乙型肝炎病毒C基因G87变异对宿主细胞HLA-I表达的影响

Impact of HBV/C gene mutation G87 on HIA-I expression of host cells

目的 探讨HBVC基因G87变异对宿主细胞表面HLA I表达的影响。方法 采用定点突变技术和亚克隆技术 ,将HBV野生型基因组质粒构建为C基因G87变异株表达载体 (EBO G87)和野生株表达载体 (EBO WT) ,以脂质体技术分别转染HepG2细胞 ,转染细胞经FITC标记的鼠抗HLA ABCmAb染色 ,流式细胞术测定细胞表面HLA I的表达。结果 构建的EBO G87和EBO WT重组载体经内切酶双酶切鉴定及核苷酸序列测定证实。HepG2细胞表面HLA I表达的平均荧光强度 ,EBO空载体转染的细胞为 2 3,EBO WT转染的细胞明显增强至 18 8,EBO G87转染的细胞增至 10 5。结论 2株HBV上调HLA I表达强度不同 ,C基因G87变异可使宿主细胞表面的HLA

Objective To study the impact of HBV/C gene G87 mutation on HLA-I expression of host cells. Methods HBV wild-type genome plasmid was reconstructed by site-directed mutagenesis technique and subcloning technique into expression vector of G87 mutant (EBO-G87) and expression vector of wild type (EBO-WT), which were transfected into HepG2 cells via liposome technique,respectively.The transfected cells were stained with murine mAb anti-HLA-ABC conjugated directly to FITC, and HLA-I expression on their membranes was analyzed by flow cytometry. Results The constructed vectors of EBO-WT and EBO-G87 were identified by restriction enzyme digestion and nucleotide sequencing. The mean fluorescence intensity of HLA-I expression on transfected cells with EBO empty vector was at low level(2.3). It was elevated remarkably to 18.8 by EBO-WT, while that of EBO-G87 was increased to 10.5. Conclusion Two strains of HBV may up-regulate the expression of HLA-I, and G87 mutation of C gene may affect the expression level of HLA-I on host cells.