Hu G J, Wang R Y, Han D S, Alter H J, Shih J W
Department of Transfusion Medicine, Warren G. Magnuson Clinical Center, National Institutes of Health, Bethesda, MD 20892-1184, USA.
Vaccine. 1999 Aug 6;17(23-24):3160-70. doi: 10.1016/s0264-410x(99)00130-9.
Hepatitis C Virus (HCV) causes most cases of posttransfusion hepatitis. Chronic HCV infection is highly related to chronic hepatitis, cirrhosis and hepatocellular carcinoma. Current therapies are only minimally effective and no vaccine has been developed. DNA-based immunization could be of prophylactic and therapeutic value for HCV infection. By intramuscular inoculation in BALB/c mice with an HCV recombinant plasmid pCI-HCV-C, we found significant levels of IgM antibody, but no significant IgG rise. After boost the immunized mice with recombinant HCV-core protein (cp1-10; 1-164aa), the anticore IgG, verified by Western-blotting, rose rapidly, which was two weeks earlier than that with control plasmid. Spleen cells from pCI-HCV-C immunized mice gave higher proliferation index (PI) than control (P < 0.05). The PI of cp1-10 boosted mice was even higher. Proliferation blocking assay with mAb proved the responding cell to be of CD4+ CD8- phenotype, supporting specific priming of T helper cells. A 51Cr-releasing CTL assay specific for HCV-core was developed, and a specific CTL response against HCV-core was demonstrated in both pCI-HCV-C immunized mice and mice boosted with cp1-10. Strong cytotoxic activity against peptide-pulsed p815 cells (H-2d), but not EL-4 cells (H-2b), suggested MHC class I restriction of the CTL activity. Blocking of CTL with mAb proved the effector cells to be of CD4- CD8+. Three CTL epitopes in HCV-core protein were demonstrated. We failed to detect CTL when immunized only with core protein. The results suggested that vaccination with HCV-core derived DNA sequences could be an effective method to induce humoral and cellular immune responses to HCV.
丙型肝炎病毒（HCV）导致了大多数输血后肝炎病例。慢性HCV感染与慢性肝炎、肝硬化及肝细胞癌密切相关。目前的治疗方法效果甚微，且尚未研发出疫苗。基于DNA的免疫接种对HCV感染可能具有预防和治疗价值。通过向BALB/c小鼠肌肉注射HCV重组质粒pCI-HCV-C，我们发现IgM抗体水平显著升高，但IgG未显著升高。用重组HCV核心蛋白（cp1-10；1-164aa）对免疫小鼠进行加强免疫后，经蛋白质印迹法验证，抗核心IgG迅速升高，比用对照质粒免疫提前两周。pCI-HCV-C免疫小鼠的脾细胞增殖指数（PI）高于对照组（P < 0.05）。cp1-10加强免疫小鼠的PI更高。用单克隆抗体进行的增殖阻断试验证明反应细胞为CD4+ CD8-表型，支持T辅助细胞的特异性启动。建立了针对HCV核心的51Cr释放CTL试验，并在pCI-HCV-C免疫小鼠和用cp1-10加强免疫的小鼠中均证明了针对HCV核心的特异性CTL反应。对肽脉冲p815细胞（H-2d）具有强烈的细胞毒活性，但对EL-4细胞（H-2b）无此活性，提示CTL活性受MHC I类限制。用单克隆抗体阻断CTL证明效应细胞为CD4- CD8+。在HCV核心蛋白中鉴定出三个CTL表位。仅用核心蛋白免疫时未检测到CTL。结果表明，用HCV核心衍生的DNA序列进行疫苗接种可能是诱导针对HCV的体液和细胞免疫反应的有效方法。
