SIRNA干扰MEKK3基因促进TRAIL诱导乳腺癌细胞凋亡

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1、上海交通大学硕士学位论文siRNA干扰MEKK3基因促进TRAIL诱导乳腺癌细胞凋亡姓名：刘寿贵申请学位级别：硕士专业：外科学指导教师：郭善禹20090501胞的生长抑制作用和诱导凋亡作用，两者联合应用存在显著的协同效应；MEKK3-siRNA单独应用可以抑制MCF-7细胞NF-kappaB信号通路的激活，MEKK3-siRNA与TRAIL联合作用也可明显抑制MCF-7细胞NF-kappaB信号通路的激活。结论：siRNA干扰MEKK3基因表达，能促进TRAIL对乳腺癌MCF-7细胞凋亡的诱导作用，

2、其机制可能是通过抑制MEKK3mRNA及蛋白表达，进而抑制NF-kappaB信号通路的激活而起作用。[关键词]乳腺癌,TRAIL,siRNA,促分裂原活化蛋白/细胞外信号调节激酶-激酶3（MEKK3）,凋亡,NF-kappaBMEKK3-TARGETEDSIRNACOMBINEDWITHTRAILINDUCESAPOPTOSISOFBREASTCANCERCELLLINEMCF-7Objective:TRAIL(tumornecrosisfactorrelatedapoptosisinducingl

3、igand)hastheapoptosisinducingeffectonbreastcancercells,buttherehasresistanceinsomecancercelllines.Mitogen-activatedprotein/extracellularsignalregulatedkinase-kinase3(MEKK3)isaupstreamregulatorofMAPK.Ithasthecellprotectingandapoptosisinhibitingeffect.Th

4、estudywasdesignedtoinhibittheexpressionofMEKK3gene.CombinedwithTRAIL,itisexpectedtoenhancetheapoptosisinducingeffectofTRAILonthebreastcancercelllineMCF-7.Methods:ChemicallysynthesizedsmallinterferingRNA(siRNA)targetingMEKK3wastransfectedintohumanbreast

5、cancercelllinesMCF-7byDharmaFECTTransfectionReagent,andtheexpressionofMEKK3mRNAandproteinwasdetectedbyRT-PCRandWesternblotting.TheproliferationandapoptosisofMCF-7cellswereanalyzedbyMTTandflowcytometry(FCM)aftertreatedwithMEKK3-siRNAcombinedwithTRAIL.Th

6、eeffectofcombinationofMEKK3-siRNAandTRAILonNF-kappaBactivitywasdetectedbyEMSA.Results:TRAILinhibitedtheproliferationofMCF-7cells,buttheinhibitoryeffectwasweak.TransfectionwithMEKK3-siRNAeffectivelyandtemporarilyinhibitedtheexpressionofMEKK3mRNAandprote

7、inexpression(P<0.01).ThecombinationofTRAILandMEKK3-siRNAcouldmarkedlyinhibittheproliferationofMCF-7cellscomparedwithTRAILalone(P<0.05);besides,thecombinedmethodalsoincreasedtheapoptosisrateofMCF-7cells(P<0.05).TheactivityofNF-kappaBsignalpathwaywasinhi

8、bitedbytheusingofMEKK3-siRNA.Conclusion:SilencingofMEKK3genewithsiRNAcangreatlypromoteTRAIL-inducedapoptosisofbreastcancercelllineMCF-7,whichmightbecontributedtothedown-regulationofMEKK3expressionandinhibitedtheactivationofNF-kappaBsign