Autophagy is a tightly regulated, programmed mechanism to eliminate damaged organelles and proteins from a cell to maintain homeostasis. Cisplatin, a chemotherapeutic agent, accumulates in the proximal tubules of the kidney and causes dose-dependent nephrotoxicity, which may involve autophagy. In the kidney, cisplatin induces the protective antioxidant heme oxygenase-1 (HO-1). In this study, we examined the relationship between autophagy and HO-1 during cisplatin-mediated acute kidney injury (AKI). In wild-type primary proximal tubule cells (PTC), we observed a time-dependent increase in autophagy after cisplatin. In HO-1-/- PTC, however, we observed significantly higher levels of basal autophagy, impaired progression of autophagy, and increased apoptosis after cisplatin. Restoring HO-1 expression in these cells reversed the autophagic response and inhibited apoptosis after treatment with cisplatin. In vivo, although both wild-type and HO-1-deficient mice exhibited autophagosomes in the proximal tubules of the kidney in response to cisplatin, HO-1-deficient mice had significantly more autophagosomes, even in saline-treated animals. In addition, ecdysone-induced overexpression of HO-1 in cells led to a delay in autophagy progression, generated significantly lower levels of reactive oxygen species, and protected against cisplatin cytotoxicity. These findings demonstrsate that HO-1 inhibits autophagy, suggesting that the heme oxygenase system may contain therapeutic targets for AKI. Copyright © 2010 by the American Society of Nephrology.