重组人p53腺病毒联合表阿霉素抑制胃癌细胞的体外研究

In Vitro Effect of Recombinant Human Adenovirus p53 Combined with Epirubicin on Human Gastric Cancer Cells

目的:研究基因治疗药重组人p53腺病毒(rAd-p53)与化疗药表阿霉素(EPI)对胃癌细胞的体外作用,探讨基因治疗联合化疗对胃癌的疗效。方法:胃腺癌MGC-803细胞分对照组、rAd-p53组、EPI组、联合用药四组,通过观察细胞生长曲线、流式细胞分析、瑞氏染色、细胞免疫化学染色,研究比较各组细胞生长、细胞周期、细胞形态学、P53蛋白表达。结果:与对照组相比,rAd-p53在用药1、2、3、6d后对MGC-803细胞抑制率为34.52%、10.53%、58.98%、79.17%,EPI为60.71%、67.29%、73.21%、95.14%,联合两药抑制率依次为59.52%、66.17%、67.15%、99.31%,联合用药抑制率显著高于单药处理(P均<0.01)。流式细胞分析发现,rAd-p53主要将细胞周期阻滞于G_2/M期(44.24%),Sub-G_1凋亡峰升高,联合EPI治疗G_2/M期(81.73%)及Sub-G_1峰增高更明显(P均<0.01)。瑞氏染色可见,rAd-p53治疗组MGC-803细胞仅有少数聚集,细胞形态不规则,胞质深染,可见凋亡细胞;EPI组细胞呈小片状聚集,形态结构较完整,与对照组无明显差别;联合用药组细胞数明显减少,形态不规则,胞质深染,凋亡细胞数更多。免疫细胞化学染色可见,P53蛋白于rAd-p53治疗组细胞核及胞质中表达,联合EPI作用效果更显著;对照组与EPI组P53蛋白于细胞核表达。结论:rAd-p53对胃癌MGC-803细胞有抑制作用,促进细胞凋亡,可提高胃癌MGC-803细胞对化疗药EPI的敏感性,二者有协同作用。

Objective： To study the effect of recombinant human adenovirus p53 injection （rAd-p53） and Epirubicin Hydrochloride （EPI） on human gastric cancer cells in vitro, and explore the therapeutic effect of gene therapy combined with chemotherapy on gastric cancer. Methods： The MGC-803 human gastric adenocarcinoma cell line was treated with rAd-p53 or/and EPI or left untreated as a blank control. Cell growth, flow cytometry, Wright＇s staining, and cyto-immunochemistry were used to analyze the therapeutic efficacy of the different treatment groups. Results： After treating MGC-803 on days 1, 2, 3 ＆ 6, the cell inhibition rates were 34.52%, 10.53%, 58.98% and 79.17%, respectively, for rAd-p53 group; 60.71%, 67.29%, 73.21% and 95.14%, respectively, for EPI group; and 59.52%, 66.17%, 67.15% and 99.31%, respectively, for the combined group. The combined group was significantly higher than that of single agent groups （P〈0.01）. Based on the flow cytometry, we found that rAd-p53 blocked the cell cycle at G2/M phase （44.243%） and a higher Sub-G1 peak was observed. In the combined group, the G2/M phase （81.734%） and Sub-G1 peak were significantly evident （P〈0.01）. Decreased cell number, abnormal cell morphology and apoptotic cells were found under Wright＇s staining, and p53 protein stained positively in the nucleus and cytoplasm by immunochemical analysis in rAd-p53 treated group, especially in the combined treatment group. The EPI group was not different from the blank control. Conclusion： The gene therapy agent tAd-p53 has an inhibiting and apoptotic effect on MGC-803 cells, thus it could enhance the chemotherapeutic effect of EPI on MGC-803 cells, suggesting synergism between gene therapy and chemotherapy.