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目的 观察野生型p5 3基因 (wtp5 3 )对体外培养人胃癌细胞系生长的抑制作用 ,探讨野生型p5 3基因在胃癌基因治疗中的重要意义。方法 以携带有野生型p5 3基因的复制缺陷型腺病毒 (pAd -p5 3 )为载体 ,转染有p5 3基因突变的人胃癌细胞系BGC -790 1。应用生化染色 ,检测外源基因的转染效率 ;应用免疫组化、原位杂交等方法检测外源基因的表达效果 ;应用细胞计数、MTT法检测pAd -p5 3转染对胃癌细胞生长的抑制效果 ;应用流式细胞仪检测细胞周期。结果 当腺病毒在 10 0MOI以上效靶比转染人胃癌细胞系时 ,转染效率达 10 0 %。转染 3d以后 ,胃癌细胞中突变性p5 3蛋白表达开始减少 ,野生型p5 3mRNA的表达开始增多 ,BGC -790 1细胞生长受到明显抑制 ,流式细胞仪检测显示G0 /G1期细胞数增加 (未转染组 5 6 47% ,转染pAd -p5 3组 79 40 % ,P<0 0 5 ) ,S期细胞数减少 (未转染组 3 0 80 % ,转染pAd -p5 3组 13 81% ,P <0 0 1)。结论 腺病毒介导的野生型p5 3基因体外转染人胃癌细胞 ,可有效抑制胃癌细胞的生长 ,可能成为胃癌基因治疗的手段之一。
Objective To observe the inhibitory effect of wild type p5 3 gene (wtp5 3) on the growth of human gastric cancer cell lines in vitro and to explore the significance of wild type p5 gene in gene therapy of gastric cancer. Methods Human gastric cancer cell line BGC-7901 mutated with p5 3 gene was transfected with the replication-deficient adenovirus (pAd-p5 3) carrying the wild type p5 3 gene. Application of biochemical staining, detection of foreign gene transfection efficiency; application of immunohistochemistry, in situ hybridization and other methods to detect the expression of foreign genes; using cell counting, MTT assay pAd-p5 3 transfected gastric cancer cell growth inhibition Effect; application of flow cytometry cell cycle. Results When the adenovirus was transfected into human gastric cancer cell line with an effective target of more than 100 MOI, the transfection efficiency reached 100%. After transfection for 3d, the expression of mutant p5 3 protein began to decrease in gastric cancer cells, the expression of wild-type p5 3 mRNA began to increase, the growth of BGC-7901 cells was significantly inhibited, and the number of G0 / G1 phase cells was increased by flow cytometry Transfected pAd-p5 3 group 79 40%, P <0 05), the number of S phase cells decreased (3080% untransfected group transfected pAd-p5 3 group 13 81%, P <0 0 1). Conclusion Adenovirus-mediated transfection of wild type p5 3 gene into human gastric cancer cells in vitro can effectively inhibit the growth of gastric cancer cells and may be one of the means of gene therapy for gastric cancer.