REGULATION OF THE IMMUNE RESPONSE TO POLYSACCHARIDES AND POLYSACCHARIDE CONJUGATE

对多糖和多糖缀合物的免疫反应的调节

• 批准号: 6293785
• 负责人: KE E STEIN
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6293785
• 关键词: Neisseria meningitidis vaccine; antibacterial antibody; antibody formation; antibody specificity; autoantibody; bacterial antigens; bacterial polysaccharides; bacterial vaccines; bactericidal immunity; gene mutation; genetic recombination; immunoconjugates; immunogenetics; immunoglobulin genes; laboratory mouse; monoclonal antibody; northern blottings; nucleic acid sequence; site directed mutagenesis; structural genes; tetanus toxoid; thymus; western blottings

The immune response to polysaccharide (PS) antigens is highly regulated and has several distinguishing features including restricted subclass, variable region gene usage, fine specificity, and avidity. Simple PS not conjugated to protein (such as bacterial Levan (BL, Neisseria meningitidis group C (MCPS)) elicit a thymus-independent (TI) response. PS conjugated to proteins (such as MCPS coupled to tetanus toxoid, (MCPS-TT)), on the other hand, elicit a different type of response, termed thymus-dependent (TD). Our earlier analysis of anti-BL antibodies had shown a germline primary response to the inulin (In) branch determinants. Two injections of BL, however, elicited IgM mAb with somatic mutations and higher affinity. We have now performed site-directed mutagenesis of the germline antibodies and identified specific sites resulting in higher or lower affinity. Analysis of the regulation of diversity in the anti-In response has led us to map the Sr1 diversity gene. Our previous data indicated linkage between Sr1 and markers on mouse chromosome 14 and this has been supported by studies using CXB recombinant inbred (RI) lines and more recently by phenotype analysis of (BALB/c x B6.C-H8)F1 mice. We are in the process of completing phenotype and genotype analysis of additional BALB/c x (BALB/c x C57BL/6)F1 backcross mice. Previous analyses of anti-MCPS and anti-MCPS TT mAb reveal that VH gene family usage is dominated by VHJ558. IgG, IgA and IgM mAbs from 3 panels were purified and tested for avidity. MAbs from mice immunized with MCPS-TT conjugate 2X(C2) and immunized with MCPS-TT and boosted with MCPS(CP), in general, had 2 orders of magnitude lower concentrations for 50% binding than anti _MCPS mAb, indicating that the TD mAb were of significantly higher avidity (10-100 fold higher) than the anti-MCPS. Sequence analysis in progress will determine if somatic mutation or different VH gene usage with in J558 or other families accounts for the increased avidity. Earlier mouse model studies in our laboratory indicated a developmental delay in the immune response to MCPS even when it was administered as TD MCPS-TT conjugates. Therefore, studies were undertaken to examine whether the delay in response TD conjugates in neonatal mice is due to defective antigen presentation. we have found that adult B cells were much more effective in presenting TT or MCPS-TT to T cells than total spleen cells or macrophages and that adult dendritic cells were the most effective antigen presenting cells in this system,the efficiency being about 2-fold higher than B cells. Of significance, both neonatal B cells and neonatal dendritic cells were defective in their ability to present antigen. The data suggest that factors that inprove the ability of neonatal cells to present antigen might be useful in stimulating neonatal responses to conjugate vaccines.

对多糖（PS）抗原的免疫应答是高度调节的，并具有几个显著特征，包括限制性亚类、可变区基因使用、良好的特异性和亲合力。未与蛋白质缀合的简单PS（例如细菌莱万（BL，脑膜炎奈瑟菌C组（MCPS））引起胸腺非依赖性（TI）应答。另一方面，与蛋白质缀合的PS（例如与破伤风类毒素偶联的MCPS（MCPS-TT））引起不同类型的应答，称为胸腺依赖性（TD）。我们早期对抗BL抗体的分析显示了对菊粉（In）分支决定簇的生殖系初级应答。然而，两次BL注射引发了具有体细胞突变和更高亲和力的IgM mAb。 我们现在已经进行了生殖系抗体的定点诱变，并确定了导致更高或更低亲和力的特定位点。在抗In反应的多样性的调节分析，使我们映射Sr 1多样性基因。我们以前的数据表明Sr 1和小鼠14号染色体上的标记之间的连锁，这已经得到了使用CXB重组近交系（RI）的研究和最近的（BALB/c x B6.C-H8）F1小鼠的表型分析的支持。 我们正在完成额外的BALB/c x（BALB/c x C57 BL/6）F1回交小鼠的表型和基因型分析。抗MCPS和抗MCPS TT mAb的先前分析揭示VH基因家族使用由VHJ 558主导。纯化来自3组的IgG、伊加和IgM mAb并测试亲合力。来自用MCPS-TT缀合物2X（C2）免疫的小鼠和用MCPS-TT免疫并用MCPS（CP）加强的小鼠的MAb通常具有比抗MCPS mAb低2个数量级的50%结合浓度，表明TD mAb具有比抗MCPS显著更高的亲合力（高10-100倍）。 正在进行的序列分析将确定体细胞突变或与J558或其他家族中不同的VH基因使用是否是增加的亲合力的原因。我们实验室的早期小鼠模型研究表明，即使以TD MCPS-TT偶联物的形式给药，对MCPS的免疫应答也存在发育延迟。因此，进行研究以检查新生小鼠中TD缀合物的响应延迟是否是由于抗原呈递缺陷。我们发现，成体B细胞在向T细胞呈递TT或MCPS-TT方面比总脾细胞或巨噬细胞有效得多，并且成体树突细胞是该系统中最有效的抗原呈递细胞，其效率比B细胞高约2倍。重要的是，新生儿B细胞和新生儿树突状细胞在呈递抗原的能力上都有缺陷。这些数据表明，提高新生儿细胞呈递抗原能力的因素可能有助于刺激新生儿对结合疫苗的反应。