In Vivo Detection And Mechanisms of Regulatory B cell Function in Transplantation

移植中调节性 B 细胞功能的体内检测和机制

• 批准号: 10393024
• 负责人: DAVID M ROTHSTEIN
• 金额: $ 47.39万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10393024
• 关键词: Acute; Address; Agreement; Allograft Tolerance; Allografting; Antibody Formation; Autoimmune; Autoimmune Diseases; Autoimmunity; B-Cell Acute Lymphoblastic Leukemia; B-Cell Development; B-Lymphocyte Subsets; B-Lymphocytes; Biology; Bone Marrow; Cell physiology; Cells; Cellular Immunity; Chronic; Consensus; Detection; Exhibits; Experimental Autoimmune Encephalomyelitis; Frequencies; Funding; Genetic Transcription; Graft Rejection; Graft Survival; Heart Transplantation; Human; Immune; Immune response; Immunity; In Vitro; Inbred Mouse; Inflammation; Interleukin-10; Kidney Transplantation; Knockout Mice; Knowledge; Mediating; Minor; Minority; Modeling; Mus; Phenotype; Plasma Cells; Play; Process; Proteins; Regulation; Regulatory T-Lymphocyte; Reporter; Reporting; Role; Self Tolerance; Signal Transduction; T-Lymphocyte; Therapeutic; Transplant Recipients; Transplantation; Vascular Diseases; Work; allograft rejection; base; cytokine; humoral immunity deficiency; improved; in vivo; infancy; insight; isoimmunity; kidney allograft; loss of function; microbiota; mouse model; mutant; novel; prevent; systemic autoimmunity; targeted treatment; transcriptome sequencing; tumor

B cells play an important role regulating immune responses. B cell deficiency or depletion in mice can worsen autoimmunity and prevent allograft tolerance. We showed that TIM-1 is a broad marker for IL-10+ regulatory B cells (Bregs) and that anti-TIM-1 induces tolerance through induction of IL-10+ Bregs. Based on this discovery, our collaborator Dr. Kuchroo, has just shown that TIM-1 regulates various inhibitory molecules on B cells in addition to IL-10, and that mice specifically lacking B cell-TIM-1 develop spontaneous systemic autoimmunity. This unequivocally demonstrates a critical role for Bregs in maintaining tolerance. In general agreement, tolerant human renal allograft recipients demonstrate a “Breg profile” and peri-transplant depletion of B cells increased acute rejection and vasculopathy after renal and cardiac transplantation, respectively. Despite these advances, our understanding of Bregs remains in its infancy. First, there is no agreement even about which B cells carry out Breg function, why Bregs in different studies belong to different subsets, or even whether plasma cells (PCs), and not B cells per se, actually carry out Breg function. Second, little is known about how Bregs are regulated in vivo, or what signals can be used to expand them. Based on our and our collaborator’s advances, this proposal, will directly address both of these major gaps. The field has been hampered because Bregs are rare and their prior best marker, IL-10, was only detected after stimulation of B cells ex vivo. Transfer of various B cell subsets can inhibit inflammation in transplant and autoimmune models. However, these subsets are suppressive because they contain a relatively high proportion of IL-10+ B cells in a given model, rather than representing a true Breg phenotype. Only 5-15% of B cells in such subsets express IL-10, and each subset comprises <20% of all IL-10+ B cells. As a result, the field has identified a myriad of different Breg "subsets” (often minor or immature), and there is no consensus as to their function. However, using IL- 10-GFP reporter mice, we recently demonstrated that IL-10+ B cells can be directly identified without in vitro culture. Moreover, on a protein level, canonical Follicular B cells (FOB), Marginal Zone B cells (MZB) and PCs each account for ~30% of all B-lineage IL-10. We hypothesize that Bregs belonging to these canonical subsets that have distinct localization in the SLO, have different functions. In AIM 1, using purified IL-10+ or TIM-1+ Bregs belonging to these 3 subsets, we will now directly determine whether they regulate the same or different aspects of the immune response (e.g. humoral vs. cellular). While PCs were reported to be essential for Breg function, we found that mice unable to generate PCs due to B cell-specific deletion of BLIMP-1 exhibit a regulated phenotype with improved allograft survival and less severe EAE, and a marked increase in both frequency and number of IL-10+ and TIM-1+ Bregs. Therefore, in Aim 2, we will identify how changes in BLIMP-1, PCs, and the microbiota interact to regulate Breg numbers in vivo. This work will greatly enhance our understanding of B cell and Breg biology and provide therapeutic insights highly relevant to allograft tolerance.

B 细胞在调节免疫反应中发挥着重要作用。小鼠 B 细胞缺乏或耗竭可能会恶化 自身免疫并防止同种异体移植耐受。我们证明 TIM-1 是 IL-10+ 调节 B 的广泛标记 细胞 (Bregs) 并且抗 TIM-1 通过诱导 IL-10+ Bregs 来诱导耐受。基于这一发现， 我们的合作者 Kuchroo 博士刚刚证明 TIM-1 在 B 细胞中调节各种抑制分子 除了IL-10之外，特别缺乏B细胞-TIM-1的小鼠会产生自发的全身性自身免疫。 这明确表明了布雷格斯在维持耐受性方面的关键作用。一般来说， 耐受性人肾同种异体移植受者表现出“Breg 特征”和移植周围 B 细胞的耗竭 肾移植和心脏移植后急性排斥反应和血管病变分别增加。尽管有这些 尽管我们已经取得了进展，但我们对布雷格斯的理解仍处于起步阶段。首先，甚至对于哪个 B 也没有达成一致。 细胞执行 Breg 功能，为什么不同研究中的 Breg 属于不同的子集，甚至是否 实际上执行 Breg 功能的是浆细胞 (PC)，而不是 B 细胞本身。其次，人们对如何做到这一点知之甚少。 Bregs在体内受到调节，或者可以使用什么信号来扩展它们。根据我们和我们的合作者的 该提案的进展将直接解决这两个主要差距。该领域受到阻碍是因为 Breg 很罕见，它们之前最好的标记物 IL-10 仅在离体刺激 B 细胞后才被检测到。 各种 B 细胞亚群的转移可以抑制移植和自身免疫模型中的炎症。然而， 这些子集是抑制性的，因为它们在给定的细胞中含有相对较高比例的 IL-10+ B 细胞 模型，而不是代表真正的 Breg 表型。这些亚群中只有 5-15% 的 B 细胞表达 IL-10， 每个子集包含<20%的所有IL-10+ B细胞。因此，该领域已经确定了无数不同的 Breg“子集”（通常较小或不成熟），并且对其功能没有达成共识。然而，使用 IL- 10-GFP报告小鼠，我们最近证明无需体外即可直接识别IL-10+ B细胞 文化。此外，在蛋白质水平上，典型的滤泡 B 细胞 (FOB)、边缘区 B 细胞 (MZB) 和 PC 每种均占所有 B 谱系 IL-10 的约 30%。我们假设 Breg 属于这些规范子集 它们在 SLO 中具有不同的定位，具有不同的功能。在 AIM 1 中，使用纯化的 IL-10+ 或 TIM-1+ Bregs属于这3个子集，我们现在将直接确定它们是否监管相同或不同 免疫反应的各个方面（例如体液与细胞）。据报道，个人电脑对于布雷格来说至关重要 功能，我们发现由于 B 细胞特异性删除 BLIMP-1，小鼠无法生成 PC 调节表型，同种异体移植物存活率提高，EAE 严重程度减轻，并且两者均显着增加 IL-10+ 和 TIM-1+ Bregs 的频率和数量。因此，在目标 2 中，我们将确定如何改变 BLIMP-1、PC 和微生物群相互作用，调节体内 Breg 数。这项工作将大大提高我们的 了解 B 细胞和 Breg 生物学，并提供与同种异体移植耐受高度相关的治疗见解。