腺病毒介导HSP70-HBsAg嵌合基因转染树突状细胞及生物学特性分析

目的 分析携带热休克蛋白70(HSP70)和乙肝表面抗原(HBsAg)嵌合基因的腺病毒表达载体感染人外周血诱导培养树突状细胞(DC)后其生物学特性的改变.方法 从正常人外周血中分离单个核细胞,经Ad-HSP70-HBsAg感染刺激后.在含细胞因子粒细胞巨噬细胞集落刺激因子(GM-CSF)、白介素-4(IL-4)、肿瘤坏死因子(TNF-α)的培养液体中体外诱导培养DC;采用荧光倒置显微镜、RT-PCR、流式细胞仪(FACS)和混合淋巴细胞反应(MLR)试验对人外周血来源的DC进行生物学特性分析.结果 Ad-HSP70-HBsAg感染后的DC能够有效表达示踪蛋白GFP和转录基因HSP70-HBsAg,病毒感染组和对照组的DC在细胞形态、DC细胞表面分子表达和刺激同种异体的人外周血幼稚型T细胞增殖能力的方面无明显差别.结论 Ad-HSP70-HBsAg可有效转染DC,感染后的病毒对DC的生长和生物学特性无显著影响.

The infection of dendritic cells by recombinant adenoviral vector carrying HBsAg-HSPT0 chimeric gene and Its biological characteristics observations

Objective To test the infeciton efficiency of recombinant adenovixal vector carrying HBsAg-HSP70 chimeric gene and to abserve its biological characteristics. Methods Peripheral blood mononuclear cells (PBMC) were separated from healthy blood donor and they were infected by Ad-HSP70-HBsAg on the first day of isolation. DCs were induced in medium with cytokines IL-4, GM-CSF and TNF-α in vitro. The biological characteristics of DC induced were analyzed by inverted fluorecent microscope, RT-PCR, flow cytometer(FACS), and mixed lymphocyte reaction (MLR). Results The traced gene-GFP were abserved in DCs by inverted fluorecent microscope and HSP70-HBsAg gene mRNA expression was detected by RT-PCR after the Ad-HSP70-HBsAg infection. FACS analysis shown that the expression of CD1a,CDSO,CD86 and HLA-DR on surfece of two groups of DCs were similar. MLR showed that there are not a statitie difference of stimulated index (SI) between two groups.Conclusion Results indicated that Ad-HSP70-HBsAg can effectively infected DCs without affecting its biological characteristics.