Toll-like Receptor Signaling in the Esophageal Epithelium

食管上皮中的 Toll 样受体信号传导

• 批准号: 8278005
• 负责人: MEI-LUN WANG
• 金额: $ 33.79万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-15 至 2015-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8278005
• 关键词: Acids; Adhesions; Affect; Animals; Barrett Esophagus; Caustics; Cell Adhesion Molecules; Cell Culture Techniques; Cell physiology; Cells; Chemotaxis; Country; Data; Development; Diagnosis; Disease; Dominant-Negative Mutation; Effector Cell; Epithelial; Epithelial Cells; Epithelium; Esophageal; Esophageal Adenocarcinoma; Esophageal Diseases; Esophageal injury; Esophagitis; Exposure to; Family; Functional disorder; Gastric Acid; Gastroesophageal reflux disease; Gene Expression; Genes; Human; ICAM1 gene; IL8 gene; Immune; Immune response; In Vitro; Individual; Inflammation; Inflammatory; Intercellular adhesion molecule 1; Lead; Leukocytes; Ligands; Link; MAP Kinase Gene; Mediating; Methods; Modeling; Molecular; Mus; Mutant Strains Mice; NF-kappa B; Necrosis; Pathogenesis; Pathway interactions; Patients; Pattern; Play; Precancerous Conditions; Process; RANTES; Receptor Signaling; Reflux; Regulation; Risk; Risk Factors; Role; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Specificity; Stomach; Stratum Spinosum; Swelling; TLR2 gene; TLR3 gene; TRAF6 gene; Testing; Toll-like receptors; Transcriptional Regulation; United States; base; cancer prevention; cell injury; chemokine; common treatment; in vivo; insight; member; mouse model; mutant; neutrophil; novel; overexpression; pathogen; public health relevance; receptor; response; socioeconomics

DESCRIPTION (provided by applicant): Gastroesophageal reflux disease (GERD) affects millions of individuals in the United States. In addition to its significant socioeconomic burden, longstanding GERD esophagitis is a risk factor for Barrett's esophagus, a premalignant condition associated with the development of esophageal adenocarcinoma. Thus, an understanding of the mechanisms which lead to acid-induced esophagitis is not only critical for the treatment of this common disease, and is also relevant to the field of cancer prevention. It is known that prolonged exposure to gastric acid leads to esophageal injury, resulting in cell necrosis in the stratum spinosum of the esophageal epithelium. In addition, the expression of proinflammatory genes is enhanced in the esophageal epithelium in both animal and human models of GERD. However, the mechanisms by which acid-induced necrosis leads to the induction of chemokines and adhesion molecules in the esophageal epithelium are completely unknown. In this proposal, we now show that primary and immortalized non-transformed human esophageal epithelial cells function autonomously as innate immune effector cells through the induction of proinflammatory genes relevant to the pathogenesis of GERD, via Toll-like receptor (TLR) signaling. Our Preliminary Data further demonstrates that substances released by necrotic esophageal epithelial cells (damage-associated molecular patterns, or DAMPs) can induce the expression of IL-8, ICAM-1, and RANTES through TLR2 and TLR3-dependent signal transduction pathways in the absence of pathogenic stimulation. Therefore, our overall hypothesis is that TLR2 and TLR3 signaling enables esophageal epithelial cells to sense molecular patterns associated with cell necrosis, activating signal transduction pathways required for neutrophil chemotaxis and adhesion. Through cell-signaling studies and novel organotypic cell culture models of GERD in vitro, we will identify the DAMPs released by necrotic esophageal epithelial cells and determine the mechanisms by which these DAMPs activate TLR2 and TLR3 signaling in human esophageal epithelial cells in vitro (Specific Aim 1). We will also identify the signal transduction pathways activated by TLR2 and TLR3 stimulation by purified endogenous DAMPs in vitro (Specific Aim 2). Finally, we will determine the role of TLR signaling in a mouse model of caustic esophageal injury using mice with targeted deletions of TLR2, TLR3, and downstream adaptor molecules TRIF, MyD88, and TRAF6, using both ex vivo and in vivo methods (Specific Aim 3). Primary esophageal epithelial cells from these mutant mice will be used to interrogate the signaling pathways studied in Aims 1 and 2. Together, the results of these studies will provide new insights into the mechanisms by which esophageal epithelial cells function as innate immune effector cells in response to esophageal damage, and may impact the diagnosis and treatment of highly prevalent human esophageal diseases. PUBLIC HEALTH RELEVANCE: Gastroesophageal reflux disease (GERD) is highly prevalent in the United States, and carries a significant socioeconomic burden as well as an increased risk for Barrett's esophagus and associated esophageal adenocarcinoma. The studies in this proposal are based upon the hypothesis that Toll-like receptors (TLRs) enable esophageal epithelial cells to sense danger signals released by dead and dying esophageal epithelial cells, thus providing a mechanistic link between acid-induced injury and esophageal inflammation. The results of our studies will elucidate the mechanisms by which acid- induced damage to the esophageal epithelium leads to esophageal inflammation, and may impact the treatment of common esophageal disorders.

描述（由申请人提供）：胃食管反流病（GERD）影响着美国数百万人。除了其显著的社会经济负担外，长期GERD食管炎是Barrett食管的风险因素，Barrett食管是一种与食管腺癌发展相关的癌前病变。因此，了解导致酸诱导性食管炎的机制不仅对治疗这种常见疾病至关重要，而且与癌症预防领域有关。众所周知，长期暴露于胃酸会导致食管损伤，导致食管上皮棘层细胞坏死。此外，在GERD动物和人类模型中，促炎基因的表达在食管上皮中增强。然而，酸诱导的坏死导致食管上皮中趋化因子和粘附分子诱导的机制完全未知。在这个提议中，我们现在表明，原代和永生化的非转化的人食管上皮细胞通过Toll样受体（TLR）信号转导诱导与GERD发病机制相关的促炎基因，自主发挥先天性免疫效应细胞的功能。我们的初步数据进一步表明，坏死食管上皮细胞释放的物质（损伤相关分子模式，或DAMP）可以诱导IL-8，ICAM-1和RANTES的表达，通过TLR 2和TLR 3依赖的信号转导途径，在没有致病性刺激。因此，我们的总体假设是，TLR 2和TLR 3信号使食管上皮细胞能够感知与细胞坏死相关的分子模式，激活中性粒细胞趋化和粘附所需的信号转导途径。通过细胞信号转导研究和新的体外GERD器官型细胞培养模型，我们将鉴定坏死食管上皮细胞释放的DAMP，并确定这些DAMP在体外激活人食管上皮细胞TLR 2和TLR 3信号转导的机制（具体目标1）。我们还将在体外鉴定通过纯化的内源性DAMP刺激TLR 2和TLR 3激活的信号转导途径（特异性目的2）。最后，我们将使用靶向缺失TLR 2、TLR 3和下游衔接分子TRIF、MyD 88和TRAF 6的小鼠，使用离体和体内方法，确定TLR信号传导在腐蚀性食管损伤小鼠模型中的作用（特异性目标3）。来自这些突变小鼠的原代食管上皮细胞将用于询问目的1和2中研究的信号传导途径。总之，这些研究的结果将为食管上皮细胞作为先天免疫效应细胞响应食管损伤的机制提供新的见解，并可能影响高度流行的人类食管疾病的诊断和治疗。 公共卫生关系：胃食管反流病（GERD）在美国非常流行，并带来了显著的社会经济负担以及巴雷特食管和相关食管腺癌的风险增加。该提案中的研究基于Toll样受体（TLR）使食管上皮细胞能够感知死亡和垂死的食管上皮细胞释放的危险信号的假设，从而提供酸诱导的损伤和食管炎症之间的机制联系。我们的研究结果将阐明酸诱导的食管上皮损伤导致食管炎症的机制，并可能影响常见食管疾病的治疗。