CCl4-induced Acute Liver Injury Research Articles

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

We attempted to elucidate how Oren-gedoku-to (Huanglian-Jie-Du-Tang) extract (TJ-15), a Chinese herbal medicine, attenuates the disruption of hepatic reactive oxygen species metabolism with the progression of carbon tetrachloride (CCl4)-induced acute liver injury in rats. TJ-15 (100, 250 or 500 mg/kg body weight) was orally administered to rats injected with CCl4 (1 ml/kg, i.p.) at 6 h after the toxicant treatment. Post-administered TJ-15 reduced progressive liver injury in CCl4-treated rats at 24 h after the toxicant treatment dose-dependently. The liver of rats treated with CCl4 alone showed increases in the concentration of thiobarbituric acid-reactive substances, an index of lipid peroxidation, and xanthine oxidase activity and decreases in reduced glutathione and ascorbic acid concentrations and superoxide dismutase, catalase, and glutathione reductase activities at 24 h after the toxicant treatment. The liver of CCl4-treated rats showed no change in Se-glutathione peroxidase activity and an increase in hepatic glucose-6-phosphate dehydrogenase activity at 24 h after the toxicant treatment. Post-administered TJ-15 attenuated the increases in hepatic thiobarbituric acid-reactive substances and xanthine oxidase activity and the decreases in hepatic reduced glutathione and ascorbic acid concentrations and superoxide dismutase, catalase, and glutathione reductase activities dose-dependently but did not affect the hepatic Se-glutathione peroxidase activity and the increased hepatic glucose-6-phosphate dehydrogenase activity. These results indicate that orally administered TJ-15 attenuates the disruption of hepatic reactive oxygen species metabolism with the progression of CCl4-induced acute liver injury in rats through its direct and indirect antioxidant actions. The results also suggest that this attenuating effect of TJ-15 could contribute to its preventive effect on the progression of CCl4-induced acute liver injury.

Chemoprotective effect of diallyl trisulfide from garlic against carbon tetrachloride‐induced acute liver injury of rats

Diallyl sulfide (DAS), diallyl disulfide (DADS) and diallyl trisulfide (DATS) are principal constituents of garlic oil. We studied the effect of these sulfides on the phase II drug-metabolizing enzymes, and on the rat model of acute liver injury induced by carbon tetrachloride (CCl4). A highly purified form of each sulfide (more than 99% purity) was administered i.p. to rats at a concentration of 10 or 100 micromol/kg body weight for 14 consecutive days. DATS (10 micromol/kg) and DADS at a 10-fold higher dose (100 micromol/kg) significantly increased the activities of glutathione S-transferase (GST) and quinone reductase (QR); whereas DAS did not. In the CCl4-induced acute liver injury model of rats, DATS (10 micromol/kg) significantly suppressed the increase in plasma lactate dehydrogenase (LDH) and aspartate aminotransferase (AST) activities. In conclusion, hepatic phase II enzymes were induced strongly by the trisulfide and weakly by the disulfide, but not by DAS. DATS significantly reduced the liver injury caused by CCl4. DATS may be one of the important factors in garlic oil that protects our body against the injury caused by radical molecules.

Melatonin prevents disruption of hepatic reactive oxygen species metabolism in rats treated with carbon tetrachloride.

We reported that melatonin prevents the progression of carbon tetrachloride (CCl4)-induced acute liver injury in rats possibly by attenuating enhanced lipid peroxidation and reduced glutathione depletion. Herein, we examined the effect of melatonin on the changes in hepatic reactive oxygen species (ROS) metabolism in rats with a single intraperitoneal injection of CCl4 (1.6 g/kg body weight); the intent was to clarify the therapeutic mechanism of the indoleamine on CCl4-induced acute liver injury. Rats with and without CCl4 treatment received a single oral dose of melatonin (10, 50 or 100 mg/kg body weight) 6 hr after CCl4 treatment. Hepatic concentrations of ascorbic acid (ASC) and vitamin E (VE) and hepatic activities of superoxide dismutase (SOD), catalase (CAT), Se-glutathione peroxidase (Se-GSH-Px), glutathione reductase (GSSG-R), glucose-6-phosphate dehydrogenase (G-6-PDH), and xanthine oxidase (XO) were determined 6 and 24 hr after CCl4 treatment. The liver of CCl4-treated rats showed reductions in ASC concentrations, and SOD activity and an increase in G-6-PDH activity at 6 hr after treatment and further decreases in ACS concentrations and SOD activity and also further increase in G-6-PDH activity in addition to decreases in CAT and GSSG-R activities and increases in VE concentrations and XO activity at 24 hr after treatment. Melatonin attenuated the reductions in hepatic ASC concentrations and SOD, CAT and GSSG-R activities and the increase in hepatic XO activity in a dose-dependent manner without affecting either hepatic Se-GSH-Px activity or the increased hepatic VE concentration and G-6-PDH activity at 24 hr after CCl4 treatment. No dose of melatonin influenced hepatic ACS and VE concentrations and SOD, CAT, Se-GSH-Px, G-6-PDH, and XO activities in CCl4-untreated rats. These results indicate that melatonin postadministered at pharmacological doses prevents the disruption of hepatic ROS metabolism associated with ASC, SOD, CAT, GSSG-R, and XO, in addition to reduced glutathione, in CCl4-treated rats.

Protective role of heme oxygenase-1 induction in carbon tetrachloride-induced hepatotoxicity

Reductive metabolism of carbon tetrachloride (CCl 4) is thought to cause lipid peroxidation which results in hepatic injury. Heme oxygenase-1 (HO-1) (EC 1.14.99.3), the rate-limiting enzyme in heme catabolism, is known to be induced by oxidative stress and to confer protection against oxidative tissue injuries. In this study, we examined the role of HO-1 induction in a rat model of CCl 4-induced acute liver injury. CCl 4 treatment (1 mL/kg, intraperitoneally) produced severe hepatic injury in rats as revealed by significant increases in serum alanine transaminase (ALT) (EC 2.6.1.2) activity and hepatic malondialdehyde (MDA) content, severe liver cell injury, and increases in hepatic tumor necrosis factor-α (TNF-α) mRNA expression and DNA binding activity of nuclear factor-κB (NF-κB). Following CCl 4 treatment, hepatic HO-1 expression was markedly increased both at transcriptional and protein levels in hepatocytes, especially around the central vein. HO-1 induction was mediated in part through a rapid increase in microsomal free heme concentration presumably derived from hepatic cytochrome P450. Inhibition of HO activity by tin-mesoporphyrin (Sn-MP), which resulted in a sustained increase in microsomal free heme concentration, exacerbated liver injury, as judged by the sustained increase in serum ALT activity, extensive hepatocytes injuries, a more pronounced expression of hepatic TNF-α mRNA and an enhanced NF-κB activation. These findings indicate that induction of HO-1 is an adaptive response to CCl 4 treatment, and it may be critical in the recovery of hepatocytes from injury. Our findings also suggest that HO-1 induction may play an important role in conferring protection on hepatocytes from oxidative damage caused by free heme.

Effects of Exogenous Growth Hormone(GH)-Treatment on Carbon Tetrachloride(CCl4)-Induced Acute Liver Injury in GH-Deficient Mini Rats.

The effects of exogenous growth hormone (GH)-treatment on the development and regeneration of CCl4-induced acute hepatic injury were examined in GH-deficient Mini rats. Alanin aminotransferase (ALT) and asparate aminotransferase (AST) activities increased significantly at 24 and 48 hours (h) after CCl4-administration in both GH-treated and non treated groups, and they tended to be suppressed by GH-treatment. The expression level of insulin-like growth factor (IGF)-1 mRNA was higher in GH (+) CCl4 (+) group than in GH (-) CCl4 (+) group, especially at 72 h, and that of hepatocyte growth factor (HGF) mRNA was also higher in some points in the former group. Histologically, degenerative changes of hepatocytes observed at 24 and 48 h was less severe in GH (+) CCl4 (+) group than in GH (-) CCl4 (+) group. At 72 h, depositions of reticular fibers and extracellular matrix materials were less severe and the number of desmin- or α-SMA-positive cells was larger in GH (-) CCl4 (+) group than in GH (+) CCl4 (+) group. Proliferating cell nuclear antigen (PCNA)-labeling index of centrilobular hepatocytes increased at 48 h in both GH (+) CCl4 (+) and GH (-) CCl4 (+) groups, and it returned to the control level at 72 h in the former group while that in the latter group showed still higher level at 72 h. In this study, it was shown that CCl4-induced acute liver injury in Mini rats can be restored by exogenous GH-treatment, and upregulation of IGF-1 and HGF mRNAs may be involved in this phenomenon.

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

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 (1ml/kg), liver injury appeared 3h after the treatment and developed by 24h, judged from the serum levels of transaminases. Hepatic and serum XOD activities and serum uric acid concentration did not change 6h after CCl4 treatment, but they all increased at 12h and further increased at 24h. 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 6h after CCl4 treatment and these changes were further enhanced at 24h. When allopurinol (50mg/kg), an XOD inhibitor, was administered to CCl4-treated rats 6h 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 24h 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.

CCl4-induced acute liver injury in mice is inhibited by hepatocyte growth factor overexpression but stimulated by NK2 overexpression

Hepatocyte growth factor (HGF) inhibits acute liver injury. NK2 acts as an antagonist to HGF in vitro, but its in vivo function has reached no consensus conclusions. We have investigated in vivo effects of HGF and NK2 on CCl4-induced acute liver injury. Elevation of the serum alanine aminotransferase level and extension of centrilobular necrosis were inhibited in HGF transgenic mice but were promoted in NK2 transgenic mice. Hepatocyte proliferation after liver injury was not inhibited in NK2 transgenic mice. Thus, this study indicates that HGF inhibits liver injury, and NK2 antagonizes HGF on liver injury, however, NK2 may not antagonize HGF on hepatocyte proliferation.

Involvement of endogenous CRF in carbon tetrachloride-induced acute liver injury in rats.

Central neuropeptides play important roles in many physiological and pathophysiological regulation mediated through the autonomic nervous system. In regard to the hepatobiliary system, several neuropeptides act in the brain to regulate bile secretion, hepatic blood flow, and hepatic proliferation. Central injection of corticotropin-releasing factor (CRF) aggravates carbon tetrachloride (CCl4)-induced acute liver injury through the sympathetic nervous pathway in rats. However, still nothing is known about a role of endogenous neuropeptides in the brain in hepatic pathophysiological regulations. Involvement of endogenous CRF in the brain in CCl4-induced acute liver injury was investigated by centrally injecting a CRF receptor antagonist in rats. Male fasted Wistar rats were injected with CRF receptor antagonist alpha-helical CRF-(9-41) (0.125-5 microg) intracisternally just before and 6 h after CCl4 (2 ml/kg) administration, and blood samples were obtained before and 24 h after CCl4 injection for measurement of hepatic enzymes. The liver sample was removed 24 h after CCl4 injection, and histological changes were examined. Intracisternal alpha-helical CRF-(9-41) dose dependently (0.25-2 microg) reduced the elevation of alanine aminotransferase and aspartate aminotransferase levels induced by CCl4. Intracisternal alpha-helical CRF-(9-41) reduced CCl4-induced liver histological changes, such as centrilobular necrosis. The effect of central CRF receptor antagonist on CCl4-induced liver injury was abolished by sympathectomy and 6-hydroxydopamine pretreatment but not by hepatic branch vagotomy or atropine pretreatment. These findings suggest the regulatory role of endogenous CRF in the brain in experimental liver injury in rats.

Evaluation of the effectiveness of Rosmarinus officinalis (Lamiaceae) in the alleviation of carbon tetrachloride-induced acute hepatotoxicity in the rat

The effect of oral administration of Rosmarinus officinalis L. (Lamiaceae) on CCl4-induced acute liver injury was investigated. Rats were daily treated with the plant extract at a dose of 200 mg/kg corresponding to 6.04 mg/kg of carnosol as determined by reverse phase high-performance liquid chromatography. The treatment was initiated 1 h after CCl4 administration and Rosmarinus officinalis fully prevented CCl4 effect on hepatic lipid peroxidation after 24 h of CCl4 administration. The increase in bilirubin level and alanine aminotransferase activity in plasma induced by CCl4 was completely normalized by Rosmarinus officinalis. The treatment also produced a significant recovery of CCl4-induced decrease in liver glycogen content. CCl4 did not modify the activity of liver cytosolic glutathione S-transferase (GST) compared with that of control groups. However, Rosmarinus officinalis increased liver cytosolic GST activity and produced an additional increment in plasma GST activity in rats treated with CCl4. Histological evaluation showed that Rosmarinus officinalis partially prevented CCl4-induced inflammation, necrosis and vacuolation. Rosmarinus officinalis might exert a dual effect on CCl4-induced acute liver injury, acting as an antioxidant and improving GST-dependent detoxification systems.

Effect of central urocortin on carbon tetrachloride-induced acute liver injury in rats

The effect of intracisternal injection of urocortin, an endogenous ligand for corticotropin-releasing factor (CRF) 2 receptor, on carbon tetrachloride (CCl 4)-induced acute liver injury was investigated in rats. Intracisternal injection of urocortin dose-dependently enhanced elevation of serum alanine aminotransferase and aspartate aminotransferase levels induced by CCl 4. Intracisternal urocortin also aggravated CCl 4-induced histological changes of the liver. The aggravating effect of central urocortin on CCl 4-induced acute liver injury was abolished by chemical sympathectomy, but not by vagotomy. These data demonstrate that urocortin acts in the brain to exacerbate acute liver injury through the sympathetic nervous system and suggest a possible involvement of the CRF2 receptor in the central CRF-induced exacerbation of acute liver injury in rats.

The roles played by crucial free radicals like lipid free radicals, nitric oxide, and enzymes NOS and NADPH in CCl 4-induced acute liver injury of mice

Mice were administered a single dose of carbon tetrachloride (CCl 4) to induce acute liver injury. We found that lactate dehydrogenase (LDH) and glutamic pyruvic transaminase (GPT) levels in serum, as well as the level of thiobarbituric acid reaction substances (TBARS) in liver homogenate increased significantly in a manner both dose dependent and time dependent after CCl 4 administration. Such results suggest that the liver is susceptible to CCl 4 treatment and that lipid peroxidation is associated with CCl 4-induced liver injury. The spin-trapping electron paramagnetic resonance (EPR) method was used to detect nitric oxide (NO) level in liver. The chemiluminescence method was also employed to measure the NO 2 −/NO 3 − concentration in serum. The NO levels in liver tissues and NO 2 −/NO 3 − concentration in serum were found to decrease significantly both in a dose-dependent manner and in time course after CCl 4 treatment. The nitric oxide synthase (NOS) II activity in the liver, in contrast, was found to increase significantly. Our study suggests that not only should the expression of NOS be analyzed but NO organ and blood concentration must be measured in the study of diseases involving nitric oxide. l-arginine treatment had no significant effect on the liver function of CCl 4-treated mice. It was found that NO donor sodium nitroprusside (SNP; 50 or 100 μg/kg) treatment resulted in decreases of LDH, GPT, and TBARS levels, leading to a protective effect on CCl 4-treated mice. On the other hand, N G-nitro- l-arginine methyl ester ( l-NAME, 100 or 300 mg/kg) treatment caused more severe liver damage. Moreover, we have found in an in vitro EPR study that SNP could scavenge lipid peroxyl radical LOO . The above results together suggest that NO may protect CCl 4-induced liver injury through scavenging lipid radical, inhibiting the lipid peroxidation chain reaction. On the basis of our analysis, we put forth two explanations for the stated discrepancy between NOS II and NO production: (i) NO was used up gradually in terminating lipid peroxidation and (ii) NADPH was depleted (on the basis of correlation evidence only).

Therapeutic effect of melatonin on carbon tetrachloride-induced acute liver injury in rats.

The therapeutic effect of melatonin on acute liver injury was examined in rats intoxicated with carbon tetrachloride (CCl4). Melatonin (10, 50, or 100 mg/kg body weight [BW]) was intraperitoneally administered to male Wistar rats 6 hr after intraperitoneal injection of CCl4 (1.6 g/kg BW) at which time an apparent liver injury occurred. This post-melatonin administration dose dependently prevented the progression of liver injury at 24 hr after CCl4 injection, judging from the levels of serum transaminases, indices of liver cell damage. Rats injected with CCl4 alone showed an increase in liver lipid peroxide (LPO) content and a decrease in liver reduced glutathione content at 6 and 24 hr after the injection. The post-melatonin administration dose dependently ameliorated both changes found at 24 hr after CCl4 injection. Rats injected with CCl4 alone showed an increase in liver triglyceride (TG) content and decreases in serum TG concentration and liver tryptophan 2,3-dioxygenase (TDO) activity, a marker of the inhibition of liver protein synthesis by CCl4, at 6 and 24 hr after the injection, and also a decrease in serum albumin concentration at 24 hr. The changes in serum TG, albumin concentration, liver TG content, and TDO activity found at 24 hr after CCl4 injection were not ameliorated by the post-administration of melatonin. The same administration of melatonin dose dependently reduced liver LPO content in CCl4-untreated rats. These results indicate that melatonin exerts a therapeutic effect on CCl4-induced acute liver injury in rats, possibly through its antioxidant action.

Effect of central corticotropin-releasing factor on carbon tetrachloride-induced acute liver injury in rats.

Central neuropeptides play important roles in many instances of physiological and pathophysiological regulation mediated through the autonomic nervous system. In regard to the hepatobiliary system, several neuropeptides act in the brain to regulate bile secretion, hepatic blood flow, and hepatic proliferation. Stressors and sympathetic nerve activation are reported to exacerbate experimental liver injury. Some stressors are known to stimulate corticotropin-releasing factor (CRF) synthesis in the central nervous system and induce activation of sympathetic nerves in animal models. The effect of intracisternal CRF on carbon tetrachloride (CCl4)-induced acute liver injury was examined in rats. Intracisternal injection of CRF dose dependently enhanced elevation of the serum alanine aminotransferase (ALT) level induced by CCl4. Elevations of serum aspartate aminotransferase, alkaline phosphatase, and total bilirubin levels by CCl4 were also enhanced by intracisternal CRF injection. Intracisternal injection of CRF also aggravated CCl4-induced hepatic histological changes. Intracisternal CRF injection alone did not modify the serum ALT level. Intravenous administration of CRF did not influence CCl4-induced acute liver injury. The aggravating effect of central CRF on CCl4-induced acute liver injury was abolished by denervation of hepatic plexus with phenol and by denervation of noradrenergic fibers with 6-hydroxydopamine treatment but not by hepatic branch vagotomy or atropine treatment. These results suggest that CRF acts in the brain to exacerbate acute liver injury through the sympathetic-noradrenergic pathways.

Preventive effect of melatonin on the progression of carbon tetrachloride-induced acute liver injury in rats.

We examined the preventive effect of melatonin on the progression of carbon tetrachloride (CCl4)-induced acute liver injury in rats. In rats injected with CCl4 (1.0 ml/kg body weight), liver injury appeared 6 h after the injection and progressed at 24 h. This liver injury progression was prevented by treatment with melatonin (50 or 100 mg/kg body weight), which was conducted 6 h after CCl4 injection. An increase in liver lipid peroxide content and a decrease in liver reduced glutathione content occurred with the liver injury progression. These changes were prevented by the post-treatment of melatonin (50 or 100 mg/kg body weight). These results indicate that melatonin can prevent the progression of CCl4-induced acute liver injury through its antioxidant action.

Effect of central urocortin on CCL4-induced acute liver injury in rats

Effect of central urocortin on CCL4-induced acute liver injury in rats

γ-Glutamylcysteinylethyl ester attenuates progression of carbon tetrachloride-induced acute liver injury in mice

We examined the effect of γ-glutamylcysteinylethyl ester ( γ-GCE), which is readily transported into hepatocytes and increases hepatocellular reduced glutathione (GSH) levels, on the progression of carbon tetrachloride (CCl 4)-induced liver injury in mice in comparison with that of GSH. Administration of more than 160 μmol/kg of γ-GCE, but not GSH, to mice at 3 h after intraperitoneal injection of CCl 4 (1 ml/kg) significantly attenuated increases in serum asparatate aminotransferase and alanine aminotransferase activities at 24 h after the CCl 4 injection. Increases in hepatic lipid peroxide (LPO) concentrations and decreases in hepatic GSH concentrations after the CCl 4 injection were significantly diminished by the γ-GCE (160 μmol/kg) administration, but not by the same dose of GSH. γ-GCE, γ-glutamylcysteine, and cysteine acted as substrates for glutathione peroxidases much less efficiently than GSH in the post-mitochondrial fraction of normal mouse liver cells. These results indicate that γ-GCE attenuates the progression of CCl 4-induced acute liver injury in mice through the maintenance of hepatic GSH levels, leading to inhibition of hepatic LPO formation, which could be due to an efficient utilization of GSH converted from γ-GCE in the liver cells.

Continuous HGF Supply from HGF-Expressing Fibroblasts Transplanted into Spleen Prevents CCl4-Induced Acute Liver Injury in Rats

Hepatocyte growth factor (HGF), first identified as a potent mitogen for mature hepatocytes, has been reported to have various activities. We investigated protective effect of continuous HGF supply on carbon tetrachloride (CCl4)-induced acute liver injury in rats. We transfected immortalized but not tumorigenic rat fibroblasts (Rat-1) with an expression plasmid containing the human HGF cDNA and established several cell lines expressing HGF. The biological activity of HGF produced by these cell lines was confirmed by its mitogenic effect on rat hepatocytesin vitro.Either one of the high-HGF-producer cell lines or parental Rat-1 cell line was transplanted into a syngenic rat spleen. Twelve days after transplantation, each rat was intraperitoneally injected with CCl4and sacrificed 48 h after CCl4injection. In rats with continuous HGF supply significantly lower serum glutamic-pyruvic transaminase (GPT) level was observed compared to its marked elevation in control rats and the degree of hepatocyte damage was slight on histological analysis. These results indicate that continuous HGF supply effectively inhibits CCl4-induced acute liver injury and may suggest the possibility that this system would be useful on various liver diseases.

Changes in DNA strand breaks in non-parenchymal cells following hepatocyte regeneration in CCl4-induced rat liver injury.

DNA strand breaks (nicks) in non-parenchymal cells (NPCs) in CCl4-induced acute or chronic liver injury in rats were detected using an in situ nick translation method; their dynamic changes were analysed in relation to the proliferation pattern of hepatocytes and NPCs, as revealed by bromodeoxyuridine (BrdU)-uptake. In acute injury, hepatocyte proliferation started before centrilobular necrosis had occurred, whereas BrdU-labeled sinusoidal NPCs markedly increased only after centrilobular necrosis was apparent. DNA breakages in NPCs paralleled the proliferation pattern of these cells, suggesting that nicks are physiological, and reflect proliferation and activated gene expression. In chronic injury, liver cirrhosis developed after 9 weeks, but BrdU-labeling of hepatocytes was almost the same level as that in untreated liver. The number of BrdU-labeled NPCs showed only a slight increase, while those with DNA breakages were much more frequent in the cirrhotic stage, suggesting a significant role for NPCs in the fibrotic process. These results indicate that DNA strand breaks in NPCs act as a marker for activation states such as proliferation, differentiation and/or activated gene expression.

急性四塩化炭素肝障害ラットにおけるＫＺ‐１０２６の予防作用 組織化学，酵素組織化学並びに超微細構造的検討

The preventive effect of KZ-1026 on acute liver injury induced by CCl4 in rats was evaluated histochemically, enzyme-histochemically and ultrastructurally. Rats that received 50% CCl4 (2 ml/kg) intraperitoneally were sacrificed at 3, 7 and 24 hr. Remarkable reductions in hepatic glycogen, RNA and G-6-Pase activity were observed in the centrilobular area at 3 hr, and ballooned cells appeared in the mid-zone at 7 hr. At 24 hr, the above histochemical parameters in the hepatocytes of the centrilobular area and mid-zone were extensively reduced, while the number of ballooned cells in the mid-zone was increased. KZ-1026 (200 mg/kg) was given orally at 24 and 4 hr before, simultaneously with or 3 hr after CCl4 treatment, and each rat was sacrificed at 24 hr after CCl4 administration. Pretreatment with KZ-1026 24 hr before CCl4 administration prevented reduction of RNA, glycogen and G-6-Phase activity, as well as disruption of rER and proliferation of sER due to CCL4 toxicity. This preventive effect of KZ-1026 was reduced by posttreatment; however, only the decrease in cytoplasmic RNA was well prevented. These results suggested that KZ-1026 is protective against CCl4-induced acute liver injury.