Human B1 Cell Immunoglobulin

人B1细胞免疫球蛋白

• 批准号: 8284733
• 负责人: THOMAS L ROTHSTEIN
• 金额: $ 20.93万
• 依托单位: FEINSTEIN INSTITUTE FOR MEDICAL RESEARCH
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-08-02 至 2014-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8284733
• 关键词: Address; Affect; Anti-DNA Antibodies; Antibodies; Antibody-Producing Cells; Appearance; Ascaridil; Autoantibodies; Autoantigens; Autoimmune Diseases; Autoimmunity; B-Lymphocytes; Binding; Biological Assay; CD27 Antigens; Cell Separation; Cells; Characteristics; Charge; DNA; Diagnostic; Disease; Enzyme-Linked Immunosorbent Assay; FarGo; Foundations; Frequencies; Functional disorder; Goals; Hand; Human; Immune response; Immunity; Immunofluorescence Immunologic; Immunoglobulin Class Switching; Immunoglobulin G; Immunoglobulins; Individual; Kidney; Length; Lupus; MS4A1 gene; Mature B-Lymphocyte; Measures; Mediating; Memory B-Lymphocyte; Mus; Mutate; Nature; Pathogenicity; Patients; Phenotype; Play; Population; Population Heterogeneity; Property; Reporting; Rheumatoid Arthritis; Role; Sampling; Somatic Mutation; Sorting - Cell Movement; Source; Structure; Structure of germinal center of lymph node; Subgroup; Testing; Time; Work; autoreactive B cell; expression cloning; in vivo; neglect; pathogen; success; therapeutic target; therapy design; tositumomab

DESCRIPTION (provided by applicant): This project concerns human B1 cells, and the immunoglobulin (Ig) produced by this B cell subpopulation. In recent years the success of B cell depletion therapies has raised the profile of B cells as key contributors to autoimmunity. However, the means by which B cells promote autoimmunity, and especially the precise origin of pathogenic autoantibodies, have not been fully worked out. For many years it has been suggested that B1 cells might be involved in lupus and related illnesses, but this has been impossible to evaluate because identifying markers for human B1 cells have been lacking. This gap has now been filled with our recent finding that human B1 cells co-express CD20, CD27 and CD43, and lack expression of CD70. With the phenotype of B1 cells in hand it is possible to examine the nature of B1 cell-generated Ig. Human B1 cells produce Ig with many features that suggest a relationship to autoimmunity. Human B1 cells produce anti-DNA autoantibody; human B1 cells express antibody that is somatically mutated; and, human B1 cell- derived antibody can be isotype-switched. And in lupus, not only is the proportion of B1 cells among all B cells increased 2-fold, but the fraction of B1 cells that express IgG is markedly increased 16-fold (to 35% of all B1 cells). Thus, human B1 cells that produce autoreactive Ig, somatically mutated Ig, and to a greater (lupus) or lesser (normal) extent, isotype-switched Ig, express the necessary attributes to be a source of autoreactive/mutated/switched autoantibody that is recognized as pathogenic. We hypothesize then that in lupus B1 cells, particularly IgG+ B1 cells, do, in fact, represent a source of pathogenic autoantibody. To address this hypothesis, we will study Ig produced by B1, naive, and memory B cells, comparing B cells from lupus patients with normal controls, and comparing IgG+ B1 cells with non-switched B1 cells. Specifically, 1) We will evaluate the structure of B1 cell Ig by single cell sorting, PCR amplification, and sequencing, focusing on VH/VD/VJ/VL usage, somatic mutation, N-region addition, and CDR3 length and charge; 2) We will evaluate the repertoire of B1 cell Ig by expression cloning, antibody purification, HEp-2 immunofluorescence, ELISA assay, and autoantigen array, focusing on the frequency of polyreactive, autoreactive antibodies; and, 3) We will evaluate the pathogenicity of B1 cell Ig by glomerular binding/immunofluorescence and histological examination of kidneys after murine in vivo autoantibody administration, focusing on the capacity of anti-DNA autoreactive/polyreactive antibodies to produce renal damage. We hypothesize that in lupus, B1 cells generate pathogenic, nephritogenic antibodies that are somatically mutated and isotype-switched, and thus represent the origin of at least some of the "classical" type of autoreactive Ig that has been thought to be purview of post-germinal center conventional B cells. The results of this work will lay the foundation for a new and discrete therapeutic target in lupus that may eliminate the need for wholesale destruction of all mature B cells. PUBLIC HEALTH RELEVANCE: The recent successful use of B cell depletion therapy shows that B cells play an important role in autoimmune diseases. Newly discovered human B1 cells are a unique subpopulation of B cells capable of making autoantibodies and thus could be directly involved in initiating autoimmunity; further, we have found that human B1 cells are expanded in patients with lupus autoimmunity. The goal of the present proposal is to gather evidence indicating that B1 cells are a source of autoantibodies in lupus and thereby contribute to lupus disease, which will provide the foundation for new targeted therapies designed to eliminate the "bad" pathogenic B1 cells without affecting the majority population of B cells needed for immunity against infectious pathogens.

描述（由申请方提供）：本项目涉及人B1细胞和该B细胞亚群产生的免疫球蛋白（IG）。近年来，B细胞耗竭疗法的成功提高了B细胞作为自身免疫的关键贡献者的形象。然而，B细胞促进自身免疫的方法，特别是致病性自身抗体的确切来源，还没有完全确定。多年来，人们一直认为B1细胞可能与狼疮和相关疾病有关，但由于缺乏人类B1细胞的识别标记，因此无法进行评估。我们最近发现，人B1细胞共表达CD 20、CD 27和CD 43，而缺乏CD 70的表达，这一发现填补了这一空白。有了B1细胞的表型，就有可能检查B1细胞产生的IG的性质。人B1细胞产生具有许多特征的IG，这些特征表明与自身免疫有关。人B1细胞产生抗DNA自身抗体;人B1细胞表达体细胞突变的抗体;并且，人B1细胞衍生的抗体可以是同种型转换的。在狼疮中，不仅B1细胞在所有B细胞中的比例增加了2倍，而且表达IgG的B1细胞的比例也显著增加了16倍（达到所有B1细胞的35%）。因此，产生自身反应性IG、体细胞突变的IG以及在更大（狼疮）或更小（正常）程度上同种型转换的IG的人B1细胞表达作为被认为是致病性的自身反应性/突变/转换的自身抗体的来源的必要属性。因此，我们假设在狼疮B1细胞中，特别是IgG+ B1细胞，实际上是致病性自身抗体的来源。为了解决这一假设，我们将研究由B1、初始和记忆B细胞产生的IG，比较狼疮患者的B细胞与正常对照，并比较IgG+ B1细胞与非转换的B1细胞。具体而言，1）我们将通过单细胞分选、PCR扩增和测序来评估B1细胞IG的结构，重点是VH/VD/VJ/VL使用、体细胞突变、N-区添加和CDR 3长度和电荷; 2）通过表达克隆、抗体纯化、HEp-2免疫荧光、ELISA检测和自身抗原芯片等方法，对B1细胞IG的库进行评估，集中于多反应性、自身反应性抗体的频率;和3）我们将通过肾小球结合/免疫荧光和在鼠体内自身抗体施用后的肾脏的组织学检查来评估B1细胞IG的致病性，集中于抗DNA自身反应性/多反应性抗体产生肾损伤的能力。我们假设在狼疮中，B1细胞产生致病性致肾炎抗体，这些抗体是体细胞突变和同种型转换的，因此代表了至少一些“经典”类型的自身反应性IG的起源 这被认为是后生发中心常规B细胞的范围。这项工作的结果将为一个新的和离散的治疗目标奠定基础，在狼疮，可能消除需要大规模破坏所有成熟的B细胞。 公共卫生相关性：最近B细胞耗竭疗法的成功应用表明，B细胞在自身免疫性疾病中发挥重要作用。新发现的人B1细胞是一种独特的B细胞亚群，能够产生自身抗体，因此可能直接参与启动自身免疫;此外，我们发现人B1细胞在狼疮自身免疫患者中扩增。本提案的目标是收集证据表明B1细胞是狼疮中自身抗体的来源，从而有助于狼疮疾病，这将为新的靶向治疗提供基础，该靶向治疗旨在消除“坏”的致病B1细胞，而不影响针对感染性病原体的免疫所需的大多数B细胞群体。