Cytokine signaling in activation of the endothelium

内皮激活中的细胞因子信号传导

• 批准号: 7659839
• 负责人: PAUL E DICORLETO
• 金额: $ 43.29万
• 依托单位: CLEVELAND CLINIC LERNER COM-CWRU
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-15 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7659839
• 关键词: Adhesions; Adult; Affect; Anti-Inflammatory Agents; Anti-inflammatory; Apolipoprotein E; Arginine; Arterial Fatty Streak; Atherosclerosis; Binding; Biochemical; Blood Vessels; Cell Nucleus; Cell Surface Receptors; Cell surface; Cells; Coronary Arteriosclerosis; Cytokine Activation; Cytokine Signaling; Data; Development; Disease; Dissociation; E-Selectin; Endothelial Cells; Endothelium; Event; Gene Activation; Gene Expression; Gene Targeting; Genes; Genetically Engineered Mouse; HOXA9 gene; HOXA9 protein; Human; In Vitro; Inflammation; Inflammatory; Inflammatory Response; Instruction; Intercellular adhesion molecule 1; Knockout Mice; Leukocyte Adhesion Molecules; Leukocyte Rolling; Leukocyte-Adhesion Receptors; Leukocytes; Mediating; Mediator of activation protein; Modeling; Molecular; Molecular Biology; Mus; Play; Post-Translational Protein Processing; Process; Proteins; Proteomics; Qualifying; Regulation; Rheumatoid Arthritis; Role; Signal Pathway; Signal Transduction; Site; Site-Directed Mutagenesis; Small Interfering RNA; Stem cells; Stimulus; System; TNF gene; TNFRSF1B gene; Technology; Testing; Therapeutic; Tibial Arteries; Tissues; Transferase; Tumor Necrosis Factor Receptor; Tumor Necrosis Factor-alpha; Tumor Necrosis Factors; Vascular Cell Adhesion Molecule-1; Vascular Endothelial Cell; angiogenesis; base; chromatin immunoprecipitation; chromatin remodeling; cremaster muscle; cytokine; deletion analysis; experience; human MPP1 protein; human tissue; in vivo; intravital microscopy; mouse model; novel; promoter; receptor; response; transcription factor; transcriptional coactivator p75; tumor necrosis factor receptor 1A

The central objective of this Project is to define signaling mediators and transcription factors of critical importance in cytokine activation of vascular endothelial cells (EC). Our specific focus is on TNF-a receptor-ll (p75) and the homeobox protein HOXA9. We have recently shown that both molecules are required for cytokine induction of the EC- leukocyte adhesion molecule E-selectin, as well as other activation genes. We showed that TNF-stimulated leukocyte firm adhesion to EC was dramatically reduced in p75-null mice, but not affected in p55 (the other TNF receptor)-null mice. Both receptors participated in TNF-induced leukocyte rolling and transmigration. Furthermore, both p75 and p55 were required for TNF induction of E-selectin and VCAM-1; whereas, p75 was sufficient for ICAM-1 induction. We also demonstrated that p75-deficiency in apoE-null mice reduced atherosclerotic lesion development. Based on these findings, we hypothesize that p75 activation in EC triggers intra-cellular signaling pathways, not induced by p55, that are essential for TNF-induced pro-inflammatory gene expression in EC. We have also recently shown that the transcription factor HOXA9 is required for TNF induction of E-selectin in EC due to its transient binding to a newly identified Abd-B-like site in the promoter. The TNF signal that leads to association of HOXA9 with the E-selectin pro- moter is triggered by p75 and not p55. We hypothesize that HOXA9 plays a novel role in the activation of EC in res- ponse to inflammatory stimuli. We propose to test our hypotheses by pursuing three aims. The first is to identify TNF- induced, p75-specific signaling pathways and novel p75-induced genes in EC using TNF-treated mouse EC isolated from p55-/-, p75-/- and p55-p75-double null mice. We will also identify TNF-induced binding partners of p75 and determine structural domains in p75 responsible for induced gene expression in EC. The second aim is to elucidate the molecular mechanism of HOXA9-mediated E-selectin expression and to identify novel target genes for HOXA9 in EC. We will pursue post-translational modifications of HOXA9 and its possible binding to other obligate transcription factors for E-selectin induction. Gene and promoter array studies will be used to identify novel target genes of HOXA9. In the third aim we will characterize the role of the HOXA9-binding partner protein arginine methyl transferase 5 (PRMT5) in activated EC using biochemical, siRNA-based and pharmacological approaches. We also plan to identify proteins methylated by PRMT5 that may qualify for novel mediators of EC activation. RELEVANCE (See instructions): EC, which line all blood vessels in the body, play a key role in inflammation - a process that underlies many diseases, including atherosclerosis. Our studies will reveal new molecular players in the regulation of inflammation. These players will represent novel targets for the development of anti-inflammatory therapeutics that may be used to treat such diseases as coronary artery disease and rheumatoid arthritis.

该项目的中心目标是确定在细胞内具有关键重要性的信号传导介质和转录因子。 血管内皮细胞（EC）的细胞因子活化。我们的具体重点是TNF-α受体-II（p75）和TNF-α受体-II（p75）。 同源框蛋白HOXA 9。我们最近已经表明，这两种分子都是细胞因子诱导EC所必需的。 白细胞粘附分子E-选择素，以及其它激活基因。我们发现TNF刺激的白细胞 在p75缺失的小鼠中，与EC的牢固粘附显著降低，但在p55（另一种TNF受体）缺失的小鼠中不受影响。 小鼠这两种受体都参与了TNF诱导的白细胞滚动和迁移。p75和p55 E-selectin和VCAM-1的TNF诱导需要p75，而ICAM-1的诱导需要p75。我们也 表明apoE基因敲除小鼠中p75缺陷减少动脉粥样硬化病变的发展。基于这些 研究结果，我们假设EC中p75激活触发细胞内信号通路，而不是由p55诱导， 是EC中TNF诱导的促炎基因表达所必需的。我们最近还表明， 转录因子HOXA 9是TNF诱导EC中E-选择素所必需的，这是由于其瞬时结合新的 在启动子中鉴定了Abd-B样位点。导致HOXA 9与E-选择素原结合的TNF信号 运动是由p75而不是p55触发的。我们假设HOXA 9在视网膜病变中EC的激活中起着新的作用。 对炎症刺激的反应。我们建议通过追求三个目标来测试我们的假设。首先是识别TNF- 诱导的，p75特异性信号通路和新的p75诱导的基因在EC中使用TNF处理的小鼠EC分离 来自p55-/-、p75-/-和p55-p75-双缺失小鼠。我们还将鉴定TNF诱导的p75结合伴侣， 确定p75中负责EC中诱导基因表达的结构域。第二个目的是阐明 HOXA 9介导的E-选择素表达的分子机制，并确定HOXA 9在EC中的新靶基因。 我们将继续研究HOXA 9的翻译后修饰及其与其他专性转录因子的可能结合 用于E-选择素诱导。基因和启动子阵列研究将用于鉴定HOXA 9的新靶基因。在 第三个目标，我们将描述HOXA 9结合伴侣蛋白精氨酸甲基转移酶5（PRMT 5）的作用。 使用生物化学、基于siRNA的和药理学方法在活化EC中进行。我们还计划识别蛋白质 甲基化的PRMT 5，可能有资格为新的调解人的EC激活。 相关性（参见说明）： EC，排列在身体的所有血管中，在炎症中起着关键作用--这是许多疾病的基础， 包括动脉粥样硬化。我们的研究将揭示炎症调节中的新分子参与者。这些球员 将代表用于开发抗炎治疗剂的新靶点， 冠心病和类风湿性关节炎。