Mechanisms of Non-Canonical NF-kB Activation in Transplant Arteriosclerosis

移植动脉硬化中非典型 NF-kB 激活机制

• 批准号: 9376977
• 负责人: DANIEL JANE-WIT
• 金额: $ 24.9万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-12-15 至 2019-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9376977
• 关键词: Alleles; Allografting; Antibodies; Arteriosclerosis; Attenuated; Binding; Biopsy; Biopsy Specimen; Blood specimen; CD4 Positive T Lymphocytes; Cardiac; Cell Compartmentation; Cell Death; Cell membrane; Cell surface; Chronic; Clinical; Collaborations; Complement; Complement Membrane Attack Complex; Complication; Coronary Vessels; Coronary artery; Deposition; Diagnostic tests; Diffuse; Elements; Endocytosis; Endothelial Cells; FDA approved; Genes; Heart Transplantation; Human; Immune system; Immunodeficient Mouse; Implant; In Vitro; Inflammation; Inflammatory; Interferon Type II; Kidney Transplantation; Label; Lasers; Legal patent; Lesion; Letters; Mediating; Mediator of activation protein; Medical; Methodology; Modeling; NF-kappa B; Organelles; Pathway interactions; Patients; Pharmaceutical Preparations; Phase; Phosphorylation Site; Phosphotransferases; Predictive Value; Process; Protein Isoforms; Protein-Serine-Threonine Kinases; Proteins; Proteomics; Protocols documentation; Publications; Publishing; RNA; Recruitment Activity; Risk Factors; Role; Sampling; Signal Transduction; Site; Small Interfering RNA; Specimen; Structure; T-Cell Activation; T-Lymphocyte; Transcript; Transplant Recipients; Transplantation; Update; Vesicle; Vitronectin; analytical method; base; clinical risk; genome-wide; graft failure; humanized mouse; immunogenicity; in vivo; membrane assembly; mouse model; novel; novel diagnostics; novel marker; peripheral blood; protein expression; renal artery; response; trafficking; trial design

Transplant arteriosclerosis (TA) is a highly-prevalent complication of heart transplantation where diffuse, irreversible stenoses form in graft coronary arteries. Donor specific antibody (DSA) binding MHC on graft endothelial cells (EC) is a strong risk factor for TA, but the mechanism(s) of how DSA causes TA are not well understood. To study this problem, DSA was modeled on EC using high panel reactive antibody (PRA) sera taken from sensitized transplant candidates. PRA sera deposited DSA on EC and activated complement, a system of immune-related proteins that when activated forms pore-like structures called membrane attack complexes (MAC) on cell surfaces. PRA caused MAC assembly on EC which elicited EC activation and enhanced EC immunogenicity of alloimmune CD4+ T cells. These processes had an overall effect of exacerbating TA lesions in human coronary arteries in a humanized mouse model. MAC mediated these effects through a novel effector pathway involving NF-κB-inducing kinase (NIK), a critical mediator of non-canonical NF-κB signaling. This proposal defines mechanisms of how MAC activates NIK in EC, explores how these mechanisms operate in a humanized mouse model for TA, and uses NIK in patient EC as a platform to develop novel diagnostic tests for TA. The Specific Aims proposed herein are unchanged from the original application. Aim 1A and Aim1C, subaims of Specific Aim 1, where endocytosis of MAC was explored as a mechanism for NIK stabilization in vitro and in vivo, were successfully completed during the K99 phase. In the R00 phase, Aim1B of Specific Aim 1, where MAC+Rab5+ subcellular compartments will be isolated to identify how Akt mediates NIK, will be completed. Additionally, in the R00 phase, Aim 2, which explores clinical correlations between NIK and TA in patient biopsy specimens, will be completed. The positive predictive value of NIK expression in two microvessel EC compartments will be assessed in patient samples, and novel markers correlating with TA will be identified using laser capture and RNA profiling of NIK.

移植动脉硬化(TA)是心脏移植的一种高度常见的并发症，其中弥漫性、 移植物冠状动脉形成不可逆的狭窄。供者特异性抗体(DSA)结合移植物上的MHC 血管内皮细胞(EC)是TA的重要危险因素，但DSA导致TA的机制(S)尚不清楚 很好理解。为了研究这一问题，在EC上用高群体反应性抗体(PRA)模拟DSA。 从致敏的移植候选人身上提取的血清。PRA血清在EC上沉积DSA并被激活 补体，一种免疫相关的蛋白质系统，当被激活时形成孔状结构，称为 细胞表面的膜攻击复合体(MAC)。PRA导致EC上的MAC组装，从而引发EC 同种异体免疫的CD4+T细胞的活化和增强EC的免疫原性。这些过程有一个总体 在人源化小鼠模型中加重人冠状动脉TA病变的效果。Mac中介 这些作用是通过一种新的效应通路实现的，它涉及到核因子-κB诱导激酶(NIK)，它是一种关键的 非典型的NF-κB信号转导。该提案定义了MAC如何在EC中激活NIK的机制， 探索这些机制如何在TA的人源化小鼠模型中运行，并在患者EC中使用Nik 作为开发TA新诊断测试的平台。 这里提出的具体目标与最初的申请保持不变。目标A和目标1C， 特定目标1的子目标，其中探讨了MAC的内吞作用作为NIK稳定的机制 体外和体内，在K99阶段成功完成。在R00阶段，Aim1B的特定 目标1，将分离MAC+Rab5+亚细胞隔膜以确定Akt如何调节Nik，将 才能完成。此外，在R00阶段，Aim 2探索了Nik和Nick之间的临床相关性 在患者的TA活检标本中，将完成。NIK蛋白表达在两种肿瘤中的阳性预测价值 将在患者样本中评估微血管EC分室，以及与TA相关的新标记物 将通过激光捕获和Nik的RNA图谱进行鉴定。