细胞因子IL-7与IL-15联合优化胃癌EBV特异性CTL细胞培养体系

摘要

Objective:To investigate the advantages of cytokine IL-7 and IL-15 in the development of gastric cancer EBV antigen-specific CTLs.Methods:DCs were generated from monocytes enriched by adherence for 2 h, and cultured in medium containing human GM-CSF and IL-4.Mature DCs were pulsed by EBV antigen peptide and then incubated with T cells to induce EBV-CTLs.To compare the amplification ability,phenotype,cellular im-mune response ability and in vitro killing ability of EBV-CTLs induced by different cytokine culture conditions.Re-sults:Compared with IL-2,IL-7/15 expanded EBV - CTLs populations contained more CD3 +T cells [(91.2 ± 1.5)% vs (80.8 ± 1.6)%,P＝0.008 4],meanwhile,IL-7/15 expanded EBV-CTLs populations contained more CD3 +CD8 +T cells as compared with IL-2[(66.8 ± 2.3)% vs (30.17 ± 6.9)%,P＝0.007 6].Mainte-nance of central memory T cell expansion,we observed higher proportion of CD8 +CD62L+T cell in culture with IL-7/15[(26.4 ± 2.3)% vs (9.6 ± 1.4)%,P＝0.003 6].Besides,we found the IL-7 combination with IL-15 led to the greater expansion of CD8 +CD27 +T cell[(38.5 ± 3.7)% vs (16.7 ± 2.9)%,P＝0.01].We found in cul-ture with IL-7/15 led to more IFN-γ secretion and CD137 expression.EBV-CTLs cultured under IL-7 combined with IL-15 showed a stronger lysis effect than other groups.Conclusion:In this study,IL-7 combined with IL-15 promotes the expansion of EBV-CTLs with more CD3 +CD8 +T cells,enhanced immune response,and superior cyto-toxic effector characteristics these results provide an objective experimental basis for the further individualized treat-ment of EBV positive gastric cancer.%目的:探讨细胞因子白细胞介素-7(IL-7)和IL-15在胃癌EB病毒(EBV)抗原特异性细胞毒性T细胞(CTLs)培养中较IL-2的优势.方法:提取胃癌患者外周血单个核细胞贴壁培养2 h,贴壁细胞加IL-4和粒细胞-巨噬细胞集落刺激因子(GM-CSF)诱导树突状细胞(DC),成熟的DC负载EBV抗原肽与T细 胞混合培养,诱导EBV-CTLs,比较在细胞因子IL-2或IL-7/15培养条件下诱导的EBV-CTLs的扩增能力、表型、免疫应答及体外杀伤能力的差别.结果:与细胞因子IL-2相比较,IL-7联合IL-15培养的EBV-CTLs细胞中含有较高比例的CD3 +T细胞[(91.2 ± 1.5)% vs (80.8 ± 1.6)%,P＝0.008 4]以及较高比例的CD3 +CD8 +T细胞[(66.8 ± 2.3)% vs (30.17 ± 6.9)%,P＝0.007 6];在维持中央记忆性T细胞扩增方面,IL-7联合IL-15扩增的EBV-CTLs细胞中含有较高比例的CD8 +CD62L+T细胞[(26.4 ± 2.3)% vs (9.6 ± 1.4)%,P＝0.003 6]及CD8 +CD27 +T细胞[(38.5 ± 3.7)% vs (16.7 ± 2.9)%,P＝0.01];在免疫应答能力方面,接触DC负载抗原肽24 h后,IL-7联合IL-15培养的EBV-CTLs能分泌更多的γ干扰素(IFN-γ),且CD137上调明显高于IL-2组;在体外杀伤实验中,IL-7联合IL-15诱导的EBV-CTLs对表达EBV抗原的靶细胞表现出更强的杀伤作用.结论:IL-7联合IL-15在扩增胃癌抗原特异性EBV-CTLs方面具有优势,扩增的细胞中含有较高比例的CD3 +CD8 +T细胞,且能维持中央记忆性T细胞的扩增,表现出有更强的免疫应答和体外抗肿瘤效果,为其进一步临床个体化治疗 EBV阳性胃癌提供客观实验依据.