以鼠尾胶为贴黏剂的小鼠淋巴管内皮细胞培养体系的建立.doc

1 以鼠尾胶为贴黏剂的小鼠淋巴管内皮细胞培养体系的建立作者:章必成,王俊,赵勇,高建飞,郭燕,陈正堂【关键词】鼠尾胶;贴黏剂;内皮, 淋巴管;细胞培养 Establishment of culture system for mouse lymphatic endothelial cells adhered to rattail collagen 【 Abstract 】 AIM: To establish a convenient, cheap and stable culture system for mouse lymphatic endothelial cells (LEC). METHODS: Mouse lymphangiomas in abdominal cavity were induced by plete Freunds adjuvant, then disrupted and digested to obtain LEC, which were cultured in the flask or plate previously coated with rattail collagen. Expressions of LEC specific markers, VEGFR3 and LYVE1, were identified by immunofluorescence. The proliferation activity of LEC was evaluated by 3HTdR incorporation efficiency and cell doubling time. The capability of LEC to form lymphatic vessellike structures was assessed by the in vitro 2 lymphatic vessel formation assay. RESULTS: Mouse lymphangiomas in abdominal cavity were induced essfully by plete Freunds adjuvant, and more than 98% living LEC were harvested. The expressions of VEGFR3 and LYVE1 were positive in LEC. In the rattail collagen coated flask or plate, LEC grew well, with 3HTdR incorporation efficiency increasing obviously and cell doubling time bEing ( ± ) h. In the gel formed by rattail collagen, LEC could form lymphatic vessellike structures. CONCLUSION: Mouse LEC culture system using rattail collagen as adhesive isa convenient, cheap and stable culture system. 【 Keywords 】 rattail collagen; adhesive; endothelium, lymphatic; cell culture 【摘要】目的: 建立一个简便、廉价和稳定的小鼠淋巴管内皮细胞( LEC )培养体系. 方法: 应用不完全弗氏佐剂诱导小鼠腹腔淋巴管瘤形成,消化法分离获得 LEC , 置于自制的鼠尾胶包被的培养瓶(板)中培养. 以免疫荧光化学法检测 LEC 特异性标记物 VEGFR3 和 LYVE1 的表达, 以 3HTd R 掺入率测定和细胞倍增时间检测 LE C 的增殖活力, 以淋巴管形成试验判定 LEC 能否形成淋巴管样结构. 结果: 3 不完全弗氏佐剂能诱导小鼠腹腔淋巴管瘤形成, 所获 LEC 活细胞数大于 98%. 免疫荧光化学检测表明, LEC 表达 VEGFR3 和 LYVE1. 在鼠尾胶包被的培养瓶(板)中, LE C 生长状况良好, 3HTdR 掺入率明显增加,倍增时间为( ± )h ;在鼠尾胶凝胶中, LEC 能形成淋巴管样结构. 结论:鼠尾胶为贴黏剂的体系是一种简便、廉价和稳定的小鼠 LEC 培养体系.【关键词】鼠尾胶; 贴黏剂; 内皮, 淋巴管; 细胞培养 0 引言淋巴管内皮细胞( lymphatic endothelial cells, LEC )是构成淋巴管壁的主要结构, 参与维