腺病毒介导胰高血糖素样肽-1受体在小鼠骨髓间充质干细胞表达的体外研究
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国家自然科学基金 (30971404, 81070621);江苏省自然科学基金(BK2007251)资助


Infection of adenovirus containing GLP-1R gene into mouse mesenchymal stem cells in vitro
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    摘要:

    目的:观察腺病毒介导胰高血糖素样肽-1受体(GLP-1R)转染小鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BM-MSCs)的表达情况及Exendin-4/GLP-1R系统对BM-MSCs细胞增殖及凋亡的影响-方法:利用Ad.Max系统构建含GLP-1R基因的腺病毒表达载体Ad.GLP-1R,感染小鼠BM-MSCs-采用RT-PCR-Western blot和免疫荧光方法检测GLP-1R的表达情况-用Exendin-4刺激感染Ad.GLP-1R的BM-MSCs,通过细胞计数和流式细胞术检测细胞生长曲线-细胞周期及细胞凋亡的变化-结果:腺病毒对小鼠BM-MSCs有较高的感染效率,病毒感染复数(MOI)为400时,感染率高达89.5%-BM-MSCs不表达GLP-1R,感染Ad.GLP-1R后有效表达GLP-1R基因和蛋白-Ad.GLP-1R感染BM-MSCs后,Exendin-4刺激组细胞生长曲线显著高于未刺激组(P < 0.05)-细胞周期检测结果显示,刺激组G0/G1期细胞数显著下降-G2/M期细胞数明显升高,增殖指数(S+G2/M)亦显著增加(P均 < 0.01)-而细胞凋亡率无明显变化-结论:Ad.GLP-1R可以有效感染BM-MSCs,获得高表达GLP-1R的BM-MSCs-Exendin-4可以促进Ad.GLP-1R感染的BM-MSCs增殖-

    Abstract:

    Objective:To investigate the expression of glucagon like peptide 1 receptor(GLP-1R) in mouse bone marrow mesenchymal stem cells(BM-MSCs) transfected by adenovirus and the effect of Exendin-4/GLP-1R on BM-MSCs proliferation and apoptosis after transfection. Methods: The adenoviral vector containing GLP-1R gene was constructed through the Ad.Max-system and infected the BM-MSCs. The expression of GLP-1R in Ad.GLP-1R-BM-MSCs was examined by RT-PCR,immunofluorescence,and Western blot,respectively. The effects of Exendin-4/GLP-1R on cell growth,cell cycle and apoptosis in Ad.GLP-1R-BM-MSCs were evaluated by cell counting and flow cytometry. Results: BM-MSCs could be effectively infected by adenovirus. When multiplicity of infection (MOI) was 400, the infection efficiency was 89.5%. Mouse BM-MSCs didn’t express the GLP-1R gene natively. After infected with Ad.GLP-1R,the BM-MSCs expressed the GLP-1R gene and protein effectively,and the cell growth could be stimulated by Exendin-4. Compared with the unstimulated Ad.GLP-1R-BM-MSCs,G0/G1 phase cells in the Exendin-4 stimulation group decreased significantly,G2/M phase cells and proliferation index (S+G2/M) increased significantly (P < 0.01). But there was no significant difference in the apoptosis rate of two groups. Conclusion: Ad.GLP-1R can infect mouse BM-MSCs effectively and induce the expression of GLP-1R. Exendin-4 can promote the proliferation of BM-MSCs infected with Ad.GLP-1R.

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张勤凤,朱 剑,徐宽枫,陈 恒,杨 涛,武晓泓.腺病毒介导胰高血糖素样肽-1受体在小鼠骨髓间充质干细胞表达的体外研究[J].南京医科大学学报(自然科学版),2012,(3):310-314

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  • 收稿日期:2011-08-31
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