Antioxidant effects and hepatoprotective activity of 2,5-dihydroxy-4,3'-di(beta-d-glucopyranosyloxy)-trans-stilbene from Morus bombycis Koidzumi roots on CCl4-induced liver damage

We investigated hepatoprotective activity and antioxidant effect of the 2,5-dihydroxy-4,3'-di(beta-D-glucopyranosyloxy)-trans-stilbene that purified from Morus bombycis Koidzumi roots against CCl4-induced liver damage in rats. The 2,5-dihydroxy-4,3'-di(beta-D-glucopyranosyloxy)-trans-stilbene displayed dose-dependent superoxide radical scavenging activity (IC50 = 430.2 microg/ml), as assayed by the electron spin resonance (ESR) spin-trapping technique. The increase in aspartate aminotransferase (AST) activities in serum associated with carbon tetrachloride (CCl4)-induced liver injury was inhibited by 2,5-dihydroxy-4,3'-di(beta-D-glucopyranosyloxy)-trans-stilbene and at a dose of 400 - 600 mg/kg samples had hepatoprotective activity comparable to the standard agent, silymarin. The biochemical assays were confirmed by histological observations showing that the 2,5-dihydroxy-4,3'-di(beta-d-glucopyranosyloxy)-trans-stilbene decreased cell ballooning in response to CCl4 treatment. These results demonstrate that the 2,5-dihydroxy-4,3'-di(beta-D-glucopyranosyloxy)-trans-stilbene is a potent antioxidant with a liver protective action against CCl4-induced hepatotoxicity.     

Effective protection of Terminalia catappa L. leaves from damage induced by carbon tetrachloride in liver mitochondria

The protective effects of chloroform extracts of Terminalia catappa L. leaves (TCCE) on carbon tetrachloride (CCl4)-induced liver damage and the possible mechanisms involved in the protection were investigated in mice. We found that increases in the activity of serum aspartate aminotransferase and alanine aminotransferase and the level of liver lipid peroxidation (2.0-fold, 5.7-fold and 2.8-fold) induced by CCl4 were significantly inhibited by oral pretreatment with 20, 50 or 100 mg/kg of TCCE. Morphological observation further confirmed the hepatoprotective effects of TCCE. In addition, the disruption of mitochondrial membrane potential (14.8%), intramitochondrial Ca2+ overload (2.1-fold) and suppression of mitochondrial Ca2+-ATPase activity (42.0%) in the liver of CCl4-insulted mice were effectively prevented by pretreatment with TCCE. It can be concluded that TCCE have protective activities against liver mitochondrial damage induced by CCl4, which suggests a new mechanism of the hepatoprotective effects of TCCE.     

Hepatoprotective and antioxidant effects of Morus bombycis Koidzumi on CCl4-induced liver damage

The antioxidant activity and liver protective effect of Morus bombycis Koidzumi were investigated. Aqueous extracts of M. bombycis Koidzumi had higher superoxide radical scavenging activity than other types of extracts. The aqueous extract at a dose of 100 mg/kg showed significant hepatoprotective activity when compared with that of a standard agent. The biochemical results were confirmed by histological observations indicating that M. bombycis Koidzumi extract together with CCl(4) treatment decreased ballooning degeneration. The water extract recovered the CCl(4)-induced liver injury and showed antioxidant effects in assays of FeCl(2)-ascorbic acid-induced lipid peroxidation in rats. Based on these results, we suggest that the hepatoprotective effect of the M. bombycis Koidzumi extract is related to its antioxidative activity.     

Hepatoprotective effects of Solanum nigrum Linn extract against CCl(4)-induced oxidative damage in rats

Solanum nigrum L. (SN) is an herbal plant that has been used as hepatoprotective and anti-inflammation agent in Chinese medicine. In this study, the protective effects of water extract of SN (SNE) against liver damage were evaluated in carbon tetrachloride (CCl4)-induced chronic hepatotoxicity in rats. Sprague-Dawley (SD) rats were orally fed with SNE (0.2, 0.5, and 1.0 g kg(-1) bw) along with administration of CCl4 (20% CCl4/corn oil; 0.5 mL kg(-1) bw) for 6 weeks. The results showed that the treatment of SNE significantly lowered the CCl4-induced serum levels of hepatic enzyme markers (GOT, GPT, ALP, and total bilirubin), superoxide and hydroxyl radical. The hepatic content of GSH, and activities and expressions of SOD, GST Al, and GST Mu that were reduced by CCl4 were brought back to control levels by the supplement of SNE. Liver histopathology showed that SNE reduced the incidence of liver lesions including hepatic cells cloudy swelling, lymphocytes infiltration, hepatic necrosis, and fibrous connective tissue proliferation induced by CCl4 in rats. Therefore, the results of this study suggest that SNE could protect liver against the CCl4-induced oxidative damage in rats, and this hepatoprotective effect might be contributed to its modulation on detoxification enzymes and its antioxidant and free radical scavenger effects.     

Protective effects of cassia seed ethanol extract against carbon tetrachloride-induced liver injury in mice

Oxidative stress has been recognized as a critical pathogenetic mechanism for the initiation and the progression of hepatic injury in a variety of liver disorders. Antioxidants, including many natural compounds or extracts, have been used to cope with liver disorders. The present study was designed to investigate the hepatoprotective effects of cassia seed ethanol extract (CSE) in carbon tetrachloride (CCl(4))-induced liver injury in mice. The animals were pre-treated with different doses of CSE (0.5, 1.0, 2.0 g/kg body weight) or distilled water for 5 days, then were injected intraperitoneally with CCl(4) (0.1% in corn oil, v/v, 20 ml/kg body weight), and sacrificed at 16 hours after CCl(4) exposure. The serum aminotransferase activities, histopathological changes, hepatic and mitochondrial antioxidant indexes, and cytochrome P450 2E1 (CYP2E1) activities were examined. Consistent with previous studies, acute CCl(4) administration caused great lesion to the liver, shown by the elevation of the serum aminotransferase activities, mitochondria membrane permeability transition (MPT), and the ballooning degeneration of hepatocytes. However, these adverse effects were all significantly inhibited by CSE pretreatment. CCl(4)-induced decrease of the CYP2E1 activity was dose-dependently inhibited by CSE pretreatment. Furthermore, CSE dramatically decreased the hepatic and mitochondrial malondialdehyde (MDA) levels, increased the hepatic and mitochondrial glutathione (GSH) levels, and restored the activities of superoxide dismutase (SOD), glutathione reductase (GR), and glutathione S-transferase (GST). These results suggested that CSE could protect mice against CCl(4)-induced liver injury via enhancement of the antioxidant capacity.     

Promethazine inhibits the formation of aldehydic products of lipid peroxidation but not covalent binding resulting from the exposure of rat liver fractions to CCl4.

Promethazine is known to have protective activity in relation to CCl4-induced liver necrosis. This hepatoprotective property has been investigated with regard to the free radical scavenging and antioxidant properties of promethazine using isolated hepatocytes and microsomal suspensions. CCl4 is activated in both systems to free radical metabolites that bind covalently to lipid and protein, and initiate lipid peroxidation. A large number of carbonyl products is produced during CCl4-induced lipid peroxidation; promethazine strongly inhibits the production of all classes of carbonyl compounds in both microsomal suspensions and isolated hepatocytes. In contrast, promethazine is a very weak inhibitor of the covalent binding of metabolites of CCl4. We conclude that promethazine acts by scavenging the trichloromethylperoxyl radical and lipid peroxyl radicals, and is a weak scavenger of the trichloromethyl radical. These data, when considered together with the hepatoprotective effects of promethazine, suggest that lipid peroxidation is of relatively more importance than covalent binding in the pathogenesis of CCl4-induced liver necrosis.     

Protective effects of melatonin against carbon tetrachloride hepatotoxicity in rats

Melatonin is an indolamine, mainly secreted by the pineal gland into the blood of mammalian species. The potential for protective effects of melatonin on carbon tetrachloride (CCl(4))-induced acute liver injury in rats was investigated in this work. CCl(4) exerts its toxic effects by generation of free radicals; it was intragastrically administered to male Wistar rats (4 g kg(-1) body weight) at 20 h before the animals were decapitated. Melatonin (15 mg kg(-1) body weight) was administered intraperitoneally three times: 30 min before and at 2 and 4 h after CCl(4) injection. Rats injected with CCl(4) alone showed significant lipid and hydropic dystrophy of the liver, massive necrosis of hepatocytes, marked increases in free and conjugated bilirubin levels, elevation of hepatic enzymes (alanine aminotransferase and aspartate aminotransferase) in plasma, as well as NO accumulation in liver and in blood. Melatonin administered at a pharmacological dose diminished the toxic effects of CCl(4). Thus it decreased both the structural and functional injury of hepatocytes and clearly exerted hepatoprotective effects. Melatonin administration also reduced CCl(4)-induced NO generation. These findings suggest that the effect of melatonin on CCl(4)-induced acute liver injury depends on the antioxidant action of melatonin.     

Protective effect of gossypitrin on carbon tetrachloride-induced in vivo hepatotoxicity

Carbon tetrachloride (CCl4) is a known environmental biohazard, which induces lipid peroxidation (LPO) and oxidative damage in rat liver. In this study, the hepatoprotective effect of Gossypitrin, a flavonoid extracted from Hibiscus elatus S.W, was investigated against the CCl4-induced in vivo hepatotoxicity. The levels of malondialdehyde (MDA) were assayed as an index of LPO and the levels of catalase (CAT) activity as a biomarker of oxidative damage. Leakage of aspartate aminotransferase (ALT) and lactate dehydrogenase (LDH), liver weight/body weight ratio as well as morphological parameters were used as signs of hepatotoxicity. CCl4 (1 ml/kg), intraperitoneally injected into rats, caused increased MDA production and CAT activity, and also a significant ALT and LDH leakage as compared to levels of these constituents in the control group. Changes in morphology, including steatosis, cells forming balloon cells and necrosis were evaluated in the hepatotoxin-induced damage. Treatment of rats with Gossypitrin (3.98, 5.97 and 8.95 mg/kg) 2 h before and 2 h after CCl4 injection, protected hepatocytes against cell injury induced by CCl4 and its efficacy as an antioxidant was similar to vitamin E (used as a reference antioxidant). These results are consistent with the conclusion that the toxicity of CCl4 is due to LPO and the generation of reactive oxygen species (ROS), and that Gossypitrin's protective effects relate to its direct radical scavenging ability and other antioxidative processes induced by its structure.     

Transgene expression of human PON1 Q in mice protected the liver against CCl4-induced injury

BACKGROUND: Oxidative stress, often in association with decreased antioxidant defenses, plays a pathogenetic role in both initiation and progression of liver injuries, leading to almost all clinical and experimental conditions of chronic liver diseases. Human paraoxonase 1 (hPON1) is a liver-synthesized enzyme possessing antioxidant properties. Here, we investigate the effects of transgene-expressed hPON1 Q on alleviating lipid peroxidation and preventing liver injury in a mouse model. METHODS: The hPON1 Q gene was cloned into pcDNA3.0 plasmid and electro-transferred into mouse skeletal muscle. After CCl4 had been administrated to induce liver injury, mice were monitored for serum levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and malonyldialdehyde (MDA). The extent of CCl4-induced liver injury was also analyzed through histopathological observations. RESULTS: After gene delivery, hPON1 mRNA expression was detected in mouse muscle and serum PON1 activity was 1.5 times higher than that of the control counterpart. In the PON1 Q gene transferred mice, protection against CCl4-induced liver injury was reflected by significantly decreased serum ALT, AST and MDA levels compared to those in control mice (P < 0.01). Histological observations also revealed that hepatocyte necrosis, hemorrhage, vacuolar change and hydropic degeneration were apparent in control mice after CCl4 administration. In contrast, the damage was significantly prevented (P < 0.01) in the hPON1 Q transferred mice. CONCLUSIONS: Intramuscular electro-transfer of the hPON1 Q gene led to efficient expression of hPON1 in mice. Elevated levels of PON1, by virtue of its potency to alleviate oxidative stress, could protect mice from suffering CCl4-induced liver damage.     

Protective effect of aminoguanidine, a nitric oxide synthase inhibitor, against carbon tetrachloride induced hepatotoxicity in mice

The present study was undertaken to evaluate the effect of aminoguanidine (AG) on carbon tetrachloride (CCl4)-induced hepatotoxicity. Treatment of mice with CCl4 (20 microl/kg, i.p.) resulted in damage to centrilobular regions of the liver, increase in serum aminotransferase and rise in lipid peroxides level 24 hours after CCl4 administration. Pretreatment of mice with AG (50 mg/kg, i.p.) 30 minutes before CCl4 was found to protect mice from the CCl4-induced hepatic toxicity. This protection was evident from the significant reduction in serum aminotransferase, inhibition of lipid peroxidation and prevention of CCl4-induced hepatic necrosis revealed by histopathology. Aminoguanidine, a relatively specific inhibitor of inducible nitric oxide synthase, did not inhibit the in vitro lipid peroxidation. Taken together, these data suggest a potential role of nitric oxide as an important mediator of CCl4-induced hepatotoxicity.     

Protective effect of L-theanine on carbon tetrachloride-induced acute liver injury in mice

We studied effects of L-theanine, a unique amino acid in tea, on carbon tetrachloride (CCl(4))-induced liver injury in mice. The mice were pre-treated orally with L-theanine (50, 100 or 200 mg/kg) once daily for seven days before CCl(4) (10 ml/kg of 0.2% CCl(4) solution in olive oil) injection. L-theanine dose-dependently suppressed the increase of serum activity of ALT and AST and bilirubin level as well as liver histopathological changes induced by CCl(4) in mice. L-theanine significantly prevented CCl(4)-induced production of lipid peroxidation and decrease of hepatic GSH content and antioxidant enzymes activities. Our further studies demonstrated that L-theanine inhibited metabolic activation of CCl(4) through down-regulating cytochrome P450 2E1 (CYP2E1). As a consequence, L-theanine inhibited oxidative stress-mediated inflammatory response which included the increase of TNF-α and IL-1β in sera, and expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in livers. CCl(4)-induced activation of apoptotic related proteins including caspase-3 and PARP in mouse livers was also prevented by L-theanine treatment. In summary, L-theanine protects mice against CCl(4)-induced acute liver injury through inhibiting metabolic activation of CCl(4) and preventing CCl(4)-induced reduction of anti-oxidant capacity in mouse livers to relieve inflammatory response and hepatocyte apoptosis.     

Hepatoprotective effect of Hibiscus hispidissimus Griffith, ethanolic extract in paracetamol and CCl4 induced hepatotoxicity in Wistar rats

Hibiscus hispidissimus Griff. is used in tribal medicine of Kerala, the southern most state of India, to treat liver diseases. In the present study, the effect of the ethanolic extract of Hibiscus hispidissimus whole plant on paracetamol (PCM)-induced and carbon tetrachloride (CCl4)-induced liver damage in healthy Wistar albino rats was studied. The results showed that significant hepatoprotective effects were obtained against liver damage induced by PCM and CCl4 as evidenced by decreased levels of serum enzymes, glutamate oxaloacetate transaminase (SGOT), glutamate pyruvate transaminase (SGPT), serum alkaline phosphatase (SAKP), serum bilirubin (SB) and an almost normal histological architecture of the liver of the treated groups compared to the toxin controls. The extract also showed significant antilipid peroxidant effects in vitro, besides exhibiting significant activity in quenching 1, 1-diphenyl-2-picryl hydrazyl (DPPH) radical, indicating its potent antioxidant effects.     

Inhibition of carbon tetrachloride-induced lipid peroxidation by novel antioxidants in rat hepatic microsomes: dissociation from hepatoprotective effects in vivo

The ability of two novel antioxidants, U-74,006F and U-78,517G, as well as the known antioxidant N,N'-diphenyl-p-phenylenediamine to inhibit lipid peroxidation induced by carbon tetrachloride (CCl4) was investigated in Aroclor 1254-induced rat hepatic microsomes. All three compounds completely inhibited lipid peroxidation in microsomes as measured by the formation of thiobarbituric acid reactive substances (TBARS). Inhibition of lipid peroxidation was not a function of decreased bioactivation of CCl4, as the compounds did not substantially inhibit benzphetamine N-demethylase activity or covalent binding of [14-C]CCl4 to lipid or protein. Parallel studies examined the hepatoprotective effects of the compounds in vivo. Rats were pretreated with antioxidant or vehicle prior to administration of CCl4 (300 or 600 microL/kg i.p.). Sera were collected 24 h postadministration of CCl4 and analyzed for alanine aminotransferase (ALT) and alkaline phosphatase (ALP) activities and total bilirubin. Administration of CCl4 produced elevations in ALT, moderate changes in bilirubin, and no change in ALP activities. Histological examination of CCl4-treated livers revealed lipidosis and centrilobular necrosis. The antioxidants partially improved the clinical chemistry parameters, but had minimal effects on the histological lesion. In contrast to the complete inhibition of lipid peroxidation observed in the in vitro studies, none of the antioxidants markedly protected against CCl4-induced toxicity in vivo.     

Antioxidant and hepatoprotective activity of tubers of Momordica tuberosa Cogn. against CCl4 induced liver injury in rats

Hydro alcoholic extract of tubers of M. tuberosa was subjected to preliminary phytochemical screening and evaluated for in vitro and in vivo antioxidant and hepatoprotective activity against CCl4 induced liver damage in rats. Pretreatment with 70% ethanolic extract of M. tuberosa reversed CCl4 induced elevation of levels of serum biomarkers to near normal levels, suggesting that the tubers of M. tuberosa possess hepatoprotective property and this property may be attributed to the antioxidant property of the plant.     

Protection against CCl4-induced injury in liver by adenovirally introduced thioredoxin gene

Antioxidation therapy is a promising strategy for treating or preventing oxidative stress-related liver diseases. The human thioredoxin (TRX) gene was inserted into an adenovirus vector (Adv-TRX), which was administered to mice. The mice were treated with 1 ml/kg CCl4 48 h after the infection. Blood samples were taken and the liver was excised 24 h after the CCl4 treatment. Serum ammonia, aspartate aminotransferase (AST), and alanine aminotransferase (ALT) levels were determined, and liver sections were stained with hematoxylin and eosin. RT-PCR analysis showed that the introduced TRX gene was expressed only in the liver. Adv-TRX decreased the serum ammonia, AST, and ALT levels. Hematoxylin-eosin staining indicated that the CCl4-induced injury was significantly prevented by the Adv-TRX infection. The gene delivery of TRX, which plays a central role in intracellular redox control, was shown to be effective in protecting the liver against oxidative stress-induced injury.     

Suppression of acute hepatic injury by a synthetic prostacyclin agonist through hepatocyte growth factor expression

Previous studies have demonstrated that mice disrupted with the cyclooxygenase-2 gene showed much more severe liver damage compared with wild-type mice after liver injury, and prostaglandins (PGs) such as PGE(1/2) and PGI(2) have decreased hepatic injury, but the mechanisms by which prostaglandins exhibit protective action on the liver have yet to be addressed. In the present study, we investigated the mechanism of the protective action of PGI(2) using the synthetic IP receptor agonist ONO-1301. In primary cultures of hepatocytes and nonparenchymal liver cells, ONO-1301 did not show protective action directly on hepatocytes, whereas it stimulated expression of hepatocyte growth factor (HGF) in nonparenchymal liver cells. In mice, peroral administration of ONO-1301 increased hepatic gene expression and protein levels of HGF. Injections of CCl4 induced acute liver injury in mice, but the onset of acute liver injury was strongly suppressed by administration of ONO-1301. The increases in serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) by CCl4 were suppressed by 10 mg/kg ONO-1301 to 39.4 and 33.6%, respectively. When neutralizing antibody against HGF was administered with ONO-1301 and CCl4, the decreases by ONO-1301 in serum ALT and AST, apoptotic liver cells, and expansion of necrotic areas in liver tissue were strongly reversed by neutralization of endogenous HGF. These results indicate that ONO-1301 increases expression of HGF and that hepatoprotective action of ONO-1301 in CCl4-induced liver injury may be attributable to its activity to induce expression of HGF, at least in part. The potential for involvement of HGF-Met-mediated signaling in the hepatotrophic action of endogenous prostaglandins generated by injury-dependent cyclooxygenase-2 induction is considerable.     

Identification of protein kinase C inhibitory activity associated with a polypeptide isolated from a phage display system with homology to PCM-1, the pericentriolar material-1 protein

L-arginine may aid in the liver detoxification and may benefit in the treatment of liver disorders such as liver injury. The present study was to investigate the possible protective and curative effects of L-arginine on carbon tetrachloride (CCl(4)) induced hepatotoxicity. Mice received a single dose of CCl(4). L-arginine treatment was given for 6 days prior or post to CCl(4) injection. CCl(4)-intoxication caused marked liver cell necrosis with inflammatory and apoptotic lesions. L-arginine treatment reduced hepatic necrosis and inflammation. CCl(4)-intoxication also enhanced hepatic lipid peroxidation, decreased hepatic GSH level and inhibited the activities of antioxidant enzymes. Pre-treatment and post-treatment with L-arginine decreased lipid peroxidation and restored the antioxidant status to near normal levels. These results suggest that L-arginine administration has hepatoprotective and hepatocurative effects against CCl(4) induced hepatotoxicity in mice.     

Effects of melatonin on carbon tetrachloride-induced changes in rat serum

Carbon tetrachloride (CCl4) is a volatile organic chemical, which causes tissue damage, especially to the liver and kidney. In experimental animals it has been shown to be carcinogenic. This study was designed to evaluate the effects of exogenous melatonin administration on the CCl4-induced changes of some biochemical parameters in rat blood. Twenty-four male Wistar rats were randomly divided into three equal groups: Control, CCl4 and CCl4 plus melatonin (CCl4+MEL). Rats in CCl4 group were injected subcutaneously with CCl4 0.5 ml/kg in olive oil while rats in CCl4+MEL group were injected with CCl4 (0.5 ml/kg) plus melatonin (25 mg/kg in 10% ethanol) every other day for one month. Control rats were treated with olive oil. Serum urea, creatinine, total protein, albumin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), total and conjugated bilirubin, alkaline phosphatase (ALP), gamma-glutamyl transferase (gamma-GT), total iron, and magnesium levels were determined. Serum AST, ALT, total and conjugated bilirubin, ALP, gamma-GT, and total iron levels were significantly higher in CCl4-treated rats than in the controls, while urea, total protein, and albumin levels were significantly lower. Melatonin treatment did not cause a significantly change in serum urea, total protein, and albumin levels. However, the elevations in AST, ALT, total and conjugated bilirubin, ALP, gamma-GT, and total iron levels induced by CCl4 injections were significantly reduced by melatonin. On the other hand, melatonin administration significantly decreased serum magnesium levels. These results indicate that melatonin could be a protective agent against the CCl4 toxicity in rats, most likely through its antioxidant and free radical scavenger effects.     

Selective protection of curcumin against carbon tetrachloride-induced inactivation of hepatic cytochrome P450 isozymes in rats

We investigated the effects of curcumin, a major antioxidant constituent of turmeric, on hepatic cytochrome P450 (CYP) activity in rats. Wistar rats received curcumin-containing diets (0.05, 0.5 and 5 g/kg diet) with or without injection of carbon tetrachloride (CCl(4)). The hepatic CYP content and activities of six CYP isozymes remained unchanged by curcumin treatment, except for the group treated with the extremely high dose (5 g/kg). This suggested that daily dose of curcumin does not cause CYP-mediated interaction with co-administered drugs. Chronic CCl(4) injection drastically decreased CYP activity, especially CYP2E1 activity, which is involved in the bioactivation of CCl(4), thereby producing reactive free radicals. Treatment with curcumin at 0.5 g/kg alleviated the CCl(4)-induced inactivation of CYPs 1A, 2B, 2C and 3A isozymes, except for CYP2E1. The lack of effect of curcumin on CYP2E1 damage might be related to suicidal radical production by CYP2E1 on the same enzyme. It is speculated that curcumin inhibited CCl(4)-induced secondary hepatic CYPs damage through its antioxidant properties. Our results demonstrated that CYP isozyme inactivation in rat liver caused by CCl(4) was inhibited by curcumin. Dietary intake of curcumin may protect against CCl(4)-induced hepatic CYP inactivation via its antioxidant properties, without inducing hepatic CYPs.