应用sirna技术下调survivin基因表达以增强sgc7901胃癌细胞对顺铂的敏感性

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1、应用siRNA技术下调survivin基因表达以增强SGC7901胃癌细胞对顺铂的敏感性【摘要】目的利用小干扰RNA(siRNA)技术特异性下调survivin基因的表达，检测SGC7901胃癌细胞转染前后对顺铂敏感性的变化。方法实验分为空白对照组(control组)、单用顺铂组(CDDP组)、脂质体组(Lip组)、单用survivinsiRNA组(siRNA组)、转染survivinsiRNA(转染组)及转染survivinsiRNA联合顺铂作用组(联合作用组)。人工合成survivinsiRNA，通

2、过Lip包裹将siRNA转染胃癌细胞SGC7901，荧光显微镜下观察转染情况。MTT法检测各组细胞的增殖抑制率，Westernblot及免疫细胞化学法检测survivin蛋白的表达，Hoechst染色观察细胞凋亡的形态改变。结果Hoechst染色荧光显微镜下control组、Lip组及siRNA组细胞核呈正常的蓝色,而CDDP组、转染组及联合作用组细胞核染色增强、核质浓缩、核碎裂,呈凋亡改变;联合作用组细胞增殖抑制率(63.93±4.22)%明显高于其余各组(P<0.05);转染组及联合作用组su

3、rvivin蛋白表达明显低于其他各组(P<0.05)。结论应用siRNA技术下调SGC7901胃癌细胞中survivin基因的表达，能够增加其对顺铂的药物敏感性。【关键词】survivin;siRNA;胃癌;顺铂　　Abstract:ObjectiveTodetectthevariationsincisplatinsensitivityofhumangastriccarcinomacellline11SGC7901beforeandaftertransfectiontospecificallydo

4、wnregulatetheexpressionofsurvivinggenewithsmallinterferingRNA(siRNA)technology.MethodsThisexperimentisdividedintothecontrolgroup,cisplatingroup(CDDPgroup),lipsomegroup(lipgroup),siRNAgroup,transfectiongroupandsiRNAcombinedCDDPgroup(combinedgroup).Survivi

5、nsiRNAwasartificiallysynthesizedandthentransfectedintogastriccancercellswithliposome.Thetransfectioneffectsweredeterminedbyfluorescencemicroscopy.TheinhibitoryrateofcellproliferationwasassayedbyMTT,andtheexpressionofsurvivinproteinwasdeterminedbyWesternb

6、lotandimmunocytochemicalstaining.ThemorphologicalchangesoftheapoptoticcellswereobservedbyHoechststaining.ResultsAsobservedunderthefluorescencemicroscopeafterHoechststaining,thenucleipresentedasnormalblueinthecontrolgroup,lipgroupandsiRNAgroup,whileCDDPgr

7、oup,siRNAtransfectiongroupandsiRNAcombinedcisplatingrouphadsuchchangesasreinforcednuclearstaining,karyopyknosisandkaryorrhexiswithapoptoticchanges.TheinhibitionrateofcellproliferationinsiRNAcombinedcisplatingroup(63.93±4.22)%wasmarkedlyhigherthantheother

8、groups(P<0.05).Comparedwithothergroups,theexpressionofsurvivinproteininthe11transfectiongroupandsiRNAcombinedcisplatingroupwassignificantlyreduced,andthedifferenceshadstatisticalsignificance(P<0.05).ConclusionTheappl