Smooth Muscle Cell Protein Serotonylation and Pulmonary Hypertension

平滑肌细胞蛋白血清素化与肺动脉高压

• 批准号: 8534244
• 负责人: Barry Fanburg
• 金额: $ 45.47万
• 依托单位: TUFTS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8534244
• 关键词: Active Biological Transport; Amines; Animal Model; Animals; BMP2 gene; BMPR2 gene; Biochemical Process; Cell Culture Techniques; Cell Proliferation; Cell Surface Receptors; Cell physiology; Clinical; Data; Development; Disease; Enzymes; Exposure to; Extracellular Matrix Proteins; Fibronectins; Human; Hypoxia; Immunoblotting; Immunoprecipitation; Integrins; Intervention; Knock-in Mouse; Knowledge; Lead; Lung; MAP Kinase Gene; Michigan; Mitogen-Activated Protein Kinase Inhibitor; Modification; Molecular; Monocrotaline; Mus; PDGFRB gene; Participant; Pathogenesis; Pathway interactions; Patients; Platelet-Derived Growth Factor; Platelet-Derived Growth Factor Receptor; Post-Translational Protein Processing; Process; Proteins; Pulmonary Hypertension; Pulmonary artery structure; Rattus; Regulation; Rodent; Rodent Model; Role; Secondary to; Serotonin; Serum; Signal Pathway; Signal Transduction; Small Interfering RNA; Smooth Muscle Myocytes; Structure of parenchyma of lung; System; Techniques; Testing; Tissues; Transfection; Transgenic Mice; Transgenic Organisms; Transglutaminases; Universities; Vascular remodeling; cell growth; cell motility; hemodynamics; in vivo; inhibitor/antagonist; migration; mutant; novel; novel therapeutic intervention; public health relevance; receptor; research study; response; rho; serotonin transporter; transglutaminase 2; uptake

DESCRIPTION (provided by applicant): We believe we have discovered a novel explanation for the mechanism by which serotonin (5- HT) produces pulmonary artery smooth muscle cell (SMC) proliferation and migration. This is important since these SMC functions participate in pulmonary hypertension (PH) and the 5- HT/5-HT transporter (SERT) system has been associated with clinical and experimental PH. From preliminary data the biochemical process of posttranslational modification of SMC protein through transamidation (serotonylation) leads to SMC proliferation and migration. Fibronectin (FN), an important extracellular matrix protein that has been associated with PH, is a major protein undergoing serotonylation. This enhanced serotonylation of SMC protein, occurring through the tissue enzyme transglutaminase (TGase), is influenced by the PDGF receptor and possibly other cell surface receptors that have been associated with PH. Furthermore, as an in vivo corollary enhanced serotonylation of FN occurs in lung tissue of mice and rats developing PH secondary to exposure to hypoxia and in rats given monocrotaline, and we have identified marked elevation of serotonylated FN in lungs and sera of some patients with PAH. We propose in this application to explore further the roles that lung tissue SERT, TGase and protein serotonylation may have in stimulation of SMC proliferation and migration and development of PH with a variety of cell culture, transfection, siRNA, immunoprecipitation and immunoblotting techniques and with the use of normal and transgenic rodent model experiments. Specifically, we will 1) better define the roles of SERT and TGase in SMC proliferation, migration and cell signaling produced by 5-HT; 2) determine the roles of serotonylation of FN and related integrins in 5-HT induced SMC proliferation and migration; 3) assess modification of SMC protein/FN serotonylation by hypoxia, PDGF/PDGFR and BMP/BMPR, known participants in PH; and 4) examine the association of lung and pulmonary artery protein/FN serotonylation in rodents developing PH secondary to hypoxia or following administration of monocrotaline, in SERT KO and "knock in" transgenic mice, in SERT KO rats and in mice deficient in TGase. The SERT "knock-in" animals that show overactivity of SERT are from Dr. Randy Blakely, an internationally recognized expert in SERT, at Vanderbilt University. Dr. Stephanie Watts from Michigan State University, another expert in SERT, will collaborate in studies related to SERT KO rats and their isolated pulmonary artery SMC's. We hope to better define any role of SERT, TGase activity and protein serotonylation in the development of PH and to consider possible new interventional strategies through these pathways that might be used for treating this disease.

描述（由申请人提供）：我们相信我们已经发现了一种新颖的解释，该机制是5-羟色胺（5-HT）产生肺动脉平滑肌细胞（SMC）增殖和迁移。这很重要，因为这些SMC功能参与肺动脉高压（pH），而5-HT/5-HT转运蛋白（SERT）系统与临床和实验性pH相关。从初步数据，通过跨增化剂（详细化）对SMC蛋白进行翻译后修饰的生化过程导致SMC增殖和迁移。纤连蛋白（FN）是一种与pH相关的重要细胞外基质蛋白，是一种经历血清素化的主要蛋白。通过组织酶转谷氨酰胺酶（TGase）发生的SMC蛋白的这种增强的血清素化受到PDGF受体以及可能与pH相关的其他细胞表面受体的影响。此外，随着体内推论增强了FN的血清素化，发生在小鼠的肺组织中，而大鼠发生了继发于暴露于缺氧的pH值以及给予单蛋白的大鼠，我们已经确定了一些PAH患者的肺和血清中血清素化的FN的明显升高。我们在此应用中提出，进一步探讨了肺组织SERT，TGASE和蛋白血清素化可能在刺激SMC增殖和迁移和pH的刺激以及通过多种细胞培养，转染，siRNA，免疫沉淀和免疫印迹技术以及使用正常和转元素的pH的技术以及使用正常和转元素模型模型实验中具有的作用。具体而言，我们将更好地定义SERT和TGase在5-HT产生的SMC增殖，迁移和细胞信号传导中的作用； 2）确定FN和相关整合蛋白在5-HT诱导的SMC增殖和迁移中的详细素化的作用； 3）评估缺氧，PDGF/PDGFR和BMP/BMPR的SMC蛋白/FN血清素化的修饰，pH中已知参与者；和4）检查在发育于缺氧继发于低氧的pH值或在单次升高后，SERT KO中，Sert KO和转基因小鼠的“敲击”，SERT KO大鼠和TGASE小鼠中的转基因小鼠中，肺和肺动脉蛋白/FN的血清素化的关联。 Sert“敲击”动物表现出Sert的过度活动，来自范德比尔特大学的国际认可的塞尔特（Sert）的国际认可的专家兰迪·布拉克利（Randy Blakely）博士。 SERT的另一位专家密歇根州立大学的Stephanie Watts博士将在与Sert KO大鼠及其孤立的肺动脉SMC有关的研究中进行合作。我们希望更好地定义SERT，TGase活性和蛋白质血清素化在pH发展中的任何作用，并通过这些可能用于治疗该疾病的途径来考虑可能的新介入策略。