Nitric oxide (NO) is an important bioregulatory molecule that was originally known as the endothelium-derived relaxing factor. Its effects in modulating vascular tone have been well documented. [1][2][3] NO is generally known to inhibit reactive oxygen species (ROS)-mediated reactions, and its protective effects under various conditions are presumably attributable to its ability to detoxify ROS such as superoxide (O 2 Ϫ · ) and hydroxyl radicals (· OH) in hepatic ischemia/reperfusion (I/R) injury.2-4) Immediately after the initiation of reperfusion, endothelial nitric oxide synthase (eNOS) becomes dysfunctional and stops generating NO, this increases ROS production by Kupffer cells and induces neutrophil recruitment. [4][5][6][7] However, the endogenous levels of eNOS during hepatic I/R injury have not been examined under conditions of ROS inhibition. In order to better understand the biological roles of NO, its levels should be measured under conditions of hepatic I/R injury with significant ROS suppression. 8)We previously reported that NO and partial pressure of oxygen (pO 2 ) levels can feasibly be monitored under ischemic conditions in rats with I/R injury of the small bowels by using electrodes selective for NO and O 2 molecules. 9) We have been studying the advanced treatment of hepatic I/R injury. 1,4-Dihydro-2,6-dimethyl-4-(3-nitrophenyl)-3,5-pyridinedicarboxylic acid methyl 6-(5-phenyl-3-pyrazolyloxy) hexyl ester (designated as CV159), which is a 1,4-dihydropyridine derivative, represents a new class of calcium antagonists that function via the selective blockade of Ca 2ϩ /Calmodulin (Ca 2ϩ /CaM).10,11) By performing in vivo and ex vivo electron paramagnetic resonance (EPR) measurements, we found that CV159 retained its organ-reducing activity against radicals in hepatic I/R injury. In ischemic liver injury, CV159 prevents intracellular Ca 2ϩ overloading and may thus effectively minimize organ damage by inhibiting ROSs.12,13) On the other hands, our previous study revealed that activation of high-mobility-group box-1 (HMGB-1) is involved in the immediate proinflammatory stress response to hepatic I/R injury and that treatment with an anti-HMGB-1 antibody significantly improves survival. We proposed that serum HMGB-1 levels can be clinically used as a marker of liver injury, especially in hepatocellular necrosis.14)The present study was designed to investigate the kinetics of HMGB-1 and the effects of CV159 treatment in hepatic I/R injury. We examined the organ-reducing ability of CV159 in rats by using a real-time system for simultaneously monitoring the NO concentrations, the dynamics of the pO 2 , and the concentration of NOx-the final metabolite of NOunder conditions of ROS suppression. Further, we performed Nitric oxide (NO) and the partial pressure of oxygen (pO 2 ) in the liver were simultaneously quantified in rats with partial hepatic ischemia/reperfusion injury (PHIRI). Real-time NO/pO 2 monitoring and immunohistochemical analysis for superoxide dismutase and inducible nitric oxide synthase (i...