MECHANISMS OF ISCHEMIC INJURY AND REPAIR

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DESCRIPTION (provided by applicant): The overall goal of the proposed studies remains to understand the mechanisms of ischemic kidney cell injury and repair with a long range goal to establish therapies that will be useful to prevent and treat acute renal failure (ARF) in man. Over the last support period we have developed and tested the hypothesis that ischemic ARF is a disease in which leukocyte-endothelial interactions, particularly in the outer medulla of the kidney, lead to congestion of the small vessels and impaired oxygen delivery with resultant tubule cell injury. The damaged tubule and activated endothelium contributes to additional vascular compromise by producing cytokines and chemokines which up regulate adhesion receptors and attract leukocytes. We have recently established three mouse models in which the kidney is protected against ischemic injury. These models will be exploited to evaluate the hypothesis and identify proteins that are candidates as important contributors to injury or endogenous processes of protection. In Specific Aim 1 we will use models of protection against ischemic injury to define the importance of outer medullary leukocyte and endothelial activation and interactions for the pathophysiology of ischemic acute renal failure. The state of differentiation and stress response of the S3 segment will be evaluated to put into context the potential importance of alterations in leukocyte-endothelial interactions relative to change in susceptibility to injury of the proximal tubule epithelial cell. Adhesion molecule and chemokine involvement will be studied as will the potential role of Kidney Injury Molecule-1 (KIM-I) in leukocyte-epithelial interactions. In Specific Aim 2 we will evaluate the role of nmb on leukocyte function and identify additional candidate genes implicated in inflammation that may be important for ischemic injury or inducible protective processes in the outer medulla of the kidney. We will determine the effect of nmb and three of its potential functional domains (the polycystic kidney [PKD] domain, and the YXXphi and di-leucine domains in the cytosolic tail) on binding of leukocytes to endothelial cells and on phagocytosis. Gene expression analyses will be performed by transcript profiling coupled to a gene database query. We will identify those genes that show coordinate up or down regulation under three conditions that we have identified to result in protection of the kidney against ischemic injury. Immunocytochemistry, in situ hybridization, laser capture microdissection and TaqMan real time RT-PCR will be used to identify that subgroup of genes whose expression pattern is differentially regulated in the postischemic outer medulla when comparing protected vs non-protected kidneys.