【摘要】 目的:探讨RNA干扰下调Burkitt淋巴瘤Daudi细胞系凋亡抑制基因survivin的表达对阿霉素敏感性的影响。方法:构建survivin发夹RNA(shRNA)真核表达载体,脂质体介导转染Daudi细胞。多重RT-PCR检测survivin mRNA表达;Western blotting检测Survivin蛋白表达;流式细胞仪(FCM)检测干扰前后细胞凋亡率;MTT法检测干扰前后细胞对化疗药物阿霉素(adriamycin,ADR)敏感性变化。结果:与无功能control-shRNA处理组和PBS处理组比较,转染survivin-shRNA细胞的survivin mRNA和蛋白表达率显著降低,抑制率分别为62.32%和61.88%(P<0.05);FCM结果示转染组细胞凋亡指数(apoptosis index,AI)显著高于两对照组(P<0.05);MTT结果显示,转染组细胞的阿霉素半数抑制浓度(IC50)为(0.25±0.43)μmol/L,显著低于无功能control-shRNA处理组(0.87±0.21)μmol/L和PBS处理组(0.91±0.36)μmol/L,P<0.05;相同剂量ADR对转染细胞的生长抑制率明显高于PBS组和control组。结论:发夹RNA干扰survivin基因可显著提高Daudi细胞凋亡率和对化疗药物阿霉素的敏感性。更多还原

【Abstract】 AIM:This study was designed to use RNA interference technique to down-regulate the expression of survivin gene in human Burkitt’s lymphoma cell line Daudi and to explore the effect on sensitivity of Daudi cells to adriamycin.METHODS:The survivin-shRNA expression vector was constructed and transfected into Daudi cells.Expression of survivin mRNA and protein were assessed by RT-PCR and Western blotting analysis,respectively.Apoptosis index of transfected Daudi cells was quantified by flow cytometry.The sensitivity of Daudi cells to adriamycin(ADR) before and after transfection was detected by MTT test.RESULTS:The mRNA and protein levels of survivin were down-regulated by 62.32% and 61.88%,respectively,compared to those in control-shRNA treated group and PBS treated group(P<0.05).Meanwhile,the apoptosis index was significantly increased(19.10%±2.15%),compared to that in control group(4.48%±1.54%) and PBS group(4.35%±1.37%,P<0.05).The 50% inhibition concentration(IC50) of ADM to Daudi cells was significantly decreased(0.25±0.43) μmol/L,compared to that in control group(0.87±0.21) μmol/L and PBS group(0.91±0.36) μmol/L,P<0.05.CONCLUSION:Down-regulation of survivin expression in Daudi cells by shRNA effectively induces apoptosis and increases the sensitivity of Daudi cells to ADR.更多还原