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目的:研究树突状细胞负载EB病毒潜伏膜蛋白(latent membrane proteins,LMPs)介导生成的LMPs特异性细胞毒性T淋巴细胞(cytotoxic T lymphocyte,CTL)LMPs-CTL的生物学特性,观察其对LMPs阳性鼻咽癌细胞SUNE的杀伤作用。方法:用淋巴细胞分离液分离人外周血单核细胞,采用贴壁分离及白细胞介素(interleukin,IL)-4、粒细胞-巨噬细胞集落刺激因子(granulocyte-macrophage colony stimulating factor,GM-CSF)等细胞因子诱导成熟树突状细胞(dendritic cell,DC),通过DC细胞负载LMPs多肽抗原并递呈给同源T淋巴细胞,制备LMPs-CTL。CCK8法检测LMPs-CTL对SUNE细胞的杀伤作用;ELISA和羟基荧光素二醋酸盐琥珀酰亚胺脂(carboxy fluoroscein succinimidyl ester,CFSE)染色法检测LMPs-CTL分泌IFN-γ及其增殖。结果:LMPsCTL对SUNE细胞的杀伤率在12 h和24 h分别为(43.47±1.93)%和(77.15±3.18)%,显著高于对照组的(11.45±3.06)%和(24.27±13.20)%(P<0.05)。SUNE细胞刺激后,LMPs-CTL增殖能力较对照组有明显增强,IFN-γ分泌量达到(613.40±121.77)pg/ml,显著高于对照组(86.90±3.70)pg/ml(P<0.05)。结论:通过DC细胞负载LMPs混合多肽可诱导生成LMPsCTL,对LMPs阳性鼻咽癌细胞有较强的杀伤作用。
Objective: To investigate the biological characteristics of LMPs-specific cytotoxic T lymphocyte (CTL) LMPs-CTL induced by dendritic cells loaded with Epstein-Barr virus latent membrane proteins (LMPs) Cytotoxicity of SUNE in LMPs positive nasopharyngeal carcinoma cells. Methods: Human peripheral blood mononuclear cells (PBMCs) were isolated by lymphocyte separation fluid. Adherent cells were treated with adherent cells and granulocyte-macrophage colony stimulating factor (GM- CSF) to induce mature dendritic cells (DCs). LMPs-CTLs were prepared by DCs loaded with LMPs polypeptide antigen and presented to homologous T lymphocytes. The cytotoxicity of LMPs-CTL on SUNE cells was detected by CCK8 assay. IFN-γ and its proliferation were detected by ELISA and carboxyfluorescein succinimidyl ester (CFSE) staining. RESULTS: The killing rates of SUNMP cells on LMPsCTL were (43.47 ± 1.93)% and (77.15 ± 3.18)% at 12 and 24 h, respectively, which were significantly higher than those in control group (11.45 ± 3.06) and (24.27 ± 13.20)% (P <0.05). After stimulated by SUNE cells, the proliferation of LMPs-CTL was significantly enhanced compared with the control group, and the secretion of IFN-γ was (613.40 ± 121.77) pg / ml, significantly higher than that of the control group (86.90 ± 3.70) pg / . CONCLUSION: LMPsCTL can be induced to produce LMPs by loading DCs with LMPs, which can kill LMPs-positive nasopharyngeal carcinoma cells.