【摘要】 目的:建立一种重复性好、传代次数多的人肾小球系膜细胞培养方法。方法:取肾脏肿瘤手术切除后远离患病部位的正常肾脏组织、水囊引产的胎儿肾,三层筛网滤过分离,取最下层筛网上组织,用胶原酶消化,培养正常人肾小球系膜细胞。同时采用形态学观察和免疫荧光技术,对鼠抗人结蛋白,鼠抗人角蛋白进行鉴定,采用MTT、流式细胞仪技术对不同来源肾脏的系膜细胞生长状况进行比较。结果:形态学、免疫荧光法鉴定表明培养细胞为肾小球系膜细胞,胎肾原代培养后8~10 d即可传代,而成年肾脏原代培养后需15~16 d方可传代。成年肾脏培养细胞至第4代细胞生长缓慢,贴壁能力减弱,出现死亡。而胎肾培养细胞则可继续培养至第7~10代。结论:三层滤网法分离人肾小球系膜细胞,方法简单、高效,培养的原代肾小球系膜细胞符合肾小球系膜细胞的生物学特性,且进一步证明胚胎组织比成年个体肾脏原代细胞培养容易成功。更多还原

【Abstract】 Objective: To isolate and identify the glomerular mesangial cells in vitro.Methods: Kidneys were isolated from the normal kidney tissues which were separated from kidney tumor patients and fetal kidneys after induction of labor with water bag,then glomerular mesangial cells were separated by using three-layer micropore fillers and they were subsequently characterized by a morphology method and an immunohistochemical method using flourescein isothiocyanate staining as a label for keratin and desmin.MTT and flow cytometry technology were used to determine the growth of mesangial cells.Results: Glomerular mesangial cells were characterized by their morphology and immunohistochemical character,and those cultured from fetal kidneys can be sub-cultured after 8 to 10 days and 7 to 10 passages in culture while from adult kidneys they can only be sub-cultured after 15 to 16 days and the fourth passage cells presented a poor growth mode.Conclusion: The isolated and cultured cells in vitro are glomerular mesangial cells and they are easier to retain in young kidneys than in adult kidneys.更多还原