摘要
目的探讨皮肤源祖细胞的体外培养、鉴定及定向诱导成脂、成骨的方法,为组织工程提供较理想的种子细胞。方法出生1～3 d的SD大鼠幼鼠皮肤,以含表皮生长因子和成纤维细胞生长因子的培养基进行培养。观察细胞生长情况,描绘生长曲线;免疫荧光鉴定细胞表达Nestin和Fibronectin情况;将第3代细胞,分别用成脂和成骨诱导液培养14 d,以油红O染色、茜素红染色和免疫荧光检测皮肤源祖细胞诱导成脂、成骨情况。结果细胞呈悬浮生长,迅速增殖形成克隆球团;细胞免疫荧光表达Nestin和Fibronectin;成脂诱导14 d后,细胞内大量致密颗粒形成,油红O染色可见红色脂滴;成骨诱导14 d后,茜素红染色可见暗红色钙盐沉积,骨桥蛋白表达阳性显示成骨细胞形成。结论皮肤源祖细胞具有干细胞的特性,能分化为成骨细胞和脂肪细胞。
To explore the methods to culture and isolate rats skin-derived precursors （SKPs） and direct them to differentiate adipocyte and osteoblast, so as to provide ideal seeded cells for tissue engineering. Methods Skin from juvenile （1-3 postnatal days） Spraque-Dawley （SD） rats was cultured with epidermal growth factor （EGF） and basic fibroblast growth factor （bFGF）. The growth of SKPs was observed and growth curves were drawn. Immunofluorescent techniques were used to detect Nestin and Fibronectin marker expression. SKPs were cultured for 14 days in revulsivum for adipocyte and osteoblast respectively. Then oil red O staining, Alizarin bordeaux staining and immunofluorescence were used to test adipocytes or osteoblasts. Results SKPs proliferated quickly and formed floated clone pellet. After induction in adipocyte revulsivum for 14 days, dense granules were found in cytolymph and lipid droplets were seen after oil red O staining. After induction in osteoblast revulsivum for 14 days, madder red calcium salts were shown in medium, and immunofluorescent expression of osteopontin revealed the formation of osteoblasts. Conclusion SKPs can be generated from juvenile rat skin and induced into adipocytes and osteoblasts in vitro.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2007年第10期1181-1182,1281,共3页
Chinese Journal of Experimental Surgery
基金
广州市科学技术局资助项目(2006Z3-E5131)
广东省科技计划项目基金(2004B35001002)
广东省自然博士启动基金(06300811)
关键词
皮肤
细胞分化
成脂细胞
成骨细胞
Skin
Cell differentiation
Adipogenic cells
Osteogenic cells
