Translational Control of Megakaryocyte and Platelet Function in Sepsis

脓毒症中巨核细胞和血小板功能的转化控制

• 批准号: 10210293
• 负责人: Matthew Thomas Rondina
• 金额: $ 38.13万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10210293
• 关键词: Abnormal Platelet; Acute; Affect; Agonist; Applications Grants; Attention; Biological Models; Blood Platelets; Cell physiology; Cells; Data; Disease; Eukaryotic Initiation Factor-4E; Event; FRAP1 gene; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genetic Translation; Goals; Health; Human; Impairment; In Vitro; Inflammation; Inflammatory; Interferons; MAP Kinase Gene; Megakaryocytes; Messenger RNA; Mitogen-Activated Protein Kinases; Modeling; Mus; Outcome; Pathogenesis; Pathway interactions; Phenotype; Phosphotransferases; Platelet Count measurement; Production; Protein Biosynthesis; Protein-Serine-Threonine Kinases; Proteins; Regulation; Ribosomes; Sepsis; Signal Pathway; Signal Transduction; Techniques; Testing; Thrombopoiesis; Translating; Translation Initiation; Translational Activation; Translational Regulation; Translations; adverse outcome; clinical effect; cytokine; human model; in vivo; in vivo Model; inflammatory milieu; inhibitor/antagonist; kinase inhibitor; mouse model; novel; platelet function; protein expression; response; ribosome profiling; septic; septic patients; side effect; systemic inflammatory response; transcriptome; virtual

Summary/Abstract Regulation of gene expression at the translational level (i.e., mRNA-protein) is poorly understood in normal megakaryocytes, and it is virtually unknown if inflammatory diseases alter translational responses in megakaryocytes and platelets. With that said, inflammation contributes to the pathogenesis of numerous diseases such as sepsis, and inflammatory agonists are considered potent triggers of mRNA translation. This grant proposal will utilize human and mouse model systems (in vitro and in vivo) and state-of-the-art sequencing (ribosomal footprinting and eIF4E-enrichment) to determine how translational control pathways regulate megakaryocyte and platelet gene expression – in the setting of health and septic situations. Preliminary data supporting this project demonstrates that the mammalian target of rapamycin (mTOR) and the MAP kinase-interacting serine/threonine-protein kinase 1 (MNK1), two translational control pathways that converge on the initiation step of translation, regulate protein synthetic events and functions in megakaryocytes and platelets. These preliminary results have led to the hypothesis that the septic milieu will significantly modify mTOR and/or MNK1-dependent translation in megakaryocytes and platelets, resulting in inappropriate formation and function of platelets during the course of sepsis. Specific Aim1 will test the hypothesis that inflammatory agonists generated during sepsis will alter protein synthetic events in megakaryocytes and the formation of platelets in an mTOR and MNK1-dependent manner. Specific Aim 2 will determine how genetic deletion of mTOR or MNK1 regulates the ex vivo and in vivo function of platelets in the presence or absence of inflammation. Specific Aim 3 will test the hypothesis that the septic milieu activates the mTOR and/or MNK1 signaling pathways and thereby increases protein synthesis by platelets. It will also determine if deletion of mTor and/or Mnk1 in mouse platelets leads to adverse outcomes in murine models of sepsis. Successful completion of these aims will 1) identify translated mRNAs in megakaryocytes that are under mTOR and/or MNK1-dependent control and whether these translational signaling pathways regulate the final steps of platelet production, 2) delineate the effects of mTOR and MNK1 on functions of megakaryocytes and platelets in health and inflammation, 3) characterize inadvertent side- effects of clinically-used mTOR and MNK1 inhibitors on megakaryocytes and platelets, and 4) determine if mTOR or MNK1 in megakaryocytes and platelets regulate outcomes in models of sepsis. Data generated will immediately increase our understanding of how translational control pathways alter megakaryocyte and platelet functions in the setting of sepsis.

摘要/摘要 正常人对翻译水平（即 mRNA-蛋白质）基因表达的调控知之甚少。 巨核细胞，并且几乎不知道炎症性疾病是否会改变巨核细胞的翻译反应 巨核细胞和血小板。话虽如此，炎症有助于许多疾病的发病机制 败血症等疾病和炎症激动剂被认为是 mRNA 翻译的有效触发因素。这 拨款提案将利用人类和小鼠模型系统（体外和体内）和最先进的 测序（核糖体足迹和 eIF4E 富集）以确定翻译控制途径 在健康和脓毒症情况下调节巨核细胞和血小板基因表达。 支持该项目的初步数据表明，雷帕霉素的哺乳动物靶点（mTOR）和 MAP 激酶相互作用丝氨酸/苏氨酸蛋白激酶 1 (MNK1)，两条翻译控制途径 集中于翻译起始步骤，调节巨核细胞中的蛋白质合成事件和功能 和血小板。这些初步结果得出这样的假设：化粪池环境将 显着改变巨核细胞和血小板中 mTOR 和/或 MNK1 依赖性翻译， 导致脓毒症过程中血小板形成和功能异常。具体的 Aim1 将检验脓毒症期间产生的炎症激动剂会改变蛋白质合成的假设 巨核细胞中的事件和血小板以 mTOR 和 MNK1 依赖性方式形成。具体的 目标 2 将确定 mTOR 或 MNK1 的基因删除如何调节离体和体内功能 存在或不存在炎症时的血小板。具体目标 3 将检验以下假设：化粪池 环境激活 mTOR 和/或 MNK1 信号通路，从而增加蛋白质合成 血小板。它还将确定小鼠血小板中 mTor 和/或 Mnk1 的缺失是否会导致不良后果 在脓毒症小鼠模型中。成功完成这些目标将 1) 识别翻译的 mRNA 巨核细胞是否处于 mTOR 和/或 MNK1 依赖性控制下，以及这些翻译是否 信号通路调节血小板生成的最后步骤，2) 描述 mTOR 和 MNK1 的作用 关于巨核细胞和血小板在健康和炎症中的功能，3）表征无意的副作用 临床使用的 mTOR 和 MNK1 抑制剂对巨核细胞和血小板的影响，以及 4) 确定是否 巨核细胞和血小板中的 mTOR 或 MNK1 调节脓毒症模型的结果。生成的数据将 立即增加我们对翻译控制途径如何改变巨核细胞和 脓毒症中的血小板功能。