牛脂肪间充质干细胞的分离、培养与鉴定

Isolation, cultivation and identification of adipose-derived stem cell in bovines

摘要:

为了给组织工程提供种子细胞，对牛间充质干细胞 (Adipose-derived stem cells，ADSCs) 进行体外分离培养。首先应用胶原酶消化法分离牛ADSCs，进行体外培养、连续传代，并观察细胞的形态变化，通过细胞计数绘制生长曲线，细胞压片进行染色体分析，采用细胞免疫荧光化学方法检测细胞表面标记，利用成骨分化和成脂分化检测其分化能力。结果显示牛ADSCs体外培养时细胞形态呈成纤维细胞样，增殖稳定；Vimentin、CD49d、CD13表达呈阳性，CD34表达呈阴性；成骨诱导条件下的细胞碱性磷酸酶活性高，茜素红染色呈阳性；成脂诱导条件下细胞周围脂滴明显，油红-O染色呈阳性。结果证明牛ADSCs体外生长稳定、增殖速度快、定向分化能力强，简易的体外分离培养及诱导方法为其在组织工程中的应用奠定了基础。

Abstract:

To obtain bovine adipose-derived stem cells (ADSCs), bovine ADSCs were digested in collagenase type I solution. The growth curve of ADSCs was checked by cell counting. Chromosome analysis was checked. The molecular markers of ADSCs were detected with immunofluoresence staining. The morphology of ADSCs was identical to fibroblast 1ike and the cells showed active proliferative ability. Vimentin, CD49d and CD13 antigens were detected, but CD34 antigen was negative. Alkaline phosphatase activity was greater in ADSCs during calcification, and Alizarin Red staining was positive. Lipid droplets were apparent around cells during adipogenesis, and Oil Red-O staining was positive. The results demonstrated that ADSCs could be used as seed cells for tissue engineering due to the simple isolation, differentiation and stable and active growth.