A Novel Role of DRA in IBD Pathogenesis

项目摘要/摘要 炎症性肠病(IBD)是一种全球性的健康负担，目前影响着全球约300万人。 美国的发病率在全球范围内呈上升趋势。根据最近的一份报告，IBD的患病率在 在过去的十年里，美国退伍军人数量迅速增加。尽管取得了所有进展，但IBD的发病机制 由于疾病的多因素性质以及治疗选择不充分，目前仍不清楚。 因此，了解IBD的分子病理生理机制对于开发有效的治疗方法至关重要。 医疗模式。几种不同的机制可能解释了IBD的发病机制。在这方面，妥协了 肠屏障是IBD肠道炎症发生的最关键的早期事件之一。 最近的研究表明，上皮离子转运体(如NHE3和CFTR)在维持 屏障完整性。在这一点上，DRA(腺瘤下调或SLC26A3)是关键的转运体 调节哺乳动物肠道对氯的吸收。DRA KO小鼠缺乏粘附性内脏 粘液层，改变了结肠隐窝的增殖动态平衡，更容易受到实验性 结肠炎。DRA在炎症中的重要作用从GWAS进一步得到证明，将DRA确定为一种新的 IBD易感基因。此外，在人IBD患者的结肠粘膜和 重现IBD的小鼠模型。然而，这背后的机制增加了对 DRA丢失引起的炎症反应尚不清楚，需要进行详细的研究。我们的 初步数据为DRA在维持上皮完整性中的新作用提供了强有力的证据。我们的 初步数据表明，DRA的丢失可能在通过多种途径打破上皮屏障方面发挥关键作用 例如，通过直接影响TJ/AJ和/或通过间接(与其Cl-/HCO3-交换有关)的机制 功能、细胞骨架相互作用和/或微生物失调)。此外，通过上调DRA来瞄准DRA 这似乎是一种恢复屏障功能和减轻炎症的新的治疗方法。然而， 在临床前模型中靶向DRA以恢复屏障功能的治疗可行性从未被 调查过了。因此，我们假设：1)DRA的丧失通过降低TJ和 AJ蛋白通过涉及RNA结合蛋白的转录后机制和/或通过直接 与细胞骨架元素的相互作用。Ii)上调DRA功能和表达可缓解 通过恢复肠道屏障功能而引起的炎症。我们的假设将利用最先进的技术进行测试 方法和人/鼠结肠样体、Caco-2细胞和KO或转基因小鼠模型。两个具体的 目的：1.阐明转录后调节肠道屏障的机制 DRA缺乏的功能和TJ/AJ蛋白；以及目的2.阐明DRA的新作用 上调对实验性肠上皮完整性和紧密连接蛋白的影响 来自健康和IBD患者的结肠炎和结肠样病变模型。这些活动的成功结果 研究应该对肠道TJ/AJ调节的分子机制产生关键的见解，并建立 DRA作为治疗IBD的新靶点。此外，这些研究将为申请者提供很好的 成为膜氯化物新作用潜在机制专家的培训工具 肠紧密连接转运蛋白的调节失调和IBD的发病机制，以及培训他在切割- 边缘入路利用肠样、结肠样病变和先进的成像技术将他进化成 独立退伍军人事务部调查员专注于IBD研究。

Project Summary/Abstract Inflammatory Bowel Disease (IBD) is a global health burden currently affecting around 3 million people in the United States with increasing incidence worldwide. According to a recent report, prevalence of IBD among US veterans rapidly increased during the past decade. Despite all the advancement, pathogenesis of IBD is still unclear due to the multifactorial nature of the disease and the treatment options are not adequate. Therefore, understanding the molecular pathophysiology of IBD is critical for developing effective therapeutic modalities. Several different mechanisms likely account for the IBD pathogenesis. In this regard, compromised Intestinal barrier is one of the most critical early event linked to the onset of intestinal inflammation in IBD. Recent studies have implicated novel roles of epithelial ion transporters (e.g. NHE3 and CFTR) in maintaining barrier integrity. In this regard, DRA (Down Regulated in Adenoma or SLC26A3) is the key transporter mediating chloride absorption in the mammalian intestine. DRA KO mice exhibit lack of an adherent inner mucus layer, altered proliferative homeostasis of the colonic crypts, and are more susceptible to experimental colitis. The important role of DRA in inflammation is further evident from GWAS, identifying DRA as a novel IBD susceptibility gene. Further, DRA levels are severely depleted in human IBD patient colonic mucosa and in mouse models recapitulating IBD. However, the mechanisms underlying this increased susceptibility to inflammation in response to loss of DRA are not known and warrant detailed investigations. Our preliminary data provide strong evidence for a novel role of DRA in maintenance of epithelial integrity. Our preliminary data suggest that loss of DRA may play a key role in breaching the epithelial barrier via multiple mechanisms e.g. either by directly affecting the TJ/AJ and/or via indirect (related to its Cl-/HCO3- exchange function, its cytoskeletal interactions and/or microbial dysbiosis). In addition, targeting DRA via its upregulation appears to be a novel therapeutic approach to restore barrier function and alleviating inflammation. However, the therapeutic feasibility of targeting DRA to restore barrier function in preclinical models has never been investigated. Therefore, we hypothesize that i) DRA loss perturbs intestinal barrier function by reducing TJ and AJ protein expression via posttranscriptional mechanisms involving RNA binding proteins and/or via direct interactions with cytoskeletal elements. ii) Upregulation of DRA function and expression can alleviate inflammation via restoration of intestinal barrier function. Our hypothesis will be tested utilizing state-of-the-art approaches and human/mouse colonoids, Caco-2 cells and KO or transgenic mouse models. Two Specific Aims have been proposed: Aim 1. Elucidate the post-transcriptional mechanisms modulating gut barrier function and TJ/AJ proteins in response to DRA deficiency; and Aim 2. Elucidate the novel role of DRA upregulation in maintenance of intestinal epithelial integrity and tight junction proteins in experimental models of colitis and colonoids derived from healthy and IBD patients. Successful outcome of these studies should yield critical insights into the molecular mechanisms of intestinal TJ/AJ regulation and establish DRA as a novel therapeutic target to manage IBD. Additionally, these studies will provide the applicant a great training vehicle for becoming an expert in mechanisms underlying the novel role of a membrane chloride transporter in intestinal tight junction dysregulation and pathogenesis of IBD, as well as train him in cutting- edge approaches utilizing enteroids, colonoids and advanced imaging techniques to evolve him into an independent VA investigator focused on IBD research.