Meprin A Metalloproteinase in Acute Kidney Injury

描述（由申请人提供）：Meprins是一种细胞表面和分泌的“astacin”家族的寡聚金属内肽酶，在肾近端小管的刷状边界膜上高度表达。meprins在急性肾损伤（AKI）中的具体作用尚不完全清楚。我们的研究发现，meprin A是大鼠肾皮质中主要的基质降解蛋白酶，能够在体外降解细胞外基质（ECM）蛋白，包括胶原IV、纤维连接蛋白、层粘连蛋白和氮原。我们最近发表的和初步研究表明，meprins也能够从其无活性的形式产生生物活性的促炎细胞因子白细胞介素1- β和蛋白水解处理趋化细胞因子MCP-1，这表明meprins在炎症中也很重要。从外周血或肾组织中分离出的白细胞表达meprin β。此外，我们的研究表明，在缺血再灌注和顺铂诱导的AKI后，meprin A向近端小管的基底外侧重新分布。这些研究表明，在急性肾小管损伤中，meprin A在根尖刷缘膜以外的其他地方的定位改变可能是有害的。初步研究表明meprin A的脱落可能与ADAM（一种崩解素和金属蛋白酶）家族成员有关。使用顺铂和缺血再灌注诱导的AKI的体内模型，我们证明了actionin（一种有效的meprin a抑制剂）在体内抑制meprin a并改善急性肾损伤，meprin a缺陷小鼠对顺铂肾毒性具有抗性。有趣的是，我们观察到正常小鼠尿液中检测不到的nidogen和meprinβ片段在顺铂肾毒性期间显著增加，而放线素显著阻止了nidogen片段的尿排泄。因此，为进一步探讨meprin a在AKI中的作用和作用机制提供了独特的机会。我们假设meprin A具有巨大的破坏潜力，由于AKI期间肾近端小管的定位改变而对肾近端小管有害，了解其作用机制对保护或减轻AKI至关重要。我们将通过以下具体目标来检验假设：具体目标是：1。利用IR和顺铂肾毒性实验模型研究AKI期间meprin A再分布、肾损伤、白细胞浸润和meprin A脱落之间的时间关系。2. 在IR和顺铂肾毒性过程中meprin a介导的ECM成分体内降解产物的鉴定。3. 使用meprin抑制剂和meprin A缺陷小鼠，确定meprin A介导的AKI炎症作用机制和meprin A的功能意义。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。

DESCRIPTION (provided by applicant): Meprins, cell-surface and secreted oligomeric metalloendopeptidases of the 'astacin' family, are highly expressed at the brush-border membranes of the kidney proximal tubules. The specific role of meprins 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-beta from its inactive proform and proteolytically processing chemotactic cytokine MCP-1, suggesting that meprins are also important in inflammation. Leukocytes isolated from the peripheral blood or the kidney tissue were found to express meprin beta. Furthermore, our studies demonstrate that, following ischemia-reperfusion- and cisplatin-induced AKI, meprin A is redistributed toward the basolateral side of the proximal tubule. These studies suggest that altered localization of meprin A in places other than the apical brush-border membranes may be deleterious in vivo in acute tubular injury. 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 ischemia- reperfusion-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 mice are resistant to cisplatin nephrotoxicity. Interestingly, we observed that nidogen and meprin-beta fragments, undetectable in the urine of normal mice, increased significantly during cisplatin nephrotoxicity and actinonin markedly prevented urinary excretion of nidogen fragments. 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. We will test the hypothesis through the following specific aims: The Specific Aims are: 1. Examine the temporal relationship between meprin A redistribution, renal injury, leukocyte infiltration, and meprin A shedding during AKI using experimental models of IR and cisplatin nephrotoxicity. 2. Identification of meprin A-mediated in vivo degradation products of the ECM components during IR and cisplatin nephrotoxicity. 3. Determine the mechanisms of meprin A-mediated inflammatory effects and functional significance of meprin A during AKI using a meprin inhibitor and meprin A-deficient mice. Understanding the underlying role of meprin A in AKI will provide insights for specific therapeutic interventions to prevent acute kidney injury. PUBLIC HEALTH RELEVANCE: Kidney disease 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 expenses related for patients with kidney failure are estimated at $10 billion per year. The risk of mortality is greater than 60% among patients who develop AKI and subsequently require hemodialysis. There has been little improvement in mortality over the last four decades. The mechanisms underlying the causes of kidney injury remain poorly defined. The studies proposed in this application are aimed directly at understanding these mechanisms, which will result in appropriate therapeutic interventions.