Xanthine oxidase released from reperfused hind limbs mediate Kupffer cell activation, neutrophil sequestration and hepatic oxidative stress in rats subjected to tourniquet shock

We have shown previously that rats subjected to tourniquet shock develop an acute form of remote organ injury of the liver that is both Kupffer cell ( KC) and polymorphonuclear (PMN) leukocyte dependent. Circulating plasma xan thine oxidase (XO) has been shown to be responsible for the development of endothelial dysfunction and for remote organ injury of the lung and intesti ne after ischemia-reperfusion protocols. We now hypothesize that XO is rele ased from rat hind limbs upon reperfusion and that it is responsible for KC and PMN leukocyte activation in this shock model. Our results show that ab out 30% of rat gastrocnemius muscle xanthine dehydrogenase (XD) is converte d to XO during the 5-h tourniquet period and that it is released into the f emoral vein within 10 min of reperfusion. Total muscle xanthine oxidoreduct ase activity (XO + XD) decreases within 30 min of reperfusion and is parall eled by a corresponding increase in femoral vein lactic dehydrogenase. In a ddition, liver tissue XO increases significantly within 30 min of reperfusi on without a corresponding conversion of endogenous XD, Conversion of hepat ic XD becomes evident 60 min after reperfusion is initiated, as does XO, an d alanine aminotransferase (ALT) release into the hepatic vein, presumably from damaged hepatocytes as a consequence of oxidative stress. Tissue myelo peroxidase activity also increases significantly after the 60-min reperfusi on period. That XO mediates KC and PMN activation is supported by the follo wing observations: a) the close relationships between plasma XO and the tim e courses of tumor necrosis factor-alpha TNF alpha release into the hepatic vein and colloidal carbon clearance by KCs; b) that colloidal carbon clear ance, TNF alpha and ALT release, loss of tissue free thiols, lipid peroxida tion (TBARS), and liver infiltration by PMN neutrophils can also be induced by the administration of exogenous XO to normal rats; and c) pretreatment of rats with allopurinol inhibits KC activation and liver leukocyte infiltr ation. These results suggest that XO, released from the ischemic limb on re perfusion, is taken up by the liver were it mediates KC and PMN neutrophil activation and thus contributes to the development of multiple system organ failure after hind limb reperfusion.