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

项目摘要/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，其原因是宿主对 感染。在美国，每年有100多万人因败血症或感染性休克而住院， 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。 适用于患有败血症的儿童。尽管大多数患上脓毒症的儿童都能通过适当的常规方法康复 CARE，大约20%会发生MODS，这些患有败血症导致的MODS的儿童中有五分之一仍然死亡。为了这个 高危人群，我们无法治愈，护理在很大程度上是支持性的。能力的大范围变化 线粒体为正常的细胞活动提供足够的能量，已被认为是一种关键的病理机制 脓毒症中导致多器官功能障碍综合征的事件。使用一种优化的方案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，长期脓毒症的儿童- 诱导性MODS更容易出现持续性线粒体功能障碍。这项建议是在我们以前的基础上提出的 关于外周血单核细胞线粒体呼吸减少是由线粒体异常引起的假说的研究 正常恢复和维护的质量控制过程(生物发生、分裂、融合和有丝分裂) 线粒体健康。由此导致的线粒体功能障碍，通过释放线粒体DNA(MtDNA) 进入胞浆，然后可以激活炎症途径，维持炎症和器官功能障碍 患有败血症的儿童。在目标1中，我们将确定线粒体质量控制的哪些变化是相关的 脓毒症儿童线粒体低呼吸的恢复期与持续性。我们将招收106人 并测量线粒体呼吸，氧化还原状态，含量，生物发生， 发病第1、3、5、7天的分裂、融合和吞丝分裂。在目标2中，我们将确定 外周血单核细胞线粒体功能障碍激活三条炎症途径(cGAS-STING、TLR-9和 炎症体)，感觉线粒体DNA释放到细胞质中。在目标3中，我们将测试线粒体 功能障碍、线粒体质量控制异常或炎症通路激活与 脓毒症患儿多器官功能障碍的严重程度或持续时间。作为目标1和目标2的一部分，我们还将进行 一组平行的体外实验，以测试内毒素和脂磷壁酸刺激血管紧张素转换酶的作用 EBV转化的淋巴母细胞作为未来治疗研究和治疗的潜在翻译平台 建立胞浆线粒体DNA作为线粒体促进脓毒症炎症的一种机制。 这些目标满足了NICHD对“以线粒体功能障碍为重点的研究 多器官功能障碍综合征…的病理生理过程具有发展到翻译和临床项目的潜力。通过 了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性 炎症和MODS，我们将为长期MODS的儿童确定新的潜在治疗靶点， 败血症的主要死亡原因，目前我们还没有治愈的方法。

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.