重组人p53腺病毒转染淋巴瘤源性树突状细胞的抗肿瘤免疫效应

目的：研究重组人p53腺病毒转染淋巴瘤源性树突状细胞的抗肿瘤免疫效应。方法：采集本科室初诊的弥漫性大B细胞淋巴瘤（DLBCL）肿大淋巴结分离单个核细胞（MNC）进行体外DC的诱导培养,分为实验组A（rAd-p53-DC）、对照组A（rAd-DC）、空白对照组A（N-DC）,同时采集患者的外周血分离单个核细胞（MNC）,进行体外DC的诱导培养,分为实验组B（rAd-p53-DC）、对照组B（rAd-DC）、空白对照组B（N-DC）,用重组人p53腺病毒（rAd-p53）转染2种来源的DCS,流式检测DCS免疫表型,用Western-blotting鉴定P53蛋白的表达,ELISA法检测上清中的细胞因子IL-12的含量,混合淋巴细胞反应（mixed lymphocyte reac-tion,MLR）测定DCS刺激同种异体淋巴细胞增殖能力,用乳酸脱氢酶（lactate dehydrogenase,LDH）释放法检测经rAd-p53转染的两种来源DCS的细胞毒性T淋巴细胞反应（CTL）。结果：DC的表型（CD1a除外）CD83、CD80、CD86和HLA-DR实验组均较对照组及空白对照组明显增高（p〈0.05）。Western-blotting可检测到实验组P53蛋白的表达。上清液中IL-12分泌水平实验组均较对照组及空白对照组明显增高（p〈0.05）。实验组具有明显的刺激自体淋巴细胞增殖的能力,且刺激能力随rAd-p53-DC与淋巴细胞比例的增加而升高。对照组及空白对照组也能刺激同种自体淋巴细胞增殖的能力,但较实验组差（p〈0.05）。对靶细胞的细胞毒作用（CTL效应）结果显示,实验组介导同种异体的淋巴细胞杀伤率显著高于对照组及空白对照组（P〈0.05）。且实验组A的CTL效应明显高于实验组B,两者之间有显著性差异（P〈0.05）。结论：rAd-p53-DC为基础的肿瘤疫苗有可能在解决淋巴瘤的MRD、DC免疫耐受等问题上发挥强大的治疗作用。

Anti-tumor Immunse Response of Dendritic Cells Derived from Lymphoma Cells Transducted with Recombinant Adenovirs Encoding Human p53

Objective：To investigate the immunological effect of modified dendritic cells（DCS） in inducing cytotoxic T cells（CTL） effect against lymphoma cells.Method：The DCS were isolated from the lymphoma node of diffuse large B cell lymphoma（DLBCL） cells and periphera blood.It was transfected with recombined adenovirus vector carrying p53 gene.The expression of DC was detected by flow cytometry.Western-blot was used to detect the expression of p53.ELISA was used to detect IL-12 level in supernatant.The mixed lymphocyte reaction（MLR） was used to detect the ability to proleferate allo-lymphocyte by DCs.The lactate dehydrogenase（LDH） release test was used to detemine the cytotoxicity of CTL.Results：The surface molecule expressions of DC（except for CD1a） CD83,CD80,CD86 and HLA-DR were significantly more high in experiment group than that in control group and blank group The secretion of IL-12 in supernatant was higher in experiment than that in control group.The autogene T lymphocyte proliferation and cytotoxic activity against the same kind of DLBL-cells increased in experiment group than that in control group and blank group（p0.05）.The ability to stimulate T lymphocyte proliferation increased with the rising of the ratio of experiment group and T lymphocyte.However,there was different for rAd-p53-DC derived from Lymph node and peripheral blood（p0.05）.Conclusion：DCs transfected with rAd-p53 can induce CTL response in vitro against lymphoma cells.