The increasing shortage of donor organs has led to the increasing use of organs from non–heart-beating donors. We aimed to assess the impact of venous systemic oxygen persufflation (VSOP) supplemented with nitric oxide (NO) gas during the cold storage (CS) of warm ischemia (WI)–damaged experimental liver grafts. Rat livers (n = 5 per group) were retrieved after 30 minutes of WI induced by cardiac arrest (the WI group) and were thereafter preserved for 24 hours by CS in histidine tryptophan ketoglutarate solution. During CS, gaseous oxygen was insufflated via the caval vein with 40 ppm NO (the VSOP-NO group) or without NO (the VSOP group). Cold-stored livers without WI served as controls. Liver viability was assessed after the preservation period by normothermic isolated reperfusion for 45 minutes with oxygenated Krebs-Henseleit buffer. After 45 minutes of reperfusion, the VSOP-NO–treated livers showed significantly lower alanine aminotransferase values than the WI-damaged livers (10.2 ± 0.2 versus 78.2 ± 14.6 IU/L), whereas the control livers showed no differences from the VSOP-NO–treated livers. The mitochondrial enzyme release was lower in the VSOP-NO group (4.0 ± 0.7 IU/L) versus the WI group (18.2 ± 4.9 IU/L). An increased portal vein pressure was observed throughout reperfusion (45 minutes) in the WI group (21.7 ± 0.2 mm Hg) versus the VSOP-NO group (12.2 ± 0.8 mm Hg) and the control group (19.9 ± 0.4 mm Hg). Furthermore, the NO concentration in the perfusate after 5 minutes of reperfusion was highest in the VSOP-NO group. The release of malondialdehyde into the perfusate was significantly reduced in the VSOP-NO group (0.9 ± 0.1 nmol/mL) versus the WI group (31.3 ± 5.3 nmol/mL). In conclusion, the resuscitation of livers after 30 minutes of WI to a level comparable to that of nonischemically damaged livers is possible with VSOP supplemented with NO gas. Moreover, the application of VSOP with NO minimizes the extent of injuries caused by oxygen free radicals during preservation. Liver Transpl 18:219–225, 2012. © 2011 AASLD.