HSP70-TKD诱导的NK细胞杀伤肝癌细胞的实验研究  

Cytolytic activity against hepatocellular carcinoma cell of natural killer cells induced by heat shock protein 70 epitope TKD

作　　者：陈蓉明[1] 郑秋红[1] 应敏刚[1] 龚福生[1] 汪相如[1] 

机构地区：[1]福建医科大学教学医院福建省肿瘤医院肿瘤研究所肿瘤分子研究室,福建福州350014

出　　处：《中华肿瘤防治杂志》2008年第15期1129-1132,共4页Chinese Journal of Cancer Prevention and Treatment

基　　金：福建省自然基金项目(C0410043);福建省科技计划重点项目(2007Y0014)

摘　　要：目的:探讨从人外周血单个核细胞(peripheral blood mononuclear cell,PBMC)中获得足够数量和高细胞毒活性的CD3-CD56+NK细胞的方法,并观察其对肝癌细胞HepG2杀伤效应。方法:以干细胞培养基(SCGM)为基础培养基,在不同浓度的抗CD3单抗、IL-2和PHA的培养条件下,从健康人PBMC中高效扩增获得CD3-CD56+NK细胞;以不同剂量的HSP70-TKD诱导NK细胞的活性,MTT法测定HSP70-TKD诱导的NK细胞对HepG2和榄香烯处理的HepG2细胞的杀伤活性。结果:建立了从人PBMC体外扩增CD3-CD56+NK细胞体系:以SCGM为基础培养基,在600U/mLIL-2、500μg/L抗CD3单抗和50mg/LPHA联合作用下,培养14d细胞扩增了116倍,含有(66.97±8.76)%CD3-CD56+NK细胞;HSP70-TKD诱导的NK细胞对HepG2杀伤活性低,与未经TKD诱导的NK细胞之间差异无统计学意义,P=0.298;而对榄香烯处理过的HepG2杀伤活性高,诱导组与未诱导组之间差异有统计学意义,P=0.008;当TKD浓度为2.0mg/L时,杀伤活性最高为(58.8±3.4)%。结论:以干细胞生长培养基为基础培养基,在抗CD3单抗和IL-2协同刺激下体外大量扩增NK细胞,HSP70-TKD诱导的NK细胞对细胞膜HSP70阳性的肿瘤细胞杀伤活性高,而对于细胞膜HSP70低表达的肿瘤细胞杀伤活性低。OBJECTIVE： To study the method of expanding enough and high eytolytie activity of CD3^- CD56^＋ natural killer （NK） cells from human peripheral blood mononuelear cells （PBMCs） and explore the eytotoxieity of NK cells against hepatoeellular carcinoma cell HepG2. METHODS： PBMCs of health adults were expanded in stem growth medium （SCGM） supplemented with anti-CD3 monoelonal antiboby, IL-2 with different concentrations and PHA . The eytolytie activity of the expanded NK cells was stimulated by HSP70-TKD at varying concentrations, and the eytotoxle activity of induced NK cells against hepatoeellular carcinoma cell HepG2 and HepG2 treated with elemene was analysed by MTT assay. RESULT： The system of expanding CD3^- CD56^＋ NK cell from PBMCs was established： PBMC expanded to 116 folds in the 14th day cultivated in SCGM supplemented with 600 U/mL IL-2, 500 μg/L anti-CD3 monoclonal antiboby and 50 mg/L PHA, and contained （66.97±8.76）% CD3^- CD56^＋ NK cells. HSP70-TKD stimulated NK cells had low cytolytic activity against HepG2 cells, and no statistical difference was observed between NK cell induced by HSP70-TKD and NK cell which was not induced by TKD, P= 0. 298. But TKD stimulated NK cell had high cytolytic against HepG2 treated with elemene, and statistical difference was observed between NK cells induced with HSP70-TKD and NK cell which were not induced by TKD, P= 0.008. The cytotoxicity was highest （58.8±3.4）% when TKD was 2.0 mg/L. CONCLUSIONS： NK cells could be expanded from PBMCs by using SCGM with anti-CD3 monoclonal antiboby and IL-2. HSP70-TKD actived natural killer cells have a high cytolytic activity against HSP70 membrane-positive tumor cells but a low cytolytic activity against HSP70 membrane-negative tumor cells.

关 键 词：肝肿瘤 肿瘤细胞 培养的 杀伤细胞 天然 

分 类 号：R735.7[医药卫生—肿瘤]