CpG-ODN2216致敏的单核细胞培养上清对HBV相关性肝癌病人的树突状细胞表型和功能的影响

HBcAg-Pulsed Monocyte-Derived Dendritic Cells from HBV-Associated Hepatocellular Carcinoma Patients Treated with Supernant from CPG-ODN2216-Pulsed PBMC Significantly Enhance HBV Specific T-Cells Response in vitro

目的:研究CpG-ODN2216致敏的外周血单核细胞(PBMC)培养上清液对HBV相关性肝癌病人的树突状细胞(DC)成熟与功能的影响,寻求一种增强DC疫苗效果的方法。方法:从9例HBV相关性肝癌患者PBMC中诱导出未成熟的单核细胞来源的DC(MoDC),经HBV核心抗原(HBcAg)负载后,用CpG-ODN2216刺激的PBMC上清液、“细胞因子鸡尾酒(IL-1β、IL-6、TNF-α和PGE2)”以及两者的联合作用促进MoDC的进一步成熟,检测MoDC表型和功能;选择其中5例HLA-A2+病人,用成熟MoDC诱导自身T细胞产生HBV特异性CD8+的细胞毒性T淋巴细胞(CTL)。结果:用细胞因子鸡尾酒和CpG-ODN2216刺激的PBMC上清液联合作用可明显增强MoDC表面的CD80、CD83和CD1a表达,其对HBcAg负载的MoDC促成熟作用大于单独用细胞因子鸡尾酒或单独用CpG-ODN2216刺激PBMC的上清液。联合作用促进MoDC分泌IL-12和IL-10的能力明显强于单独应用PBMC上清液或细胞因子鸡尾酒,其刺激自体T细胞分泌IFN-γ、TNF-α、IL-6的能力也明显增高。联合作用促成熟的MoDC诱导HLA-A2+病人的自体T细胞产生HBVcore18-27特异性CD8+CTL的频率明显高于细胞因子鸡尾酒单独促成熟的MoDC。结论:CpG-ODN2216刺激PBMC的上清液和细胞因子鸡尾酒联合作用可以明显促进HBcAg负载的HBV相关性肝癌病人的MoDC成熟,增强MoDC分泌细胞因子、刺激自体特异性T淋巴细胞应答、诱导HBV特异性细胞毒性T细胞的能力。为提高HBV特异性树突状细胞疫苗的效果提供了一种可行方案。

Objective： To investigate the roles of supernant from CpG-ODN2216-pulseded peripheral blood mononuclear celIs（PBMC） potentiating HBcAg-pulsed dendritic cells（DC） from HBV-associated hepatocellular carcinoma（HCC） patients maturation and function. Methods： Monocyte-derived DC（MoDC） from 9 HBV-associated HCC patients were pulsed with HBcAg, and promoted maturation with ＂cytokine cocktails＂ composed of TNF-α, IL-6, IL-1β, PGE2 only（G1）, supernatant of PBMC stimulated with CpG-ODN2216（G2）, and cytokine cocktails combined with supernatant of PBMC stimulated with CpG-ODN2216（G3）. Phenotype and function of mature MoDC were characterized. HBV-specific CD8^＋ T cells were induced by mature MoDC in autologous T cells from 5 cases of HLA-A2^＋ patients. Results： CD80/CD83/CDla expressions on HBcAg-pulsed MoDC were significantly higher in G3 than those in G1 and G2. The levels of IL-12 and IL-10 produced by mature HBcAg-pulsed MoDC in G3 were higher than those in G1 and G2, and same pattern was in IFN-γ/TNF-α/ IL-6 secretion by autologous T cells co-cultured with MoDC. HBV core18-27-epitope specific CD8^＋ T cells could be induced in 3 cases of 5 patients in vitro, and the frequencies of specific CD8^＋ T cells induced by mature MoDC in G3 were higher than those in G1. Conclusion： cytokine cocktails combined with supernatant of PBMC stimulated with CpGODN2216 can significantly promote HBcAg-pulsed MoDC maturation and cytokines secretion, and capacity of autologous T cells stimulation, inducing HBV-specific CD8^＋ T cells in HBV-associated HCC patients in vitro, and may be a feasible method to improve the HBV-specific DC vaccine.