Effects of neonatal microbial exposure on anti-polysaccharide B cell development

摘要 天然样B细胞衍生的抗T-非依赖性多糖（PS）的天然抗体（NAb）和 磷脂表位普遍存在于哺乳动物中，并在保护免受 许多细菌病原体。然而，在感染后针对PS抗原的稳健抗体应答或 新生儿缺乏免疫接种。我们之前的研究表明新生儿接触PS和磷脂 抗原通过免疫接种或与微生物群的自然定殖，影响 抗原特异性先天性B样细胞导致天然抗体（Na B）水平的定量增加。这些 影响成人对感染的易感性，以及对过敏原和自身抗原的免疫反应性 具有相似的B细胞表位。虽然微生物群影响其他淋巴谱系的发育， 在成人先天性B样细胞和NA B谱系发育中的作用还未被探索。我们 假设除了在B细胞发育期间发生的充分描述的选择过程之外， 新生儿期自体抗原和外源性胎盘来源表位联合收割机可及性 发展以塑造自然剧目。这些拟议研究的目标是阐明 使用三种模型B细胞抗原- 特异性代表相关病原体和寄生虫生物体的表位， 抗原潜力：磷酸胆碱（PC）、N-乙酰-D-葡糖胺（GlcNAc）和葡聚糖（DEX）。目标1， 这些研究将用确定的微生物群定殖野生型无菌小鼠， 外源性抗原信号传导对克隆B细胞发育和NA B库形成的影响。骨髓 嵌合体系统，由抗原特异性免疫球蛋白（IG）重链转基因小鼠品系组成， 和谱系追踪将能够评估B淋巴细胞上的内源性和外源性抗原信号传导 选择和受体编辑过程。在目标2中，我们将分析频率，细胞表面表型， 亚群和同种型分布以及对这些特异性人B细胞的免疫球蛋白基因使用 抗原从脐带血、扁桃体/腺样体分离的B细胞和外周血B细胞的检查 不同年龄的供体将提供新的见解抗原经验的作用，在先天性， 就像个体内部和个体之间的B细胞库多样性一样。将克隆抗原特异性IG基因， 以确定抗原驱动的成熟对这些重组抗体的功能的影响。 人类抗体总的来说，这些研究将确定微生物对小鼠NAb发育的影响， 确定人NAb发育的发育动力学，并比较和对比NAb库 两个物种之间的发展。这种对人类婴儿对感染和殖民的反应的深入了解 将推进我们开发有效的新生儿疫苗策略和介入治疗的长期目标 提供对传染性病原体、过敏性和自身免疫性疾病的保护。

ABSTRACT Innate-like B cell-derived natural antibodies (NAbs) against T-independent polysaccharide (PS) and phospholipid epitopes are universally present in mammals and serve important roles in the protection against many bacterial pathogens. However robust antibody responses against PS antigens following infection or immunization are absent in neonates. We previously showed that neonatal exposure to PS and phospholipid antigens by immunization or natural colonization with the microbiota, influences the clonal distribution of antigen-specific innate-like B cells leading to quantitative increases in natural antibody (Nab) levels. These effects impact adult susceptibility to infection, and immune-responsiveness to allergens and auto antigens bearing similar B cell epitopes. Although the microbiota impacts the development of other lymphoid lineages its role in the development of the adult innate-like B cell and NAb repertoires is largely unexplored. Our hypothesis is that in addition to the well-described selection processes that occur during B cell development, the accessibility to autologous antigens and exogenous commensal-derived epitopes combine during neonatal development to shape the natural repertoire. The goal of these proposed studies is to elucidate mechanisms of exogenous antigen-directed development of the innate-like B cell repertoire using three model B cell antigen- specificities that represent epitopes for relevant pathogenic and commensal organisms with a spectrum of auto antigenic potential: phosphorylcholine (PC), N-acetyl-D-glucosamine (GlcNAc), and dextran (DEX). In Aim 1 of these studies wild-type gnotobiotic mice will be colonized with defined microbiota to assess the contribution of exogenous antigen signaling on clonal B cell development and formation of the NAb repertoire. Bone marrow chimera systems, composed of antigen-specific immunoglobulin (Ig) heavy chain-transgenic mouse strains, and lineage tracking will enable assessment of endogenous and exogenous antigen signaling on B lymphocyte selection and receptor editing processes. In Aim 2 we will analyze the frequency, cell surface phenotype, subset and isotype distribution as well as immunoglobulin gene usage of human B cells specific for these antigens. Examination of B cells isolated from cord blood, tonsils/adenoids, and peripheral blood B cells of donors of different ages will provide novel insight into the role of antigen experience in the formation of innate- like B cell repertoire diversity within and across individuals. Antigen-specific Ig genes will be cloned and expressed as recombinant antibody to determine effects of antigen-driven maturation on functions of these human antibodies. Collectively, these studies will define the microbial influences on NAb development in mice, determine developmental kinetics of human NAb development, and compare and contrast the NAb repertoire development between both species. This insight into the human infant responses to infection and colonization will advance our long-term goal of developing effective neonatal vaccine strategies and interventional therapies that provide protection from infectious pathogens and allergic and autoimmune diseases.