The goals of this project are to develop and validate an in vitro testing battery which mechanistically predicts the nephrotoxic potential of chemical agents. This in vitro testing battery is designed to detect direct cytotoxicity, hemodynamic, and pH dependent solubility mechanisms of injury. Direct cytotoxicity is assessed by measuring changes in integrated cellular functions (gluconeogenesis; oxygen consumption; transport of sugars, organic acids and bases), redox status and compartmental integrity (mitochondrial, lysosomal, plasma membrane integrity, oxidant stress, etc) in rat and rabbit renal slices or tubular suspension. The feasibility of monitoring hemodynamic induced cytotoxicity in vitro is novel and will be assessed by measuring calcium dependent mesangial cell contractility and locally released hemodynamic mediators (arachidonic acid, renin, prostaglandins) from isolated glomeruli and medullary slices. Hormone stimulated activation of cAMP in slices and pH dependent solubility determinations in urine will be assessed as models for predicting distal tubular injury. Sulfanilamides, cyclosporine and cysteine conjugates will be used (+ liver S9 as a metabolism system) as model compounds which should discriminately effect this nephrotoxicity battery. In vitro technologies capable of predicting toxic mechanisms would markedly accelerate the product development process in the pharmaceutical industry while markedly reducing the costs associated with animal usage.
