转染HPV16E6基因人树突状细胞疫苗的制备及生物学特性

Vaccine preparation of dendritic cell transfected with HPV16E6 antigen gene and its biological characteristics

目的制备来源于人外周血并转染HPV16E6基因的树突状细胞(DC)疫苗,检测其细胞形态、分子表型及诱导的免疫效应。方法细胞因子扩增人外周血DC,Lipofectamine转染HPV16E6制备DC疫苗。动态形态学观察,免疫细胞化学及流式细胞术检测分子表达,体外诱导并测定CTL活性。结果转染DC呈形态迥异的多突起状,其E6蛋白、CD80、CD86和CD83分子的表达率依次为47.3%、82.5%、79.8%和85.7%,诱导杀伤Caski细胞的活性明显高于对照组(P<0.01)。结论转染DC疫苗保持了功能成熟DC的形态特征,且内源性表达E6蛋白,能诱导高效的特异性抗肿瘤免疫应答。

Objective：To prepare the vaccine of DC derived from human peripheral blood and transfected with HPV16E6 antigen gene, and to detect its morphological character,surface marker and immunological effect. Methods：DC-enriched populations were prepared from human peripheral blood mononuclear cell（PBMC） with the combination of rhGM-CSF,rhlL-4 and rhTNF-α. The plasmid containing HPV16E6 gene was transfected into DC with lipofectamine. The morphology of DC was observed dynamically, and the expression of surface markers of DC vaccine could be detected using immuno-cytochemical staining and flow cytometry. MTT assay was applied to detect the activity of CTL in vitro. Results：Tile transfected DC had typical morphologic and phenotypic characteristics, and expressed E6 protein 47.3% , CD80 82.5% , CD86 79. 8% and CD83 85.7%. The killing activities of CTL to Caski cells induced by transfeeted DC were higher evidently than that of control groups（ P 〈 0. 01 ）. Conclusion： DC transfected with HPV16E6 gene can sustain the morphology and phenotype of mature DC, endogenously express E6 protein, and induce specific antitumor immune CTL response in vitro.