Ⅱ型胶原酶/弹性蛋白酶消化法培养大鼠血管平滑肌细胞

Enzymatic Dispersion Method of Primary Culture of Rat Aortic Vascular Smooth Muscle Cells

为快速、成功培养大鼠胸主动脉血管平滑肌细胞 ,在无菌条件下分离大鼠胸主动脉 ,并剥离外膜及内皮 ,然后用Ⅱ型胶原酶 弹性蛋白酶消化血管中层。 2h左右完全消化成单个细胞时 ,离心收集细胞 ,然后重悬、接种细胞。 2 4h后便可见细胞贴壁 ,4～ 5天便可传代。传至第 3代时 ,用台盼蓝检查细胞存活率为 96 % ,光镜和免疫组织化学鉴定培养细胞 ,培养细胞纯度为 97%。与国内报道的贴块法及消化法相比 ,用酶解法培养血管平滑肌细胞的培养周期缩短 4～ 5天 ,能更好的保护细胞 ,维持细胞形状。

Aim To culture rat vascular smooth muscle cells by novel enzymatic dispersion method. Methods Rat aorta was removed from left subclavian origin to diaphragmatic insertion in sterile conditions. The vascular media was digested,then the cells were centrifuged and plated down in prepared flask. Results The primary vascular smooth muscle cells of rat aorta was performed successfully by using enzymatic dispersion method. The cell viability of third generation is 96% by typan blue,and the cell purity is 97% tested by light microscope and immunocytochemical stains. Conclusions The growth period of enzyme-dispersed vascular smooth muscle cells is shorter by 4～5 days than that of explant method and enzymatic dispersion method reported in domesticity.