摘要
目的探究慢病毒介导分泌型糖蛋白神经生长导向因子2(Slit2)过表达载体转染对骨髓间充质干细胞(BMSCs)向神经细胞转化的作用。方法利用基因过表达技术,构建Slit2过表达载体;慢病毒转染BMSCs;利用流式细胞仪检测BMSCs的表面蛋白,RT-PCR和Western-blot方法检测空白对照组、空载体病毒组和过表达载体病毒组神经细胞表面蛋白胶质纤维酸性蛋白(GFAP)和神经元特异性烯醇化酶(NSE)的表达。结果第2代BMSCs表面CD44、CD29、CD34和CD45的表达率分别为60.2%、58.3%、3.4%和2.6%,慢病毒转染效率为90%以上。空白对照组、空载体病毒组和过表达载体病毒组的GFAP、NSE mRNA和蛋白相对表达量比较,差异有统计学意义(F=9.089～13.893,P<0.05);其中过表达载体病毒组的GFAP、NSE mRNA和蛋白相对表达量明显高于空载体病毒组(P<0.05),空载体病毒组和空白对照组比较差异无显著性(P>0.05)。结论 Slit2过表达质粒可以提高BMSCs向神经细胞转化的效率,为神经损伤和脊髓损伤提供种子细胞。
Objective To explore the effect of lentivirus-mediated overexpression of Slit2 on the transformation of bone marrow mesenchymal stem cells (BMSCs) into neural cells. Methods A Slit2 overexpression vector was constructed and used to transfect BMSCs via lentivirus. Flow cytometry was used to detect surface proteins of BMSCs. RT-PCR and Western blot were used to determine the expression of glial fibrillary acidic protein (GFAP) and neuron-specific enolase (NSE) on the surface of neural cells in blank control group, empty lentiviral vector group, and overexpression lentiviral vector group. Results The expression of CD44, CD29, CD34, and CD45 was 60.2%, 58.3%, 3.4%, and 2.6%, respectively, on the surface of the second generation BMSCs. The transfection efficiency of lentivirus was over 90%. There were significant differences in relative mRNA and protein expression of GFAP and NSE between the blank control group, the empty lentiviral vector group, and the overexpression lentiviral vector group ( F =9.089-13.893, P <0.05). The overexpression lentiviral vector group had significantly higher relative mRNA and protein expression of GFAP and NSE than the blank control group and the empty lentiviral vector group ( P <0.05), while there were no significant differences between the blank control group and the empty lentiviral vector group ( P >0.05). Conclusion The Slit2 overexpression plasmid can promote the efficiency of the transformation of BMSCs into neural cells, which provide seed cells for nerve injury and spinal cord injury.
作者
刘文忠
蔡静
李军
荣春
范磊
张海宁
LIU Wenzhong;CAI Jing;LI Jun;RONG Chun;FAN Lei;ZHANG Haining(Department of Joint Surgery, the Affiliated Hospital of Qingdao University, Qingdao 266003, China)
出处
《青岛大学学报（医学版）》
CAS
2019年第2期137-140,共4页
Journal of Qingdao University(Medical Sciences)
基金
国家自然科学基金资助项目(81171774
81272056
81672197)
作者简介
刘文忠(1982-),男,硕士研究生,主治医师;通讯作者:张海宁(1976-),男,博士,主任医师,硕士生导师。E-mail:zhanghaining@163.com。
