Age-dependent mechanisms of metabolic recovery in hemorrhagic shock

 描述(申请人提供)：多器官衰竭(MOF)是创伤患者死亡的主要原因。即使通过液体复苏和器官支持方案成功地处理了急性创伤，许多在最初创伤中幸存下来的患者仍面临着发展为持续性器官损伤的高风险。临床证据表明，高龄是MOF持续性和死亡率的危险因素。多器官功能不全持续的分子机制和年龄的影响尚不明确。此外，促进器官功能恢复的治疗策略仍然不可用。科学证据表明，线粒体功能障碍是多器官功能衰竭的重要致病事件。在初步研究中，我们观察到自噬是使细胞能够处理有缺陷的线粒体的一个重要过程，与年轻动物(2-3个月)相比，老年大鼠(18-24个月)的肾脏失血性休克时自噬受到了损害。这种与年龄相关的自噬损害与肾脏损伤和炎症反应的严重程度相关。在分子分析方面，我们发现AMP激活的蛋白激酶(AMPK)是调节失血性休克后器官功能和潜在恢复的关键信号分子。该激酶是一种重要的能量状态感受器，它负向调节自噬控制器雷帕霉素复合体1(MTORC1)的哺乳动物靶标。相反，AMPK激活了过氧化体增殖物激活受体(PPAR)辅助激活因子1-a(PGC-1a)，它是线粒体生物发生的主要调节因子，即功能线粒体的恢复过程。与临床相关的是，我们还观察到二甲双胍的治疗，一种用于治疗2型糖尿病的常见药物，通过上游肝激酶B1(LKB1)激活AMPK，减少失血性休克老年大鼠的肾脏炎症和改善心血管功能。因此，我们的初步数据提出了一个新的假设，即AMPK的年龄相关性失调会导致自噬功能受损和线粒体生物生成减少，从而导致失血性休克期间器官损伤的持续。该项目的长期目标是通过维持受损细胞器的适当处理和促进器官恢复来确定靶途径和评估治疗器官功能障碍的药物治疗。为了验证这些新概念，本文提出了三个目标。目的1研究不同年龄大鼠失血性休克时主要器官自噬和线粒体生物发生的增龄变化及其与AMPK激活的关系。在目标2中，我们将采用药理学研究来确定AMPK对不同年龄大鼠失血性全身炎症反应、MOF和致死率的作用。在目标3中，我们将使用AMPKA基因敲除小鼠来确定AMPK通过调节mTORC1、PGC-1a和SIRT1激活自噬和线粒体生物发生的分子机制，SIRT1是一种被描述为抗衰老特性的脱乙酰酶。这些研究可能揭示影响器官代谢恢复的新的调节范例，并可能导致失血性休克的新疗法。

 DESCRIPTION (provided by applicant): Multiple organ failure (MOF) is a leading cause of death in trauma patients. Even with success of acute trauma management with fluid resuscitation and organ support protocols, many patients who survive their initial trauma are at high risk to progress towards a persistent organ injury. Clinical evidence suggests that advanced age is a risk factor for persistence of MOF and mortality. The molecular mechanisms and the impact of age in the persistence of MOF are yet to be defined. Furthermore, therapeutic strategies to promote recovery of organ function remain unavailable. Scientific evidence suggests that mitochondrial dysfunction is a critical pathogenetic event of MOF. In preliminary studies, we have observed that autophagy, an important process which enables the cells to dispose defective mitochondria, is impaired during hemorrhagic shock in the kidney of old rats (18-24 months) when compared to younger animals (2-3 months). This age-dependent impairment of autophagy correlates with severity of kidney injury and inflammatory response. At molecular analysis, we have identified AMP-activated protein kinase (AMPK) as a key signaling molecule regulating organ function and potential recovery after hemorrhagic shock. This kinase is a crucial energy status sensor, which is known to negatively regulate the autophagy controller the mammalian target of rapamycin complex 1 (mTORC1). Conversely, AMPK activates the peroxisome proliferator-activated receptor (PPAR)¿ coactivator 1-a (PGC-1a), the master regulator of mitochondrial biogenesis, i.e. the process of restoration of functional mitochondria. Of clinical relevance, we have also observed that treatment with metformin, a common drug used for type 2 diabetes that activates AMPK through the upstream liver kinase B1 (LKB1), reduces kidney inflammation and ameliorates cardiovascular function in old rats subjected to hemorrhagic shock. Thus, our preliminary data raise the novel hypothesis that an age-dependent dysregulation of AMPK causes impairment of autophagy and reduction of mitochondrial biogenesis, leading to persistence of organ injury during hemorrhagic shock. The long-term goal of this project is to identify target pathways and to evaluate pharmacological treatments for organ dysfunction by maintaining proper disposal of damaged organelle and promoting organ recovery. Three aims are proposed to validate these novel concepts. In aim 1 we will determine the age-dependent alterations of autophagy and mitochondrial biogenesis and their correlation with AMPK activation during hemorrhagic shock in major organs of rats of different ages. In aim 2 we will adopt pharmacological studies to determine the role of AMPK on hemorrhage-induced systemic inflammatory response, MOF and lethality in rats of different ages. In aim 3 we will use AMPKa knockout mice to define the molecular mechanisms by which AMPK activates autophagy and mitochondrial biogenesis through regulation of mTORC1, PGC-1a, and SIRT1, a deacetylase described for its anti-aging properties. These studies may unveil novel regulatory paradigms that impact organ metabolic recovery and may lead to new therapies in hemorrhagic shock.