Role of Resident Mesenchymal Stem Cells in Lung Allograft Rejection

驻留间充质干细胞在肺同种异体移植排斥反应中的作用

• 批准号: 8974429
• 负责人: Vibha N Lama
• 金额: $ 42.91万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2018-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8974429
• 关键词: Allografting; Animals; Binding; Biological Assay; Bronchiolitis Obliterans; Bronchoalveolar Lavage; Cells; Clinical Trials; Collagen; Complex; Data; Defect; Deposition; Development; Disease; Drug Targeting; Effectiveness; Embryo; Ethers; Exhibits; FOXF1 gene; FRAP1 gene; Fibrosis; Funding; Future; G-Protein-Coupled Receptors; Gene Expression; Gene Targeting; Grant; Health; Histology; Human; Investigation; Ions; Label; Laboratories; Lead; Ligation; Lung; Lung Transplantation; MEKs; Mediating; Mesenchymal; Mesenchymal Stem Cells; Mesenchyme; Modeling; Mus; ORC1L gene; Organ Transplantation; Outcome; Pathogenesis; Pathway interactions; Patients; Peptide Initiation Factors; Phenotype; Phosphorylation; Phosphotransferases; Play; Protein Kinase; Proteins; Proto-Oncogene Proteins c-akt; Role; Sampling; Scheme; Signal Transduction; Sirolimus; Small Interfering RNA; Solid; Sting Injury; Syndrome; Testing; Therapeutic; Therapeutic Intervention; Transgenic Organisms; Translational Activation; Translations; Transplant Recipients; Transplantation; Work; allograft rejection; autocrine; bindin; cell growth; extracellular; fibrogenesis; genome-wide; in vivo; inhibitor/antagonist; knock-down; lung allograft; lysophosphatidic acid; mouse model; pre-clinical; preclinical study; prevent; research study; ribosome profiling; transcription factor; vzg-1 Receptor

DESCRIPTION (provided by applicant): Long term survival post-lung transplantation is limited by development of progressive obstructive ventilatory defect termed bronchiolitis obliterans syndrome (BOS). BOS results from fibrotic obliteration of the small airways or bronchiolitis obliterans (BO). Studies from our laboratory have demonstrated that graft-resident mesenchymal cells (MCs) play an important role in fibroproliferation of BO. Exciting new preliminary data suggest that the evolutionary conserved protein kinase mTOR (mechanistic target of rapamycin) within both mTOR complex 1 (mTORC1) and mTORC2 contributes to the augmented collagen synthetic function of mesenchymal cell in BOS. MCs derived from patients with BOS were found to possess constitutive activation of both mTORC1 and mTORC2. siRNA-mediated knockdown of rictor, an mTORC2 binding partner, inhibited mTORC1 substrate phosphorylation and reversed the fibrotic phenotype of MCs, suggesting an important role for mTORC2 upstream of mTORC1 in activated fibrotic MCs. Selective ATP competitive mTOR inhibitors which target both mTORC1 and mTORC2 effectors were found to be potent inhibitors of collagen expression in BOS MCs. These data lead us to hypothesize that mTOR complex 1 (mTORC1) and mTORC2 play critical roles in BO pathogenesis by mediating translational activation of graft-resident MCs. In this application we propose experiments investigating in vivo role of mTOR pathway in pathogenesis of BO and aim to determine if ATP competitive mTOR inhibitors can be a potential therapeutic option in this disease. To achieve these aims we have established a whole lung mouse orthotopic lung transplant model where a moderate MHC mismatch leads to development of airway fibrosis by day 28. In Aim 1 we propose to investigate mTORC1/2 signaling in the resident MCs in vivo using this murine model. We will utilize mice with BAC florescent labeling of Foxf1, an embryonic lung mesenchyme-associated transcription factor which is highly expressed in lung mesenchymal cells, to specifically identify and study graft-resident MCs. Upstream signaling mechanism(s) that contribute to constitutive mTOR1/2 activation of MCs in BOS will be studied. We will focus on the role of G-protein coupled receptor, lysophosphatidic acid (LPA)1, signaling in a proposed autocrine mechanism of activation. Aim 2 will dissect the role of downstream mTORC1 effectors and dysregulated translation in sustaining fibrotic functions of MCs in BOS. Furthermore, we will utilize state-of-the-art ribosomal profiling (Ribo-seq) to identify genome wide differences in translation between BOS and non-BOS MCs and to delineate the mTORC1 translation controlled gene expression network by studying BOS cells upon complete or partial mTOR inhibition. In Aim 3 we will test if ATP competitive mTOR inhibitors can prevent onset or progression of allograft fibrosis in the murine orthotopic single lung transplant model. The proposed work will be the first investigation of role of mTORC1/2 signaling in lung allograft fibrogenesis and will provide important rationale for future clinical trials targeting this pathway in lung transplantation.

描述（由申请人提供）：肺移植后的长期生存受到称为闭塞性细支气管炎综合征（BOS）的进行性阻塞性肺功能缺损的限制。BOS由小气道的纤维化闭塞或闭塞性细支气管炎（BO）引起。我们实验室的研究表明，移植物驻留间充质细胞（MCs）在BO的纤维增殖中起重要作用。令人兴奋的新的初步数据表明，在mTOR复合物1（mTORC 1）和mTORC 2中的进化保守的蛋白激酶mTOR（雷帕霉素的机制靶标）有助于增强BOS中间充质细胞的胶原合成功能。来自BOS患者的MCs被发现具有mTORC 1和mTORC 2的组成性激活。siRNA介导的mTORC 2结合伴侣rictor的敲除抑制mTORC 1底物磷酸化并逆转MC的纤维化表型，表明mTORC 1上游的mTORC 2在活化的纤维化MC中起重要作用。发现靶向mTORC 1和mTORC 2效应物的选择性ATP竞争性mTOR抑制剂是BOS MC中胶原蛋白表达的有效抑制剂。这些数据使我们假设mTOR复合物1（mTORC 1）和mTORC 2通过介导移植物驻留MC的翻译激活在BO发病机制中起关键作用。在本申请中，我们提出了研究mTOR通路在BO发病机制中的体内作用的实验，旨在确定ATP竞争性mTOR抑制剂是否可以成为这种疾病的潜在治疗选择。为了实现这些目标，我们建立了全肺小鼠原位肺移植模型，其中中度MHC错配导致第28天发生气道纤维化。在目的1中，我们建议使用这种小鼠模型在体内研究驻留MC中的mTORC 1/2信号传导。我们将利用小鼠BAC荧光标记Foxf 1，胚胎肺间充质相关的转录因子，这是在肺间充质细胞中高度表达，专门确定和研究移植驻留MC。将研究有助于BOS中MC的组成性mTOR 1/2激活的上游信号传导机制。我们将集中在G蛋白偶联受体，溶血磷脂酸（LPA）1，信号在一个拟议的自分泌机制的激活作用。目的2将剖析下游mTORC 1效应子和失调的翻译在维持BOS中MC纤维化功能中的作用。此外，我们将利用最先进的核糖体分析（Ribo-seq）来鉴定BOS和非BOS MC之间的全基因组翻译差异，并通过研究完全或部分mTOR抑制后的BOS细胞来描绘mTORC 1翻译控制的基因表达网络。在目的3中，我们将测试ATP竞争性mTOR抑制剂是否可以在鼠原位单肺移植模型中预防同种异体移植物纤维化的发作或进展。这项工作将是首次研究mTORC 1/2信号在肺移植纤维化中的作用，并将为未来针对肺移植中这一途径的临床试验提供重要的理论基础。