Contribution of B cells to human cardiac allograft vasculopathy

B 细胞对人心脏同种异体移植血管病的作用

• 批准号: 9114457
• 负责人: Emmanuel Zorn
• 金额: $ 40.25万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-08-01 至 2020-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9114457
• 关键词: Alloantigen; Antibodies; Antigen Presentation; Antigen Targeting; Antigen-Antibody Complex; Antigen-Presenting Cells; Antigens; Apoptotic; Autoantigens; B cell repertoire; B-Lymphocytes; Biological Assay; Biopsy Specimen; Blood; Blood specimen; Canada; Cardiac; Cell physiology; Cells; Cellular Assay; Clonality; Complement; Complication; Coronary artery; Development; Enzyme-Linked Immunosorbent Assay; Fc Immunoglobulins; Flow Cytometry; Functional disorder; Graft Rejection; Health; Heart Transplantation; Heavy-Chain Immunoglobulins; Human; Humoral Immunities; Immunofluorescence Immunologic; Immunoglobulin G; In Situ; In Vitro; Inflammation; Inflammatory; Location; MS4A1 gene; Molecular; Monoclonal Antibodies; Morbidity - disease rate; Multi-Institutional Clinical Trial; Mus; Patients; Phenotype; Preventive; Process; Property; Protein Microchips; Research; Rheumatoid Factor; Role; Sampling; Sequence Analysis; Series; Somatic Mutation; Source; Specificity; Specimen; T cell response; T-Lymphocyte; Techniques; Testing; Therapeutic Agents; Time; Tissue Grafts; Tissues; Transplant Recipients; Transplantation; United States National Institutes of Health; Vascular Diseases; base; cytokine; deep sequencing; graft function; heart allograft; macrophage; mortality; nonhuman primate; prevent; research study; response; rituximab; uptake

 DESCRIPTION (provided by applicant): CAV is almost invariably associated with B cell infiltrates in or around coronary arteries as well as in endomyocardial tissue (quilty effect). It s assumed that these cells participate in the rejection process, although a formal demonstration of such contribution is still lacking. On the whole, infiltrating B cells are loosely defined. Their eact phenotype, antigen specificity and possible function are currently unknown. Understanding how B cells contribute to mechanisms of graft rejection would undoubtedly facilitate the development of therapeutic agents to target them and treat rejection. Studies in humans are challenging due to the limited source of samples and the difficulty of setting up techniques to study primary B cells retrieved directly from tissue. For these reasons, the assessment of human graft infiltrating B cells has been limited thus far and most studies have focused on humoral immunity using blood samples. We have begun collecting fresh cardiac graft specimens rejected because of CAV and explanted for re-transplantation through a collaborative network of transplant centers in the USA and Canada. Preliminary experiments using deep sequencing to analyze rearranged immunoglobulin heavy chain repertoire in situ demonstrated the massive expansion and somatic mutation of B cell clones in 4 cardiac allografts. We have also started immortalizing B cell clones directly from these cardiac allograft specimens. Our proposed research will characterize these cells, uncover their main function and evaluate their participation in the pathophysiology of CAV. Aim 1. To characterize the phenotype, clonality and specificity of graft infiltrating B cells during CAV: We will first assess the distribution and phenotype of B cells within the rejected graft tissue. B cell repertoire analyses will then identify predominant B cell clones expanded in situ. In parallel, B cells isolated from explanted grafts will be immortalized and cultured in vitro. Selected clones corresponding to B cells expanded in situ will be further characterized. Aim 2. To identify the function of antibodies secreted by graft infiltrating B cells during CAV: Monoclonal antibodies secreted by immortalized clones found to be expanded in situ in aim 1, will be assessed for their capacity to form immune complexes and induce cytokine secretion by macrophages in situ as well as facilitate antigen presentation to T cells. Aim 3. To determine the function of graft infiltrating B cells during CAV: Aside from their capacity to secrete pathogenic antibodies, we will examine whether graft-infiltrating B cells during CAV can uptake and present antigens to T cells. We will also investigate whether B cells can polarize T cell responses in situ. Lastly, we will also examine the role of the complement to modulate these responses.

 描述（由适用提供）：CAV几乎与冠状动脉和内膜组织中的B细胞浸润​​几乎总是与B细胞浸润​​相关（Quilty效应）。它假设这些细胞参与了拒绝过程，尽管仍然缺乏对这种贡献的形式证明。总体而言，浸润的B细胞是松散定义的。目前，他们的EACT表型，抗原特异性和可能的​​功能尚不清楚。了解B细胞如何促进移植排斥的机制无疑将支持治疗剂的发展以靶向并治疗排斥。由于样本的来源有限以及建立直接从组织中检索到的原代B细胞的技术的困难，对人类的研究受到了挑战。由于这些原因，人类移植的评估浸润 到目前为止，B细胞受到限制，大多数研究都集中在使用血液样本的体液免疫学上。我们已经开始收集由于CAV而拒绝的新鲜心脏移植标本，并通过在美国和加拿大的移植中心的协作网络进行了重新转移。初步实验使用深度测序来分析重排的免疫球蛋白重链曲目原位证明了4个心脏同种异体移植物中B细胞克隆的大规模扩张和体细胞突变。我们还直接从这些心脏同种异体移植规格中直接开始使B细胞克隆永生。我们提出的研究将表征这些细胞，发现它们的主要功能并评估其参与CAV的病理生理学。目的1。要表征CAV期间移植物浸润B细胞的表型，克隆性和特异性：我们将首先评估被拒绝的移植组织内B细胞的分布和表型。然后，B细胞库分析将确定原位扩展的主要B细胞克隆。同时，从扩张的移植物中分离出的B细胞将在体外永生和培养。与原位扩展的B细胞相对应的选定克隆将进一步表征。目标2。确定移植物浸润B细胞分泌的抗体的功能 在CAV期间：被发现在AIM 1的原位扩展的永生克隆分泌的单克隆抗体，将评估其形成免疫复合物并诱导巨噬细胞的细胞因子分泌的能力，并由原位的设施抗原表现为T细胞。目的3。确定在CAV期间移植物浸润B细胞的功能：除了它们具有秘密致病性抗体的能力外，我们将检查CAV期间移植物浸润的B细胞是否可以摄取并将抗原摄取对T细胞。我们还将研究B细胞是否可以在原位偏振T细胞反应。最后，我们还将研究完成这些响应的完成的作用。