负载肿瘤抗原的DC疫苗体外诱导的特异性抗膀胱癌效应研究

Study on the specific anti-bladder cancer effect of DC vaccine loaded with tumor antigen in vitro

目的探讨负载膀胱癌抗原成分树突状细胞(dendritic cells,DC)疫苗的制备和体外诱导T淋巴细胞特异性杀伤膀胱癌细胞的作用。方法冻融法制备EJ细胞裂解物抗原成分,体外培养的人外周血单个核细胞(hu-PBMC)在rhGM-CSF、rhIL-4、TNF-α诱导下分化出DC,负载EJ细胞裂解物抗原后制备膀胱癌DC疫苗;免疫磁珠分离法从人免疫重建Balb/c裸小鼠脾脏组织中分离CD3+T淋巴细胞,3H-TdR掺入试验测定DC疫苗刺激自体T淋巴细胞增殖的能力,51Cr释放试验检测DC疫苗诱导的T细胞对EJ细胞的杀伤作用。结果 Hu-PBMC在细胞因子rhGM-CSF、rhIL-4和TNF-α的刺激下分化为成熟DC,负载EJ抗原的DC疫苗体外可使同源T淋巴细胞活化,增殖指数增加,活化的T淋巴细胞对EJ细胞的杀伤率为(62.58±6.13)%,和对照组比较差异有统计学意义(P<0.05)。结论负载膀胱癌冻融抗原的DC疫苗体外可诱导人T淋巴细胞活化增殖,对EJ细胞有明显的杀伤作用。

Objective To investigate the way to prepare dendritic cells（DC） vaccine loaded with bladder cancer antigen and the activation of homologous T lymphocytes in vitro,to observe the specific killing effect of bladder cancer cells induced by T lymphocytes.Methods EJ cell lysate antigen components were obtained by freeze-thaw method.The DC were induced by rhGM-CSF,rhIL-4,TNF-α from human peripheral blood mononuclear cells（hu-PBMC） cultured in vitro.EJ cell lysate antigen pulsed DC.DC vaccine loaded with EJ cell lysate antigen was generated.CD3＋ T lymphocytes in spleen tissue in Balb/c-nu mouse with reconstituted human immune system were isolated by T-immunomagnetic beads.The proliferation ability of autologous T cells and T lymphocytes＇ killing activity towards EJ cells induced by DC vaccine were detected using 3H-TdR and 51Cr release assay.Results The DC were induced by rhGM-CSF,rhIL-4 from hu-PBMC.TNF-α promoted immature DC to mature DC.T lymphocytes were activated by DC vaccine loaded with EJ cell lysate antigen.The proliferation index was higher,and the killing rate of lymphocytes activated by DC vaccine against EJ cells was（62.58±6.13）%.Compared with the control group,these differences were statistically significant（P〈0.05）.Conclusions DC loaded with freeze-thawed bladder cancer antigen can induce the proliferation of autologous T cells in vitro,and have obviously killing effect on EJ cells.