Heme oxygenase-1 (HO-1), a 32-kd microsomal enzyme, is induced as an adaptive response to a wide variety of injurious stimuli. We examined the possible role of HO-1 in cold storage of renal proximal tubular epithelial (RPTE) cells. Hemin, a potent HO-1-inducer, caused a time-dependent increase in HO-1 mRNA and protein expression. Hemin pretreatment of human RPTE cells before cold storage conferred cytoprotection. Increased HO-1 protein was associated with a brisk and early increase in catalytically active iron and a robust increase in cellular ferritin. Deferoxamine, an iron sequestrating antioxidant, prevented hemin-induced iron release and the increase in ferritin, suggesting iron release as an antecedent mechanism for ferritin induction. To verify that the proximate cause of hemin cytoprotection was due to HO-1 induction, we transiently transfected LL-CPK1 porcine kidney cells with a HO-1 expression vector before cold storage. HO-1 transfection resulted in increased expression of HO-1 protein and reduced cell injury during cold storage. The novel observation that prior induction of HO-1 prevents cold storage-induced cell injury suggests that a similar strategy may prove efficacious in preventing cold storage-induced organ damage during transplantation.