组织块贴壁法扩增兔脂肪干细胞  

Amplification of rabbit adipose-derived stem cells using explants culture method

作　　者：刘琴[1] 王丽平[1] 喻晶[1] 陈芳[1] 刁波[1] 张宜[1] 

机构地区：[1]解放军广州军区武汉总医院医学实验科,湖北省武汉市430070

出　　处：《中国组织工程研究》2014年第1期88-93,共6页Chinese Journal of Tissue Engineering Research

摘　　要：背景:研究显示兔脂肪干细胞的体外分离方法大多数为Ⅰ型胶原酶消化法,采用组织块贴壁法扩增兔脂肪干细胞尚不多见。目的:采用组织块贴壁法从兔脂肪组织中分离培养兔脂肪干细胞,并进行成脂、成骨的诱导分化。方法:采用组织块贴壁法分离出兔脂肪干细胞,进行体外培养,观察其形态特征。取对数生长期的第3代细胞,用MTT法绘制其生长曲线;流式细胞仪检测其表面抗原CD29、CD44、CD45的表达情况;分别用成脂和成骨诱导培养液诱导其向脂肪细胞和成骨细胞分化,油红O、茜素红染色定性鉴定。结果与结论:体外培养的兔脂肪干细胞呈梭形纤维样细胞形态,生长力旺盛。流式细胞术分析结果显示,第3代兔脂肪间充质干细胞强表达CD29,CD44,阴性表达CD45。兔脂肪干细胞经成脂诱导14 d后,油红O染色阳性;成骨诱导培养14 d后,茜素红染色阳性。提示组织块贴壁法可以分离培养出兔脂肪干细胞。BACKGROUND：The rabbit adipose-derived stem cells are mostly isolated by type I col agenase digestion, but rarely by explants culture method. OBJECTIVE：To isolate rabbit adipose-derived stem cells for adipogenic and osteogenic differentiation. METHODS：The rabbit adipose-derived stem cells were isolated from rabbit adipose by explants culture method, and cultured in vitro fol owed by morphological observation. The grow curve and cellsurface markers CD29, CD44, CD45 of passage 3 cells were analyzed respectively by 3-（4,5-dimethylthiazol-2-yl）-2, 5-diphenyltetrazolium bromide and flow cytometry;cells from the third passages were induced for adipogenic and osteogenic differentiation by different revulsants, and cells were examined by oil red O staining and alizarin red staining . RESULTS AND CONCLUSION：The rabbit adipose-derived stem cells cultured in vitro exhibited a spindle-shaped appearance and could rapidly expand. Flow cytometry analysis revealed that the third passage of rabbit adipose-derived stem cells was positive for CD29, CD44, but negative for CD45. Rabbit adipose-derived stem cells were positive for oil red O staining at 14 days of adipogenic induction, and positive for alizarin red staining at 14 days of osteogenic induction. In conclusion, we could successful y isolate rabbit adipose-derived stem cells using explants culture method.

关 键 词：干细胞 脂肪干细胞 培养 组织块贴壁培养 细胞鉴定 诱导分化 成脂诱导 成骨诱导 

分 类 号：R394[医药卫生—医学遗传学]