白血病细胞来源胞外体的生物学特性及其致敏DC的抗白血病效应  

Biological properties of leukemia cell-derived exosome and anti-leukemia effects of its sensitized DCs

作　　者：姚烨[1] 王椿[2] 沈畅[2] 陈琳军[1] 邓晓辉[1] 郝思国[1] 

机构地区：[1]上海交通大学医学院附属新华医院血液科,上海200092 [2]上海交通大学医学院附属第一人民医院血液科,上海200080

出　　处：《中国肿瘤生物治疗杂志》2013年第1期13-19,共7页Chinese Journal of Cancer Biotherapy

基　　金：国家自然科学基金资助项目(No.81070432);上海市浦江人才计划资助项目(No.08PJ1407600)~~

摘　　要：目的:探索白血病细胞来源的胞外体(leukemia cell-derived exosome,LEX)的生物学特性及其致敏DC的抗白血病免疫效应。方法:超速离心分离纯化BCR-ABL阳性的白血病K562细胞来源的胞外体(LEXK562),采用免疫电镜及Westernblotting检测LEXK562中热激蛋白70(heat shock protein 70,HSP70)及BCR-ABL的表达,采用激光扫描共聚焦显微镜及流式细胞术检测LEXK562靶向结合DC的动力学,采用LDH释放法以及小鼠模型体内实验检测LEX及其致敏DC的抗白血病免疫效应。结果:与其他细胞来源的胞外体相似,K562细胞来源的胞外体LEXK562为直径50～100 nm的囊状结构,且表达K562细胞特异性的BCR-ABL和HSP70分子。LEXK562在体外可靶向结合DC,3～4 h到达高峰,并能在DC中稳定存在72 h以上。LEXK562致敏DC(DC/LEXK562)诱导的细胞毒性T淋巴细胞(cytotoxic T lymphocyte,CTL)可有效杀伤K562靶细胞,效靶比为50∶1时,其杀伤活性显著高于LEXK562诱导的CTL[(68.6±5.7)%vs(22.5±2.9)%,P<0.01];体内实验进一步证实,白血病L1210细胞来源的胞外体(LEXL1210)免疫后小鼠接种L1210细胞的成瘤率明显高于LEXL1210致敏DC(DC/LEXL1210)免疫小鼠的成瘤率[(54.17±8.33)%vs(16.67±4.18)%,P<0.05]。结论:LEX表达白血病细胞相关抗原,LEX体外可靶向结合DC,其致敏的DC能诱导更强的抗白血病免疫效应。Objective：To explore the biological properties of leukemia cell-derived exosomes （LEXs） and the anti-leukemia immunological effect of LEX-sensitized dendritic cells （DCs）. Methods： LEX from BCR-ABL positive leukemia K562 cells was separated and purified by ultracentrifugation. The expressions of heat shock protein 70 （HSP70） and BCR-ABL were investigated by immuno-electron microscopy and Western blotting. The kinetics of LEXK562 targeted binding to DCs was examined by laser scanning confocal microscopy and flow cytometry. The anti-leukemia immunological effect of LEX and its sensitized DCs was explored by LDH release assay and in vivo mouse model studies. Results： K562 cell-derived exosomes （LEXK562） had cystic structures between 50 and 100 nm diameter as the other cell-derived exosomes, and expressed K562 cell specific proteins, HSP70 and BCR-ABL. LEXK562 could target and bind DCs in vitro, and reached a plateau after 3 to 4 h of co-culturing. LEXK562 uptaken in DCs was quite stable for over 72 h. Cytotoxic T lymphocytes （CTLs） induced by LEXK562-sensitized DCs （DC/LEXK562） could effectively kill K562 target cells, and their cytotoxicity was significantly higher than that of CTLs induced by LEXK562 （／[68.6±5.7／]% vs ／[22.5±2.9／]%, P〈0.01） at an effector to target ratio of 50∶1. Furthermore, the in vivo study demonstrated that the incidence of tumor in mice incubation with leukemia L1210 cells after being immunized with L1210-derived exosomes （LEXL1210） was significantly higher than that of LEXL1210-sensitized DCs （／[54.17±8.33／]% vs ／[16.67±4.18／]%, P〈0.05）. Conclusion： LEX expresses antigens associated with leukemia cells and can target bind DCs in vitro. LEX-sensitized DCs can induce a stronger anti-leukemia immunological effect.

关 键 词：白血病细胞 K562细胞 L1210细胞 胞外体 树突状细胞 免疫治疗 

分 类 号：R733.7[医药卫生—肿瘤] R730.54[医药卫生—临床医学]