Regulation of Immune Response to Polysaccharides

总结： 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。数据显示芳香族和碱性氨基酸在抗PS抗体的CDR中的重要性。在我们的分析中的多样性的调节，在抗-在响应中，我们已经映射Sr 1多样性基因小鼠染色体14。最近在TCR KO小鼠中的研究表明，对In的反应不仅是TI，而且当加回时不受T细胞的影响，尽管血清IgG和IgM大量增加。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。抗MCPS单克隆抗体的序列分析表明，即使是具有相同特异性的单克隆抗体，在这种反应中也使用了几个不同的生殖系基因。在对MCPS的反应中观察到很少的体细胞突变，但当观察到时，与亲和力的增加相关。最近的模拟研究表明，结合位点中碱性氨基酸的存在也与亲和力的增加相关。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。作为这些研究的一部分，从用MCPS TT免疫的小鼠产生T细胞克隆。这些克隆已被证明包括MCPS的特异性和PS反应性克隆现在已被表征为它们的MHC和辅助细胞的要求。这些数据表明，不仅B细胞，而且PS反应性T细胞可能有助于新生儿对缀合物疫苗应答的保护性刺激。儿童疫苗倡议的目标之一是减少为儿童进行全面免疫所需的接触次数。为了实现这一目标，已经测试了几种组合疫苗在婴儿中的安全性和免疫原性。我们已经在SW远交系小鼠中开发了一种模型，以检查组合疫苗对单个组分的反应的影响。 我们最初观察到DTaP干扰了Hib-TT的抗Hib反应随后的研究证实了早期的发现，但DTaP的干扰并不显着。我们还观察到，Hib-TT干扰了对所有三种类型脊髓灰质炎（IPV）的反应。在近交系BALB/c小鼠中重复了这些研究，并证实了Hib-TT对脊髓灰质炎病毒的干扰。初步研究表明，另一种Hib结合物Hib-CRM 197不会引起这种干扰。

Summary: 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. Data show the importance of aromatic and basic amino acids in the CDRs of anti-PS antibodies. In our analysis of the regulation of diversity in the anti-In response we have mapped the Sr1 diversity gene to mouse chromosome 14. Recent studies in TCR KO mice have shown that the response to In is not only TI, but is not influenced by T cells when added back, despite a large increase in serum IgG and IgM. Previous analyses of anti-MCPS and anti-MCPS TT mAb reveal that VH gene family usage is dominated by VHJ558. Sequence analysis of anti MCPS mAb shaows that several different germline genes are used in this response, even by mAb of the same fine specificity. Few somatic mutations are seen in response to MCPS, but when observed, correlate with an increase in affinity. Recent modeling studies suggest that the presence of basic amino acids in the combining site aslo correlate with an increase in affinity. 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. As part of these studies, T cell clones were generated from mice immunised with MCPS TT. These clones have been shown to include specificities for MCPS and the PS reactive clones have now been characterized as to their requirements for MHC and for accessory cells. The data suggest that not only B cells, but also PS reactive T cells may contribute to the protective stimulation of neonatal responses to conjugate vaccines. One of the goals of the Children's Vaccine Initiative is to reduce the number of contacts required to immunize a child fully. To meet this goal several combined vaccines have been tested for their safety and immunogenicity in infants. We have developed a model in SW outbred mice to examine the effects of combining vaccines on the response to individual components. We initially observed that DTaP interferes with the anti-Hib response to Hib-TT Subsequent studies have confirmed the earlier findings but the interference from DTaP is not significant. We also observed that Hib-TT interfered with the response to all three types of polio, given as IPV. These studies have been repeated in inbred BALB/c mice and confirm the interference with poliotiters by Hib-TT. Preliminary studies suggest that anothe Hib conjugate, Hib-CRM197, does not cause this interference.