目的: 探讨exosomes(Exo)联合卡介苗（bacillus CalmetteGuérin vaccine,BCG）的体内抗肿瘤效应。方法：通过密度梯度离心法分离和纯化E.G7OVA肿瘤细胞来源的Exo,Western blotting检测其蛋白成分。分别以Exo、BCG 、Exo+BCG、PBS免疫小鼠，以E.G7OVA细胞攻击，观察各组免疫保护效应；建立E.G7OVA荷瘤小鼠模型，观察各组免疫治疗效应。LDH法检测4组免疫小鼠脾细胞CTL细胞毒性。结果：Western blotting检测显示，Exo含有HSP60、OVA、HSC70和CD63分子；免疫保护实验结果显示，Exo+BCG组免疫小鼠90 d无瘤率显著高于Exo组和BCG组（60% vs 20%、0%，P<0.01）；免疫治疗实验结果显示，Exo+BCG对小鼠移植瘤的抑制显著强于Exo组和BCG组（P<0.01）。CTL检测结果显示， Exo+BCG免疫小鼠的CTL细胞特异性杀伤E.G7OVA细胞的活性显著高于其他各组（P<0.01）。结论： BCG作为免疫佐剂能显著增强exosomes的体内抗肿瘤效应。

objective: To study the in vivo antitumor effect of exosomes (Exo) combined with bacillus CalmetteGuérin vaccine（BCG）. Methods:Exo was isolated and purified from culture supernatant of E.G7OVA tumor cells by density gradient centrifugation. Protein components of Exo were detected by Western blotting. Exo, BCG, Exo combined with BCG (Exo+BCG) or PBS were preinjected into mice before injection of E.G7OVA cells, and the antitumor effects were observed in each group. Mouse model bearing E.G7OVA cells was established to examine the immunotherapy effects of Exo with or without BCG. Cytotoxity of spleen CTL was measured by LDH in different groups. Results: Exo derived from E.G7OVA cells contained HSP60, OVA, HSC70 and CD63 as detected by Western blotting. Tumorfree rate at 90 d was significantly higher in Exo+BCG vaccinated mice than those in Exo or BCG vaccinated mice as measured by immunoprotective assay (60% vs 20% or 0%，P<0.01). Immunotherapy assay showed that tumor inhibitory effect in Exo+BCG group was significantly higher than those in Exo or BCG groups (P<0.01). CTL results showed that CTL of Exo+BCG vaccinated mice had significantly enhanced ability to specifically kill target E.G7OVA cells compared with those of Exo and BCG groups (P<0.01). Conclusion: BCG as an immunoadjuvant can significantly enhance the antitumor effect of exosomes in vivo.

浙江省医药卫生科研基金资助项目(No.2008B023)