Meprin Metalloproteases in Kidney Injury

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ABSTRACT Acute kidney injury (AKI) is associated with high morbidity and mortality rates, and significantly impacts the quality of life for patients and their families. The cost of treating acute kidney disease in the United States runs in the tens of billions of dollars annually. However, treatment outcomes for AKI are poor in part because underlying mechanisms are not fully understood. Ischemia/reperfusion (IR) is the leading cause of AKI and is associated with inflammation in the initiation, tissue repair, and chronic phases. However, the genes which modulate inflammation in IR are not fully defined. The proximal kidney tubules are the most susceptible to AKI IR-induced injury. Meprins, zinc metalloproteases of the astacin family, are the most abundantly expressed proteins in the brush border membranes (BBMs) of proximal kidney tubules. Meprins are made up of two subunits, α and β, which form two highly similar protein isoforms; meprin A (a homooligomer of α-α subunits or a heterooligomer of α-β subunits) and meprin B (a homooligomer of β-β subunits). Disruption of the meprin genes and administration meprin inhibitors protect mice from IR-induced AKI, suggesting that meprins exacerbate kidney injury. The mechanisms by which meprins enhance IR-induced kidney injury are not fully understood. The meprin protein isoforms have common and distinct substrates. Studying the interactions between meprins and their targets in the kidney will increase understanding of how they impact cell function and the pathology of kidney disease. Known meprin substrates include several mediators of inflammation such as (i) proinflammatory cytokines e.g. interleukins (IL-1β, IL-6, IL-18) and chemokines e.g. monocyte chemo- attractant protein-1 (MCP-1). Proteolytic processing by meprins inactivates IL-6, but activates IL-1β and IL-18. Additional support for a role for meprins in inflammation came from recent studies demonstrating that meprin α mediates the release of the anti-inflammatory peptide N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) from thymosin β4. It is not known if the Ac-SDKP release plays a role in modulating IR-induced kidney injury. A long-term consequence of inflammation is fibrosis, suggesting that there is an imbalance in extracellular matrix (ECM) protein metabolism. Since meprins cleave and/or degrade several ECM proteins (e.g. nidogen-1, laminin, fibronectin, and collagen), they could alleviate the fibrosis associated with IR-induced kidney injury. Meprins also proteolytically process proteins that mediate cell signaling pathways involved in ECM metabolism (e.g. the protein kinase A pathway). Proteolytic processing by meprins reduces the kinase activity of three isoforms of the catalytic subunit of PKA (PKA Cα, Cβ1, and Cβ2). The proposed studies will utilize meprin knock out mice to determine the mechanisms by which meprins modulate inflammation and the progression of IR-induced renal injury in the initiation, reparative, and chronic phases. The goals will be achieved by pursuing three closely related specific aims; (i) determine how and which meprin isoforms impact the inflammatory response in the initiation, reparative, and chronic phases of IR-induced kidney injury, (ii) determine how proteolytic processing of isoforms of the catalytic subunit of PKA (PKA C) by meprins impacts downstream targets of the PKA signaling pathway in IR-induced acute kidney injury, (iii) determine fibrosis-associated genes impacted by meprin activity in IR-induced renal injury. A combination of proteomics, real-time PCR, flow cytometry, and immunohistochemical analysis will be used. Data from these studies will enhance understanding of the mechanisms underlying the progression of IR-induced kidney injury and inform the development of more effective therapies.