Preservation of mucosal barrier in surgical diseases

外科疾病中粘膜屏障的保护

• 批准号: 8499291
• 负责人: Xiao-Di Tan
• 金额: $ 29.11万
• 依托单位: LURIE CHILDREN'S HOSPITAL OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-12-15 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8499291
• 关键词: 3' Untranslated Regions; Abbreviations; Achievement; Actins; Acute; Affect; Animals; Attenuated; Binding; Binding Proteins; Biological Preservation; Critical Illness; Data; Development; Disease; Down-Regulation; EGF gene; EMSA; Electrolyte Disorder; Electrophoretic Mobility Shift Assay; Elements; Enterocytes; Epididymis; Epithelial; Epithelial Cells; Epithelium; Functional disorder; Gastrointestinal Injury; Gene Expression; Genes; Glycoproteins; Goals; Human; Immune; Impairment; Individual; Inflammation; Inflammation Mediators; Inflammatory; Injury; Integrins; Intensive Care Units; Intestinal Mucosa; Intestines; Lead; Lipopolysaccharides; Liquid substance; Luciferases; MAPK8 gene; Maintenance; Malnutrition; Mediating; Mediator of activation protein; Medical; Messenger RNA; MicroRNAs; Molecular; Molecular Biology; Morbidity - disease rate; Multiple Organ Failure; Mus; N-terminal; Operative Surgical Procedures; Pathogenesis; Patients; Phosphatidylserines; Phosphotransferases; Physiological; Play; Process; Protein Kinase; RNA-Binding Proteins; Regulation; Repression; Research; Rodent; Role; Sepsis; Septic Shock; Shock; Signal Pathway; Signal Transduction; Signaling Molecule; Site-Directed Mutagenesis; Small Intestines; Stimulus; Systemic infection; Technology; Testing; Tissues; Transglutaminases; Transplantation; Trauma; Villus; Work; Wound Healing; attenuation; body system; cell motility; effective therapy; follow-up; gastrointestinal; heat injury; in vivo; intestinal epithelium; macrophage; migration; milk fat globule; mortality; myristoylated alanine-rich C kinase substrate; novel; novel strategies; novel therapeutics; promoter; public health relevance; repaired; response; septic; translational medicine

DESCRIPTION (provided by applicant): Surgical patients with critical disorders such as thermal injury, sever trauma, shock, and sepsis are commonly complicated with the gastrointestinal mucosal injury, which can lead to the development of systemic infection, septic shock, and multiple organ failure. Restitution is an important process for repair of disrupted intestinal lining. Previous studies have suggested that intestinal epithelial restitution is delayed in critically ill individuals. Our recent work has revealed that milk fat globule-EGF factor 8 (MFG-E8) is a macrophage-derived intestinal epithelial restitution factor. MFG-E8 promotes intestinal epithelial wound-healing via a PKC5 dependent manner. In sepsis, intestinal injury is associated with downregulation of MFG-E8, which contributes to the impairment of restitution. In this proposal, therefore, we will test a central hypothesis that under the physiological state, MFG-E8 maintains intestinal epithelial cell migration via a typical signal pathway, whereas under the septic condition, inflammatory mediators down-regulate MFG-E8 expression, which in turn impairs intestinal epithelial cell restitution. In Specific Aim 1, we will define the molecular and signaling mechanism through which MFG-E8 enhances intestinal epithelial restitution. Our preliminary data suggest that MFG-E8 activates several PKC5-associated intracellular signal molecules. Therefore, we will investigate the role of these molecules in MFG-E8-induced intestinal epithelial cell migration using molecular biology and gene knockdown approaches. In Specific Aim 2, we will focus on understanding the cause of down-regulation of intestinal MFG-E8 gene expression and impairment of MFG-E8-dependent intestinal restitution in sepsis. Our preliminary data suggest that a sepsis-derived inflammatory mediator mimics septic effect on intestinal MFG-E8 gene expression and restitution. Thus, we will test the hypothesis that this mediator inhibits intestinal MFG-E8 gene expression, which subsequently results in impairment of intestinal restitution. In Specific Aim 3, we will investigate the molecular mechanism underlying the inhibition of MFG-E8 expression in macrophages by septic insult-derived inflammatory stimulus. Our preliminary data show that repression of MFG-E8 gene expression by an inflammatory stimulus is involved in affecting the activities of promoter and mRNA 3'-untranslated region (3'-UTR) of MFG-E8. To follow up on these findings, we will characterize binding motifs in the MFG-E8 gene promoter and their binding proteins, and study their role in suppression of MFG-E8 gene expression during inflammation. The standard approach for analysis of the gene promoter function will be applied. In addition, we will use the microRNA and site-directed mutagenesis technologies to study the role of 3'-UTR and microRNAs in regulation of MFG-E8 gene expression by an inflammatory stimulus. Achievement of these specific aims and confirmation of our hypotheses will provide the novel mechanistic information that allows us to understand the crucial role of MFG-E8 in the gut and its physio/pathological implications. Our studies will ultimately lead to development of novel strategies for management and treatment of intestinal injury in critically ill patients.

描述（由申请人提供）：患有严重疾病的手术患者，例如热损伤，严重创伤，休克和败血症，通常会因胃肠道粘膜损伤而变得复杂，这可能导致全身感染，败血性休克和多器官衰竭的发展。恢复原状是修复干扰肠壁的重要过程。先前的研究表明，重症患者肠上皮恢复原状延迟。我们最近的工作表明，牛奶脂肪球-EGF因子8（MFG-E8）是巨噬细胞衍生的肠上皮恢复因子。 MFG-E8通过PKC5依赖性方式促进肠上皮伤口愈合。在败血症中，肠道损伤与MFG-E8的下调有关，这有助于恢复损害。因此，在该提案中，我们将检验一个中心假设，即在生理状态下，MFG-E8通过典型的信号途径维持肠上皮细胞的迁移，而在化粪池条件下，炎症介体下调MFG-E8的表达，这在转发上会损害肠道上皮细胞的恢复。在特定的目标1中，我们将定义MFG-E8通过该机制增强肠上皮恢复原状的分子和信号传导机制。我们的初步数据表明，MFG-E8激活了几个与PKC5相关的细胞内信号分子。因此，我们将使用分子生物学和基因敲低方法研究这些分子在MFG-E8诱导的肠上皮细胞迁移中的作用。在特定的目标2中，我们将重点放在理解肠道MFG-E8基因表达下调的原因和MFG-E8依赖性肠道衰落中的损害。我们的初步数据表明，败血症衍生的炎症介质模仿了化粪池对肠道MFG-E8基因表达和恢复的影响。因此，我们将检验以下假设：该介体抑制肠道MFG-E8基因表达，这随后导致肠道恢复损害。在特定的目标3中，我们将通过化粪池损伤的炎症刺激研究巨噬细胞中MFG-E8表达抑制MFG-E8表达的基本机制。我们的初步数据表明，通过炎症刺激对MFG-E8基因表达的抑制参与了MFG-E8的启动子和mRNA 3'-非翻译区（3'-UTR）的活性。为了跟进这些发现，我们将表征MFG-E8基因启动子及其结合蛋白中的结合基序，并研究它们在炎症过程中抑制MFG-E8基因表达中的作用。将采用分析基因启动子功能的标准方法。此外，我们将使用microRNA和定向诱变技术来研究3'-UTR和microRNA通过炎症刺激调节MFG-E8基因表达的作用。实现这些特定目的并确认我们的假设将提供新的机械信息，使我们能够了解MFG-E8在肠道中的关键作用及其生理/病理意义。我们的研究最终将导致危重患者肠道损伤的管理和治疗的新型策略。