Isaguliants M G, Kadoshnikov Y P, Kalinina T I, Smirnov V D, Wahren B
Ivanovsky Institute of Virology, Moscow, 123098, Russia.
Biochemistry (Mosc). 1998 May;63(5):551-8.
Nucleocapsid (core) protein of hepatitis B virus (HBcAg) induces potent cellular and humoral responses that have a clear protective potential. Rabbits were immunized by particles formed by recombinant molecules of HBcAg carrying N-terminally inserted heterologous sequences. Specificity of humoral and cellular immune response against HBcAg and selection of HBcAg epitopes was surveyed. Immunological properties of the recombinant particles were similar to those of the original HBcAg. Recombinant particles were not toxic to the peripheral blood mononuclear cells (PBMC) of non-immune or HBcAg-immunized animals ex vivo. Proliferative response of PBMC (T-lymphocytes) to HBcAg in immunized animals increased in a concentration-dependent manner in the broad interval of HBcAg concentrations (10-104 ng/ml). On the contrary, a narrow bell-shaped HBcAg dose-dependence curve was earlier observed for T-lymphocytes of donors immune to HBV after natural infection that was probably due to the cytotoxic effect of HBcAg on the expressing cells. Specificity of humoral and cellular immune response against HBcAg particles in the immunized animals and in natural infection with hepatitis B virus (HBV) was compared. Immunization with recombinant HBcAg particles induced potent anti-HBcAg antibody responses: high (up to 2.107) titers of anti-HBcAg antibodies were reached. Appearance of anti-HBcAg antibodies was in every case preceded by an increasing T-cell response to the whole protein and HBcAg-derived peptides, thus mimicking immune responses during acute HBV infection in humans. A predominant universal (haplotype-independent) T-helper cell epitope (amino acid residues (aa) 61-85 of HBcAg (p61-85)) was recognized by T-cells of all animals. Transient antibody response against p61-85 was recorded during the early stages of immunization in spite of the fact that a major B-cell epitope localized in this region is supposed to be purely conformational. A sequence representing another cluster of immunodominant T-cell epitopes of mice and HBV infected humans, aa 121-140 (p121-140), was not immunogenic on the T-cell level. However, it appeared to be a potent B-cell immunogen, despite a common assumption that HBcAg and p121-140 are not cross-reactive at the B-cell level. A possibility that anti-p121-140 antibodies were induced by an exposed region of the native particulate HBcAg and not by the denatured protein molecules, was confirmed by recognition of the particulate HBcAg by antibodies specific to synthetic peptides representing aa 120-140 of HBcAg. The data point to the exposition of aa 121-140 on the surface of the particles.
乙型肝炎病毒的核衣壳（核心）蛋白（HBcAg）可诱导产生具有明确保护潜力的强大细胞免疫和体液免疫反应。用携带N端插入异源序列的HBcAg重组分子形成的颗粒对兔子进行免疫。检测了针对HBcAg的体液免疫和细胞免疫反应的特异性以及HBcAg表位的选择。重组颗粒的免疫学特性与原始HBcAg相似。重组颗粒在体外对未免疫或经HBcAg免疫动物的外周血单核细胞（PBMC）无毒。在较宽的HBcAg浓度区间（10 - 104 ng/ml）内，免疫动物中PBMC（T淋巴细胞）对HBcAg的增殖反应呈浓度依赖性增加。相反，对于自然感染HBV后具有免疫力的供体的T淋巴细胞，较早观察到呈窄钟形的HBcAg剂量依赖性曲线，这可能是由于HBcAg对表达细胞的细胞毒性作用。比较了免疫动物和自然感染乙型肝炎病毒（HBV）时针对HBcAg颗粒的体液免疫和细胞免疫反应的特异性。用重组HBcAg颗粒免疫可诱导产生强大的抗HBcAg抗体反应：抗HBcAg抗体的滴度高达2.107。在每种情况下，抗HBcAg抗体的出现之前，T细胞对全蛋白和HBcAg衍生肽的反应都会增加，从而模拟了人类急性HBV感染期间的免疫反应。所有动物的T细胞都识别出一个主要的通用（单倍型无关）T辅助细胞表位（HBcAg的氨基酸残基（aa）61 - 85（p61 - 85））。尽管有观点认为位于该区域的主要B细胞表位纯粹是构象性的，但在免疫早期仍记录到针对p61 - 85的短暂抗体反应。代表小鼠和HBV感染人类的另一组免疫显性T细胞表位的序列，aa 121 - 140（p121 - 140），在T细胞水平上没有免疫原性。然而，尽管普遍认为HBcAg和p121 - 140在B细胞水平上没有交叉反应，但它似乎是一种强大的B细胞免疫原。通过代表HBcAg的aa 120 - 140的合成肽特异性抗体对颗粒状HBcAg的识别，证实了抗p121 - 140抗体是由天然颗粒状HBcAg的暴露区域而非变性蛋白分子诱导产生的可能性。这些数据表明aa 121 - 140暴露在颗粒表面。
