Role of Meprin A in Acute Kidney Injury

描述(由申请人提供)： Meprin A在急性肾损伤中的作用Gur P.Kaushal，Ph.D.项目摘要Meprin A是一种膜相关的中性金属内切酶，在皮质髓质交界处近端小管的刷缘膜上高度表达。Meprin A在急性肾损伤(AKI)中的具体作用尚不完全清楚。我们的研究证实，Meprin A是大鼠肾皮质中主要的基质降解酶，能够在体外降解细胞外基质(ECM)蛋白，包括IV型胶原、纤维连接蛋白、层粘连蛋白和Nidogen。我们最近发表的初步研究表明，MEPRIPs还能够从其非活性的ProForm中产生具有生物活性的促炎细胞因子IL-2，并能够蛋白水解性地处理趋化细胞因子MCP-1，这表明MEPRIPs在炎症中也是重要的。在缺血再灌注(IR)和顺铂诱导的AKI后，Meprin A重新分布在细胞质中，细胞外黏附于基底侧质膜，并在AKI期间从尿中排出裂解形式的Meprin A。这些研究表明，在急性肾小管损伤中，除根尖刷状缘膜外，Meprin A在体内的脱落和定位改变可能是有害的。膜相关的meprin A是如何重新分布和脱落的尚不清楚。初步研究表明，Meprin A的脱落可能涉及ADAM(一种去整合素和金属蛋白酶)家族的成员。利用顺铂和IR诱导的AKI体内模型，我们证明了MEPRIN A的有效抑制剂放线宁在体内抑制了MEPRIN A并改善了急性肾损伤，并且MEPRIN A缺陷的C3H/He小鼠对AKI具有抵抗力。有趣的是，我们观察到，在IR和顺铂诱导的AKI进展过程中，在正常小鼠的尿液中检测不到的Nidogen和meprin-beta片段在血清肌酐上升之前显著增加。因此，有一个独特的机会来进一步探讨meprin A在AKI中的作用和作用机制。我们推测，梅普林A具有巨大的破坏力，在急性肾损伤期间由于定位改变而对肾近端小管有害，了解其作用机制对于保护或减少急性肾损伤具有重要意义。此外，我们假设在AKI过程中排出的甲氨蝶呤和Nidogen片段将作为AKI早期检测的重要生物标志物。我们将通过以下具体目标来验证这些假说：1.利用IR和顺铂肾毒性的实验模型，研究急性肾损伤期间梅普林A再分布、肾脏损伤、白细胞渗透、梅普林A脱落和尿中梅普林亚单位排泄之间的时间关系。2.检测甲孕酮在IR和顺铂肾毒性中的体内降解产物。3.探讨急性心肌梗死后MEPRIN A介导炎症反应的机制及抑制MEPRIN A的作用意义。了解meprin A在AKI中的潜在作用将为预防急性肾损伤的特定治疗干预提供见解。 公共卫生相关性： 急性肾损伤(AKI)是一种常见而严重的疾病，占所有住院患者的5-7%，ICU住院患者的30%，死亡率高，在发展为AKI并随后需要血液透析的患者中，死亡风险大于60%。与肾衰竭相关的费用估计为每年100亿美元。在过去的40年里，死亡率几乎没有改善。目前建议的主要目标是研究meprin A作为预防或减少AKI的新的治疗靶点，最重要的是，研究其在AKI过程中分布改变的机制。此外，该研究还将确定梅普林A介导的尿标志物用于AKI的早期检测。因此，由于退伍军人中有很大一部分人患有AKI，该提案在临床上与VA患者群体相关。

DESCRIPTION (provided by applicant): Role of Meprin A in acute kidney injury Gur P. Kaushal, Ph.D. Project Summary Meprin A, a membrane-associated neutral metalloendoprotease, is highly expressed at the brush- border membranes of proximal tubules in the corticomedullary junction. The specific role of meprin A during acute kidney injury (AKI) is not fully understood. Our studies identified that meprin A is the major matrix-degrading protease in the rat kidney cortex capable of degrading the extracellular matrix (ECM) proteins including collagen IV, fibronectin, laminin, and nidogen in vitro. Our recently published and preliminary studies demonstrated that meprins are also capable of producing biologically active proinflammatory cytokine interleukin 1-2 from its inactive proform and proteolytically processing chemotactic cytokine MCP-1, suggesting that meprins are also important in inflammation. Following ischemia-reperfusion (IR)- and cisplatin-induced AKI, meprin A is redistributed throughout the cytoplasm and extracellularly adhering toward the basolateral plasma membrane and the cleaved form of meprin A is excreted in the urine during AKI. These studies suggest that shedding and altered localization of meprin A in places other than the apical brush-border membranes may be deleterious in vivo in acute tubular injury. How membrane-associated meprin A is redistributed and shed is not known. Preliminary studies suggest that meprin A shedding may involve a member of the ADAM (a disintegrin and metalloproteinase) family. Using in vivo models of cisplatin- and IR-induced AKI, we demonstrated that actinonin, a potent inhibitor of meprin A inhibits meprin A in vivo and ameliorates acute kidney injury and meprin A-deficient C3H/He mice are resistant to AKI. Interestingly, we observed that nidogen and meprin-beta fragments, undetectable in the urine of normal mice, increased significantly before the rise in serum creatinine during the progression of IR- and cisplatin- induced AKI. Thus, a unique opportunity exists to further explore the role and mechanism of action of meprin A in AKI. We hypothesize that meprin A, with its enormous destructive potential, is detrimental to renal proximal tubules due to altered localization during AKI and that understanding its mechanism of action is important in protecting or reducing AKI. In addition, we hypothesize that meprin and nidogen fragments excreted during AKI will serve as important biomarkers for early detection of AKI. We will test these hypotheses through the following specific aims: 1. Examine the temporal relationship between meprin A redistribution, renal injury, leukocyte infiltration, meprin A shedding, and urinary excretion of meprin subunits during AKI using experimental models of IR and cisplatin nephrotoxicity. 2. Examine meprin A-mediated in vivo degradation products of nidogen during IR and cisplatin nephrotoxicity. 3. Determine the mechanisms of meprin A-mediated inflammatory effects and functional significance of meprin A inhibition after the onset of AKI. Understanding the underlying role of meprin A in AKI will provide insights for specific therapeutic interventions to prevent acute kidney injury. PUBLIC HEALTH RELEVANCE: NARRATIVE Kidney disease due to ischemia and toxic acute kidney injury (AKI) is a frequent and serious disease, which affects 5-7% of all hospitalized patients and 30 % of ICU admissions with a high mortality rate and the risk of mortality is greater than 60% among patients who develop AKI and subsequently require hemodialysis. The expenses related with kidney failure are estimated at $10 billion per year. There has been little improvement in mortality over the last four decades. The primary goal of the current proposal is to study meprin A as a novel therapeutic target to prevent or reduce AKI and most importantly, to investigate the mechanism of its altered distribution during AKI. In addition the study will identify meprin A-mediated urinary markers for early detection of AKI. Therefore, the proposal is clinically relevant to the VA patient population since a large segment of veterans suffer from AKI.