Contribution of Xanthine Oxidase-Derived Oxygen Free Radicals to the Development of Carbon Tetrachloride-Induced Acute Liver Injury in Rats

Abstract

Summary We examined how oxygen free radicals derived from xanthine oxidase (XOD) contribute to the development of carbon tetrachloride (CCl4)-induced acute liver injury in rats. In rats treated with CCl4 (1 ml/kg), liver injury appeared 3 h after the treatment and developed by 24 h, judged from the serum levels of transaminases. Hepatic and serum XOD activities and serum uric acid concentration did not change 6 h after CCl4 treatment, but they all increased at 12 h and further increased at 24 h. An increase in the concentration of thiobarbituric acid reactive substances (TBARS), an index of lipid peroxidation, and decreases in reduced glutathione concentration and Cu,Zn-superoxide dismutase (SOD) activity, but not Mn-SOD activity, were observed in the liver at b h after CCl4 treatment and these changes were further enhanced at 24h. When allopurinol (50 mg/kg), an XOD inhibitor, was administered to CCl4-treated rats 6 h after CCl4 treatment, liver injury progression was prevented with attenuation of increased hepatic and serum XOD activities, serum uric acid concentration, and hepatic TBARS concentration and decreased hepatic reduced glutathione concentration and Cu,Zn-SOD activity at 24 h after CCl4 treatment. These results indicate that XOD-derived oxygen free radicals contribute to the progression of CCl4-induced acute liver injury in rats both by stimulating hepatic lipid peroxidation and by disrupting hepatic antioxidant defense system.

Attenuation by Oren-gedoku-to Extract (TJ-15) of Disruption of Hepatic Reactive Oxygen Species Metabolism with Progression of Carbon Tetrachloride-Induced Acute Liver Injury in Rats

“…Hepatic XO activity in CCl4-treated rats increased 24 h after the toxicant treatment. These time-related changes observed in the liver of CCl4-treated rats were well consistent with those reported previously [9,13,14]. We have reported that XO-derived ROS contributes to stimulation of hepatic lipid peroxidation and disruption of hepatic antioxidant defense systems at a progressed stage of CCl4-induced acute liver injury in rats [13].…”

“…Rats received a single i.p. injection of CCl4 at a dose of 1 ml/kg body weight as a 50% olive oil solution between 9:00 and 10:00 am, as described previously [9,11,13,14]. The CCl4-untreated rats received an equal volume of olive oil in the same manner.…”

“…This activity was assessed by measuring the increase in absorbance at 292 nm following the formation of uric acid. For this enzyme assay, livers were homogenized in 9 volumes of an ice-cold buffered solution (pH 7.8) containing 0.05 M potassium phosphate, 0.1 M EDTA, 0.5 mM dithiothreitol, 0.5 mg/mL of leupeptin, and 0.2 mM phenylmethylsulfonylfluoride as described previously [13,14]. One unit (U) of XO activity is defined as the amount of enzyme forming 1 µmol uric acid per min.…”

“…It has been shown that hepatic lipid peroxidation increases at a progressed stage of liver injury as well as at an early stage of the injury in rats treated once with carbon tetrachloride (CCl4) [7][8][9][10][11][12][13][14]. It has also been shown in rats with a single CCl4 treatment that hepatic antioxidant defense systems associated with antioxidants such as reduced glutathione (GSH) and ascorbic acid and antioxidant enzymes such as superoxide dismutase (SOD), an enzyme to scavenge O2 ·− , catalase (CAT), an enzyme to decompose hydrogen peroxide (H2O2), and glutathione reductase (GSSG-R), an enzyme to regenerate GSH from oxidized glutathione (GSSG) using NADPH, are disrupted at a progressed stage of liver injury [8,9,[12][13][14][15][16], although the activity of glucose-6-phosphate dehydrogenase (G-6-PDH), an enzyme to generate NADPH, increases at a progressed stage of the injury [9,14,15,17].…”

“…It has also been shown in rats with a single CCl4 treatment that hepatic antioxidant defense systems associated with antioxidants such as reduced glutathione (GSH) and ascorbic acid and antioxidant enzymes such as superoxide dismutase (SOD), an enzyme to scavenge O2 ·− , catalase (CAT), an enzyme to decompose hydrogen peroxide (H2O2), and glutathione reductase (GSSG-R), an enzyme to regenerate GSH from oxidized glutathione (GSSG) using NADPH, are disrupted at a progressed stage of liver injury [8,9,[12][13][14][15][16], although the activity of glucose-6-phosphate dehydrogenase (G-6-PDH), an enzyme to generate NADPH, increases at a progressed stage of the injury [9,14,15,17]. In addition, our previous report has shown that O2 ·− and H2O2 derived from xanthine oxidase (XO) contribute to the progression of CCl4-induced acute liver injury in rats by stimulating lipid peroxidation and by disrupting antioxidant defense systems in the liver tissue [13]. We have reported that a single oral administration of TJ-15 (500 mg/ kg body weight) to rats treated once with CCl4 after the appearance of liver injury prevents progressive liver injury with attenuation of decreased hepatic GSH level and SOD activity and increased hepatic lipid peroxide level [9].…”