白藜芦醇对喉癌Hep-2细胞顺铂敏感性的增强作用及其机制

Enhancement effect of resveratrol on sensitivity of laryngeal carcinoma Hep-2 cells to cisplatin and its mechanism

目的:探讨白藜芦醇提高喉癌Hep-2细胞株对顺铂(DDP)敏感性的作用,并阐明其作用机制。方法:Hep-2喉癌细胞随机分为空白对照组和DDP组(终浓度分别为3、6、12和24μmol·L-1)培养6、12、24及48h。CCK-8法检测各组细胞生存率;选出DDP的最佳作用浓度6μmol·L-1和作用时间24h。Hep-2喉癌细胞株经培养后随机分为对照组、DDP组、白藜芦醇组、sirtinol组、白藜芦醇+sirtinol组、白藜芦醇+DDP组、sirtinol+DDP组和白藜芦醇+sirtinol+DDP组。CCK-8法检测各组细胞生存率;采用免疫荧光法观察各组细胞中Sirt1蛋白的表达强度;AnnexinⅤ-FITC Kit法检测各组细胞凋亡率。结果:CCK-8检测,与对照组比较,DDP组和白藜芦醇组细胞生存率明显下降(P<0.01);与DDP组比较,白藜芦醇+DDP组细胞生存率明显下降(P<0.01);与白藜芦醇+sirtinol+DDP组比较,白藜芦醇+DDP组细胞生存率明显下降(P<0.05)。细胞免疫荧光法检测,与DDP组比较,白藜芦醇+DDP组细胞中Sirt1蛋白的平均免疫荧光强度明显升高(P<0.01);与白藜芦醇+sirtinol+DDP组比较,白藜芦醇+DDP组细胞中Sirt1蛋白的平均免疫荧光强度明显升高(P<0.01)。AnnexinⅤ-FITC Kit检测,与DDP组比较,白藜芦醇+DDP组细胞凋亡率均明显升高(P<0.01);与白藜芦醇+sirtinol+DDP组比较,白藜芦醇+DDP组细胞凋亡率明显升高(P<0.01)。结论:白藜芦醇提高Hep-2喉癌细胞株对DDP的敏感性,其机制可能与提高喉癌细胞中Sirt1蛋白表达有关联。

Objective To investigate the enhancement effect of resveratrol on the sensitivity of laryngeal carcinoma Hep-2cells to cisplatin（DDP）and its mechanism.Methods The Hep-2cells were randomly divided into blank control group and different doses（3,6,12,and 24μmol·L-1）of DDP groups and the cells were treated for 6,12,24,and 48 h.The suitable concentration of DDP（6μmol·L-1）and culture time（24h）were chosen.The Hep-2cells treated under the suitable condition were randomly divided into control group,DDP group,resveratrol group,sirtinol group,resveratrol＋ sirtinol group,resveratrol ＋ DDP group,sirtinol ＋ DDP group and resveratrol＋ sirtinol ＋ DDP group.The survival rates of the Hep-2cells in various groups were detected by CCK-8assay;the expression levels of Sirt1 protein in the Hep-2cells in various groups were determined with immunofluorescence method;the apoptotic rates of the Hep-2cells in various groups were detected by AnnexinⅤ-FITC Kit.Results The CCK-8assay results showed that the survival rates of the Hep-2cells in resveratrol group and DDP group were decreased significantly compared with control group（P〈0.01）;compared with DDP group,the survival rates of the Hep-2cells in resveratrol ＋ DDP group were decreased（P〈0.01）;compared with resveratrol＋DDP＋sirtinol group,the survival rates of the Hep-2cells in resveratrol＋ DDP group were decreased（P〈0.05）.The immunofluorescence results showed that the mean fluorescence intensity of Sirt1 protein of the cells in resveratrol＋ DDP group were increased significantly compared with DDP group（P〈0.01）;compared with resveratrol＋ sirtinol＋DDP group,the mean fluorescence intensity of Sirt1 protein of the cells in resveratrol＋DDP group was increased significantly（P〈0.01）.The Annexin Ⅴ-FITC Kit results showed the apoptotic rate of the cells in resveratrol＋ DDP group was higher than those in DDP group and resveratrol＋ sirtinol＋DDP group（P〈0.01）.Conclusion Resveratrol can improve the sensitivity of Hep-2cells to cisplatin,and its mechanism may be related to increasing the expression of Sirt1 protein.