人永生化骨髓基质干细胞单克隆细胞系分化差异性的研究

Clonal heterogeneity in differentiation potential of immortalized human mesenchymal stem cells in vitro and in vivo

目的研究单克隆永生化骨髓基质干细胞在体内和体外多向分化的差异性。方法对挑取的单克隆细胞系和亲代细胞系分别在体外进行脂肪、骨、神经样细胞诱导分化；接种联合免疫缺陷性鼠，对移植物进行免疫组化染色；比较体内分化的不同。结果在体外诱导分化中，单克隆细胞系丢失成骨分化的能力；体内分化中，人永生化骨髓基质细胞（HMSC-TERT）-2较其他细胞系在人白细胞共同抗原，神经胶质酸性蛋白，波形蛋白表达上明显增强；HMSC—TERT-C19较HMSC-TERT-2，HMSC-TERT-20以及MSC-H在角蛋白表达上明显增强；HMSC—TERT—C2比HMSC-TERT-2在肌动蛋白表达上明显增强。结论人永生化骨髓基质细胞单克隆细胞系在体内和体外的分化存在差异性。提示标准化体外培养的条件和体内接种的程序，对早日将干细胞应用于临床有重要的意义。

Objective To study the clonal heterogeneity in differentiation potential of immortalized mesenchymal stem cells in vitro and in vivo. Methods The monoclonal cell lines were performed with limiting dilution cloning, and were induced to adipocytic, asteogenic and neuronal differentiation in vitro. After transplanted the monoclonal cell lines into SCID mice,the xenotransplants were removed and evaluated by immunohistochemistry. Results From the parental HMSC-TERT,32 single-cell derived clones were established, of which the differentiation properties varied considerably in vitro. The ceils grow in different plating densities during expansion in culture： HMSC- TERT-2 expressed more strongly in LCA ,GFAP and vimentin; HMSC-TERT-C19 expressed more strongly in keratinose than HMSC-TERT-2, HMSC-TERT-20, and MSC-H; HMSC-TERT-C2 expressed more strongly in actin than HMSC-TERT-2. Conclusions The HMSC-TERT monoclone cells are heterogeneity in differentiation potential in vitro and in vivo, suggesting that standard in vitro culture and in vivo inoculate procedure play an important role in the clinical application of stem cells.