Estrogen-mediated immune regulation in human and experimental inflammatory bowel disease

项目摘要/摘要 调节性T细胞(Treg)免疫抑制对于维持对不同种类的 肠粘膜中的潜在抗原。炎症性肠病(IBD)、慢性肠病 炎症压倒了局部Treg功能，使炎症持续存在。我们以前的工作和 另一些研究发现17β-雌二醇(雌激素，E_2)信号在促进Treg分化中起重要作用 和功能。E2信号通过两个核受体，α和β(ERα，ERβ)来调节基因 靶细胞中的转录。尽管ERα和ERβ具有很高的序列同源性，但它们介导不同的 经常在基因调控上起相反的作用。在之前发表的研究中，我们证明了平衡的转移 向ERα传递的E2信号通常是促炎的，而向ERβ转变通常是保护性的。 在最近对炎症性肠病患者样本的初步研究中，我们观察到ERβ的表达显著降低 在活动性克罗恩病(CD)女性的肠道活检组织和外周T细胞中。我们还发现 ERβ缺陷T细胞对体外转化生长因子β依赖的Treg分化和ERβ缺失具有抵抗力 在自发性回肠炎模型(SAMP/YitFC，“SAMP”小鼠)中，Treg显著受损 转录和功能反应，加剧炎症。因此，这一目标是 项目是确定改变的E2信号影响的分子和细胞机制(S) Treg的分化和功能，有助于肠道炎症。雌二醇组的作用机制 信号与炎症信号的相互作用影响免疫细胞功能的机制还知之甚少。这 一项提案试图通过我们的中心假设来解决这一知识差距，即调控失调的E2信号 有助于Treg转录重塑和功能丧失，促进持续的肠道 炎症性肠病。在目标1中，我们建议描述ERα-和ERβ的分子机制 与原代T细胞转化生长因子β下游信号的相互作用，影响转化生长因子β依赖的foxp3的表达 和功能。目标2将确定重新平衡Treg特异性E2信号对肠道的功能影响 体内炎症，验证了我们的假设，即增强Treg特异性ERβ信号可能预防和/或 挽救肠道炎症。实验将包括将表达ERβ的Tregs过继转移到SAMP 小鼠，以及使用T细胞依赖的结肠炎模型的体内试验。在目标3中，我们计划确定 新型MaxCyte在CD患者树突中重新平衡E2信号的转录和功能效应 转染技术，评估ERα和ERβ对(I)转化生长因子β依赖的foxp3诱导的作用 幼稚T细胞和(Ii)Tregs的体外抑制功能。圆满完成我们提出的目标 将为E2信号在Tregs中的功能影响提供关键的机制洞察，一个正在研究中的 对多种表现出ER表达和/或失调的疾病具有广泛适用性的区域 Treg功能的激活和随后的减少。

Project Summary/Abstract Regulatory T cell (Treg) immunosuppression is critical for maintaining immune tolerance to a diverse array of potential antigens in the intestinal mucosa. In patients with inflammatory bowel disease (IBD), chronic intestinal inflammation overwhelms local Treg function, allowing inflammation to persist. Our previous work and that of others have identified important roles for 17β-estradiol (estrogen, E2) signaling in promoting Treg differentiation and function. E2 signals through two nuclear receptors, alpha and beta (ERα, ERβ), to modulate gene transcription in target cells. Although they share high sequence homology, ERα and ERβ mediate distinct and often opposing functions on gene regulation. In previously published work, we showed that shifting the balance of E2 signaling towards ERα is generally pro-inflammatory, whereas shifting towards ERβ is generally protective. In recent preliminary studies using IBD patient samples, we observed significantly diminished ERβ expression in intestinal biopsy tissues and peripheral T cells from females with active Crohn’s disease (CD). We also found that ERβ-deficient T cells are resistant to ex vivo, TGFβ-dependent Treg differentiation, and that deletion of ERβ in a spontaneous ileitis model (SAMP/YitFC, “SAMP” mice) results in significant impairment of Treg transcriptional and functional responses, contributing to exacerbated inflammation. Therefore, the goal of this project is to determine the molecular and cellular mechanism(s) by which altered E2 signaling impacts Treg differentiation and function, contributing to intestinal inflammation. The mechanisms by which E2 signaling cross-talks with inflammatory signals to influence immune cell function are poorly understood. This proposal seeks to address this knowledge gap through our central hypothesis that dysregulated E2 signaling contributes to Treg transcriptional remodeling and loss-of-function, facilitating sustained intestinal inflammation in IBD. In Aim 1, we propose to delineate the molecular mechanisms by which ERα- and ERβ cross-talk with signaling downstream of TGFβ in primary T cells, influencing TGFβ-dependent Foxp3 expression and function. Aim 2 will determine the functional impact of rebalancing Treg-specific E2 signaling on intestinal inflammation in vivo, testing our hypothesis that augmenting Treg-specific ERβ signaling may prevent and/or rescue intestinal inflammation. Experiments will include adoptive transfer of ERβ-expressing Tregs to SAMP mice, as well as in vivo assays using a T cell-dependent colitis model. In Aim 3, we plan to determine the transcriptional and functional effects of rebalancing E2 signaling in CD patient Tregs using novel MaxCyte transfection technology, assessing ERα- and ERβ-specific effects on (i) TGFβ-dependent Foxp3 induction in naïve T cells and (ii) ex vivo suppressive function of Tregs. Successful completion of our proposed Aims will provide key mechanistic insight into the functional impact of E2 signaling in Tregs, an under-studied area with broad applicability to numerous diseases exhibiting dysregulation of ER expression and/or activation and subsequent reductions in Treg function.