GDNF基因修饰的BMSCs对大鼠缺氧复氧神经细胞凋亡的实验研究

Effects of GDNF gene modified BMSCs on apoptosis of neurons induced by hypoxia reoxygenation*

目的研究腺病毒介导的外源性胶质细胞源性神经营养因子（GDNF）基因在大鼠骨髓间充质干细胞（BMSCs）中的表达及其对缺氧复氧神经细胞凋亡的影响。方法将GDNF基因重组腺病毒感染大鼠BMSCs,采用酶联免疫吸附试验（ELISA）技术检测外源性GDNF基因在BMSCs中的表达水平;将GDNF基因修饰的BMSCs与缺氧复氧神经元共培养,观察神经细胞的凋亡情况。结果 GDNF基因感染组在感染后3~12 d能稳定表达GDNF蛋白,空质粒载体感染组和未感染组基本上无GDNF蛋白的表达;与BMSCs共培养组相比,BMSCs/GDNF分泌的GDNF能显著降低缺氧复氧神经细胞的凋亡率（复氧12 h至5 d,P〈0.05）。结论腺病毒介导的基因感染技术能够有效地将GDNF基因转至BMSCs中,表达和分泌有生物学活性的GDNF蛋白,这为进一步研究GDNF基因修饰的BMSCs应用于中枢神经系统疾病的治疗研究奠定了基础。

Objective To study the expression of exogenous glia cell line derived neurotrophic factor（GDNF） gene mediated by adenovirus in bone marrow mesenchymal stem cells（BMSCs） of rats,and the effection of GDNF on apoptosis of neurons induced by hypoxia reoxygenation. Methods Theexpression level of exogenous GDNF gene in BMSCs was detected by Enzyme linked immu- nosorbent assay（ELISA） technology after GDNF gene recombinant adenovirus transfected BMSCs of rats;To observe apoptosis of hypoxia reoxygenation neurons when cocultured with GDNF gene modified BMSCs. Results GDNF gene infection group could ex- press exogenetic GDNF protein during the following 3--12 days, empty plasmid vector infection group and not infection group basi- cally has not the expression of GDNF protein;compared to BMSCs group,GDNF secreted by BMSCs/GDNF could significantly reduce the apoptosis rate of hypoxia reoxygenation neurons（12 b to 5 d,P〈0.05）. Conclusion The gene infection technology media- ted by adenovirus could effectively transfect GDNF gene to gMSCs,express and secrete bioactive GDNF protein,which will provide foundation for the further research GDNF gene modified BMSCs therapy of Central nervous system diseases.