High mobility group box 1 protein (HMGB1), a cytokine actively secreted by phagocytes and passively released from necrotic cells, is an inflammatory mediator in experimental hepatic ischemia/reperfusion injury. We characterized its expression in human liver transplantation. In 20 patients, in addition to systemic samples, blood was drawn from portal and hepatic veins during and after reperfusion to assess changes within the graft. Plasma HMGB1, tumor necrosis factor ␣ (TNF-␣), and interleukin-6 (IL-6) levels were measured, and HMGB1 immunohistochemistry was performed on biopsies taken before and after reperfusion. Plasma HMGB1 was undetectable before reperfusion, and levels in systemic circulation peaked after graft reperfusion. At portal declamping, HMGB1 levels were substantially higher in the caval effluent [188 (80-371) ng/mL] than in portal venous blood [0 (0-3) ng/mL, P Ͻ 0.001]. HMGB1 release from the graft continued thereafter. HMGB1 levels were not related to TNF-␣ or IL-6 levels. HMGB1 expression was up-regulated in biopsies taken after reperfusion (P ϭ 0.020), with intense hepatocyte and weak neutrophil staining. HMGB1 levels in hepatic venous blood correlated with graft steatosis (r ϭ 0.497, P ϭ 0.03) and peak postoperative alanine aminotransferase levels (r ϭ 0.588, P ϭ 0.008). Our results indicate that HMGB1 originates from the graft and is a marker of hepatocellular injury in human liver transplantation.