Effect of Mitochondrial Quality Control on Systemic Inflammation and Organ Dysfunction in Pediatric Sepsis

项目概要/摘要 脓毒症是一种危及生命的器官功能障碍的医疗紧急情况，由于宿主对 感染在美国，每年有超过一百万人因败血症或败血性休克住院， 其中包括75，000名儿童。多器官功能障碍综合征（MODS）是最常见的死亡原因 治疗儿童败血症虽然大多数患败血症的儿童在适当的常规治疗下可以康复， 护理，约20%发展为MODS，并且这些败血症诱导的MODS儿童中有五分之一仍然死亡。为此 高危亚群，我们无法治愈，护理主要是支持性的。大范围的改变， 线粒体为正常细胞活动提供足够的能量已经被认为是一种关键的病理学机制。 败血症中导致MODS的事件。使用优化的协议来测量线粒体呼吸和含量 在外周血单核细胞（PBMC）中，我们的研究小组发现， 诱导的MODS更可能具有持续的线粒体功能障碍。这项提议是建立在我们先前的 我们的工作是检验PBMCs中低线粒体呼吸是由异常的线粒体呼吸引起的这一假设。 质量控制过程（生物发生，裂变，融合和线粒体自噬），通常恢复和维持 线粒体健康通过释放线粒体DNA（mtDNA）导致线粒体功能障碍 进入胞质溶胶，然后可以激活维持炎症和器官功能障碍的炎症途径， 败血症的孩子在目标1中，我们将确定线粒体质量控制的哪些变化与 脓毒症患儿线粒体呼吸降低的恢复与持续。我们将招收106名 脓毒症诱导的MODS儿童，并测量线粒体呼吸，氧化还原状态，含量，生物发生， 在患病的第1、3、5和7天进行核分裂、核融合和线粒体自噬。在目标2中，我们将确定 PBMC线粒体功能障碍激活三种炎症途径（cGAS-STING、TLR-9和TLR-10）。 炎性小体），其感知线粒体DNA释放到胞质溶胶中。在目标3中，我们将测试线粒体是否 功能障碍、异常线粒体质量控制或炎症通路的激活与 脓毒症患儿MODS的严重程度或持续时间。作为目标1和2的一部分，我们还将进行一次 一组平行的体外实验，以测试脂多糖和脂磷壁酸刺激的效用， EBV转化的淋巴母细胞作为未来治疗研究的潜在转化平台， 确立胞浆mtDNA是线粒体在脓毒症中促进炎症的一种机制。 这些目标解决了NICHD的兴趣"研究集中在线粒体功能障碍， MODS的病理生理过程......具有推进转化和临床项目的潜力。通过 理解为什么线粒体功能障碍在一些儿童中持续存在，以及这如何导致持续的 炎症和MODS，我们将为长期MODS的儿童确定新的潜在治疗靶点， 败血症的主要死因，目前我们还无法治愈。

PROJECT SUMMARY / ABSTRACT Sepsis is a medical emergency of life-threatening organ dysfunction due to a dysregulated host response to infection. In the United States, over one million people are hospitalized with sepsis or septic shock every year, including >75,000 children. Multiple organ dysfunction syndrome (MODS) is the most common cause of death for children with sepsis. Although most children who develop sepsis recover with appropriate conventional care, ~20% develop MODS, and one in five of these children with sepsis-induced MODS still die. For this high-risk subset, we have no cure and care is largely supportive. Widespread alterations in the ability of mitochondria to supply sufficient energy for normal cellular activities has been implicated as a key pathologic event leading to MODS in sepsis. Using a protocol optimized to measure mitochondrial respiration and content in peripheral blood mononuclear cells (PBMC), our group discovered that children with prolonged sepsis- induced MODS are more likely to have persistent mitochondrial dysfunction. This proposal builds on our prior work to test the hypothesis that low mitochondrial respiration in PBMCs is caused by abnormal mitochondrial quality control processes (biogenesis, fission, fusion, and mitophagy) that normally restore and maintain mitochondrial health. The resulting mitochondrial dysfunction, through release of mitochondrial DNA (mtDNA) into the cytosol, can then activate inflammatory pathways that sustain inflammation and organ dysfunction in children with sepsis. In Aim 1, we will determine which changes in mitochondrial quality control are associated with recovery versus persistence of low mitochondrial respiration in children with sepsis. We will enroll 106 children with sepsis-induced MODS and measure mitochondrial respiration, redox state, content, biogenesis, fission, fusion, and mitophagy on days 1, 3, 5, and 7 of illness. In Aim 2, we will determine if the persistence of PBMC mitochondrial dysfunction activates three inflammatory pathway (cGAS-STING, TLR-9, and inflammasome) that sense release of mtDNA into the cytosol. In Aim 3, we will test whether mitochondrial dysfunction, abnormal mitochondrial quality control, or activation of inflammatory pathways are associated with the severity or duration of MODS in children with sepsis. As part of Aims 1 and 2, we will also conduct a parallel set of in vitro experiments to test the utility of lipopolysaccharide- and lipoteichoic acid-stimulation of EBV-transformed lymphoblasts as a potential translational platform for future therapeutic investigation and to establish cytosolic mtDNA as one mechanism through which mitochondria can promote inflammation in sepsis. These aims address the NICHD’s interest in “studies focused on mitochondrial dysfunction as a pathophysiologic process of MODS…with potential to advance to translational and clinical projects.” By understanding why mitochondrial dysfunction persists in some children and how this can lead to sustained inflammation and MODS, we will identify new potential therapeutic targets for children with prolonged MODS, the leading cause of death in sepsis for which we currently have no cure.