【摘要】 目的:观察bcl-2基因脂质体法转染人胃上皮细胞系GES-1及其转染后对GES-1细胞增殖的影响.方法:以脂质体法转染构成bcl-2基因表达的人胃上皮细胞系GES-1细胞模型,空载体质粒pcDNA3转染GES-1细胞及正常GES-1细胞为对照;经酶标免疫组织化学法鉴定bcl-2基因表达阳性细胞;用HE染色法进行形态学观察;并采用细胞计数及MTT法分析bcl-2转染后对GES-1细胞增殖的影响.结果:脂质体法转染bcl-2基因后,细胞高表达Bcl-2蛋白;HE染色后形态学观察无明显改变;细胞计数法绘制细胞生长曲线结果显示bcl-2基因转染6d后细胞生长速度明显超过对照组,MTT法结果显示bcl-2基因转染第24,48,72h,A490nm值分别为4.15±0.31,5.98±0.56,8.94±0.79,对照组分别为3.01±0.20,4.76±0.52,7.69±0.84,两组相比较具有显著性差异(P<0.05).结论:bcl-2基因转染使GES-1细胞稳定且高表达Bcl-2蛋白,并促进了细胞增殖.更多还原

【Abstract】 AIM:To observe the effect of bcl-2 gene transfection on the proliferation of human gastric epithelial cell GES-1. METHODS:The eukaryotic vector carrying the full length of bcl-2 gene and the neomycin resistance gene (pcDNA3/bcl-2) and the one only carrying the neomycin resistance gene (pcDNA3) were amplified and purified. The plasmid of pcDNA3/bcl-2 was used to transfect GES-1 cells by using the lipofectamine, and the empty vector pcDNA3 to transfect GES-1 cells as controls. And then, the cells were cultured in 1640 culture medium containing an appropriate concentration of G418 for selection. The method of HRP label immunohistochemical staining was used to identify the cells expressing bcl-2 gene positively. The morphologic changes of GES-1 cells were observed under an optical microscope by HE staining. The cell proliferation affected by the transfection of bcl-2 was measured by cell counting and MTT assay. RESULTS: bcl-2 gene and the vector pcDNA3 were transfected separately into GES-1 cells by lipofectamine. After G418 selection, the cells were transfected steadily. Compared with controls, HRP label immunohistochemical staining showed that bcl-2 gene transfected cells expressed Bcl-2 protein much more obviously. No morphologic changes were observed by HE staining. The growth curve was drawn by the method of cell counting,which showed that the growth of bcl-2 gene transfected cells was faster than that of empty vector transfected cells and untransfected cells after 6 d culture. And MTT assay showed that the growth of bcl-2 gene transfected cells were 4.15±0.31,5.98±0.56 and 8.94±0.79, respectively. While MTT assay showed that the growth of control cells were 3.01± 0.20 , 4.76±0.52 and 7.69±0.84, respectively. Significant differences were found between the 2 groups (P< 0.05). CONCLUSION: The transfected of bcl-2 gene makes the cells express Bcl-2 protein steadily and highly, and enhances the cells’ proliferation and malignancy.更多还原