Raft/Caveolar Mechanisms in Pulmonary Hypertension

描述(由申请人提供)： 这项翻译研究计划试图将最近对质膜水平细胞信号机制的新见解(小窝/RAFT信号假说和IL-6-RAFT-STAT3信号模型)应用于理解PH的发病机制。含有Caveolin-1的抗洗涤剂质膜筏现在被认为是包括细胞因子信号在内的专门的信号细胞器。现在有越来越多的证据表明细胞因子在肺部疾病的发病机制中发挥了作用。例如，在原发性肺动脉高压(PH)以及与自身免疫性疾病和艾滋病相关的PH中，已观察到血清IL-6水平升高。在大鼠模型中，单次注射植物生物碱野百合碱(MCI)可在48h内引起内皮细胞损伤、膜渗漏、IL-6mRNA和生物活性上调，但显著下调肺内小窝蛋白-1的表达，并在10-14天后发展为PH。 建议的研究重点是双管齐下的：(A)评估MCT对肺内皮细胞RAFT/小窝破坏是PH发病机制中的始动事件的假说(特异性目标I)，以及(B)研究膜筏和新发现的含有胞浆小窝蛋白的Palade复合体在IL-6诱导的肺特异性细胞STAT3信号转导中的作用(特异性目标II和III)。目的研究MCT治疗大鼠肺血管和肺旁组织小窝蛋白和基因表达下调的时间进程、组织学定位、细胞和分子机制，以及小窝/RAFT功能的完整性。目的研究STAT3在肺内皮细胞、肺泡II型上皮细胞和肺成纤维细胞中与小窝蛋白-1结合的分子机制以及STAT3在质膜筏上的激活机制。目的研究含STAT3的胞质Palade复合体的蛋白质组分及其将信号分子从质膜筏输送到细胞内部的功能。 从这个项目中获得的机械学见解可能会为肺动脉高压的管理提供新的治疗方法。此外，所提出的研究具有特别广泛的意义，因为对RAFT-STAT信号涉及的分子机制的深入了解可能适用于细胞因子介导的STAT转录因子在所有细胞类型中的激活，以及定位于RAFT微域的其他信号通路(eNOS和血管紧张素II信号)。

DESCRIPTION (provided by applicant): This translational research proposal seeks to apply recent novel insights into the mechanisms of cell signaling at the level of the plasma membrane (the caveola/raft signaling hypothesis and the interleukin-6-raft-STAT3 signaling model) to an understanding of the pathogenesis of PH. Caveolin-1-containing detergent-resistant plasma membrane rafts are now recognized as specialized signaling organelles, including cytokine signaling. There is now growing evidence for a role of cytokines in the pathogenesis of lung diseases. As examples, elevated serum levels of IL-6 have been observed in primary pulmonary hypertension (PH) and in PH associated with autoimmune diseases and AIDS. In a rat model, a single injection of the plant alkaloid monocrotaline (MCI) results within 48 hrs in endothelial cell damage, membrane leakage, upregulation of IL-6 mRNA and bioactivity but a marked downregulation of caveolin-1 in the lung, followed by development of PH 10-14 days later. The focus of the proposed studies is two-pronged: (a) to evaluate the hypothesis that pulmonary endothelial-cell raft/caveolar disruption by MCT is an initiating event in the pathogenesis of PH (Specific Aim I), and (b) to investigate the function of membrane rafts and of the newly discovered cytosolic caveolin-containing Palade complexes in IL-6-induced STAT3 signaling in lung-specific cells (Specific Aims II and III). Aim I will include investigations of the time-course, histologic location, and cellular and molecular mechanisms for the downregulation of caveolin proteins and gene expression, and of the integrity of caveolar/raft function in pulmonary vascular and parenehymal tissues of MCT-treated rats. Aim II includes molecular studies of the mechanisms of association of STAT3 with caveolin-1 and of STAT3 activation in plasma membrane rafts in pulmonary endothelial cells, alveolar type II-like epithelial cells and lung fibroblasts. Aim Ill includes studies of the protein components of STAT3-containing cytosolic Palade complexes and their function in ferrying signaling molecules from the plasma membrane rafts to the cell interior. Mechanistic insights derived from this project are likely to suggest novel therapeutic approaches in the management of pulmonary hypertension. Moreover, the proposed studies are of particularly broad significance in that insights into the molecular mechanisms involved in raft-STAT signaling are likely to be applicable to cytokine-mediated activation of STAT transcription factors in perhaps all cell types, as well as to other signaling pathways localized in raft microdomains (eNOS and angiotensin II signaling).