Expression of SSAT, a novel biomarker of tubular cell damage, increases in kidney ischemia-reperfusion injury

Abstract

Ischemia-reperfusion injury (IRI) is the major cause of acute renal failure in native and allograft kidneys. Identifying the molecules and pathways involved in the pathophysiology of renal IRI will yield valuable new diagnostic and therapeutic information. To identify differentially regulated genes in renal IRI, RNA from rat kidneys subjected to an established renal IRI protocol (bilateral occlusion of renal pedicles for 30 min followed by reperfusion) and time-matched kidneys from sham-operated animals was subjected to suppression subtractive hybridization. The level of spermidine/spermine N 1-acetyltransferase (SSAT) mRNA, an essential enzyme for the catabolism of polyamines, increased in renal IRI. SSAT expression was found throughout normal kidney tubules, as detected by nephron segment RT-PCR. Northern blots demonstrated that the mRNA levels of SSAT are increased by greater than threefold in the renal cortex and by fivefold in the renal medulla at 12 h and returned to baseline at 48 h after ischemia. The increase in SSAT mRNA was paralleled by an increase in SSAT protein levels as determined by Western blot analysis. The concentration of putrescine in the kidney increased by ∼4- and ∼7.5-fold at 12 and 24 h of reperfusion, respectively, consistent with increased functional activity of SSAT. To assess the specificity of SSAT for tubular injury, a model of acute renal failure from Na+depletion (without tubular injury) was studied; SSAT mRNA levels remained unchanged in rats subjected to Na+ depletion. To distinguish SSAT increases from the effects of tubular injury vs. uremic toxins, SSAT was increased in cis-platinum-treated animals before the onset of renal failure. The expression of SSAT mRNA and protein increased by ∼3.5- and >10-fold, respectively, in renal tubule epithelial cells subjected to ATP depletion and metabolic poisoning (an in vitro model of kidney IRI). Our results suggest that SSAT is likely a new marker of tubular cell injury that distinguishes acute prerenal from intrarenal failure.