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ID:33165874
大小:8.37 MB
页数:50页
时间:2019-02-21
《parg引起的肿瘤局部免疫状态变化在大肠癌浸润转移中的作用及可能机制》由会员上传分享,免费在线阅读,更多相关内容在学术论文-天天文库。
1、重庆医科大学硕士学位论文PARG引起的肿瘤局部免疫状态变化在大肠癌浸润转移中的作用及可能机制姓名:王洁琼申请学位级别:硕士专业:病理学与病理生理学指导教师:王娅兰201205重庆医科大学硕士研究生学位论文PARG引起的肿瘤局部免疫状态变化在大肠癌浸润转移中的作用及可能机制摘要目的:探讨PARG基因沉默后引起的肿瘤局部免疫状态的变化在大肠癌浸润转移中的作用及其可能机制。方法:以未作任何处理的CT26细胞(即未转染组)和慢病毒空载体转染的CT26细胞(即空载体组)作为对照组,转染PARG.shRNA慢病毒载体的CT26细胞(即PARG基因沉默组
2、)作为实验组,将三组细胞分别在小鼠脾包膜下接种形成大肠癌肝转移模型;观察各组小鼠体内肝脏转移瘤结节及荷瘤小鼠生存时间的变化;免疫荧光双标记法分别标记各组脾脏及肝脏中的B220+DEC205+树突状细胞(B220+DEC205+DC)和CDllc+CDllb+树突状细胞(CDllc+CDllb+DC),免疫荧光单标记各组脾脏及肝脏中CD4+T细胞及CD8+T细胞,激光共聚焦显微镜观察;Westernblot法检测各组脾脏移植瘤及肝脏转移瘤中PARG、PARP和NF.v.B的表达;ELISA法检测各组小鼠血清及各组肿瘤细胞培养液上清中的IL.1
3、0、TGF一6的变化。结果:PARG基因沉默后,肝脏转移瘤结节数目及分级降低,荷瘤小鼠生存时间延长,与对照组相比差异显著(尸<0.05);脾脏及肝脏组织中B220+DEC205+DC较对照组明显降低,而CDllc+CDllb+DC重庆医科大学硕士研宄生学位论文较对照组明显增多,CD4+/CD8+比值较对照组明显增加(P<0.05);脾脏移植瘤及肝脏转移瘤中PARG、PARP和NF.KB蛋白的表达量较对照组明显降低(尸4、可抑制小鼠大肠癌CT26细胞肝转移,可能与PARG沉默后影响PARP的表达,并下调NF.KB的活性,进而影响NF.vJ3依赖性因子IL.10、TGF.6的分泌,从而最终影响DC及T细胞的增殖分化有关。关键词:PARG;DC;T细胞:免疫机能;大肠癌THEACTIONOFPARGRESULTEDINIMMUNERESPONSECHANGESOFLOCALTUMORINMETASTASISOFCOLONCARCINOMAANDITSMECHANISMABSTRACTObjective:ToinvestigatetheactionofPARGre5、sultedinimmuneresponsechangesoflocaltumorinmetastasisofcoloncarcinoma肌d“spossiblemechanism.Methods:BoththeCT26cellswithoutanytreatmentandCT26cellswhichweretreatedwithLentivirusemptyvectorservedascontrolgroupandCT26cellstransfectedwithLentivirusPARG—shRNA(ShorthairpinRNA)se6、rvedasexperimentgroup.AnimalmodelsforlivermetastasesofcoloncarcinomawereestablishedbysplenicsubcapsularinoculationoftumorcellsinBALB/cmice.Thechangesoflivermetastasescarcinomanoduleswereobservedandthealterationofsurvivaltimesofmicebometumor.ThenumbersofB220+DEC205+dendriti7、ccell(B220+DEC205+DC)andCDlc+CDllb+dendriticcell(CDllc+CDllb+DC)inthespleenandliverweremeasuredbyimmunofluorescencedoublelabelingassayCD4+TcellsandCD8+Tcellsinthespleenandliverweretestedbyilnmunofluorescencesinglestaining.Allofthesecellswereobservedby4confocallaserscanning8、microscope.TheexpressionofPARG,PARPandnuclearfactor-kappaB(NF一1cB)proteinsinspleentranspl
4、可抑制小鼠大肠癌CT26细胞肝转移,可能与PARG沉默后影响PARP的表达,并下调NF.KB的活性,进而影响NF.vJ3依赖性因子IL.10、TGF.6的分泌,从而最终影响DC及T细胞的增殖分化有关。关键词:PARG;DC;T细胞:免疫机能;大肠癌THEACTIONOFPARGRESULTEDINIMMUNERESPONSECHANGESOFLOCALTUMORINMETASTASISOFCOLONCARCINOMAANDITSMECHANISMABSTRACTObjective:ToinvestigatetheactionofPARGre
5、sultedinimmuneresponsechangesoflocaltumorinmetastasisofcoloncarcinoma肌d“spossiblemechanism.Methods:BoththeCT26cellswithoutanytreatmentandCT26cellswhichweretreatedwithLentivirusemptyvectorservedascontrolgroupandCT26cellstransfectedwithLentivirusPARG—shRNA(ShorthairpinRNA)se
6、rvedasexperimentgroup.AnimalmodelsforlivermetastasesofcoloncarcinomawereestablishedbysplenicsubcapsularinoculationoftumorcellsinBALB/cmice.Thechangesoflivermetastasescarcinomanoduleswereobservedandthealterationofsurvivaltimesofmicebometumor.ThenumbersofB220+DEC205+dendriti
7、ccell(B220+DEC205+DC)andCDlc+CDllb+dendriticcell(CDllc+CDllb+DC)inthespleenandliverweremeasuredbyimmunofluorescencedoublelabelingassayCD4+TcellsandCD8+Tcellsinthespleenandliverweretestedbyilnmunofluorescencesinglestaining.Allofthesecellswereobservedby4confocallaserscanning
8、microscope.TheexpressionofPARG,PARPandnuclearfactor-kappaB(NF一1cB)proteinsinspleentranspl
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