Following heterogeneous forms of experimental renal injury (e.g., nephrotoxins, ischemia, urinary tract obstruction), proximal tubules undergo adaptive changes which protect them from further attack. This can either prevent a worsening of established acute renal failure (ARF), or block recurrent episodes. Characteristics of this """"""""acquired cytoresistance"""""""" (AC) are that: 1) it requires about 18 hrs to develop; 2) it is expressed against diverse forms of secondary insults; 3) it can protect for at least 30 days; 4) it does not require de novo protein synthesis; and 5) it is mediated by a fundamental change within the proximal tubular plasma (and possibly mitochondrial) membrane(s), conferring cellular protection against further attack. Recent work from this laboratory has identified increased tubular membrane cholesterol expression following renal injury, and this is critical to the AC state. Given the broad based biologic relevance of AC (e.g., it can also contribute to cancer cell resistance to chemotherapy), this proposal has the following 3 Specific Aims: 1. Determine the mechanisms for, and nature of. the cholesterol elevations in the AC state: a) do the cholesterol elevations arise from increased synthesis, increased uptake, or decreased efflux? b) do the cholesterol increments reflect elevations of free vs. esterifled cholesterol pool(s)? and c) what subcellular cholesterol distribution patterns result? 2. Determine mechanisms by which cholesterol mediates AC: a) which cholesterol moieties (e.g. free vs. esterifled) confer AC? b) do they, how do they, increase membrane resistance to attack?; and c) is there a mechanistic link between increased cell cholesterol and improved mitochondrial performance in the AC state? 3. Define whether AC is impacted by two cholesterol- associated signaling pathways: Specifically, do the """"""""upstream"""""""" mevalonate pathway and/or the """"""""downstream"""""""" cholesterol/sphingomyelin microdomain (raft/caveolae) pathway impact the emergence, or expression, of the AC state? The ultimate goal of these experiments is to better define cellular adaptations to injury in an attempt to devise prophylactic strategies to either increase (or, in the case of cancer chemotherapy, decrease) cellular resistance to superimposed attack.
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