Non-platelet P2Y Receptor in Vascular Inflammation and Thrombogenesis

非血小板 P2Y 受体在血管炎症和血栓形成中的作用

基本信息

批准号：
9237082
负责人：
Jianzhong Shen
金额：
$ 37万
依托单位：
AUBURN UNIVERSITY AT AUBURN
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-01-01 至 2021-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9237082
关键词：
3&apos Untranslated Regions ATF2 gene Acute Address Affect Apyrase Arterial Fatty Streak Aspirin Binding Binding Proteins Binding Sites Biological Assay Blood Platelets Blood Vessels Blood coagulation Blood flow CYP2C19 gene Cardiovascular Diseases Cause of Death Cell Line Cells Cessation of life Chronic Clinical Complication Coronary Coronary artery Disease Disseminated Intravascular Coagulation Drug Targeting EMSA Elements Endothelial Cells Endothelium Expenditure Family Fos-Related Antigens Gene Expression Genes Genetic Polymorphism Genetic Transcription Goals Growth Factor Healthcare Hemorrhage Hormones Human Infiltration Inflammation Inflammatory Knockout Mice Knowledge LDL-Receptor Related Protein 1 Leukocytes Link Luciferases MAPK14 gene MAPK8 gene MK-2 Mediating Mediator of activation protein Messenger RNA MicroRNAs Modeling Molecular Target Monitor Mus Myocardial Infarction Necrosis Northern Blotting Nucleotidases Nucleotides P2Y2 receptor PF4 Gene Pathologic Pathway interactions Performance Pharmaceutical Preparations Phosphotransferases Plavix Population Post-Transcriptional Regulation Prevention Process Prophylactic treatment Receptor Activation Receptor Signaling Regulation Reporting Research Resistance Role Sepsis Series Site Solid Source Stents Stimulus Stroke Testing Therapeutic Agents Thromboplastin Thrombosis Time Time Factors Transcription Factor AP-1 United States Up-Regulation Vascular Endothelial Cell Western Blotting Wild Type Mouse atherothrombosis base cell type clopidogrel combat cytokine extracellular ferric chloride in vivo liver injury mRNA Stability mRNA Transcript Degradation member monocyte mouse model new therapeutic target novel nucleotide receptor prevent receptor response rho transcription factor vascular inflammation

3&apos Untranslated Regions ATF2 gene Acute Address Affect Apyrase Arterial Fatty Streak Aspirin Binding Binding Proteins Binding Sites Biological Assay Blood Platelets Blood Vessels Blood coagulation Blood flow CYP2C19 gene Cardiovascular Diseases Cause of Death Cell Line Cells Cessation of life Chronic Clinical Complication Coronary Coronary artery Disease Disseminated Intravascular Coagulation Drug Targeting EMSA Elements Endothelial Cells Endothelium Expenditure Family Fos-Related Antigens Gene Expression Genes Genetic Polymorphism Genetic Transcription Goals Growth Factor Healthcare Hemorrhage Hormones Human Infiltration Inflammation Inflammatory Knockout Mice Knowledge LDL-Receptor Related Protein 1 Leukocytes Link Luciferases MAPK14 gene MAPK8 gene MK-2 Mediating Mediator of activation protein Messenger RNA MicroRNAs Modeling Molecular Target Monitor Mus Myocardial Infarction Necrosis Northern Blotting Nucleotidases Nucleotides P2Y2 receptor PF4 Gene Pathologic Pathway interactions Performance Pharmaceutical Preparations Phosphotransferases Plavix Population Post-Transcriptional Regulation Prevention Process Prophylactic treatment Receptor Activation Receptor Signaling Regulation Reporting Research Resistance Role Sepsis Series Site Solid Source Stents Stimulus Stroke Testing Therapeutic Agents Thromboplastin Thrombosis Time Time Factors Transcription Factor AP-1 United States Up-Regulation Vascular Endothelial Cell Western Blotting Wild Type Mouse atherothrombosis base cell type clopidogrel combat cytokine extracellular ferric chloride in vivo liver injury mRNA Stability mRNA Transcript Degradation member monocyte mouse model new therapeutic target novel nucleotide receptor prevent receptor response rho transcription factor vascular inflammation

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项目摘要

Thrombosis is a fatal complication of major diseases, such as stroke, sepsis, and heart attack. In addition to aspirin, current medication for thrombosis prevention/treatment mainly focus on platelet P2Y12 receptor with plavix as an antagonist in routine clinical use. However, plavix has showed unstable performance, either failing to avoid thrombosis or causing bleeding. The long-term goal of this project is to establish non-platelet P2Y receptors, e.g. P2Y2 receptor (P2Y2R), as a new molecular target that controls inflammation-induced vascular thrombogenesis. Although it is well established that tissue factor (TF) is the initiator of thrombosis, very little is known on the contribution of P2Y receptors in regulation of various inflammatory stimuli-induced TF expressions. This is an important question because many pro-inflammatory stimuli promote cellular nucleotide release and inactivate ectonucleotidases, leading to accumulation of extracellular nucleotides, which in turn amplify original receptors’ signaling via P2Y receptor co-activation. The PI reported previously that P2Y2R is up-regulated in stented coronary arteries and it is the predominant subtype of all P2Y receptors in human coronary artery endothelial cells (HCAEC). They also reported that activation of P2Y2R dramatically induces TF expression and activity in HCAEC. The PIs’ preliminary studies showed that TF induction by P2Y2R is also applicable to human primary blood monocytes and mouse coronary endothelium in vivo, and it involves both transcriptional and post-transcriptional mechanisms. Further, LPS-induced TF induction is significantly decreased when extracellular nucleotides are removed by apyrase, and importantly, P2Y2R-null mice are protected from endotoxic death. Based on these original findings, the PI proposes the hypothesis: The non-platelet P2Y2R is a previously unrecognized key mediator in inflammation-induced thrombosis via TF induction. This hypothesis will be tested by the pursuit of three specific aims: 1) Determine the transcriptional mechanism underlying P2Y2R activation of the TF gene in vascular endothelial cells and blood monocytes; 2) Define the post-transcriptional mechanism by which P2Y2R activation leads to increased TF mRNA stability in vascular endothelial cells and blood monocytes; 3) Assess the role of P2Y2R/TF axis in mouse models of inflammation-induced thrombosis. Aim 1 will determine the role of a new AP-1 binding site with new AP-1 components Fra-1/ATF2 in TF mRNA induction by P2Y2R. Aim 2 will determine the roles of the AU-rich elements in TF 3’UTR along with their binding proteins and the miRNA mechanisms contribute to P2Y2R-mediated TF mRNA stabilization. Aim 3 will verify if deletion of P2Y2R prevents LPS-induced disseminated intravascular coagulation and reduces atherothrombosis. The proposed study will be performed in primary cultured human cells and cells isolated from P2Y2R-null mice, followed by a series of in vivo studies. The proposed research is significant, because it is expected to advance and expand understanding of how inflammation leads to increased thrombogenicity of blood vessels via the new P2Y2R-TF axis. Ultimately, such knowledge has the potential to produce new strategies in combating thrombogenic diseases.

血栓形成是主要疾病的致命并发症，例如中风，败血症和心脏病发作。此外阿司匹林，目前预防血栓形成/治疗的药物主要集中在血小板P2Y12受体上 Plavix是常规临床用途的对手。但是，Plavix表现出不稳定的性能，要么无法避免血栓形成或引起出血。该项目的长期目标是建立非平台P2Y受体，例如P2Y2受体（P2Y2R），作为控制注射诱导的血管的新分子靶标血栓形成。尽管已经确定组织因子（TF）是血栓形成的发起者，但很少已知P2Y受体在调节各种炎症刺激诱导的TF表达方面的贡献。这是一个重要的问题，因为许多促炎性刺激会促进细胞核释放和灭活的经济核苷酸酶，导致细胞外核苷酸酶的积累，进而扩大原始受体通过P2Y受体共激活的信号传导。 PI先前报告说P2Y2R已上调支架冠状动脉，是人类冠状动脉中所有P2Y受体的主要亚型内皮细胞（HCAEC）。他们还报告说，P2Y2R的激活显着诱导TF表达和 HCAEC的活动。 PIS的初步研究表明，P2Y2R的TF诱导也适用于人类原发性血单核细胞和小鼠冠状动脉内皮体体内，涉及转录和转录后机制。此外，LPS诱导的TF诱导在细胞外显着降低核苷酸通过apyrase去除，重要的是，P2Y2R-NULL小鼠免受内毒性死亡的保护。基于这些原始发现，PI提案假设：非静脉P2Y2R是先前的通过TF诱导，无法识别的炎症引起的血栓形成的关键介体。该假设将进行检验通过追求三个特定目的：1）确定P2Y2R激活的转录机制血管内皮细胞和血单核细胞中的TF基因； 2）通过 P2Y2R激活导致血管内皮细胞和血液单核细胞中TF mRNA稳定性增加。 3）评估P2Y2R/TF轴在炎症引起的血栓形成的小鼠模型中的作用。 AIM 1将确定新的AP-1结合位点具有新的AP-1成分FRA-1/ATF2在P2Y2R诱导TF mRNA诱导中的作用。目的 2将确定富含Au元素在TF 3'UTR中的作用以及它们的结合蛋白和miRNA 机制有助于P2Y2R介导的TF mRNA稳定。 AIM 3将验证是否删除P2Y2R的删除 LPS诱导的散布血管内凝结并减少了动脉粥样硬化。拟议的研究将是在原代培养的人类细胞和从P2Y2R-NULL小鼠中分离出来的细胞，然后进行一系列体内研究。拟议的研究很重要，因为它有望提高和扩大对炎症如何通过新的P2Y2R-TF轴导致血管的血管血管增加。最终，这种知识有可能制定对抗血栓形成疾病的新策略。