重组血管生成素-Ⅰ基因在大鼠骨髓间充质干细胞中的转染和表达

摘要

目的 获得能稳定表达重组血管生成素-Ⅰ基因(Ang-Ⅰ)的骨髓间充质干细胞(MSCs),为进一步观察其对急性肺损伤的保护作用奠定实验基础.方法 提取人胎盘组织总RNA,RT-PCR扩增出Ang-I基因片段,构建Ang-Ⅰ真核表达载体pEGFP-Ang-Ⅰ,利用脂质体介导转染大鼠MSCs,Western blot法检测转基因MSCs的Ang-Ⅰ蛋白表达情况.结果 成功构建Ang-Ⅰ真核表达载体,并成功地将其转入MSCs中;MSCs细胞在转染pEGFP-Ang-Ⅰ质粒后,其培养上清中Ang-Ⅰ蛋白表达量显著升高.结论 Ang-Ⅰ基因成功转染至MSCs中,并可进行有效表达,此为进一步观察Ang-Ⅰ对急性肺损伤的保护作用奠定了实验基础.%Objective To obtain marrow mesenchymal stem cells(MSCs) stablely expressed Ang- Ⅰ ,and to establish the experimental foundation to further observe its protective function for acute lung injury. Methods The total RNA of person placenta was extracted,RT-PCR was used to amplify the Ang- Ⅰ gene fragment, the Ang- Ⅰ eukaryon expression vector pEGFP-Ang- Ⅰ was constructed, and the rat MSCs were transfected using liposome-mediated method, the protein expression of the transgenic MSCs Ang-Ⅰ was detected by Western blot. Results Ang- Ⅰ eukaryotic expression vector was successfully constructed and transfected into the MSCs;the Ang-Ⅰ protein expression in MSCs cell supernatant increased obviously after transfected with the pEGFP-Ang- Ⅰ plasmid. Conclusions Ang- Ⅰ gene can be successfully transfected into MSCs and expressed effectively,this study establish experimental foundation for further observation on its protective function to acute lung injury.