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: 8462241
• 负责人: MEI-LUN WANG
• 金额: $ 32.61万
• 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/8462241
• 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.

描述（由申请人提供）：胃食管反流病 (GERD) 影响着美国数百万人。除了其重大的社会经济负担外，长期的 GERD 食管炎还是巴雷特食管的危险因素，巴雷特食管是一种与食管腺癌发展相关的癌前病变。因此，了解导致酸诱发食管炎的机制不仅对于治疗这种常见疾病至关重要，而且与癌症预防领域相关。众所周知，长期接触胃酸会导致食管损伤，导致食管上皮棘层细胞坏死。此外，在动物和人类 GERD 模型中，食管上皮中促炎基因的表达均增强。然而，酸诱导的坏死导致食管上皮中趋化因子和粘附分子的诱导的机制完全未知。在这项提议中，我们现在证明，原代和永生化的非转化人食管上皮细胞通过 Toll 样受体 (TLR) 信号传导诱导与 GERD 发病机制相关的促炎基因，作为先天免疫效应细胞自主发挥功能。我们的初步数据进一步表明，在没有病原体刺激的情况下，坏死食管上皮细胞释放的物质（损伤相关分子模式，或 DAMP）可以通过 TLR2 和 TLR3 依赖性信号转导途径诱导 IL-8、ICAM-1 和 RANTES 的表达。因此，我们的总体假设是，TLR2 和 TLR3 信号传导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。通过细胞信号传导研究和新型胃食管反流病体外器官细胞培养模型，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中TLR2和TLR3信号传导的机制（具体目标1）。我们还将在体外鉴定纯化的内源性 DAMP 刺激 TLR2 和 TLR3 所激活的信号转导途径（具体目标 2）。最后，我们将使用体外和体内方法，使用靶向删除 TLR2、TLR3 和下游接头分子 TRIF、MyD88 和 TRAF6 的小鼠，确定 TLR 信号传导在小鼠腐蚀性食管损伤模型中的作用（具体目标 3）。来自这些突变小鼠的原代食管上皮细胞将用于探究目标 1 和 2 中研究的信号通路。总而言之，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。