TNF-alpha Regulation of Intestinal Paracellular Transport

摘要： 肠上皮细胞间的细胞旁转运或细胞旁渗透受到调控 通过紧密的交汇点。 克罗恩病(CD)患者的肠道紧密连接(TJ)屏障有缺陷，表现为 细胞旁通透性增加。肠道TJ屏障缺陷是导致肠梗阻的重要致病因素 CD使有毒的管腔抗原在细胞旁的渗透增加，从而导致肠道炎症。 肿瘤坏死因子是一种促炎细胞因子，已被证明在 CD的肠道炎症过程。肿瘤坏死因子的一个重要的促炎作用是引起功能性的 肠道TJ屏障开放，导致有害肠腔的细胞旁通透性增加 抗原。肿瘤坏死因子-β诱导的肠道细胞旁通透性增加被认为是一种 CD致肠TJ屏障缺陷及其他炎性状态的重要机制 我的直觉。这项拨款提案的主要目标是阐明细胞和分子机制。 介导肿瘤坏死因子诱导的肠道TJ通透性增加，并测定其潜在的 预防肿瘤坏死因子诱导的TJ屏障缺陷和肠道后续发育的治疗靶点 发炎。为了实现这些目标，我们打算使用体外(由过滤生长的Caco-2组成) 肠上皮细胞)和活体(小鼠肠道灌流系统)模型系统。 我们的初步研究表明，肿瘤坏死因子诱导的肠上皮通透性增加。 部分受ERK 1/2和p38信号级联诱导肌球蛋白轻链激酶增加的调节 (MLCK)基因活性和microRNA诱导occludin基因表达下调。基于我们的 初步研究，我们提出了一个新的假说，即肿瘤坏死因子-β诱导了肠道TJ的增加 通透性部分由ERK1/2和p38信号级联诱导MLCK基因激活和 对microRNA表达的调控。提出的具体目标是：1)阐明细胞内和 介导肿瘤坏死因子诱导的MLCK基因活性和肠道TJ增加的分子过程 通透性；2)阐明了介导肿瘤坏死因子-β诱导的细胞和分子机制。 阻滞素基因和蛋白表达的调控；3)阐明了肿瘤坏死因子的作用机制。 体内诱导的肠通透性增加及靶向治疗意义的确定 TJ屏障功能在肿瘤坏死因子诱导的肠炎中的保护作用

Abstract: Intestinal paracellular transport or paracellular permeation in-between intestinal epithelial cells is regulated by tight junctions. Patients with Crohn's disease (CD) have a defective intestinal tight junction (TJ) barrier, manifested by an increase in paracellular permeability. The defective intestinal TJ barrier is an important pathogenic factor of CD that allows increased paracellular permeation of toxic luminal antigens, leading to intestinal inflammation. Tumor necrosis factor-¿ (TNF-¿), a pro-inflammatory cytokine, has been shown to play a central role in the intestinal inflammation process of CD. An important pro-inflammatory action of TNF-¿ is to cause a functional opening of the intestinal TJ barrier, leading to an increase in paracellular permeation of noxious luminal antigens. The TNF-¿ induced increase in intestinal paracellular permeability has been postulated to be an important mechanism contributing to the intestinal TJ barrier defect in CD and other inflammatory conditions of the gut. The broad objectives of this grant proposal are to elucidate the cellular and molecular mechanisms that mediate the TNF-¿ induced increase in intestinal TJ permeability and to determine the potential therapeutic targets to prevent the TNF-¿ induced defect in TJ barrier and subsequent development of intestinal inflammation. To achieve these objectives, we intend to use both in-vitro (consisting of filter-grown Caco-2 intestinal epithelial cells) and in-vivo (mouse intestinal perfusion system) model systems. Our preliminary studies suggested that the TNF-¿ induced increase in intestinal epithelial TJ permeability was regulated in part by ERK 1/2 and p38 signaling cascade induced increase in myosin light chain kinase (MLCK) gene activity and microRNA induced down-regulation of occludin gene expression. Based on our preliminary studies, we advance a novel hypothesis that the TNF-¿ induced increase in intestinal TJ permeability is mediated in part by ERK1/2 and p38 signaling cascade induced activation of MLCK gene and modulation of microRNA expression. The proposed specific aims are to: 1) elucidate the intracellular and the molecular processes that mediate the TNF-¿ induced increase in MLCK gene activity and intestinal TJ permeability; 2) delineate the cellular and molecular mechanisms that mediate the TNF-¿ induced down- regulation of occludin gene and protein expression; and 3) delineate the mechanisms involved in TNF-¿ induced increase in intestinal permeability in-vivo and determine the therapeutic implications of targeted preservation of TJ barrier function in TNF-¿ induced intestinal inflammation.