Mechanisms of Acceptance in Mouse Renal Allografts

小鼠肾同种异体移植物的接受机制

• 批准号: 7985705
• 负责人: Robert B Colvin
• 金额: $ 44.25万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-15 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7985705
• 关键词: Acute; Address; Allografting; Behavior; Blood Vessels; CD4 Positive T Lymphocytes; Cell physiology; Cells; Characteristics; Chimerism; Clinical; Complement; Computer Assisted; Custom; Dendritic Cells; Dependence; Development; Diphtheria Toxin; Employee Strikes; Eragrostis; Event; Excision; Funding; Generations; Genotype; Goals; Heart; Human; Image; Immune response; Immunosuppression; Inbred BALB C Mice; Infiltration; Institution; Kidney; Kidney Transplantation; Lead; Life; Lymphoid; Measurement; Mediating; Mediator of activation protein; Minor Histocompatibility Antigens; Molecular; Motion; Mus; Neonatal; Nephrectomy; Organ; Organ Transplantation; Outcome; Pathologic; Pathologic Processes; Patients; Pharmaceutical Preparations; Phenotype; Physiology; Predisposition; Process; Production; Progress Reports; Protocols documentation; Regulatory T-Lymphocyte; Research Design; Role; Skin Transplantation; Skin graft; System; T-Lymphocyte; Testing; Therapeutic; Time; Transplantation; Transplantation Tolerance; Tryptophan 2,3 Dioxygenase; Tubular formation; Video Recording; base; cell motility; cell type; chemokine; clinically relevant; cytokine; enzyme linked immunospot assay; follow-up; in vivo; insight; kidney allograft; latency-associated protein; novel; public health relevance; research study; trafficking

DESCRIPTION (provided by applicant): This proposal addresses the need for definitive evidence on the role of Foxp3+/T regulatory cells in the acceptance of physiologically relevant full MHC incompatible organ grafts in various settings of operational tolerance. In the initial year of funding (ARRA), spontaneous acceptance of MHC-mismatched mouse renal allografts was shown to depend exquisitely on Foxp3 cells using diphtheria toxin (DT) to transiently deplete circulating and intragraft Foxp3 cells in B6.Foxp3DTR mice bearing life sustaining DBA/2 renal allografts. The accepted grafts had distinctive nodular perivascular infiltrates of Foxp3 cells and dendritic cells (DC). Deletion of foxp3 cells caused dissolution of the nodular aggregates and widespread infiltration of the cortex, tubules and blood vessels by T cells, typical of acute rejection, and manifested by a rise in BUN over 7 days. Similar nodular aggregates of Foxp3 cells occur in grafts of patients who have accepted MHC mismatched kidneys on mixed chimerism tolerance induction protocols. Specific aim 1 will extend these observations to other forms of induced tolerance, mixed chimerism and neonatal tolerance, some of which are expected to be Foxp3 dependent; those with deletional mechanisms are expected to be Foxp3 independent. We will seek distinctive pathologic features in the graft, indicating Treg activity, such as the co-expression of latency associated protein (LAP) and Foxp3 or DC expression of indoleamine 2,3-dioxygenase (IDO), that might be used to distinguish foxp3 dependent from foxp3 independent forms of graft acceptance. The donor reactivity and regulatory potential of the cells in grafts and in lymphoid organs will be tested by ELISPOT and compared between the different forms of acceptance and acute rejection. The dependence of systemic tolerance on the intragraft infiltrates will be tested by graft nephrectomy in recipients with accepted kidney and skin grafts that have a remaining native kidney. Specific aim 2 will seek to identify the molecular and cellular basis for the dramatic strain differences in susceptibility to spontaneous graft acceptance, comparing BALB/c (rejected) and DBA/2 (accepted) kidneys (both H-2d) in B6 recipients. We will test for strain differences in DC function, cytokine/chemokine/complement mediators, minor histocompatibility antigens and ability to generate Foxp3 cells. Follow-up experiments seek to alter identified factors to promote acceptance of BALB/c kidneys. Specific aim 3 visualize interaction of Foxp3+ cells with other T cells, recipient and donor dendritic cells and tubular cells in accepting and rejecting renal grafts using a custom real time confocal, multiphoton endomicroscope. This system permits observations of the same graft over time, live video recording, delineation of 3-4 different cell types and computer assisted measurement of cell motility and interactions. These studies are designed to reveal the significance and in vivo function of Foxp3+ Treg in murine renal allograft acceptance and provide insights potentially applicable in clinical renal transplantation. PUBLIC HEALTH RELEVANCE: The mouse kidney transplantation system offers an optimal combination of fidelity to human pathological processes, a rich spectrum of outcomes without drugs, and the ability to manipulate and observe the components of the immune response for mechanistic studies. These studies will provide insights into Treg physiology in renal allografts that will clarify their therapeutic relevance and reveal features that distinguish pathologic from beneficial intragraft infiltrates, of considerable importance in clinical transplantation and tolerance induction studies.

描述（由申请人提供）：该提案解决了对 Foxp3+/T 调节细胞在各种操作耐受性环境下接受生理相关的完全 MHC 不相容器官移植物中的作用的明确证据的需求。在资助的第一年 (ARRA)，MHC 不匹配的小鼠肾同种异体移植物的自发接受被证明完全依赖于 Foxp3 细胞，使用白喉毒素 (DT) 暂时耗尽 B6 中的循环和移植内 Foxp3 细胞。具有维持生命的 DBA 的 Foxp3DTR 小鼠/2同种异体肾移植物。接受的移植物具有独特的结节状血管周围 Foxp3 细胞和树突状细胞 (DC) 浸润。 Foxp3细胞的缺失引起结节聚集物的溶解以及T细胞对皮质、肾小管和血管的广泛浸润，这是典型的急性排斥反应，并表现为BUN在7天内升高。类似的 Foxp3 细胞结节聚集体出现在接受混合嵌合耐受诱导方案 MHC 不匹配肾脏的患者移植物中。具体目标 1 将这些观察扩展到其他形式的诱导耐受、混合嵌合和新生儿耐受，其中一些预计是 Foxp3 依赖性的；具有删除机制的那些预计将独立于 Foxp3。我们将寻找移植物中独特的病理特征，表明Treg活性，例如潜伏相关蛋白（LAP）和Foxp3的共表达或吲哚胺2,3-双加氧酶（IDO）的DC表达，这可能用于区分foxp3依赖于foxp3 独立的移植接受形式。移植物和淋巴器官中细胞的供体反应性和调节潜力将通过 ELISPOT 进行测试，并在不同形式的接受和急性排斥之间进行比较。系统耐受性对移植物内浸润的依赖性将通过移植肾切除术在接受了接受的肾脏和具有剩余天然肾脏的皮肤移植的受者中进行测试。具体目标 2 将寻求通过比较 B6 受者中的 BALB/c（拒绝）和 DBA/2（接受）肾脏（均为 H-2d）来确定自发移植物接受易感性的巨大菌株差异的分子和细胞基础。我们将测试 DC 功能、细胞因子/趋化因子/补体介质、次要组织相容性抗原和生成 Foxp3 细胞的能力方面的菌株差异。后续实验试图改变已确定的因素以促进 BALB/c 肾脏的接受。具体目标 3 使用定制的实时共焦、多光子内窥镜可视化 Foxp3+ 细胞与其他 T 细胞、受体和供体树突状细胞以及肾小管细胞在接受和排斥肾移植物时的相互作用。该系统允许随着时间的推移观察同一移植物、实时视频记录、描绘 3-4 种不同的细胞类型以及计算机辅助测量细胞运动和相互作用。这些研究旨在揭示 Foxp3+ Treg 在小鼠肾同种异体移植接受中的重要性和体内功能，并提供可能适用于临床肾移植的见解。 公共健康相关性：小鼠肾移植系统提供了对人类病理过程的保真度、无需药物即可获得丰富结果的最佳组合，以及操纵和观察免疫反应组成部分以进行机制研究的能力。这些研究将深入了解同种异体肾移植物中的 Treg 生理学，阐明其治疗相关性，并揭示区分病理性和有益移植物内浸润的特征，这在临床移植和耐受诱导研究中具有相当重要的意义。