Cellular Pathophysiology of Acute Renal Failure

描述（由申请人提供）：缺血性和毒性急性肾衰竭仍然是住院患者发病率和死亡率的重要原因，并大大增加了护理费用。在肾近端小管，急性肾功能衰竭期间损伤的主要部位，ATP的产生是特别敏感的线粒体功能障碍，因为，取决于节段，糖酵解是不存在或最小的近端小管细胞。当分离的近端小管在体内缺血/再灌注相关条件下经受缺氧/复氧时，它们会出现严重的能量缺陷，其特征是持续的ATP耗尽和复氧期间线粒体膜电位恢复受损，这在小管细胞存活和恢复中发挥着关键作用。从损伤中恢复。能量不足可以通过特定的补充柠檬酸循环代谢物来改善。在目前的资助期间的工作表明，赤字不能很容易地解释为异常的线粒体电子传递，腺嘌呤核苷酸移位酶或F1 FO-ATP酶。相反，它似乎主要归因于由非酯化脂肪酸（NEFA）产生的线粒体脱髓鞘。降低NEFA的可用性恢复线粒体膜电位和ATP的产生，是补充底物提供益处的主要机制。为了进一步验证这一假设，并研究其机制和对肾小管细胞损伤的影响，我们计划：1）阐明NEFA穿梭于线粒体内膜阴离子载体在NEFA诱导的线粒体膜电位耗散中的作用。2)确定UCP 2是否参与NEFA效应。3)评估恢复线粒体膜电位的药物降低NEFA的功效。4)定量介导线粒体膜电位耗散的游离脂肪酸水平。5)评估NEFA诱导的质子泄漏的程度和NEFA在呼吸抑制中的作用。6)测试NEFA诱导的线粒体膜电位耗散是否解释了能量不足的基质凝聚特征，以及凝聚本身是否进一步损害线粒体功能。7)进一步研究线粒体活性氧产生的作用和NEFA对其的修饰在能量缺乏中的作用。8)表征肾小管中线粒体通透性转换的表达、NEFA对其的修饰及其对线粒体功能障碍进展的贡献。这些研究与理解和治疗缺血性急性肾衰竭和保存肾脏和其他器官移植和线粒体的关键作用，现在被认为在所有细胞类型的坏死和凋亡细胞死亡中发挥的基本理解。

DESCRIPTION (provided by applicant): Ischemic and toxic acute renal failure remain important causes of morbidity and mortality in hospitalized patients and greatly increase the expense of care. ATP production in the kidney proximal tubule, a major site of injury during acute renal failure, is especially sensitive to mitochondrial dysfunction because, depending on the segment, glycolysis is absent or minimal in proximal tubule cells. When isolated proximal tubules are subjected to hypoxia/reoxygenation under conditions relevant to ischemia/reperfusion in vivo, they develop a severe energetic deficit characterized by persistent ATP depletion and impaired recovery of mitochondrial membrane potential during reoxygenation, which plays a pivotal role in tubule cell survival and recovery from the insult. The energetic deficit can be ameliorated by specific, supplemental citric acid cycle metabolites. Work during the current funding period has shown that the deficit cannot be readily explained by abnormalities of mitochondrial electron transport, the adenine nucleotide translocase or the F1FO-ATPase. Instead, it appears to be primarily attributable to mitochondrial de-energization produced by nonesterified fatty acids (NEFA). Lowering NEFA availability restores mitochondrial membrane potential and ATP production and is a major mechanism for the benefit provided by the supplemental substrates. To further test this hypothesis and investigate the mechanisms involved and the implications for tubule cell injury, we plan to: 1) Clarify the role of NEFA shuttling on mitochondrial inner membrane anion carriers in the mediation of NEFA-induced dissipation of mitochondrial membrane potential. 2) Determine whether UCP2 is involved in the NEFA effects. 3) Assess the NEFA lowering efficacy of agents that restore mitochondrial membrane potential. 4) Quantify the free fatty acid levels mediating dissipation of mitochondrial membrane potential. 5) Assess the magnitude of the NEFA- induced proton leak and the role of NEFA in respiratory inhibition. 6) Test whether NEFA-induced dissipation of mitochondrial membrane potential accounts for the matrix condensation characteristic of the energetic deficit and whether condensation itself further impairs mitochondrial function. 7) Further investigate the role of mitochondrial reactive oxygen species production and its modification by NEFA in the energetic deficit. 8) Characterize expression of the mitochondrial permeability transition in the tubules, its modification by NEFA, and its contribution to progression of mitochondrial dysfunction. These studies are relevant to understanding and treating ischemic acute renal failure and preserving kidneys and other organs for transplantation and to the basic understanding of the critical role that mitochondria are now recognized to play during both necrotic and apoptotic cell death in all cell types.