论文部分内容阅读
目的探讨人促肝细胞生长因子(hHGF)对人骨髓间充质干细胞(hBMSCs)向肝样细胞分化的影响。方法克隆hHGF基因,构建携带hHGF基因的真核表达质粒(pcDNA3.1(+)-hHGF),用脂质体法将pcDNA3.1(+)-hHGF转染hBMSCs,分别通过激光共聚焦、反转录聚合酶链反应(RT-PCR)、免疫蛋白印迹(Western blot)检测hBMSCs中表达hHGF的水平;用酒精性肝硬化患者血清诱导hHGF上调后的hBMSCs,采用Western blot分别检测诱导细胞内甲胎蛋白(AFP)及白蛋白(ALB)的表达变化。结果成功构建pcDNA3.1(+)-hHGF表达质粒;激光共聚焦、RT-PCR、Western blot结果均表明转染后hBMSCs中hHGF分子呈高表达,Western blot结果发现诱导7 d,14 d转染后hBMSCs中AFP、ALB表达明显高于对照组。结论转染pcDNA3.1(+)-hHGF可以明显上调hHGF在hBMSCs中的表达;hBMSCs高表达hHGF后,经酒精性肝硬化血清诱导可明显促进其向肝样细胞分化,为基因治疗终末期肝病提供了一种新方法。
Objective To investigate the effect of human hepatocyte growth factor (hHGF) on differentiation of human bone marrow mesenchymal stem cells (hBMSCs) into hepatocyte-like cells. Methods The hHGF gene was cloned and the eukaryotic expression plasmid carrying hHGF gene (pcDNA3.1 (+) - hHGF) was constructed. The hBMSCs were transfected with pcDNA3.1 (+) - hHGF by lipofectamine respectively. The expression of hHGF in hBMSCs was detected by RT-PCR and Western blot. The hBMSCs induced by hHGF were induced by serum of patients with alcoholic cirrhosis. Western blot was used to detect the expression of hHGF. Fetal protein (AFP) and albumin (ALB) expression changes. Results The recombinant plasmid pcDNA3.1 (+) - hHGF was constructed successfully. The results of confocal laser scanning microscopy, RT-PCR and Western blot showed that hHGF was highly expressed in hBMSCs after transfection. After hBMSCs AFP, ALB expression was significantly higher than the control group. Conclusion Transfection of pcDNA3.1 (+) - hHGF can upregulate the expression of hHGF in hBMSCs. After hBMSCs are overexpressed, serum induced by alcoholic cirrhosis can obviously promote the differentiation of hHGF into hBMSCs. Provide a new method.