目的:探讨SIRT1蛋白在胃癌组织和细胞中的表达,分析SIRT1蛋白表达与耐药相关蛋白P-gp和Top-Ⅱα的相关性.方法:免疫组化SP法检测SIRT1、P-gp及Top-Ⅱα蛋白在15例胃癌组织和15例正常胃黏膜组织中的表达.Western blot检测正常人胃黏膜上皮细胞株GES-1和人胃癌细胞株SGC7901、MKN45、MKN28、HGC27、AGS、MGC803中SIRT1蛋白的表达;检测GES-1、SGC7901及MKN45中的SIRT1、P-gp和Top-Ⅱα的蛋白表达;siRNA-SIRT1/脂质体转染胃癌细胞SGC7901后,Western blot检测SIRT1、P-gp和Top-Ⅱα的蛋白表达变化;MTT-检测SGC7901细胞的体外增殖能力变化及对化疗药物5-Fu的敏感性变化.结果:15例胃癌组织中SIRT1蛋白表达阳性15例,P-gp蛋白表达阳性13例,Top-Ⅱα蛋白表达阳性3例;15例正常胃黏膜中SIRT1蛋白表达阳性6例,P-gp蛋白表达阳性5例,Top-Ⅱα蛋白表达阳性0例.GES-1、SGC7901、MKN45、MKN28、HGC27、AGS、MGC803中SIRT1相对蛋白表达量平均值分别为0.385、0.827、0.009、0.232、0.275、0.159、0.117;GES-1、SGC7901、MKN45中的P-gp相对蛋白表达量平均值分别为0.339、0.526、1.03;Top-Ⅱα的蛋白相对表达量分别为0.093、0.889、0.158. siRNA-SIRT1转染SGC7901 8 h,培养72 h后检测control、BC-V、negative、siRNA-1、siRNA-2、siRNA-3的SIRT1相对蛋白表达量平均值分别为0.965、0.937、0.958、0.567、0.253、0.083;P-gp相对蛋白表达量平均值分别为1.893、1.905、1.932、0.465、0.006、0.005;Top-Ⅱα的蛋白相对表达量分别为0.09、0.07、0.073、0.085、0.168、0.195.M-TT结果显示抑制SIRT1蛋白表达后,SGC7901体外增殖能力无明显变化,对化疗药物5-Fu的敏感性显著增加.结论:SIRT1在胃癌组织中过表达,且过表达的胃癌组织、细胞中SIRT1扮演促癌因子角色;SIRT1蛋白过表达能促进P-gp蛋白表达,降低Top-Ⅱα蛋白表达,导致胃癌细胞化疗多重耐药性的增加.%Objective:Explore the expression of SIRT1 protein in gastric cancer tissues and cells.Analyze the correlation of SIRT1 protein expression and multidrug resistance protein P-gp and Top-Ⅱ alpha.Methods:Immunohistochemical to detect SIRT1,P-gp and Top-Ⅱ alpha protein in 15 cases of gastric cancer tissue and 15 cases of normal gastric mucosa tissues.Western blot test the expression of SIRT1 protein in normal gastric mucosa epithelial cell line GES-1 and human gastric cancer cell line SGC7901,MKN45,MKN28,HGC27,AGS and MGC803.Western blot test the expression of SIRT1,P-gp and Top-Ⅱ alpha protein in normal gastric mucosa epithelial cell line GES-1 and human gastric cancer cell line SGC7901,MKN45.After siRNA-SIRT1/liposome transfection gastric cancer cells SGC7901,western blot test the changes of expression of SIRT1,P-gp and Top-Ⅱ alpha protein and MTT test the changes of SGC7901 cells proliferation in vitro and the sensitivity of chemotherapy drugs 5-Fu.Results:In 15 cases of gastric cancer tissue have the positive expression of 15 cases of SIRT1 protein,13 cases of P-gp protein and 3 cases of Top-Ⅱ alpha protein.In 15 cases of normal gastric mucosa tissues have the positive expression of 6 cases of SIRT1 protein,5 cases of P-gp protein and 0 cases of Top-Ⅱ alpha protein.The relative quantity of SIRT1 protein expression average were 0.385,0.827,0.009,0.232,0.275,0.159,0.275 in GES-1,SGC7901,MKN45,MKN28,HGC27,AGS and MGC803,respectively.The relative quantity of P-gp protein expression average were 0.339,0.526,1.03 in GES 1,SGC7901 and MKN45 and Top-Ⅱ alpha protein were 0.093,0.889,and 0.158,respectively,siRNA-SIRT1 transfected SGC7901 for 8 hours and after 72 hours to test the relative quantity of SIRT1 protein expression average were 0.965,0.937,0.958,0.567,0.253,0.083 in control,BC-V,negative,siRNA-1,2,3 SIRT1,P-gp were 1.893,1.905,1.932,0.465,0.006,0.465 and Top-Ⅱ alpha were 0.09,0.07,0.073,0.085,0.168,0.085.MTT results showed that after SIRT1 protein expression was inhibited the SGC7901 proliferation has no obvious change in vitro and cell sensitivity to the chemotherapy drugs 5-Fu was increased significantly.Conclusion:SIRT1 expression in gastric cancer tissues and the expression of SIRT1 in gastric cancer tissue and cell carcinoma factors role,SIRT1 protein overexpression can promote P-gp protein expression lower Top-Ⅱ alpha protein expression in gastric cancer cells to chemotherapy of multiple drug resistance increase.
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