Aging lymphocytes: senescence programs in lymphocytes relevant to human inflammatory bowel disease

项目总结/摘要 FOXP 3 + CD 4+细胞在克罗恩病（CD）患者的发炎粘膜内扩增;然而， 肠道炎症的持续存在表明这些CD相关的CD 4 + FOXP 3+细胞缺乏典型的调节性， T细胞（Treg）抑制功能。FOXP 3 + Treg中衰老的功能和治疗意义 细胞未知。这项应用的长期目标是剖析驱动遗传学的表观遗传机制。 CD 4 + FOXP 3+细胞在肠道炎症中的分化和功能。因此 本提案的总体目标是确定导致FOXP 3+细胞衰老的机制 胃肠道炎症和测试治疗的机会，以恢复调节功能。核心假设是， CD相关的FOXP 3+细胞代表衰老状态并促进肠道炎症。此外，委员会认为， 这种新的细胞表型（衰老FOXP 3+细胞，snFOXP 3）代表了潜在的治疗靶点。这 通过对来自板层的CD 4 + FOXP 3+细胞的单细胞转录组学分析， 由本申请人的实验室生产的CD患者的固有。基本原理是， 对于snFOXP 3细胞在炎症环境中的发育和功能至关重要， TREG导向疗法，包括用于过继细胞疗法试验的人类TREGS的先进工程。 在强有力的初步数据的指导下，这一假设将通过追求三个具体目标来检验：1）为什么 FOXP 3+细胞在缺乏BMI 1？的情况下变得衰老; 2)为什么FOXP 3+细胞在克罗恩病中衰老 疾病？为什么snFOXP 3细胞是促炎性的？根据第一个目标，申请人将利用 表观遗传测定，其在申请人手中已被确定为可行的，以确定BMI 1 抑制FOXP 3+细胞中ETS 1调节的衰老程序。抗体和测定试剂， 将被使用。根据第二个目标，申请人将利用生物化学方法， 处理培养的TREGS，以确定TNFα诱导的对BMI 1介导的抑制的破坏， 衰老最后，在第三个目标下，申请人将使用一种新型的体外共培养系统来建立 snFOXP 3细胞的促炎能力，并建立治疗靶点。方法是创新的 因为它偏离了利用单个细胞的CD 4 + FOXP 3+细胞的功能表征的现状 方法，和一种机制的方法来定义表观遗传事件，导致致病snFOXP 3 cell.结论后，申请人将了解导致以下疾病发展的表观遗传机制 snFOXP 3细胞在肠道炎症环境中的作用。这一贡献是重要的，因为snFOXP 3是 在克罗恩病病变中扩展，是促炎性的，并且代表重要的表观遗传药物靶标。这 该项目是第一个阐明调节衰老激活的标志性和精确的表观遗传事件的项目 在肠道炎症中的TREGS，因此推进了工程细胞疗法的治疗领域， 自身免疫性疾病

PROJECT SUMMARY/ABSTRACT FOXP3+ CD4+ cells are expanded within the inflamed mucosa of Crohn’s disease (CD) patients; yet the persistence of intestinal inflammation suggests these CD associated CD4+FOXP3+ cells lack typical regulatory T cell (TREG) suppressive function. The functional and therapeutic implications of senescence in FOXP3+ TREG cells is unknown. The long-term goal of this application is to dissect the epigenetic mechanisms driving the differentiation and function of CD4+FOXP3+ cells in the setting of intestinal inflammation. Consequently, the overall objective of this proposal is to identify the mechanisms leading to senescent FOXP3+ cells in the setting of GI inflammation and test therapeutic opportunities to restore regulatory function. The central hypothesis is that CD associated FOXP3+ cells represent a senescent state and contribute to intestinal inflammation. Furthermore, this new cellular phenotype (senescent FOXP3+ cell, snFOXP3) represents a potential therapeutic target. This hypothesis was formulated through single cell transcriptomic analysis of CD4+FOXP3+ cells from the lamina propria of CD patients, produced by this applicant’s laboratory. The rationale is that defining the mechanisms critical to the development and function of snFOXP3 cells in the setting of inflammation will uncover targets for TREG-directed therapies, including the advanced engineering of human TREGS for adoptive cell therapy trials. Guided by strong preliminary data, this hypothesis will be tested by pursuing three specific aims: 1) Why do FOXP3+ cells become senescent in the absence of BMI1?; 2) Why are FOXP3+ cells senescent in Crohn’s disease?; and 3) Why are snFOXP3 cells proinflammatory? Under the first aim, the applicant will utilize epigenetic assays, which have been established as feasible in the applicant’s hands, to establish BMI1 repression on an ETS1-regulated senescence program in FOXP3+ cells. Antibodies and assay reagents that are already on hand will be used. Under the second aim, the applicant will utilize a biochemistry approach with treatment of cultured TREGS to determine a TNFα-induced disruption of BMI1-mediated repression of senescence. Finally, under the third aim, the applicant will use a novel in vitro co-culture system to establish the proinflammatory capacity of snFOXP3 cells and establish therapeutic targets. The approach is innovative because it departs from the status quo of functional characterization of CD4+FOXP3+ cells utilizing single cell approaches, and a mechanistic approach to define the epigenetic events that lead to a pathogenic snFOXP3 cell. Upon conclusion, the applicant will understand the epigenetic mechanisms leading to the development of snFOXP3 cells in the setting of intestinal inflammation. This contribution is significant as the snFOXP3 is expanded in the Crohn’s lesion, is proinflammatory, and represents an important epigenetic drug target. This project is the first to articulate both signature and precise epigenetic events that regulate activation of senescence in TREGS in intestinal inflammation, therefore advancing the field of therapeutics in engineered cell therapies in the setting of autoimmune diseases.