悬浮组织块法分离培养SD大鼠脂肪干细胞的实验研究  

An experimental study of isolation of SD rat adipose-derived stem cells using suspended explant culture method

作　　者：刘琴[1] 王丽平[1] 陈芳[1] 李晓明[1] 朱以良[1] 张宜[1] 

机构地区：[1]中国人民解放军武汉总医院医学实验科,武汉430070

出　　处：《中国免疫学杂志》2017年第8期1197-1200,共4页Chinese Journal of Immunology

摘　　要：目的:建立SD大鼠脂肪干细胞悬浮组织块分离培养法。方法:取正常SD大鼠股沟处脂肪组织,用悬浮组织块法和组织块贴壁法分离培养SD大鼠脂肪干细胞,观察细胞的生长状况及形态特征。取第3代对数期细胞,CCK-8法绘制其生长曲线;免疫荧光法检测其表面抗原CD29、CD44、CD45的表达情况;分别用成脂、成骨诱导培养液进行诱导分化,油红O、茜素红染色定性鉴定。结果:两种方法体外培养得到的SD大鼠脂肪干细胞形态为梭形纤维样,生长力旺盛,生长曲线为典型的"S"型;免疫荧光法检测显示第3代细胞强表达CD29、CD44,CD45表达阴性;细胞成脂诱导培养14 d后,油红O染色为阳性;成骨诱导培养14 d后,茜素红染色为阳性。结论:悬浮组织块法可以分离培养出SD大鼠脂肪干细胞,方法简便,为体外分离培养SD大鼠脂肪干细胞提供了一种新的方法。Objective：To isolate SD rat adipose-derived stem cells（ASCs）by suspended explant culture method . Methods： The healthy rat inguinal fat pads were obtained. The SD rat ASCs were isolated by suspended explant culture method and explant adherent culture method . The growth status and morphology were observed. The growth curve and cell surface markers CD29, CD 44, CD45 of the 3rd passage cells were analyzed respectively by CCK- 8, immunocytochemistry; the 3rd passage cells were induced ndividually by adipogenic differentiation medium and osteogenic differentiation medium,and cells were examined by oil red 0 staining and alizarin red staining. Results ： The SD rat ASCs obtained by the two methods exhibited a spindle-shaped appearance and could apidly expand. The cell growth curves were typical of ” S” type. Immunocytochemistry analysis revealed that the third passage of SD rat ASCs were positive for CD29, CD 44,but were negative for CD 45. SD rat ASCs were positive for oil red 0 staining at 14 days after adipogenic induction,and positive for alizarin red staining at 14 days after osteogenic induction. Conclusion ： Isolation of SD rat ASCs by suspended explant culture method is successfully established. The method is simple. It provides a new method for the isolation of SD at ASCs in vitro .

关 键 词：悬浮组织块法 SD大鼠脂肪干细胞 分离 培养 

分 类 号：R392.33[医药卫生—免疫学]