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目的:研究肿瘤冻融抗原及弱酸洗脱提取的肿瘤抗原肽分别冲击致敏树突状细胞(dendritic cells,DCs)体外诱导前列腺癌特异性细胞毒性T淋巴细胞(CTL)的效应。方法:采用反复冻融法和弱酸洗脱法,分别获取前列腺癌细胞株PC-3的肿瘤抗原,应用重组人GM-CSF、IL-4体外培养诱导外周血单个核细胞获得DC,并分别负载两种前列腺癌肿瘤抗原,检测其体外激活肿瘤特异性CTL的能力,用LDH释放法检测其对PC-3的体外杀伤效应。结果:用枸橼酸-磷酸盐缓冲液洗脱或反复冻融PC-3(2×107)后获得的肿瘤抗原蛋白含量分别为(312.2±7.9)μg和(963.0±25.3)μg,负载两种前列腺癌细胞抗原的DCs致敏的CTL对PC-3细胞有明显的杀伤作用,而且负载弱酸洗脱提取的肿瘤抗原肽的DCs致敏的特异性CTL具有更明显的杀伤作用[(60.4±5.52)%vs(43.7±4.11)%,P<0.01)]。结论:弱酸洗脱和反复冻融方法都可以有效获取前列腺癌抗原,用于体外致敏DCs后,均具有活化CTL的能力,而且弱酸洗脱提取的肿瘤抗原肽更为有效。
OBJECTIVE: To study the effect of tumor freeze-thaw antigen and weakly acid-eluting tumor antigen peptide on the in vitro induction of prostate cancer-specific cytotoxic T lymphocytes (CTLs) by dendritic cells (DCs). Methods: Tumor antigens of prostate cancer cell line PC-3 were obtained by repeated freeze thawing and weak acid elution. DCs were induced from peripheral blood mononuclear cells by recombinant human GM-CSF and IL-4 in vitro. Two kinds of prostate cancer tumor antigens were tested for their capability of activating tumor-specific CTLs in vitro. LDH release assay was used to detect the killing effect of PC-3 in vitro. Results: The tumor antigen protein content obtained after citrate-phosphate buffer elution or repeated freeze-thawing of PC-3 (2 × 107) was (312.2 ± 7.9) μg and (963.0 ± 25.3) μg, respectively DCs sensitized with prostate cancer cell antigen showed obvious killing effect on PC-3 cells, and DCs sensitized with tumor antigen peptides extracted with weak acid elution had a more obvious killing effect [(60.4 ± 5.52)% vs (43.7 ± 4.11)%, P <0.01)]. CONCLUSION: Both weak acid elution and repeated freezing and thawing methods can effectively obtain prostate cancer antigens, which have the ability to activate CTL after in vitro sensitization of DCs. Moreover, the tumor antigen peptide extracted by weak acid elution is more effective.