摘要
目的观察依达拉奉联合表皮生长因子(EGF)诱导人骨髓间充质干细胞(hBMSCs)神经元样细胞分化过程中端粒酶活性的变化。方法依达拉奉联合EGF诱导hBMSCs,观察其形态变化。免疫细胞化学法检测神经元特异性烯醇化酶(NSE)、胶质纤维酸性蛋白(GFAP)表达,采用TRAP-ELISA和Western blotting分别检测诱导后hBMSCs端粒酶活性及端粒酶逆转录酶(TERT)变化。结果经诱导后,hBMSCs呈典型神经元样细胞分化,NSE阳性率为81.7%,GFAP阳性率为8.5%。端粒酶活性和TERT蛋白诱导4h后均无明显变化,但诱导6h后细胞端粒酶活性明显降低,诱导至24h时,细胞端粒酶活性已近阴性。Western blotting试验的结果与细胞端粒酶活性的改变一致。结论依达拉奉联合表皮生长因子体外可以诱导hBMSCs分化为神经元样细胞,端粒酶活性在诱导分化早期呈高表达,分化过程中逐渐降低直至消失。
Objective To evaluate the telomerase activity during the differentiation of human bone marrow mesenchymal stem cells（hBMSCs）into neuron-like cells in vitro,and the relation between the telomerase reverse transcriptase（TERT）and telomerase.Methods hBMSCs were isolated and purified by means of density-gradient centrifugation and the differential time adherent methods,and expanded in different culture medium in vitro.The changes of morphological characteristics of neuron-like cells differentiation from hBMSCs induced by Edaravone and EGF were observed;the expression of NSE,GFAP in treated and untreated hBMSCs were performed with immunocytochemistry,and TRAP-ELISA was used to detect telomerase activity before and after hBMSCs induction.The TERT protein expression were detected respectively by western blot.Results The treated hBMSCs exhibited lots of neurites and extended to weave into a network.The induced cells presented classical neuron-like morphology.The expressions of NSE,GFAP in control group was negative,while NSE expression was rich and weak expression for GFAP in treated groups.TRAP-ELISA showed telomerase activity of hBMSCs was no significant after 4 hours of induction,and then decreased after 6 hours of induction,and negative after 24 hours of induction（P〈0.05）.The expression of TERT protein was decreased correspondingly during the induction.The results were similar with the telomerase activity of hBMSCs.Conclusions The hBMSCs could differentiate into neuron-like cells induced by Edaravone and EGF with enhancement of telomerase activity in the early,decrease and even disappearance during the induction.
出处
《现代实用医学》
2010年第7期732-734,747,共4页
Modern Practical Medicine
基金
广东省自然科学基金资助项目
编号:06028967
关键词
依达拉奉
表皮生长因子
人骨髓间充质干细胞
端粒酶
Edaravone
Epidermal growth factor
Human bone marrow mesenchymal stem cells
Telomerase
