The teleost melano-macrophage center response to immunization and helminth-mediated immunosuppression

硬骨鱼黑素巨噬细胞中心对免疫和蠕虫介导的免疫抑制的反应

• 批准号: 10399548
• 负责人: Natalie C. Steinel
• 金额: $ 38.56万
• 依托单位: UNIVERSITY OF MASSACHUSETTS LOWELL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-10 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10399548
• 关键词: Address; Animal Model; Antibody Affinity; Antibody Formation; Antigens; B Cell Proliferation; B-Cell Activation; B-Lymphocytes; Behavioral; Biological Assay; Biological Markers; Biological Models; Biology; Bromodeoxyuridine; CRISPR/Cas technology; Candidate Disease Gene; Cells; Cellular Immunity; Cestoda; Comparative Study; Complement; Data; Dextrans; Discipline; Disease; Ecology; Evolution; Exhibits; Fishes; Foundations; Future; Gasterosteidae; Gene Expression; Gene Expression Profile; Gene Expression Profiling; Gene Mutation; Genes; Genetic Screening; Genomics; Genotype; Helminthiasis; Helminths; Histologic; Human; Humoral Immunities; Immune; Immune system; Immunity; Immunization; Immunize; Immunobiology; Immunoglobulin Genes; Immunoglobulin Somatic Hypermutation; Immunological Models; Immunologics; Immunology; Immunosuppression; In Vitro; Individual; Infection; Investigation; Knowledge; Label; Laboratories; Link; Mammals; Maps; Mediating; Methodology; Modeling; Molecular; Mus; NON Mouse; Osteichthyes; Parasites; Persons; Physiologic pulse; Play; Poikilotherms; Population; Proliferating; Public Health; Quantitative Trait Loci; Refractory; Reporting; Resistance; Role; Site; Structure of germinal center of lymph node; Study models; System; T-Independent Antigens; Taxes; Testing; Vaccines; Variant; Work; adaptive immunity; co-infection; experimental study; genomic locus; helminth infection; immune activation; immune function; immunoregulation; insight; macrophage; novel; novel strategies; pathogen; response; teleost; tool; transcriptome sequencing; vaccine development; vaccine efficacy; vaccine response

Abstract Non-mouse models can be fertile ground for the discovery of previously unappreciated immunologic paradigms and provide new perspectives of vertebrate immunity. The proposed work addresses a fundamental question of bony fish (teleost) immunity: are melano-macrophage centers (MMC) the sites where fish B cells proliferate and generate high affinity antibody, homologous to germinal centers (GC) in mammals? To answer this long- standing question, we will characterize the threespine stickleback (Gasterosteus aculeatus) MMC response. Clarifying MMC function will yield important insights into the basis of humoral adaptive immunity in fish, and the evolutionary origins of vertebrate adaptive immunity. The first Aim of this project will investigate MMC function in response to immunization. To determine if MMCs are the site of B cell proliferation, immunized fish will be pulse labeled with BrdU so that proliferating BrdU+ cells can be localized. Next, to clarify whether somatic hypermutation occurs in MMCs, we will quantify immunoglobulin gene mutations in MMC-adjacent and non- adjacent B cells. To resolve if MMC function is indeed GC-like, we will compare the MMC response to immunization with T-dependent (NP-CGG) and T-independent (NP-dextran) antigens. The results of this first Aim will determine the suitability of the MMC as a biomarker of teleost humoral adaptive immunity, and whether the MMC is an “evolutionarily primitive” GC. Preliminary data strongly support our hypothesis that MMCs are GC-like. Consequently, in our second Aim we will apply our knowledge of MMC function to investigate the immuno-modulatory effects of a helminth parasite, Schistocephalus solidus. By combining experimental infection and immunization we will determine whether this tapeworm suppresses teleost adaptive immunity. We find that some stickleback genotypes are refractory to this immunosuppression. Using a combination of Quantitative Trait Locus (QTL) mapping, population genomics, and RNAseq analysis of gene expression we will locate the host loci responsible for variable parasite-mediated immune-modulation. We will then use the CRISPR/Cas9 system to modify these genes in cultured MMCs and evaluate their role in MMC function in vitro. In the final Aim, we will identify the helminth-derived factors that dampen MMC activity. Through QTL mapping we will identify the tapeworm-specific loci responsible for MMC suppression. To complement this approach, we will assess whether helminth excretory/secretory products directly modulate MMC gene expression. Clarifying the mechanistic basis of helminth-mediated immunomodulation in fish may reveal new approaches to regulate vertebrate immune function. This work will also generate a new and broadly applicable assay of teleost immunity, which has the potential to be used in the study of diverse fish (and other poikilotherm) species. Beyond expanding our view of vertebrate immunobiology, this work has implications for the disciplines of disease ecology, host-pathogen co-evolution, and vaccine development.

摘要 非小鼠模型可以为发现以前不受重视的免疫学范例提供肥沃的土壤 为脊椎动物免疫研究提供了新的视角。拟议的工作解决了一个根本问题 硬骨鱼（硬骨鱼）免疫：黑素巨噬细胞中心（MMC）是鱼类B细胞增殖的场所吗 并产生高亲和力抗体，与哺乳动物的生发中心（GC）同源？为了回答这个问题- 长期存在的问题，我们将描述三棘鱼（Gasterosteus aculeatus）MMC反应。 阐明MMC的功能将有助于深入了解鱼类体液适应性免疫的基础， 脊椎动物适应性免疫的进化起源。本课题的第一个目的是研究MMC功能 来应对免疫接种。为了确定MMC是否是B细胞增殖的位点，将免疫的鱼接种到免疫系统中。 用BrdU标记脉冲，以便可以定位增殖的BrdU+细胞。接下来，为了澄清躯体是否 在MMC中发生超突变，我们将定量MMC相邻和非MMC中的免疫球蛋白基因突变。 相邻的B细胞。为了确定MMC功能是否确实类似GC，我们将MMC响应与 用T依赖性（NP-CGG）和T非依赖性（NP-葡聚糖）抗原免疫。这个结果首先 目的是确定MMC作为硬骨鱼体液适应性免疫生物标志物的适用性， MMC是否是“进化上原始的”GC。初步数据有力地支持了我们的假设， MMC类似GC。因此，在我们的第二个目标中，我们将把MMC函数的知识应用于 研究寄生蠕虫Schistocephalus solidus的免疫调节作用。通过组合 实验感染和免疫，我们将确定这种绦虫是否抑制硬骨鱼适应性 免疫力我们发现，一些棘鱼基因型是难治性的这种免疫抑制。使用 结合数量性状基因座（QTL）作图，群体基因组学和基因的RNAseq分析， 表达，我们将定位负责可变寄生虫介导的免疫调节的宿主基因座。我们将 然后使用CRISPR/Cas9系统修饰培养的MMC中的这些基因，并评估它们在MMC中的作用， 体外功能在最后的目标，我们将确定蠕虫衍生的因素，抑制MMC活动。 通过QTL定位，我们将确定负责MMC抑制的绦虫特异性位点。到 作为这种方法的补充，我们将评估蠕虫排泄/分泌产物是否直接调节 MMC基因表达。阐明蠕虫介导的鱼类免疫调节的机制基础， 揭示了调节脊椎动物免疫功能的新方法。这项工作还将产生一个新的和广泛的 适用的硬骨鱼免疫测定，这有可能被用于研究不同的鱼类（和其他 变温动物）物种。除了扩大我们对脊椎动物免疫生物学的看法，这项工作还对 疾病生态学、宿主-病原体共同进化和疫苗开发等学科。