摘要
目的探讨IL-12基因修饰树突状细胞(DC)体外诱导免疫杀伤肝癌细胞的效能及其机制。方法IL-12基因修饰肝癌病人外周血DC(DC-IL-12),ELISA法检测DC-IL-12培养上清中IL-12和IFN-γ水平,以冻融HepG2所获得的肿瘤相关抗原(TAA)刺激DC-IL-12,MTT法检测TAA负载的DC-IL-12刺激同源T淋巴细胞增殖分化能力,细胞毒试验检测DC-IL-12诱导的细胞毒T淋巴细胞(CTL)及其上清液对HepG2肝癌细胞株的杀伤作用,ELISA法检测CTL上清液IFN-γ水平。结果DC经IL-12基因修饰后48h分泌高水平IL-12(24·35±5·4)pg/ml及IFN-γ(725±30)pg/ml,均显著高于未转染DC组(P<0·01)。DC-IL-12诱导的CTL及其上清液对HepG2均有显著杀伤作用,杀伤率显著高于未转染DC组,分别为(82·43±8·7)%vs(57·4±4·3)%和(76·45±8·5)%vs(18·23±5·3)%(P<0·01),DC-IL-12诱导的CTL上清液IFN-γ水平显著高于未转染DC组,分别为(1125·0±32·7)pg/ml、(281·3±14·7)pg/ml(P<0·01)。结论IL-12基因修饰增强DC体外诱导自体T淋巴细胞产生特异性抗肝癌免疫,其机制与IL-12基因修饰促进DC分泌IL-12、增强T淋巴细胞活化及分泌IFN-γ密切相关。
Objective To investigate the efficacy and mechanism of IL-12 gene modified DC in inducing specific anti-tumoral immunity against hepatocellular carcinoma (HCC) in vitro. Methods DC isolated from the peripheral blood of HCC patients was transfected by recombinant IL-12 retrovirus (DC-IL-12). IL-12 and IFN-γ levels in culture supernatant of DC-IL-12 were determined by ELISA. DC-IL-12 was then pulsed with tumor associated antigen (TAA) obtained from cooling and thawing of HepG2 cells. The capacity of TAAS-pulsed DC-IL-12 in inducing proliferation and differentiation of Autologous T lymphocytes was evaluated by MTT test. The killing efficacy of cytotoxic T lymphocytes (CTL) induced by DC-IL-12 and its culture supernatant against HepG2 was assessed. IFN-γ level in culture supernatant of CTL was determined by ELISA. Results Forty-eight hours after IL-12 gene transfection, DC IL 12 produced markedly higher levels of IL 12 and IFN-γ (24.35±5.4 pg/ml and 725±30 pg/ml) as compared with DC (P〈0.01). CTL induced by DC-IL-12 and its supernatant had a remarkably greater killing efficacy to HepG2 as compared with DC (82.43±8.7% vs. 57.4±4.3%, 76.45±8.5% vs. 18.23±5.3%, P〈0.01). IFN-γ level in culture supernatant of CTL in duced by DC-IL 12 was significantly higher than that by DC (1125.0±32.7pg/ml vs. 281.3±14.7 pg/ml, P〈0.01). Conclusions IL-12 gene modification can enhance the capacity of DC in inducing autologous T lymphocytes to generate specific anti tumoral immunity against HCC. The possible mechanism of which might be related to the increased secretion of IL 12 and INF-γ and enhanced in duction of CTL.
出处
《中华肝胆外科杂志》
CAS
CSCD
2005年第12期824-827,共4页
Chinese Journal of Hepatobiliary Surgery
基金
国家自然科学基金(No.30100180)
广东省自然科学基金(No.001344)资助项目
关键词
癌
肝细胞
树突状细胞
IL-12
基因修饰
Carcinoma, hepatocellular
Dendritic cell
IL-12
Gene Modification