EARLY RENAL ISCHEMIA, WITH OR WITHOUT REPERFUSION, ACTIVATES NFκB AND INCREASES TNF-α BIOACTIVITY IN THE KIDNEY

Abstract

Acute tubular necrosis (ATN) and the ensuing renal failure induced by ischemia and reperfusion injury (I/R) remain a major cause of morbidity and mortality among patients in the intensive care unit. Although it is well established that exogenous tumor necrosis factor-α (TNF) induces renal injury, it remains unknown whether ischemia and/or reperfusion activates the signaling mechanisms required for renal TNF production. We hypothesized that ischemia and/or reperfusion would activate the oxidant sensitive TNF transcription factor, nuclear factor kappa B (NFκB), and thereby lead to renal TNF production. Male Sprague-Dawley rats were anesthetized with sodium pentobarbital, after which various periods of renal ischemia, with or without reperfusion, were induced in rats. At different time intervals, kidneys were harvested and NFκB activation (electrophoretic mobility shift assay), TNF mRNA content (RT-PCR), and TNF bioactivity (WEHI-164 cell clone cytotoxicity assay) were determined. Results indicate that 15 minutes of ischemia alone activates NFκB, whereas peak activation occurred at 30 minutes of ischemia alone. NFκB remained activated through 60 minutes reperfusion. Thirty minutes of ischemia is required to induce renal TNF mRNA production; however, renal TNF protein expression and bioactivity peaked following 1 hour of ischemia and 2 hours reperfusion. These results are the initial demonstration that renal ischemia, with or without reperfusion, activates the TNF transcription factor NFκB and increases TNF bioactivity in the kidney.