Background: Carbon monoxide (CO) is a cytoprotective and homeostatic molecule with important signaling capabilities in physiological and pathophysiological situations. CO protects cells/tissues from damage by free radicals or oxidative stress. NAD(P)H:quinone oxidoreductase (NQO1) is a highly inducible enzyme that is regulated by the Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway, which is central to efficient detoxification of reactive metabolites and reactive oxygen species (ROS). Methods: We generated NQO1 promoter construct. HepG2 cells were treated with CO Releasing Molecules-2 (CORM-2) or CO gas and the gene expressions were measured by RT-PCR, immunoblot, and luciferase assays. Results: CO induced expression of NQO1 in human hepatocarcinoma cell lines by activation of Nrf2. Exposure of HepG2 cells to CO resulted in significant induction of NQO1 in dose- and time-dependent manners. Analysis of the NQO1 promoter indicated that an antioxidant responsible element (ARE)-containing region was critical for the CO-induced Nrf2-dependent increase of NQO1 gene expression in HepG2 cells. Conclusion: Our results suggest that CO-induced Nrf2 increases the expression of NQO1 which is well known to detoxify reactive metabolites and ROS.