摘要
目的探讨人脂肪干细胞(ASCs)多能性分子Oct4和Sox2的表达情况。方法利用胶原酶将人脂肪组织消化后,分离培养得到ASCs,绘制细胞生长曲线;应用流式细胞仪检测不同代数的P2、P3、P5的ASCs的表面标志物;应用诱导分化培养基诱导ASCs朝成脂、成软骨和成骨方向分化,分化后分别行油红O、阿尔辛蓝和茜素红特殊染色鉴定;应用免疫荧光技术分析P1、P2、P5、P6细胞中Oct4和Sox2的表达情况。结果原代及传代后各代ASCs呈成纤维样贴壁生长,形态均一,细胞群体倍增时间为31 h。各代ASCs均表达CD29、CD44、CD90(>90%)和CD105(>50%),不表达CD34和CD45(<1%);成脂、成软骨和成骨诱导后染色鉴定结果均为阳性;Oct4在P1、P2细胞中的表达位于细胞核内,而在P5、P6细胞的表达则位于胞质;Sox2在P1、P2细胞的胞质中表达,而在P5、P6细胞的胞质中几乎不表达。结论 ASCs具有间充质来源干细胞的一些共同特性及间充质谱系的多向分化潜能。多能性分子Oct4和Sox2在不同代数ASCs中的表达量及表达位置有一定差异,可能反应了各代ASCs分化潜能有所不同。
Obejective To study and analyze the expression features of pluripotency markers Oct4 and Sox2 in adiposederived stem cells（ASCs）. Methods After digesting subcutaneous fat tissues with collagenase P, cells were cultured with medium containing DMEM and 10% FBS, cell growth curve were drawn to evaluate the population doubling times, P2, P3, P5 of ASCs were identified by Flow Cytometry ; when induced into adipocytic, chondrogenic, Osteoblastic lineage, Oil red O, Alclan Blue and Alizarin Red staining were conducted respectively. The expression of Oct4 and Sox2 in P1, P2, P5, P6 ASCs were detected by immunofluorescence. Results ASCs were spindle-like and passaged stably in vitro, population doubling time were 31 h. The phenotype of ASCs were CD29 ＋ , CD90＋ , CD44 ＋ （ 〉 95% ）, CD105 ＋ （ 〉 50% ）, CD34 -, CD45 - （ 〈 1% ）. After induction, ASCs could differentiate into adipocytes, chondrogenic cells and osteoblast. The P1 and P2 ASCs expressed Oct4 in the nucleus,but the expression of Oct4 in P5 ,P6 ASCs were located in the cytoplasm;Sox2 in P1 ,P2 ASCs were expressed in the cytoplasm, but hardly expressed in P5 and P6 ASCs. Conclusion ASCs have some common features of mesenchymal stem cells and the potential of multi-leneage differention. Different passaged ASCs express pluripotent markers Oct4 and Sox2 in different positions, reflecting that ASCs after multiple times passaging have different differentiation potential.
出处
《中国输血杂志》
CAS
CSCD
北大核心
2014年第7期692-696,673,共5页
Chinese Journal of Blood Transfusion
