Novel Role of Pre-B-Cell Colony Enhancing Factor in Acute Lung Injury (ALI)

前 B 细胞集落增强因子在急性肺损伤 (ALI) 中的新作用

• 批准号: 7204218
• 负责人: Shui Qing Ye
• 金额: $ 30.71万
• 依托单位: UNIVERSITY OF MISSOURI-COLUMBIA
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7204218
• 关键词: Acute; Acute Lung Injury; Affect; Animals; Arteries; Biological Assay; Biological Markers; Blood Vessels; Candidate Disease Gene; Cell Count; Cells; Cytochrome c Reductase; Data; Development; Electrical Resistance; Electrophoretic Mobility Shift Assay; Endothelial Cells; Endotoxins; Environmental air flow; Epithelial Cells; Evaluation; Evans blue stain; Extravasation; Figs - dietary; Functional disorder; Gene Expression; Genes; Genetic; Genetic Polymorphism; Genetic Transcription; Genomics; Goals; Haplotypes; Histone Deacetylase; Human; IL8 gene; In Vitro; Inflammatory; Knock-out; Lead; Lung; Mechanics; Mediating; Modality; Modeling; Molecular; Morphology; Mus; Myosin Light Chain Kinase; Myosin Light Chains; Nicotinamide adenine dinucleotide; PBEF Gene; Pathogenesis; Pathway interactions; Patients; Permeability; Phosphorylation; Physiological; Predisposition; Promoter Regions; Protein Overexpression; Proteins; Publishing; Pulmonary Edema; Pulmonary artery structure; Rate; Recombinants; Regulation; Regulatory Element; Reporter Genes; Research Personnel; Risk; Risk Factors; Role; Signal Transduction Pathway; Single Nucleotide Polymorphism; Small Interfering RNA; Stimulus; Stress Fibers; Structure of parenchyma of lung; System; Therapeutic; Thrombin Receptor; Tidal Volume; Transcriptional Regulation; Tumor Necrosis Factor-alpha; Up-Regulation; Variant; Vascular Permeabilities; Weight; Work; cell type; cytokine; genetic variant; human TNF protein; in vivo; insight; lung injury; macrophage; novel; novel diagnostics; pre-B-cell colony-enhancing factor protein; promoter; research study; response; tissue culture; translational study

DESCRIPTION (provided by applicant): Pre-B-cell colony enhancing factor (PBEF) is a unique, little-studied cytokine, whose relevance to the lung pathophysiology is unknown. Recent work by the PI has provided the first evidence of PBEF expression in lung tissues and overexpression in acute lung injury (ALI) as well as the association of PBEF genetic variants with susceptibility to ALI. The haplotype weighted analysis of single nucleotide polymorphism (SNP) T-1001G and C-1543T in the PBEF gene promoter revealed a susceptible haplotype GC with a 7.7-fold higher risk of ALI. Reporter gene assays indicated that theses variants affected transcription rate. Our recent data suggest that PBEF affects pulmonary endothelial cell permeability in vitro and increased pulmonary edema in C57 BL/6 mice in vivo. In addition, stealth PBEF siRNA significantly inhibited TNF-alpha mediated increase of IL-8 secretion, a known edematogenic factor in ALI, in pulmonary cells. All these results support PBEF as a potential novel candidate gene and biomarker in ALI. To further detail underlying molecular mechanisms of PBEF in the pathogenesis of ALI, we hypothesize that patients with the genetically- determined susceptible haplotype in the PBEF promoter region are more susceptible to the upregulation by mechanical forces and inflammatory stimuli, two most inciting risk factors to ALI; increased PBEF expression contribute to the increased susceptibility to ALI by stimulating expression of other inflammatory cytokines such as IL-8, which leads to the pulmonary artery endothelial and epithelial cell barrier dysfunction and increased pulmonary permeability, resulting in the pathogenesis of ALI. The goal of this application is to define the physiologic and molecular significance of the identified PBEF polymorphisms. Furthermore, we will determine the pathophysiological role of PBEF in inflammatory lung injury with a specific focus on a role of PBEF in increased lung vascular leak/permeability, a pathophysiological feature of ALI, both in tissue culture system in vitro and in Pbef gene knock out C57 BL/6 mice in vivo. Specific Aim 1 will functionally characterize PBEF gene promoter SNPs utilizing the reporter gene and electrophoretic mobility shift assays. Specific Aim 2 will examine the role of PBEF gene expression in endothelial cell barrier regulation and contractility in vitro by evaluating endothelial cell transendothelial electric resistance, stress fiber formation, and phosphorylation of myosin light chains. Specific Aim 3 will define the signal transduction pathways of PBEF-mediated endothelial barrier regulation by examining PBEF- regulated genes and interacting partners. Specific Aim 4 will examine the in vivo role of PBEF in a murine model of ALI using Pbef knock out and over-expressing animals. Together, these novel and highly translational studies using the genomic and genetic approaches will provide new insight into molecular mechanisms of PBEF function in ALI, which may lead to the development of novel diagnostic modalities and therapeutic strategies in ALI.

描述（由申请方提供）：前B细胞集落增强因子（PBEF）是一种独特的、研究较少的细胞因子，其与肺病理生理学的相关性尚不清楚。PI最近的工作提供了肺组织中PBEF表达和急性肺损伤（ALI）过度表达以及PBEF遗传变异与ALI易感性相关的第一个证据。单倍型加权分析PBEF基因启动子区T-1001 G和C-1543 T单核苷酸多态性（SNP）显示GC单倍型易感，其ALI风险高7.7倍。报告基因分析表明，这些变异影响转录速率。我们最近的数据表明，PBEF在体外影响肺内皮细胞的通透性，并在体内增加C57 BL/6小鼠的肺水肿。此外，隐形PBEF siRNA显著抑制肺细胞中TNF-α介导的IL-8分泌增加，IL-8是ALI中已知的致水肿因子。这些结果支持PBEF作为一个潜在的新的候选基因和ALI的生物标志物。为了进一步阐明PBEF在ALI发病机制中的潜在分子机制，我们假设在PBEF启动子区域具有遗传决定的易感单倍型的患者更容易受到机械力和炎症刺激的上调，这两种是ALI的最激发危险因素;增加的PBEF表达通过刺激其它炎性细胞因子如IL-8的表达而有助于增加对ALI的易感性，导致肺动脉内皮和上皮细胞屏障功能障碍，肺通透性增加，从而导致ALI的发病。本申请的目的是确定所鉴定的PBEF多态性的生理和分子意义。此外，我们将确定PBEF在炎性肺损伤中的病理生理作用，特别关注PBEF在肺血管渗漏/通透性增加中的作用，这是ALI的病理生理特征，无论是在体外组织培养系统中还是在体内Pbef基因敲除C57 BL/6小鼠中。具体目标1将利用报告基因和电泳迁移率变动分析功能表征PBEF基因启动子SNP。具体目标2将通过评估内皮细胞跨内皮电阻、应力纤维形成和肌球蛋白轻链磷酸化来研究PBEF基因表达在体外内皮细胞屏障调节和收缩性中的作用。具体目标3将通过检查PBEF调节的基因和相互作用伴侣来定义PBEF介导的内皮屏障调节的信号转导途径。具体目标4将使用Pbef敲除和过表达动物来检查PBEF在ALI的鼠模型中的体内作用。总之，这些新的和高度翻译的研究，使用基因组和遗传学的方法将提供新的见解PBEF功能的分子机制在ALI，这可能会导致新的诊断模式和治疗策略在ALI的发展。