EBV-LMP2A肽诱导的特异性CTL对EBV阳性胃癌细胞的体外杀伤作用

目的:探讨HLA限制性EB病毒(Epstein-Barr virus,EBV)表位肽EBV-潜伏膜蛋白2A (EBV-latent membrane protein 2A,EBV-LMP2A)诱导的细胞毒性T淋巴细胞(CTL)对EBV阳性胃癌细胞的体外杀伤作用.方法:选用南京大学医学院附属肿瘤医院肿瘤中心HLA-A2阳性胃癌患者外周血单个核细胞(PBMC)、人胃腺癌细胞株(AGS)和人EBV阳性胃腺癌细胞株(AGS-EBV),通过改良的体外细胞培养技术用HLA-A2限制性的EBV-LMP2A抗原肽从人PBMC中诱导扩增出特异性CTL,采用四聚体技术和流式细胞术检测抗原肽诱导产生的特异性CTL的含量,通过FITC-PI标记流式术检测EBV-CTL对EBV阳性和EBV阴性的胃癌细胞的体外杀伤作用.结果:经改良的细胞培养技术和抗原肽双重刺激后EBV-LMP2A抗原肽可以诱导产生出高比例的抗原特异性CTLEBV-LMP2A-356特异性T细胞占CD8+T细胞的(47.1±5.2)％];EBV-CTL对EBV阳性胃癌细胞的体外杀伤作用较EBV阴性的胃癌细胞明显增强(45.1±9.3)％s(19.4±2.5)％,P＜0.05].结论:通过改良的细胞培养技术使用EBV-LMP2A抗原肽能诱导产生高比例的EBV特异性CTL,其对EBV阳性胃癌细胞有较强的特异性杀伤作用.

Cytotoxicity of specific CTL induced by EBV-LMP2A peptide on human EBV associated gastric carcinoma cell in vitro

Objective:To study the cytotoxicity of specific cytotoxic T lymphocyte (CTL) induced by EBVLPM2A peptide on human EBV associated gastric cancer cells in vitro.Methods:The peripheral blood mononuclear cells (PBMC) of HLA-A2 positive gastric cancer patients from Cancer Center of Drum Tower Hospital Affiliated to Medical School of Nanjing University,human gastric adenocarcinoma cell line (AGS) and human EBV positive adenocarcinoma cell line (AGS-EBV) were selected.By modified in vitro cell culture technique,specific CTL was amplified from human PBMCs with the induction of HLA-A2 restricted EBV-LMP2A peptide;the content of specific CTL induced by peptide was detected by the method of tetramer and Flow cytometry;the in vitro cytotoxicity of EBV-CTL on both EBV+ and EBV human gastric cancer cells was detected by FITC-PI.Results:Modified cell culture technique and EBV-LMP2A peptide can induce a high proportion of antigen specific CTL (EBV-LMP2A-356 specific T cells accounted for 47.1 ±5.2]％ of CD8+T cells);the cytotoxicity of EBV-CTL on EBV+ gastric cancer cells was significantly stronger than that of EBV gastric cancer cells (45.1 ± 9.3]％ vs 19.4± 2.5]％,P＜0.05).Conclusion:Using the modified cell culture technique,EBV-LMP2A antigen peptide can induce a high proportion of EBV specific CTL,which has high specific cytotoxicity on EBV+ gastric cancer cells.