Shen Jian-guo, Cheong Jae-ho, Noh Sung-hoon, Wang Lin-bo
Department of Surgical Oncology, Sir Run Run Shaw Hospital, Zhejiang University College of Medicine, Hangzhou 310016, China.
Zhonghua Zhong Liu Za Zhi. 2007 May;29(5):338-41.
To investigate the effects of hepatocyte growth factor (HGF) on adriamycin-induced apoptosis of gastric cancer cells in vitro.
An eukaryotic expression plasmid (pIRES2-EGFP) containing HGF (pIRES2-EGFP-HGF) was constructed. Human gastric cancer cell line MKN-45 cells were transfected in vitro with pIRES2-EGFP containing HGF or not. RT-PCR and Western blot were used to determine the target gene expression. Function of HGF was determined by MDCK cell scattering assay. Cell viability was tested by MTT assay. Apoptosis was evaluated by DNA fragmentation assay as well as flow cytometry using PI staining.
The HGF transfected MKN45 cells could stably express HGF mRNA, and secrete HGF protein to the cell culture median which was detected to exhibit normal function. The cell inhibition rate induced by adriamycin in HGF-transfected cells was decreased as compared to that of parental and mock transfected cells. When treated with adriamycin at 0.1 microg/ml, the parental and mock transfected cells present typical apoptotic ladder on DNA electrophoresis while HGF transfected cell did not. The apoptotic rate was decreased in HGF transfected cells as compared with that of parental and mock transfected cells.
HGF gene transfection may suppress adriamycin-induced apoptosis of gastric cancer cells.
探讨肝细胞生长因子（HGF）对阿霉素诱导的胃癌细胞体外凋亡的影响。
构建含HGF的真核表达质粒（pIRES2-EGFP）（pIRES2-EGFP-HGF）。人胃癌细胞系MKN-45细胞在体外转染含或不含HGF的pIRES2-EGFP。采用RT-PCR和Western印迹法检测目的基因表达。通过MDCK细胞散射试验确定HGF的功能。采用MTT法检测细胞活力。通过DNA片段化分析以及使用PI染色的流式细胞术评估细胞凋亡。
转染HGF的MKN45细胞可稳定表达HGF mRNA，并向细胞培养基中分泌HGF蛋白，检测显示其具有正常功能。与亲本细胞和mock转染细胞相比，阿霉素诱导的转染HGF细胞的细胞抑制率降低。当用0.1μg/ml阿霉素处理时，亲本细胞和mock转染细胞在DNA电泳上呈现典型的凋亡梯带，而转染HGF的细胞则没有。与亲本细胞和mock转染细胞相比，转染HGF的细胞凋亡率降低。
HGF基因转染可能抑制阿霉素诱导的胃癌细胞凋亡。
