Background Ischemia/reperfusion injury (I/R injury) of the liver remains a significant problem during liver surgery and transplantation. I/R injury is associated with liver apoptosis, which is mediated by death receptors such as Fas and tumor necrosis factor alpha (TNF-α), and/or mitochondrial dysfunction induced by cellular stress. Caspase-8 is presumed to be the apex of the death-mediated apoptosis pathway, whereas caspase-3 belongs to the "effector" proteases in the apoptosis cascade. Synthetic small interfering RNAs (siRNAs) specifically suppress gene expression by RNA interference. Therefore, we evaluated the therapeutic efficacy of caspase-8 and caspase-3 siRNA in a murine model of liver I/R injury. Methods In C57BL/6 mice, 45% or 70% of the liver mass was clamped for 90 minutes. For survival analysis, total hepatic ischemia was induced for 45 minutes. In vivo delivery of siRNA was performed via the portal vein by high-volume injection (0.5 nmol of siRNA in 1 mL containing 10% lipiodol) 60 minutes before ischemia. As a control, animals received either vehicle or non-sense siRNA (siRNA-scrambled). Results Liver uptake of siRNA was analyzed in transgenic mice who express β-galactosidase (β-gal) (C57BL/6J-TgN(MTn-LacZ)204Bri) after administration of siRNA-LacZ. A 3- to 4-fold decrease in β-gal activity was accomplished at 0.5 nmol. No significant change in β-gal activity was demonstrated in mice receiving non-sense siRNA. Immunohistochemical studies found that 60% of the liver cells efficiently took up siRNA. Significant reduction in serum aspartate transaminase was found in animals treated with siRNA caspase-8 or caspase-3 compared with siRNA-scrambed or vehicle-treated controls. More than a 60% reduction in caspase-8 and caspase-3 gene expression and activities was accomplished after siRNA administration. Animals treated with siRNA presented lower infiltration of polymorphonuclear leukocytes and better preservation of the liver architecture compared with controls. All of the control mice subjected to total liver ischemia died within 5 days. In contrast, 30% of the animals given siRNA caspase-8 and 50% of those treated with siRNA caspase-3 survived indefinitely (>30 days). Conclusions Small interfering RNA targeted to caspase-8 and caspase-3 provided significant protection against I/R injury to the liver. This approach could be therapeutic in liver transplantation and other conditions associated with I/R injury to the liver.