Curative and protective effects of l-arginine on carbon tetrachloride-induced hepatotoxicity in mice

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Abstract

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 (CCl4) induced hepatotoxicity. Mice received a single dose of CCl4. l-arginine treatment was given for 6 days prior or post to CCl4 injection. CCl4-intoxication caused marked liver cell necrosis with inflammatory and apoptotic lesions. l-arginine treatment reduced hepatic necrosis and inflammation. CCl4-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 CCl4 induced hepatotoxicity in mice.

Highlights

l-arginine has protective/curative effects against hepatotoxicity in mice. ► l-arginine decreases hepatic lipid peroxidation induced by CCl4-intoxication. ► l-arginine enhances the activities of hepatic antioxidant enzymes. ► l-arginine improves liver pathological changes induced by CCl4-intoxication.

Introduction

Amino acid l-arginine is a precursor for the synthesis of protein, nitric oxide (NO), creatine, agmatine, and polyamines, and is an intermediate in the detoxification of ammonia. l-arginine has been linked to enhanced immunity, the release of the human growth hormone, greater muscle mass, rapid healing from injury, increased sexual potency, and helping to reverse atherosclerosis [1], [2]. l-arginine may benefit in the treatment of liver disorders such as liver injury, hepatic cirrhosis, and fatty liver degeneration [3].

CCl4 is a well-known hepatotoxin induces oxidative stress and causes liver injury by the formation of free radicals [4]. CCl4 causes liver damage following the cleavage by cytochrome P450 to form trichloromethyl free radical (CCl3). This radical quickly adds molecular oxygen to form the trichloromethyl peroxyl radical (CCl3OO) [5]. Abstraction of hydrogen atoms from unsaturated lipids by such radicals creates carbon-centered lipid radicals [6]. These lipid radicals quickly add molecular oxygen to form lipid peroxyl radicals, thereby initiating the process of lipid peroxidation. Unless scavenged by radical scavengers, these lipid peroxyl radicals in turn abstract hydrogen atoms from other lipid molecules, thereby propagating the process of lipid peroxidation [7]. In a study of plasma amino acid levels after CCl4 induced acute liver damage in rats, at 16 and 24 h after CCl4 treatment, an increase in blood plasma amino acid levels and positive correlations with the dose of CCl4 were observed for most individual amino acids. The only exception was l-arginine which decreased in a dose dependent manner [8].

Cellular protection against free radicals involves an elaborate antioxidant defense system, including several antioxidant enzymes and low molecular weight substances such as glutathione. These biochemical defenses serve to lower the steady state concentrations of free radicals, which might otherwise cause excessive damage to cell components [9]. l-arginine pre-treatment of rats 5 days before and concomitantly with cyclosporine prevented the significant increase in MDA, improved the activity of GPx enzyme and ameliorated the depletion of the GSH content. These findings may indicate a possible protective effect of l-arginine against nephrotoxicity induced by cyclosporine treatment [10]. Depletion in the antioxidant enzymes and thiol status was observed in ethylene glycol-treated rats (hyperoxaluric rats). l-arginine co-supplementation to these rats prevents the retention of calcium oxalate crystals by way of protecting the renal cells from oxidative injury [11]. Pretreatment of ischemic rats with molsidomine and l-arginine markedly attenuated renal dysfunction, morphological alterations, improved the tissue as well as urine NO contents, reduced elevated thiobarbituric acid reactive substances levels and restored the depleted renal antioxidant enzymes [12].

The present study has been conducted to evaluate the possible hepatoprotective and hepatocurative roles of l-arginine administration against CCl4-induced hepatotoxicity in mice through investigation of lipid peroxidation and antioxidant enzymes.

Section snippets

Chemicals

CCl4 and l-arginine were obtained from Sigma chemicals, Saint Louis, MO, USA. All other chemicals and solvents used were of the highest purity grade available.

Maintenance of animals

Forty female Swiss albino mice (25 ± 2 g) obtained from Theodor Bilharz Research Institute, Cairo (Egypt) were used. The animals were housed for about one month prior to experimental use. All mice were housed under constant conditions of a 12-h light/dark cycle in a temperature and humidity controlled room, and were maintained according to

Histopathological analysis

As represented in Table 1, liver sections of control mice showed no pathological changes (0); hepatic cells are arranged in cord separated by widened sinusoid, hepatic cells show low nucleo/cytoplasmic ratio, nuclei are small round and densely stained, and sinusoids are lined by simple flat endothelium (Fig. 1A). Injection with CCl4 caused severe pathological changes (+++) (Table 1); widespread necrosis with acute inflammation and apoptotic changes in liver are present (Fig. 1B). l-arginine at

Discussion

The significant impairment of hepatic GSH status associated with a substantial hepatocellular damage induced by CCl4 suggested the determinant role of hepatic GSH in the development of CCl4 toxicity [24]. Bhadauria et al. observed that, administration of CCl4 caused a significant depletion in hepatic GSH with significantly enhanced hepatic lipid peroxidation in rats [25], [26]. In the present study and in accordance with Hewawasam et al. [27] and with Jiang et al. [28], a significant decrease (P

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