Complement Induces Inflammasome Assembly in Human Endothelium: Mechanisms and Consequences for Graft Rejection

补体诱导人内皮细胞炎症小体组装：移植物排斥的机制和后果

基本信息

批准号：
9609855
负责人：
Catherine Bingchan Xie
金额：
$ 2.93万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-06-01 至 2022-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9609855
关键词：
Acute Alleles Allogenic Allografting Antibodies Arteries Binding Biological Blood Vessels CASP1 gene Cell Adhesion Molecules Chronic Clathrin Cleaved cell Complement Complement Activation Complement Membrane Attack Complex Complex Data Deposition Development Disease Endocytosis Endosomes Endothelial Cells Endothelium Event Failure Family member Forms Controls Foundations Gene Expression Graft Rejection Heart failure Human Immune Immunodeficient Mouse Immunologics Immunology In Vitro Incidence Infiltration Inflammasome Inflammatory Injury Interferon Type II Interleukin-1 Interleukin-1 Receptors Interleukin-1 beta Isoantibodies Kidney Lead Link Liver Lung Measures Mediating Messenger RNA Modeling Organ Transplantation Pathogenesis Pathway interactions Patient-Focused Outcomes Phenotype Phosphorylation Phosphotransferases Physicians Process Production Protein Biosynthesis Protein Kinase Risk Role Savings Scaffolding Protein Scientist Secondary to Signal Pathway Signal Transduction Solid T cell response T memory cell T-Cell Activation T-Cell Proliferation T-Lymphocyte Therapeutic Uses Training Transplantation Up-Regulation Vascular Diseases Work allograft rejection base career cell fixing chemokine clinically relevant cytokine deep sequencing humanized mouse immune function immunogenicity improved in vitro Model in vivo mouse model new therapeutic target novel prevent receptor binding recruit response transplantation medicine

项目摘要

Solid organ transplantation, the best available treatment for end-stage kidney, liver, lung and heart failure, may fail due to chronic immunological rejection, often taking the form of allograft vasculopathy. Allograft vasculopathy results from recruitment to and activation within the graft vessel wall of IFN-γ-producing graft- reactive host T cells by graft ECs. The development of donor specific antibodies (DSA) that recognize non-self alleles of MHC molecules expressed by graft endothelial cells (ECs) and that fix complement is a major risk for allograft vasculopathy. Complement enhances this process by activating immune functions of the ECs that mediate both recruitment and activation of alloreactive T cells. Binding of high titer panel reactive antibody (PRA) from allo-sensitized transplant candidates, used to model DSA, deposits complement membrane attack complex (MAC) on human ECs, both in culture or in vessel grafts in immunodeficient mouse hosts, and induces gene expression of adhesion molecules and chemokines in a manner dependent upon MAC internalization and non-canonical NF-κB signaling. The mechanism(s) by which MAC potentiates T cell activation, measured as effector memory T cell proliferation and cytokine production in vitro or augmented vasculopathic changes in vivo is unknown. My preliminary data show that MAC induces formation of an active inflammasome in ECs, a previously undescribed phenomenon. Activated caspase-1 in the inflammasome processes pro-IL-1β to active IL-1β and mediates its release. Blocking caspase-1 activity or inhibiting IL-1 signaling with IL-1 receptor antagonist (IL-1Ra) blocks both downstream inflammatory gene expression in ECs secondary to canonical NF-κB activation and also blocks the augmentation of allogeneic T cell response in vitro. I hypothesize that complement activation of the inflammasome in ECs intensifies the host T cell response to graft arterial ECs by increasing local production of IL-1, potentiating allograft vasculopathy. In Specific Aim 1, I will characterize the MAC-induced inflammasome and determine the mechanisms linking MAC to inflammasome assembly. I will examine if kinases of non-canonical NF-κB signaling are linked to inflammasome assembly or IL-1 mRNA and protein synthesis. In Specific Aim 2, I will investigate the biological consequences of alloantibody-induced inflammasome activation in ECs on the alloreactive T cell response that drives allograft vasculopathy. I will determine if this response is also MAC-dependent and if mature IL-1 released from MAC-induced inflammasomes acts on ECs, T cells or both. I will characterize IL-1 dependent changes of the clonal repertoire and subsets of activated alloreactive effector memory T cells using TCR deep sequencing and FACS-analytic phenotyping and the effect of IL-1Ra or caspase-1 inhibition on the alloreactive T cell response in vitro using cultured human ECs. I will use well-developed humanized mouse models of allograft vasculopathy to assess the role of the EC inflammasome in vivo. Successful completion of this project may reveal novel targets for therapies to improve patient outcomes in transplant medicine.

实体器官移植是终末期肾脏、肝脏、肺和心脏的最佳治疗方法失败，可能是由于慢性免疫排斥反应而失败，通常表现为同种异体移植血管病的形式。血管病变是由产生 IFN-γ 的移植物血管壁募集和激活引起的通过移植 EC 产生反应性宿主 T 细胞识别非自身的供体特异性抗体 (DSA) 的开发。移植物内皮细胞 (EC) 表达的 MHC 分子等位基因和固定补体是主要风险补体通过激活 EC 的免疫功能来增强这一过程。介导同种异体反应性 T 细胞的募集和激活。 (PRA) 来自同种异体致敏移植候选者，用于模拟 DSA，沉积补体膜攻击人类 EC 上的复合物（MAC），无论是在培养物中还是在免疫缺陷小鼠宿主的血管移植物中，以及以依赖于 MAC 的方式诱导粘附分子和趋化因子的基因表达 MAC 增强 T 细胞的机制。激活，以效应记忆 T 细胞增殖和细胞因子产生体外或增强来衡量体内血管病变的变化尚不清楚。我的初步数据表明，MAC 会诱导活性物质的形成。 EC 中的炎症小体，这是炎症小体内激活的 caspase-1 现象。将 pro-IL-1β 加工成活性 IL-1β 并介导其释放，阻断 caspase-1 活性或抑制 IL-1。 IL-1 受体拮抗剂 (IL-1Ra) 信号传导可阻断 EC 中下游炎症基因的表达继发于典型的 NF-κB 激活，也可阻断同种异体 T 细胞反应的增强我认为 EC 中炎症小体的补体激活会增强宿主 T 细胞的反应。通过增加局部 IL-1 的产生来移植动脉 EC，从而增强同种异体移植血管病变。 1，我将描述 MAC 诱导的炎症小体的特征，并确定将 MAC 与炎症小体联系起来的机制我将检查非典型 NF-κB 信号传导激酶是否与炎症小体组装有关。在特定目标 2 中，我将研究炎症小体组装或 IL-1 mRNA 和蛋白质合成。同种抗体诱导的 EC 炎症小体激活对同种异体反应性 T 细胞反应的影响我将确定这种反应是否也依赖于 MAC，以及是否成熟的 IL-1。 MAC 诱导的炎症小体释放的 I 会作用于 EC、T 细胞或两者，我将描述 IL-1 依赖性。使用 TCR Deep 改变激活的同种异体反应性效应记忆 T 细胞的克隆库和子集测序和 FACS 分析表型以及 IL-1Ra 或 caspase-1 抑制对同种异体反应的影响使用培养的人类 EC 进行体外 T 细胞反应我将使用成熟的人源化小鼠模型。同种异体移植血管病评估 EC 炎症小体在体内的作用该项目成功完成。可能会揭示改善移植医学患者预后的治疗新靶标。