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: 9197259
• 负责人: DAVID M ROTHSTEIN
• 金额: $ 37.74万
• 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 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9197259
• 关键词: Acute; Address; Allograft Tolerance; Allografting; Anatomy; Antigen Presentation; Autoimmune Diseases; Autoimmunity; B-Lymphocyte Subsets; B-Lymphocytes; Biological; Cell physiology; Cells; Collagen-Induced Arthritis; Computer Assisted; Data; Detection; Eragrostis; Exhibits; Experimental Autoimmune Encephalomyelitis; Graft Rejection; Graft Survival; Hour; Human; Humoral Immunities; IgG3; Image; Imagery; Immune response; Immunobiology; Immunofluorescence Immunologic; In Situ; In Vitro; Inflammation; Insulin-Dependent Diabetes Mellitus; Interleukin-10; Knockout Mice; Knowledge; Lead; Locales; Location; Lymphoid; Mediating; Minority; Modeling; Mus; Organ; Phenotype; Plasma Cells; Play; Population; Regulation; Reporting; Role; Shapes; Source; Spleen; Splenic Red Pulp; System; T-Lymphocyte; T-Lymphocyte Subsets; Therapeutic; Time; Tissue imaging; Transplantation; Work; allograft rejection; base; cell motility; chemokine; clinically relevant; cytokine; falls; humoral immunity deficiency; improved; in vivo; insight; kidney allograft; mouse model; novel; prevent; programs; public health relevance; response; simulation

 DESCRIPTION (provided by applicant): It is now clear that B cells play an important role regulating immune responses. B cell deficiency or depletion in mice can worsen autoimmunity and prevent allograft tolerance by various agents thought to target T cells. Moreover, we showed that regulatory B cells (Bregs) can be induced and are responsible for prolonged graft survival mediated by anti-TIM-1. These studies suggest that Bregs play an important role in allograft tolerance. This is echoed by the "B cell profile" observed in tolerant human renal allograft recipients. However, our understanding of Bregs is hampered because they are rare and their only specific marker is IL-10, which thus far, has only been detected after stimulation ex vivo. Transfer of various B cell subsets can inhibit inflammation and prevent allograft rejection in an IL-10 dependent fashion. However, these subsets suppress because they contain the highest proportion of IL-10+ B cells in a given model, rather than representing a true Breg phenotype. Of note, these subsets are generally small (each making up 2-15% of total B cells), and IL-10+ cells are still only 5-15% in each subset. Thus, current studies only account for a minority of IL-10+ B cells. In contrast, <1% of follicular (FO) B cells express IL-10, but because of 50-70% of all B cells are FO B cells, these are a major source of IL-10+ B cells. This has made study of Bregs by transfer of B cell subpopulations challenging, and the solution lies in identifying IL-10+ B cells directly without the need for ex vivo culture and simulation. These same issues have contributed to a remarkable lack of understanding of how Bregs function in vivo. Mice receiving Bregs have largely been treated as a "black box". We do not know which IL-10 + B subsets function as Bregs, where they arise, and how and where they effect suppressor function. Such understanding is key to developing clinically relevant approaches to enhance Breg number and function, and to avoid their inadvertent depletion. We now show that IL-10 expression by B cells can in fact be identified without in vitro simulation, and that most IL-10+ B cells are actually, B1a, MZ, and FO B cells, and even Plasma Cells. Moreover, IL-10+ B cells can be directly visualized in situ. Our lead data confirm that IL-10+ B cells populate the splenic MZ, follicle, and red pulp. Based on these data we now aim to determine how Bregs actually function in vivo to prevent rejection in Breg-dependent allograft models. Different subsets of B cells have different roles and localization within the secondary lymphoid organs (SLO). Therefore in Aim 1 we will determine which IL-10+ B cell subpopulations exhibit the Breg activity and whether different subsets have subspecialized functions. In Aim 2, we will determine where within the SLO Bregs actually act to inhibit the alloimmune response. In Aim 3, we will directly determine whether Bregs suppress T cells through direct cognate interactions. We utilize state of the art imaging and inducible KO mice not previously applied to Breg studies. This work will greatly enhance our understanding of Breg immunobiology and provide therapeutic insights highly relevant to allograft tolerance.

 描述（由申请人提供）：现在已经清楚，B 细胞在调节免疫反应中发挥着重要作用。小鼠 B 细胞缺乏或耗竭会恶化自身免疫，并阻止各种被认为针对 T 细胞的药物的同种异体移植耐受。此外，我们发现调节性 B 细胞 (Bregs) 可以被诱导，并负责由抗 TIM-1 介导的延长移植物存活。这些研究表明 Bregs 在同种异体移植耐受中发挥着重要作用。这与在耐受性人类同种异体肾移植受者中观察到的“B细胞谱”相呼应。然而，我们对 Bregs 的理解受到阻碍，因为它们很罕见，而且它们唯一的特异性标记物是 IL-10，迄今为止，只有在离体刺激后才能检测到。 各种 B 细胞亚群的转移可以抑制炎症并以 IL-10 依赖性方式防止同种异体移植排斥。然而，这些子集会受到抑制，因为它们在给定模型中含有最高比例的 IL-10+ B 细胞，而不是代表真正的 Breg 表型。值得注意的是，这些亚群通常很小（每个亚群占 B 细胞总数的 2-15%），而 IL-10+ 细胞在每个亚群中仍然仅占 5-15%。因此，目前的研究仅考虑了少数 IL-10+ B 细胞。相比之下，<1% 的滤泡 (FO) B 细胞表达 IL-10，但由于所有 B 细胞中 50-70% 是 FO B 细胞，因此它们是 IL-10+ B 细胞的主要来源。这使得通过转移 B 细胞亚群来研究 Bregs 具有挑战性，解决方案在于直接识别 IL-10+ B 细胞，而不需要离体培养和模拟。这些相同的问题导致人们对 Bregs 在体内的功能缺乏了解。接受 Bregs 的小鼠在很大程度上被视为“黑匣子”。我们不知道哪些 IL-10 + B 子集充当 Bregs，它们出现在哪里，以及它们如何以及在何处影响抑制功能。这种理解对于开发临床相关方法来增强 Breg 数量和功能并避免其无意耗尽至关重要。 我们现在表明，B 细胞的 IL-10 表达事实上可以在不进行体外模拟的情况下进行鉴定，并且大多数 IL-10+ B 细胞 细胞实际上是 B1a、MZ 和 FO B 细胞，甚至是浆细胞。此外，IL-10+ B 细胞可以直接在原位可视化。我们的主要数据证实，IL-10+ B 细胞存在于脾 MZ、滤泡和红髓中。基于这些数据，我们现在的目标是确定 Breg 在体内实际上如何发挥作用，以防止 Breg 依赖性同种异体移植模型中的排斥反应。不同的 B 细胞亚群在次级淋巴器官 (SLO) 中具有不同的作用和定位。因此，在目标 1 中，我们将确定哪些 IL-10+ B 细胞亚群表现出 Breg 活性，以及​​不同的亚群是否具有亚特化功能。在目标 2 中，我们将确定 SLO Bregs 中的哪些位置实际上起到抑制同种免疫反应的作用。在目标 3 中，我们将直接确定 Bregs 是否通过直接同源相互作用抑制 T 细胞。我们利用了之前未应用于 Breg 研究的最先进的成像和诱导型 KO 小鼠。这项工作将极大地增强我们对 Breg 免疫生物学的理解，并提供与同种异体移植耐受高度相关的治疗见解。