论文部分内容阅读
目的构建肝细胞生长因子(HGF)真核细胞表达质粒pEGFP-HGF-C1,探讨HGF对大鼠血管内皮细胞(VECs)的作用。方法通过亚克隆技术,构建含有人HGF cDNA全长的真核细胞表达质粒pEGFP-HGF-C1并测序;向大鼠原代骨骼肌细胞转染质粒pEGFP-HGF-C1,测定转染效率;ELISA法测定细胞上清液中HGF蛋白表达浓度;MTT法测定HGF表达产物对大鼠VECs促增殖作用。结果成功构建正向表达HGF的真核细胞表达质粒pEGFP-HGF-C1,该质粒可有效转染原代培养的骨骼肌细胞并表达HGF,且其表达产物HGF对大鼠VECs的促增殖作用具有量效和时效关系(P<0.05或P<0.01)。结论成功构建的真核表达质粒pEGFP-HGF-C1能有效转染大鼠骨骼肌细胞,分泌的HGF对大鼠VECs有促增殖作用。
Objective To construct eukaryotic expression plasmid pEGFP-HGF-C1 of hepatocyte growth factor (HGF) and explore the effect of HGF on rat vascular endothelial cells (VECs). Methods The full-length eukaryotic expression plasmid pEGFP-HGF-C1 containing human HGF cDNA was constructed by subcloning technique and sequenced. Plasmid pEGFP-HGF-C1 was transfected into primary rat skeletal muscle cells to determine the transfection efficiency. ELISA Method to determine the concentration of HGF protein in supernatant of cells. MTT assay was used to determine the effect of HGF on proliferation of rat VECs. Results The eukaryotic expression plasmid pEGFP-HGF-C1 expressing HGF was successfully constructed. The recombinant plasmid could effectively transfect primary cultured skeletal muscle cells and express HGF, and the effect of its expression product HGF on the proliferation of rat VECs was The dose-effect and time-effect relationship (P <0.05 or P <0.01). Conclusion The successfully constructed eukaryotic expression plasmid pEGFP-HGF-C1 can effectively transfect rat skeletal muscle cells, and the secreted HGF can promote the proliferation of rat VECs.