Mechanisms of NaCl Absorption in the Human Colon

描述(由申请人提供)： 与炎症性肠病(IBD)或食源性病原体感染相关的腹泻仍然是我们退伍军人患者群体的主要健康问题。尽管医学取得了重大进展，但腹泻病在世界范围内仍导致非常高的发病率和死亡率。因此，更好地了解与这些疾病相关的腹泻的病理生理学对于设计新的和更好的干预策略至关重要。腹泻结果 由于肠道分泌物增加和/或吸收减少。人体肠道电解质吸收的一个主要途径是Na+/H+和Cl-/HCO3-交换器的耦合作用。基因Slc26a3的突变与先天性氯化物腹泻有关，它编码DRA(腺瘤下调)，这是一种已知在介导肠道氯吸收方面发挥关键作用的蛋白质。最近，DRA表达的显著降低被证明是导致IBD患者腹泻的主要机制之一。然而，到目前为止，人们对腹泻疾病中降低DRA表达的分子机制知之甚少。为了阐明DRA在IBD中表达下调的机制，我们的研究集中在肝脏核因子(HNFs，许多转运蛋白表达所必需的转录因子)和miRNAs(已知在IBD患者和IBD动物模型中上调)的作用。我们广泛的初步数据为HNF及其激活剂地塞米松(DEX)和共轭亚油酸(Clas)以及miRNAs对DRA表达的调控提供了强有力的证据。基于这些数据，我们假设炎症肠道中HNFS的下调和某些miRNAs的上调可能分别通过转录和转录后机制导致DRA表达的下降。因此，目前的应用旨在利用体外(目标1和2)和体内模型(目标3)来研究HNFS和miRNAs对DRA的调节如下：目的1.阐明HNFS对DRA启动子调控的详细机制，降低IBD中DRA的表达，并建立DEX和Clas上调DRA的分子基础；目的2：广泛分析miRNAs对DRA转录后调控的机制；以及目标3：在野生型和IL-10基因敲除的小鼠模型中分析HNFS和miRs对DRA的调节。这些研究的结果将确定HNFS在基础、炎症或CLA/DEX介导的DRA上调中的作用，并将首次确定MIR在正常和炎症条件下肠道盐吸收调节中的作用。这些研究还应该确定干预IBD相关性腹泻的新靶点。

DESCRIPTION (provided by applicant): Diarrhea associated with inflammatory bowel diseases (IBD) or infections by food-borne pathogens remains a major health problem in our veteran patient population. Despite major medical advances, diarrheal diseases still result in very high morbidity and mortality worldwide. Therefore, a better understanding of the pathophysiology of diarrhea associated with these diseases is critical for designing novel and superior strategies for intervention. Diarrhea results from increased intestinal secretion and/or decreased absorption. A major route of electrolyte absorption in the human intestine involves coupled operation of Na+/H+ and Cl-/HCO3- exchangers. The gene Slc26a3, whose mutations are associated with congenital chloride diarrhea, encodes DRA (Down-Regulated in Adenoma), a protein known to play a critical role in mediating intestinal chloride absorption. Recently, significant reduction of DRA expression has been shown to be one of the major mechanisms responsible for diarrhea in IBD patients. To date, however, very little is known about the molecular mechanisms involved in decreasing DRA expression in diarrheal diseases. In order to elucidate the mechanisms underlying the downregulation of DRA expression in IBD, our studies focused on the role of hepatic nuclear factors (HNFs, transcription factors essential for the expression of many transport proteins) and miRNAs (known to be upregulated in IBD patients and animal models of IBD). Our extensive preliminary data provided strong evidence for the regulation of DRA expression by both HNFs and their activators dexamenthasone (DEX) and conjugated linoleic acids (CLAs), as well as miRNAs. Based on these data, we hypothesize that a down-regulation of HNFs and upregulation of certain miRNAs in inflamed gut may underlie the decrease in DRA expression via transcriptional and post-transcriptional mechanisms, respectively. The current application is, therefore, designed to investigate the regulation of DRA by HNFs and miRNAs utilizing both in vitro (Aims 1 & 2) and in vivo models (Aim 3) as follows: Aim 1. Elucidate detailed mechanisms underlying basal regulation of DRA promoter by HNFs, in decreased DRA expression in IBD and to establish the molecular basis for the upregulation of DRA by DEX and CLAs; Aim 2: Extensively analyze the mechanisms underlying post-transcriptional regulation of DRA by miRNAs; and Aim 3: Analyze the regulation of DRA by HNFs and miRs in wild type and IL-10 knock out mouse model for IBD. The outcome of these studies should define the role of HNFs in basal, inflammatory or CLA/DEX-mediated upregulation of DRA, and will also establish for the first time, the role of miRs in the modulation of intestinal NaCl absorption under normal and inflammatory conditions. These studies should also identify novel targets for intervention in IBD associated diarrhea.