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（LEXK562），采用免疫电镜及 Western blotting检测LEXK562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LEXK562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LEXK562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。LEXK562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。LEXK562致敏DC（DC/LEXK562）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LEXK562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（LEXL1210）免疫后小鼠接种L1210细胞的成瘤率明显高于LEXL1210致敏DC(DC/LEXL1210）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.

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）