A novel model of ischemia in renal tubular cells which closely parallels in vivo injury

Purpose. Renal ischemia-reperfusion (IR) injury is a devastating clinical p roblem. While effective animal models have been developed to investigate th is condition, they are limited by differential renal cell inflammatory medi ator production and heterogeneous cell sensitivity to ischemia. We therefor e developed an in vitro model of renal tubular cell ischemia that simulates the cellular injury observed in animal models of renal IR injury. Materials and methods. Using the established renal tubular cell line, LLC-P K1, simulated ischemia was induced by immersing the cellular monolayer in m ineral oil. The effect of simulated ischemia on renal tubular cells was the n determined by measuring the time course of TNF-alpha protein expression ( ELISA), TNF-alpha mRNA induction (RT-PCR), and renal tubular cell apoptosis (TUNEL). Results. Maximal TNF-alpha protein expression occurs following 60 min of si mulated ischemia and 2 h of substrate replacement (reimmersion in media), a nd maximal TNF-alpha mRNA induction occurs following 60 min of simulated is chemia. Cellular apoptosis peaks following 60 min of simulated ischemia and 24 h of reperfusion. Conclusion. The time course of TNF-alpha production and apoptosis induction in this model closely parallels the time course for these markers in vivo. This study constitutes the initial demonstration that an in vitro oil imme rsion model of ischemia simulates the cellular injury (TNF-alpha production and apoptosis) observed in animal models of renal ischemia-reperfusion. Th is model may be used to study cellular mechanisms of IR In the absence of t he systemic confounding variables. (C) 2001 Academic Press.