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: 8071240
• 负责人: Robert B Colvin
• 金额: $ 43.81万
• 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/8071240
• 关键词: Acute; Address; Allografting; Behavior; Blood Vessels; CD4 Positive T Lymphocytes; Cell Communication; 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）：本提案解决了Foxp3+/T调节细胞在各种操作耐受环境下接受生理相关的完全MHC不相容器官移植中的作用的明确证据的需求。在资助的最初一年（ARRA）， mhc错配的小鼠肾异体移植的自发接受被证明完全依赖于Foxp3细胞，使用白喉毒素（DT）在B6中短暂地消耗循环和植入的Foxp3细胞。Foxp3DTR小鼠移植维持生命的DBA/2同种异体肾。接受的移植物有明显的结节状血管周围浸润的Foxp3细胞和树突状细胞（DC）。foxp3细胞的缺失导致结节聚集体溶解，T细胞广泛浸润皮质、小管和血管，这是典型的急性排斥反应，表现为7天内BUN升高。在混合嵌合耐受诱导方案下接受MHC错配肾脏的患者移植物中也出现类似的Foxp3细胞结节聚集。特异性目标1将把这些观察结果扩展到其他形式的诱导耐受性、混合嵌合和新生儿耐受性，其中一些预计是Foxp3依赖性的；而那些具有删除机制的基因则与Foxp3无关。我们将在移植物中寻找独特的病理特征，表明Treg活性，如潜伏期相关蛋白（LAP）和Foxp3的共表达或吲哚胺2,3-双加氧酶（IDO）的DC表达，可以用来区分Foxp3依赖和Foxp3独立的移植物接受形式。将通过ELISPOT检测供体反应性和细胞在移植物和淋巴器官中的调节潜能，并比较不同形式的接受和急性排斥。系统耐受对移植物浸润的依赖性将通过移植肾和皮肤移植者的移植肾切除术来测试。特异性目标2将通过比较B6受体的BALB/c（拒绝）和DBA/2（接受）肾脏（均为H-2d），寻求确定对自发移植接受易感性的巨大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肾的接受。使用定制的实时共聚焦多光子内窥镜观察Foxp3+细胞与其他T细胞、受体和供体树突状细胞和小管细胞在接受和排斥肾移植中的相互作用。该系统可以随时间观察同一移植物，实时视频记录，描绘3-4种不同细胞类型，并通过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在小鼠肾异体移植接受中的意义和体内功能，并为临床肾移植提供潜在的应用见解。