Nucleophosmin Centered Diagnostics and Treatment of Ischemic Acute Kidney Injury

项目总结/摘要 缺血性急性肾损伤（阿基）是一种常见的，往往是一个危及生命的后果减少， 流向这一关键器官的血液导致严重的发病率和死亡率，并且缺乏有效的治疗。 这是我们第一次在缺血鼠和人中检测到相同的生化事件。 肾脏介导肾细胞死亡，导致阿基。在本建议中，我们研究了 核磷蛋白（NPM），一种存在于所有哺乳动物细胞中的蛋白质，它从“朋友”转化为“朋友”。 “敌人”在缺血性应激。在肾缺血期间，NPM经历生化变化， 促进其在肾上皮细胞中的毒性，这是阿基期间器官衰竭的主要原因。这些步骤 包括：NPM从核到胞质区室的易位，NPM去寡聚化， 与Bax相互作用，Bax是线粒体外膜损伤的主要原因， 细胞凋亡和坏死，这两种形式的细胞死亡在缺血性人类中一直被检测到。 肾脏使用质谱的初步数据已经确定了5个磷酸化变化， 小鼠和人类近端小管细胞，肾组织和尿液，调节NPM毒性， 肾缺血在三个AIMS中，我们将把翻译后磷酸化事件与NPM毒性联系起来， 使用库存肾活检组织确定NPM在人类肾脏疾病中的作用，测试NPM 磷酸突变蛋白，复制毒性修饰，并开发新的肽试剂， 改善NPM毒性。一旦在体外鉴定，我们将测试最有效的肽， 用于治疗体内缺血性阿基的药物。这种体外试验最大限度地减少了动物数量 要求进行我们提出的研究，将检查性别在阿基中的生物学效应 诊断和治疗。我们已经开发了一种新的尿NPM检测方法， 早期阿基生物标志物，也用于测量我们治疗的疗效。评估 尿液和组织中总的和位点特异性磷酸化的NPM将触发早期治疗， 最终使我们能够预测阿基的严重程度及其恢复。人肾脏中NPM的检测 活组织检查组织将识别潜在的阿基原因（例如，缺血或肾毒素诱导阿基）， 适合抗NPM疗法。由于NPM在小鼠缺血期间受到相同的调节， 对于人类，在小鼠中的有效干预可能会转化为人类临床试验。

Project Summary/Abstract Ischemic acute kidney injury (AKI) is a common and often a life-threatening consequence of reduced blood flow to this critical organ causes major morbidity and mortality, and lacks an effective treatment. For the first time, we have detected identical biochemical events in both ischemic murine and human kidneys that mediate renal cell death leading to AKI. In this proposal, we study the role of nucleophosmin (NPM), a protein present in all mammalian cells that is converted from “friend” to a “foe” during ischemic stress. During renal ischemia, NPM undergoes biochemical changes that promote its toxicity in renal epithelial cells, a major contributor to organ failure during AKI. These steps include: NPM translocation from the nuclear to the cytosolic compartment, NPM de-oligomerization, interaction with Bax, a major cause of outer mitochondrial membrane injury, and renal cell death by both apoptosis and necrosis, the two forms of cell death consistently detected in ischemic human kidneys. Preliminary data using mass spectroscopy have identified 5 phosphorylation changes in murine and human proximal tubule cells, kidney tissue, and urine that regulate NPM toxicity during renal ischemia. In three AIMS, we will link post-translational phosphorylation events with NPM toxicity, identify the role of NPM in human renal disease using banked kidney biopsy tissue, test NPM phospho-mutant proteins that replicate toxic modifications, and develop novel peptide reagents for ameliorating NPM toxicity. Once identified in vitro, we will test the most effective peptides and pharmaceuticals to treat ischemic AKI in vivo. This in vitro testing minimizes the number of animals required to perform our proposed studies that will examine the biologic effects of gender in AKI diagnosis and treatment. We have developed a novel urinary NPM assay that will be used to as an early AKI biomarker and also for measuring the therapeutic efficacy of our treatment. Assessment of total and site-specific phosphorylated NPM in urine and tissue will trigger early treatment, and may ultimately permit us to predict the severity of AKI and its recovery. Detection of NPM in human kidney biopsy tissue will identify potential AKI causes (e.g., ischemia or nephrotoxin-induced AKI) that are amenable to anti-NPM therapeutics. Since NPM is identically regulated during ischemia in mice and humans, it is likely that effective interventions in mice will translate to human clinical trials.