Vancomycin (VAN) is a cationic glycopeptide antibiotic used to treat a variety of gram-positive infections. VAN is regarded as a potentially ototoxic and nephrotoxic agent. The reported incidence of nephrotoxicity in humans is highly variable and ranges from 5% to 17%. The incidence of nephrotoxicity increases to as high as 35% when VAN is used concomitantly with an aminoglycoside antibiotic. Animal studies evaluating the mechanisms of VAN-induced nephrotoxicity are incomplete and mostly descriptive in nature. Little is known regarding the mechanisms for renal accumulation of VAN within the kidney and nothing is known about the mechanisms involved in the production of nephrotoxicity. this laboratory has developed a model for VAN-induced nephrotoxicity in the female Sprague-Dawley (SD) rat. Nephrotoxicity is demonstrated by a number of functional and structural alterations. These changes include elevations in blood urea nitrogen (BUN) concentrations, decreased gluconeogenic capacity and decreased ability to accumulate organic ions. Increases in kidney weight due to increases in total renal water, protein and phospholipid are also observed. Pretreatment of rats with polyaspartic acid (PAA), prior to VAN administration, prevents nephrotoxicity and appears to be linked to renal VAN accumulation. The premise on which this proposal is based is that VAN must be accumulated within renal cells in order to produce nephrotoxicity. Once inside the cells, VAN interacts with renal proteins and phospholipids at the level of critical membrane systems such as the plasma membrane and mitochondrial membrane. These alterations result in dysregulation of renal cell transport properties and energy production, which are then translated into a nephrotoxic response. The purpose of this proposal is to delineate the mechanisms for VAN accumulation within renal cells and then determine the mechanisms by which VAN produces nephrotoxicity. These mechanisms will be addressed in terms of site-specific responses of proximal & distal tubular cells to VAN. This goal will be accomplished by taking an integrated, mechanistic approach to the problem. Renal VAN accumulation will be determined in vivo and in vitro. Primary cultures of proximal and distal tubular cells grown on membrane supports will be used to determine the apical vs basolateral uptake of VAN. The mechanisms for VAN-induced phospholipidosis and increased protein content will be determined by evaluating synthesis and degradation. The effects of VAN on mitochondrial and plasma membrane phospholipid profiles will be evaluated by HPLC separation of key phospholipids. These alterations will be correlated to changes in plasma membrane (apical and basolateral) transport ability and mitochondrial energy production. Further examination on the nature of the interaction between VAN and PAA may shed some more light into the mechanisms for VAN interactions with renal tissue. the mechanism for PAA interference with renal VAN accumulation will be evaluated by competitive binding studies on whole cells and membrane fractions. The data gathered from this proposal should provide significant information regarding the mechanisms of VAN-induced nephrotoxicity and lay the groundwork for future studies on differential susceptibility of proximal vs distal tubular cells to toxicant-induced injury.
