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1、DC-CIK細(xì)胞對人肝癌HEPG2細(xì)胞周期、凋亡的影響項穎秦波田小利(重慶醫(yī)科大學(xué)附屬第一醫(yī)院感染科重慶400030)摘要目的探討細(xì)胞因子從肝癌患者外周血貼壁單個核細(xì)胞(peripheralbloodmononuclearcells,PBMCs)誘導(dǎo)的樹突狀細(xì)胞（dendriticcells，DCs）與殺傷（cytokineinducedkiller,CIK）細(xì)胞對人肝癌HEPG2細(xì)胞系周期、凋亡的影響。方法用肝癌患者肝癌組織裂解物（腫瘤抗原）致敏經(jīng)GM-CSF、TNF-ａ及IL-4等誘導(dǎo)該患者PBMC產(chǎn)生的DCs。用IFN-γ、
2、IL-2和人CD3單克隆抗體（humanCD3monoclonalantibody,hCD3mAb)等誘導(dǎo)該PBMC的懸浮細(xì)胞產(chǎn)生CIK細(xì)胞。用Tanswell培養(yǎng)小室將DC-CIK細(xì)胞與HEPG2細(xì)胞在同一培養(yǎng)體系內(nèi)經(jīng)該小室膜分隔培養(yǎng)24、48h。流式細(xì)胞儀檢測該HEPG2細(xì)胞的周期；AnnexinV-PI雙染法檢測其凋亡。RT-PCR、Westernblot分別檢測凋亡相關(guān)基因Bax及其編碼蛋白的表達(dá)。結(jié)果DC-CIK細(xì)胞可明顯影響人HEPG2細(xì)胞生長周期，兩者共培養(yǎng)，可將HEPG2細(xì)胞系阻滯于G1期。DC-CIK細(xì)胞能顯著誘
3、導(dǎo)HEPG2細(xì)胞的凋亡(與對照相比，其凋亡誘導(dǎo)率差異顯著，P<0.01)?；蚺c蛋白水平檢測表明，DC-CIK細(xì)胞可致HEPG2細(xì)胞系的凋亡相關(guān)基因Bax表達(dá)明顯上調(diào)。結(jié)論肝癌患者DC-CIK細(xì)胞可明顯抑制肝癌HEPG2細(xì)胞系的生長，顯著誘導(dǎo)該細(xì)胞凋亡，且該凋亡誘導(dǎo)具“時間-依賴”與“細(xì)胞數(shù)量-依賴”特性。關(guān)鍵詞DC-CIK細(xì)胞肝癌HEPG2細(xì)胞細(xì)胞周期細(xì)胞凋亡TheeffectofCIKcellscombinedwithdendriticcellsofpatientwithlivercanceronthecycleandapop
4、tosisofhumanHEPG2celllineXIANGYingQINBoTIANXiaoli(DepartmentofInfectiousDiseases,TheFirstAffiliatedHospitalofChongqingMedicalUniversity,Chongqing400030,China)Abstract:ObjectiveToexploratetheeffectofcytokineinducedkiller(CIK）cellsanddendriticcells（DCs）fromperipheralblo
5、odmononuclearcell(PBMC)ofpatientwithlivercanceronthecycleandapoptosisofhumanHEPG2celllineoflivercancer.MethodsTheadherentPBMCswereinducedbyGM-CSF、TNF-ａandIL-4inordertoproduceDCs,which9基金項目：重慶市衛(wèi)生局醫(yī)學(xué)科學(xué)技術(shù)研究項目(2010-2-301)作者簡介：項穎，男，重慶市人，主任醫(yī)師。主要從事腫瘤發(fā)病機(jī)制及其治療方面的研究。E-mail:xian
6、g519@hotmail.com通信作者：秦波，電話：15923261188，E-mail：cqqinbo@126.comweresensitizedwithantigenofautologoustissueoflivercancer.Meanwhile,CIKcellswereobtainedfromthesuspendedPBMCsinducedbyIFN-γ、IL-2andhumanCD3monoclonalantibody（hCD3mAb).TheDCs,CIKcellsandHEPG2celllinewerecultur
7、edinasamecircumstancewithaculturingcabinnamedTranswell，buttheDCs-CIKcellswereculturedatonesideofthemembraneoftheTranswell,HEPG2celllineatanothersideofthesamemembranefor24hand48hrespectively.SubsequentlythecycleofhumanHEPG2celllineculturedwithDCs-CIKcellsof24hand48hwer
8、etestedthroughflowcytometry(fcm)respectively.TheAnnexinV-PIstainingwasemployedtoanalysistheapoptosisofthesamecellline,andthe
