肌源性干细胞分离培养及诱导分化为平滑肌细胞的研究  

Isolation and characterization of muscle-derived stem cells and their differentiation into smooth muscle cells

作　　者：张金明[1] 何涛[1] 黄红军[1] 

机构地区：[1]中山大学附属第二医院整形外科,广州510120

出　　处：《中华实验外科杂志》2006年第8期1003-1004,F0004,共3页Chinese Journal of Experimental Surgery

摘　　要：目的探讨兔肌源性干细胞分离、鉴定及诱导分化为平滑肌细胞的方法。方法取1.5个月龄新西兰兔大腿肌肉,采用Preplate技术分离培养肌源性细胞,并进行流式细胞仪(FCM)、免疫细胞化学、Western blot等确定细胞表型。采用添加血管内皮生长因子(VEGF)和与兔阴茎海绵体平滑肌细胞(CCSMC)共培养的方法诱导分离而得的细胞分化为平滑肌细胞并运用免疫细胞化学和Western blot检测特异性α-平滑肌肌动蛋白(α-SMA)的表达。结果经过连续6d的差速贴壁分离得到小圆形或短梭形的小体积细胞,FCM结果显示这些细胞＞80%为desmin+,＞70%为bcl-2^+,＞95%为CD45^-。免疫细胞化学定性结果显示desmin+。高汇合度(＞50%)或低血清培养时容易融合形成肌管或肌细胞核链。诱导处理后分离的细胞表达α-SMA。结论采用Preplate技术能很好地分离出肌源性干细胞,并能在体外诱导分化为平滑肌细胞,为组织工程提供种子细胞。Objective To investigate the methods for the isolation and characterization of musclederived stem cells （MDSCs） of rabbits and their differentiation into smooth muscle cells. Methods MDSCs were isolated from thigh muscle biopsies of normal 1.5-month-old New Zealand rabbits using Preplating technique. The isolated cells were characterized by flow cytometry （FCM）, immunocytochemistry and Western blotting, and were induced to differentiate into smooth muscle lineage by adding vascular endothelial growth factor （VEGF） and coculturing with corpus cavernosal smooth muscle cells （CCSMCs）. Immunocytochemistry and Western blotting were performed to detect the expression of specific α-SMA. Results The isolated cells were small round or short spindle-shaped, and significantly smaller than skeletal muscle cells. FCM revealed that these cells were 〉80% desmin＋, 〉70% bcl-2 ＋, 〉95% CD45-. Immunocytochemistry showed that these cells were desmin ＋ . Cells grown at high confluence or cultured with low concentration of serum tended to fuse into myotubes. After induction, the isolated cells expressed the specific α-SMA. Conclusion MDSCs could be effectively isolated by using Preplating technique, and they could be induced into smooth muscle cells in vitro, thus providing sufficient amount of seed cells to tissue engineering.

关 键 词：干细胞 平滑肌细胞 分化 共培养 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]