HepG2细胞外泌体冲击树突状细胞介导的抗肿瘤作用  

Stimulation of dendritic cellmediated antitumor tumor effect from HepG2 cell exosomes

作　　者：章菊 叶书来 张昌龙 周倩 ZHANG Ju;YE Shulai;ZHANG Changlong;ZHOU Qian(Department of Clinical Laboratory,the First Affiliated Hospital of University of Science and Technology of China/Anhui Provincial Hospital,Hefei 230001,Anhui,China)

机构地区：[1]中国科技大学附属第一医院/安徽省立医院检验科,安徽合肥230001

出　　处：《癌变．畸变．突变》2019年第1期29-34,共6页Carcinogenesis,Teratogenesis & Mutagenesis

基　　金：安徽省自然科学基金(1808085MH262)

摘　　要：目的:分析来源于肝癌HepG2细胞外泌体的免疫原性物质,为基于树突状细胞(DCs)的肝癌免疫治疗寻找合适的肿瘤抗原提供思路。方法:用透射电镜和Western blot方法鉴定肝癌HepG2细胞外泌体的形态和蛋白表达;用终浓度为8μg/mL的外泌体冲击DCs,流式细胞术检测DCs表面分子的变化;将外泌体冲击的DCs与T淋巴细胞共培养5 d,采用羧基荧光素二乙酸盐琥珀酰亚胺酯(CFSE)荧光染色法检测淋巴细胞增殖,Annexin-V/PI双染法检测淋巴细胞对肿瘤的杀伤效应。结果:肝癌HepG2细胞的外泌体表达CD63、CD81标志性蛋白,同时携带大量的肿瘤抗原甲胎蛋白(AFP)和热休克蛋白HSP70、HSP90,不表达细胞蛋白calnexin。与未成熟DCs组比较,外泌体冲击DCs后上调其表面分子如CD83、CD80、CD86的表达(P<0.01);且可促进T淋巴细胞增殖(P<0.01),增加T细胞介导的肿瘤特异性和非特异性杀伤效应(P<0.01)。结论:肝癌HepG2细胞外泌体可能携带大量肿瘤抗原,刺激DCs成熟,可能是基于DCs的肝癌或其他肿瘤免疫治疗潜在的抗原谱。OBJECTIVE:To investigate the liver cancer cell HepG2-dervied immunogenic substances as tumor antigens for dendritic cell-(DC)based immunotherapy of liver cancer.METHODS:Changes in morphology and protein expression of HepG2 cells were determined by transmission electron microscopy and western blot,respectively;and in surface molecules of DCs by flow cytometry using exosomes with a final concentration of 8μg/mL.In addition,exosome-pulsed DCs were co-cultured with T lymphocytes for 5 days.Proliferation of lymphocytes was detected by CSFE staining,and their killing of tumors was detected by Annexin-V/PI double staining.RESULTS:The hepatoma cell HepG2 exosomes expressed the marker proteins CD63 and CD81,carried a large number of tumor antigens AFP and heat shock proteins HSP70 and HSP90,but did not express the cellular protein calnexin.Compared with the DC-MOCK group,exosome-pulsed DCs up-regulated the expression of surface molecules such as CD83,CD80,and CD86;promoted T lymphocyte proliferation(P<0.01),and increased T cell-mediated tumor-specific and non-specific killing effects(P<0.01).CONCLUSION:Hepatoma cell HepG2 exosomes carried a large number of tumor antigens and stimulated the maturation of DCs,which may serve as a potential antigenic spectrum of DCs-based immunotherapy for liver and other cancers.

关 键 词：外泌体 肝癌 树突状细胞 HEPG2细胞 

分 类 号：R730.5[医药卫生—肿瘤]