恒河猴骨髓间充质干细胞的分离、培养及传代扩增

目的 建立成年恒河猴骨髓间充质干细胞(rMSC)体外分离、培养及传代扩增的有效体系,为探索新型人工生物骨替代材料提供种子细胞.方法 从恒河猴肋骨抽取骨髓,使用淋巴细胞分离液(Ficoll-Hypaque)密度梯度离心法分离骨髓细胞.用含10%胎牛血清(FCS)的低糖DMEM培养基贴壁培养、胰酶消化传代扩增.通过形态学及流式细胞技术对rMSC进行细胞周期分析及表型特征鉴定.结果 成年恒河猴骨髓来源的rMSC为成纤维样细胞群体,以梭形的成纤维样细胞为主.流式细胞技术结果 显示,rMSC表达CD29、CD44表面分子,不表达CD34、CD45等造血细胞特异性表面分子.结论 成年恒河猴骨髓来源的rMSC的分离、培养及传代扩增的细胞群中无造血系细胞污染且符合干细胞增殖特性,为探索新型人工生物骨替代材料的种子细胞提供了一个可靠来源.

Isolation and ex-expansion of mesenchymal stem cells derived from adult rhesus bone marrow

Objective To establish method system for isolating and ex-expansion of rhesus marrow mesenchymal stem cells(rMSCs) derived from adult rhesus bone marrow in order to provide seed cells for exploration of new artificial biological bone.Methods Marrow was extracted from healthy adult rhesus rib. Marrow cells were separated by density grade acentric method with Ficoll-Hypaque liquid and then cultured in low glucose DMEM containing 10% fetal cattle serum. After full growth,cells were expended used steapsin. According to morphology and flow cytometer technique,cell cycle and cell-surface antigen profiles of rMSCs were identified.Results rMSCs from adult rhesus bone marrow exhibited fibroblast-like morphology with spindle shape and were positive for antigens of CD29 and CD44,while negative for CD34,CD45 and other haematopoietic cellular specific surface molecular with flow cytometer identification. Conclusion rMSCs derived form adult rhesus bone marrow can be successfully isolated and ex-expanded,with multiplication characteristic of stem cell but without hematopoietic origin,which can provide a reliable source of seed cells for new artificial biological bone research.