目的： 研究肿瘤抗原负载的树突状细胞（DCs）对原发性肝癌肿瘤浸润淋巴细胞(TILs)体外增殖和抗瘤作用的影响。 方法： 将自体肿瘤细胞匀浆粗提物(Tuly)与原发性肝癌患者外周血来源的DCs共培养以制备肿瘤抗原负载的DC(DC-Tuly)，将DC-Tuly与自体TILs在低浓度IL-2(100 U/ml)中共培养，用FCM测定DC-Tuly 的表型以及与DC-Tuly混合培养前后的TILs表型，并计数TILs。用LDH法测定TILs的细胞毒作用。ELISA法检测TILs培养上清中IFN-γ和TNF-α的含量。 结果： DC-Tuly表面CD1a，CD83，CD80，CD86，CD40和HLA－DR分子表达水平增高（P<0.05）；DC-Tuly明显诱导TILs的增殖(P<0.01)及对自体肝癌细胞的细胞毒作用 (P<0.01), CD3+TILs增多（P<0.05）， CD4+TILs比例较混合前升高(P<001)，但仍以CD8+TILs为主。与DC-Tuly混合培养后第1,2天，TILs分泌IFN-γ和TNF-α的量明显提高(P<0.01)。 结论： 负载肿瘤抗原的DCs可促进TILs的增殖，增强TILs的特异性抗肿瘤活性。

Objective:To investigate the effects of dendritic cells(DCs) loaded with tumor antigens on the proliferation and antitumor activity of tumor-infiltrating lymphocytes(TILs) from hepatocellular carcinoma (HCC) in vitro. Methods:DCs derived from the peripheral blood of patients with HCC were loaded with autologous tumor lysate(Tuly) to generate DC-Tuly and then the DC-Tuly was co-cultured with autologous TILs in low concentration of IL-2. TILs were counted, and the phenotypes of DCs and TILs were assayed by FCM and the cytotoxicity of TILs was determined with LDH method. The concentration of cytokines in TILs culture supernatant such as IFN-γ and TNF-α was detected by ELISA . Results:The expression levels of CD1a, CD80, CD83,CD86, CD40, HLA-DR molecules on the DC-Tuly are notably increased（P<005）. The proliferation of TILs which were induced by DC-Tuly was markedly enhanced (P<0.01) and the cytotoxicity of the TILs against autologous tumor cells was remarkably increased (P<0.01). After co-cultured with DC-Tuly, CD3+TILs were increased（P<0.05）and the percentage of CD4+ TILs was enhanced(P<0.01), however, the percentage of CD8+ TILs was still higher than that of CD4+ TILs. Both concentrations of IFN-γ and TNF-α in the culture supernatant of the TILs distinctly increased on the 1st and 2nd days of co-culturing (P<0.01). Conclusion: DCs loaded with tumor antigens can obviously promote the proliferation and the specific antitumor activity of TILs.

江苏省卫生厅资助课题(H9923)