Carbon monoxide (CO), a product of heme degradation by heme oxygenases (HO), has been shown to provide cytoprotection in various tissue injury models. This study examined the efficacy and molecular mechanisms of exogenously delivered inhaled CO in protecting liver grafts from cold ischemia/reperfusion (I/R) injury associated with liver transplantation. Orthotopic syngenic liver transplantation (OLT) was performed in Lewis rats with 18 hrs cold preservation in UW solution. Recipients were exposed to air or different concentrations of CO (20-250 ppm) for 1 hr before and 24 hrs after OLT and sacrificed 1-48 hrs posttransplant. CO inhalation significantly decreased serum ALT levels and suppressed hepatic necrosis and neutrophil accumulation at 24-48 hrs after OLT in a dose dependent manner. Reduced hepatic injury with inhaled CO associated with marked downregulation of early mRNA expression for TNF- and IL-6. Expression in liver grafts of mRNA and protein of stress-responding enzyme, iNOS, was significantly reduced by CO, while HO-1 was only marginally suppressed. Cold hepatic I/R injury associated with prompt MAPK phosphorylation in liver grafts at 1 hr after OLT, and CO significantly inhibited phosphorylation of ERK1/2 MAPK and its upstream MEK1/2 and downstream transcriptional factor c-Myc. CO also significantly inhibited I/R injury-induced STAT1 and STAT3 activation. In contrast, CO did not inhibit p38 or JNK MAPK pathways during hepatic I/R injury. Results demonstrate that exogenous CO suppresses early proinflammatory and stress-response gene expression and efficiently ameliorates hepatic I/R injury. The possible mechanism may include the downregulation of MEK/ERK1/2 signaling pathway with CO.