MOLECULAR ASPECTS OF ALPORT RENAL DISEASE PROGRESSION

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DESCRIPTION: Alport syndrome is a relatively common (1 in 5000) genetic disorder that results in progressive renal disease with hearing loss and retinal flecks. In the past decade, the genes responsible for the syndrome have been identified, and significant progress has been made towards understanding the molecular mechanisms underlying the progressive glomerular and tubulointerstitial disease. This proposal is a focused approach aimed at defining specific aspects that may contribute towards mechanisms underlying glomerular pathogenesis. In the preliminary results section we present evidence that MMP-2, MMP-9, MMP-12 (metalloelastase) and MMP-14 (MT1-MMP) may contribute to glomerular pathogenesis in distinct ways. The role of MMPs in glomerular pathogenesis has been only marginally explored. We suggest dual contributory roles where elevated MMP-2 and MT-1 MMP are protective, while metalloelastase plays a destructive role. Real time PCR analysis of RNA from isolated glomerular preparations will be used for in situ hybridization and immunofluorescence studies to identify the temporal and spatial regulation of these MMPs in glomerular cells in vivo. Cell culture studies will be employed to probe the mechanism underlying elevated MMP expression in mouse model systems. Double and triple knockout mouse models will be used in an attempt to functionally define the roles of these MMPs in Alport glomerular pathogenesis. In the third aim we explore the mechanism of MMP dysregulation in Alport and integrin alpha1-null Alport mice. We provide evidence that the MAP kinase-signaling pathway is activated in integrin alpha1-null mice. This signaling pathway has been linked to MMP dysregulation in other systems. We will employ specific inhibitors of pp38 and pERK as well as integrin-specific neutralizing antibodies to dissect the role of integrins and MAP kinase activation in MMP dysregulation in glomeruli from integrin a1-null Alport mice. In the preliminary results, we show MMP-12 inhibition arrests the progression of renal disease in Alport mice. We have identified markedly elevated expression of GM-CSF and MCP- 1 in Alport glomeruli, cytokines known to induce expression of MMP-12 in other systems. We show that the receptors for these cytokines (alpha-GMR and CCR2, respectively) are expressed both in Alport glomeruli and on cultured glomerular podocytes. We will employ in vivo and cell culture systems to determine whether this pathway underlies MMP-12 activation in Alport glomeruli. These studies will likely form the basis for related studies in other renal disease models where these same systems have been implicated.