,即ITS组)和促黄体素(LH)处理组(加入ITS和LH,即ITS+LH组),采用Ed U标记技术检测睾丸间质干细胞增殖情况,放射免疫分析技术及3β-HSD染色检测睾酮分泌及睾丸间质干细胞的分化情况。取雄性成年小鼠睾丸并分离生精小管,分别以不同浓度的EDS处理,其后再进行分组(ITS组和ITS+LH组)处理连续培养,采用放射免疫法检测培养基上清液中睾酮的分泌情况。结果与ITS组比较,大鼠生精小管ITS+LH组睾丸间质干细胞增殖较为明显。睾酮的分泌呈现与体内睾丸间质细胞发育分化趋势一致;且该模型在LH作用下,睾酮产生量大,持续时间长,表明该模型对LH敏感。1.72 mmol/L EDS可有效杀死小鼠生精小管上的成熟睾丸间质细胞;该处理模型在经过培养后又有睾酮的产生,且ITS+LH组生精小管培养上清液中睾酮含量显著高于ITS组(P<0.01),表明EDS可以杀死成熟睾丸间质细胞,同样该模型也表现出对LH敏感。结论雄性大鼠和小鼠生精小管体外培养模型中,睾丸间质干细胞的发育分化过程与体内睾丸间质干细胞的发育分化相似,该模型是一个研究睾丸间质细胞发育分化的合适模型。Objective To establish the culture model of seminiferous tubules treated by ethane dimethanesulfonate（ EDS） in vitro. Methods Male SD rats were intraperitioneally injected with EDS 7 d before experiment.Seminiferous tubules of rats were removed and cultured in vitro, and divided into the basic treatment group [treated by insulin-transferin-selenium（ ITS）, i. e. the ITS group] and luteinizing hormone（ LH） treatment group（ treated by ITS and LH, i. e. the ITS ＋ LH group）. Ed U staining was adopted to detect the proliferation of stem Leydig cells.Radioimmunoassay and 3β-HSD staining were employed to detect the secretion of testosterone and differentiation of stem Leydig cells. Seminiferous tubules were isolated from testicles of adult male mouse and treated by EDS of different concentrations. Then seminiferous tubules were treated by ITS（ ITS group） and ITS ＋ LH（ ITS ＋ LH group）. The secretion of testosterone of culture supernatant was detected by the radioimmunoassay. Results Compared with the ITS group, the proliferation of stem Leydig cells of seminiferous tubules of the ITS ＋ LH group wasmore significant. The secretion of testosterone was consistent with the development and differentiation of Leydig cells.The production of testosterone of the ITS ＋ LH group was large and the duration was long after being treated by LH,which showed that the culture model of seminiferous tubules was sensitive to LH. EDS of 1. 72 mmol / L effectively killed mature Leydig cells of seminiferous tubules. Testosterone was produced after the model was cultured and the testosterone level of culture supernatant of seminiferous tubules of the ITS ＋ LH group was significantly higher than that of the ITS group（ P 0. 01）, which indicated that EDS could kill mature Leydig cells and the model was also sensitive to LH. Conclusion The development and differentiation processes of stem Leydig cells of the male rat and mouse models and the culture model of seminiferous tubules in vitro are similar. The cultur

关 键 词：生精小管 睾丸间质细胞 二甲磺基乙烷 促黄体素 

分 类 号：Q492.4[生物学—生理学]