乙型肝炎病毒C蛋白对NK细胞功能影响的体外研究

Effect of Hepatitis B virus C protein on function of natural killer cell in NK-92 cells

目的研究乙型肝炎病毒C蛋白在NK细胞中的表达及其对NK细胞功能的影响。方法用脂质体转染法将HBVC基因真核表达载体pHBI-CMV-HBC转入NK-92细胞，通过Western Blot检测C基因在NK-92细胞中表达．用ELISA法检测NK细胞分泌IFN-γ水平，MTT法检测NK细胞对HepG2细胞的细胞毒作用。结果Western Blot证实转染有pHBI-CMV-HBC的NK-92细胞能表达HBVC蛋白。转染了重组真核表达质粒的NK-92细胞IFN-γ水平均高于空质粒对照组和空白对照组（P〈0．01）。与两对照组相比，重组真核表达质粒转染组NK-92细胞对于HepG2细胞的细胞毒活性明显提高，差异具有统计学意义（P〈0．01）。结论HBc基因瞬时高表达可影响NK-92细胞分泌IFN-γ和细胞毒功能。

Objective To investigate the influence of hepatitis B virus C protein on the function of natural killer cell. Methods Recombinant eukaryotic expression plasmid pHBI-CMV-HBC was constructed and confirmed by double restrictive enzyme digestion and DNA sequencing analysis. Then the recombinant plasmid was tmnsfected into NK-92 cells with lipofectamine encapsuled. The transfected NK-92 cells containing expressive HBV C protein was confirmed by Western Blot analysis. ELISA was employed to determine the IFN-γ level secreted by NK-92 cells. And finally the cytotoxicities of NK cells were analysed by MTT colorimetry, with the hepatoblastoma cell line （HepG2） as target cell. Results Western blotting confirmed the expression of HBV C protein in the NK-92 cells transfected with pHBI-CMV-HBC. NK cytotoxicities and IFN-7 secretion level of NK-92 cells transfected with recombinant plasmid significantly increased compared to control NK-92 cells transfected with blank plasmid （ P 〈 0.01 ） and untransfected NK-92 cells（ P 〈 0.01 ）. Conclussion Transient expression of HBC can increase IFN-γ secretion and cytotoxicities of NK-92 cells.