人脐带静脉血管内皮细胞的体外培养及其凋亡的研究

CELL CULTURE OF HUMAN UMBILICAL VEIN ENDOTHELIAL CELLS IN VITRO AND STUDY ON APOPTOSIS OF THIS TYPE OF CELLS

本文从提取生长因子入手 ,建立人脐带静脉血管内皮细胞系 ,用电镜进行了细胞鉴定 .然后用去除生长因子 (FGF和胎牛血清 )的方法诱导该细胞凋亡 ,利用荧光显微技术、DNA凝胶电泳、电镜技术和流式细胞分光光度技术 ,研究了血管内皮细胞凋亡过程中超微结构和细胞周期的变化 .发现去除生长因子 3h后 ,在细胞发生明显的DNA片段化和形成凋亡小体的同时 ,细胞核与细胞质的超微结构发生了明显的变化 ,S期细胞显著减少 ,G1期无明显变化 ,G2细胞显著增多 ,说明用去除生长因子的方法诱导血管内皮细胞凋亡时 ,细胞从G2期脱离细胞周期进入凋亡程序 .本文的人血管内皮细胞在体外的成功培养和对该细胞凋亡的研究对深入研究其凋亡的分子调控机制具有重要的参考价值 .

Human umbilical vein endothelial cells (HUVEC) were cultured in vitro and identified by observing the Weibel Palade body (W P body) which only exists in endothelial cells with electron microscope. HUVEC apoptosis, which induced by deprivation of growth factors (fibroblast growth factor and serum), was investigated by the methods of fluorescence microscopy, DNA electrophoresis, electron microscopy and flow cytometry. We found that, companing with nuclear DNA fragmentation, the ultrastructures of nucleus and cytoplasm dramatically changed. We also found that the number of cells in S phase declined dramatically while the number of cells in G2 phase increased during HUVEC apoptosis, which suggest that the cells which deprived of growth factors enter apoptosis from G2 phase of cell cycle. These researches give us basis to study the molecular mechanism of HUVEC apoptosis in the immediate future.