k562细胞survivin基因表达对淋巴细胞增殖和功能的影响

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1、K562细胞Survivin基因表达对淋巴细胞增殖和功能的影响罗颖，温贤浩，徐酉华（重庆医科大学附属儿童医院血液科，重庆400014）提摘要：目的探讨K562细胞中Survivin基因的表达对淋巴细胞增殖和功能的影响。方法将融合重组质粒pEGFP–C1–survivin和靶向survivin基因的RNAi的重组质粒pTZU6＋1–survivin分别转染K562细胞，得到K562/survivin+和K562/survivin-2种细胞，用G418筛选K562/survivin+细胞，用半定量逆转录-聚合酶链反应(RT－－PCR)方法及免疫细胞化学法检测K562/survivi

2、n+和K562/survivin-2种细胞中SurvivinmRNA及蛋白水平的表达，并与K562细胞作对照。将这3种细胞与健康人外周血单个核细胞（PBMC）混合培养（MLTC），检测混合培养体系中淋巴细胞增殖能力、NK细胞活性及培养上清中IFN-γ水平。结果K562/survivin+组的淋巴细胞增殖指数明显低于其余两组。Survivin基因的表达水平与NK细胞活性呈负相关。混合培养上清中分泌的IFN-γ，K562/survivin+组是K562组的1/4.5，是K562/survivin-组的1/8。结论高表达的Survivin可以抑制淋巴细胞的增殖、NK细胞活性及IFN-

3、γ的分泌。关键词：Survivin；淋巴细胞增殖；K562细胞；RNA干扰中图法分类号：R730.231文献标识码：ATheeffectofsurvivingeneexpressiononthelymphocyteproliferationandfunctioninK562cellsLUOYing,WENXian-hao,XUYou-hua.（DepartmentofHematology，Children＇’sHospital，ChongqingUniversityofMedicalSciences，Chongqing400014，China）Abstract：Objectiv

4、eTostudytheeffectofsurvivingeneexpressiononthelymphocyte’sproliferationandfunctioninK562cellsinvitro.MethodsThevectorpEGFP-C1withsurvivingeneandtheplasmidpTZU6+1containingshorthairpinRNAofsurvivinweretransfectedintoK562cellsrespectivelytogenerateK562/survivin+cellsandK562/survivin-cells.K562

5、/survivin+cellsweresievedbyG418.ThesurvivinmRNAandproteinlevelsinthethreekindsofcells(K562/survivin+，K562andK562/survivin-)weredetectedbysemi-quantitativeRT-PCRandimmunohistochemistrymethods.PBMCofhealthysubjectswasco-culturedwithK562/survivin+，K562andK562/survivin-cells(mixedlymphocyte-tumo

6、rcellculture,MLTC).LymphocyteproliferationinthesupernatantwasevaluatedbyMTTassay.NaturekilleractivitywasdetectedbyflowcytometryandIFN-γlevelwasdetectedbyELISAassay.ResultProliferationIndexinK562/survivin+groupwasmuchsmallerthanthatintheothertwogroups(P＜0.05).Survivingeneexpressionlevelwasofn

7、egativecorrelationwithNKactivity.IFN-γsecretionintheMLTCwithK562/survivin+groupwaslowerthanthatinK562group(1/4.5)andK562/survivin-group(1/8).ConclusionsThehighexpressionofsurvivinmayinhibitlymphocyteproliferation,naturekilleractivityandthesecretion