Liliana Zaidel

The hydroxyl radical scavengers dimethylsulfoxide and dimethylthiourea protect rats against thioacetamide-induced fulminant hepatic failure

BACKGROUND/AIMS Reactive oxygen species, proinflammatory cytokines, glutathione depletion and nitric oxide have all been implicated in the pathogenesis of fulminant hepatic failure. The aim of the present study was to examine the respective roles of these factors in the pathogenesis of thioacetamide-induced fulminant hepatic failure in rats. METHODS Fulminant hepatic failure was induced by 3 consecutive intraperitoneal injections of thioacetamide (400 mg/kg) at 24-h intervals. Rats were pretreated with one of the following agents: the free radical scavengers dimethylsulfoxide (4 g/kg every 6 h) or dimethylthiourea (200 mg/kg every 12 h), the glutathione donor, N-acetylcysteine (130 or 200 mg/kg every 6 h), or the anti-tumor necrosis factor-alpha agents pentoxifylline (100 and 200 mg/kg) and soluble tumor necrosis factor receptor (100 or 1000 microg/rat). The nitric oxide synthase inhibitor N-mono-methyl arginine ester (L-NAME, 0.1 mg/ml) was administered in the drinking water, starting 7 days prior to thioacetamide administration. RESULTS Serum levels of liver enzymes, blood ammonia and prothrombin time and the stage of hepatic encephalopathy were significantly improved in rats treated with dimethylsulfoxide or dimethylthiourea compared to the other treatment groups (p<0.001). Liver histology and the survival rate in these rats were not adversely affected by thioacetamide administration (p<0.001), while in all the other treatment groups those parameters were similar to control rats with fulminant hepatic failure. Furthermore, dimethylsulfoxide ameliorated liver damage and improved survival even when its administration was initiated 8 and 16 h after the first thioacetamide injection. The hepatic concentration of methanesulfinic acid, which is produced after direct interaction of dimethylsulfoxide with hydroxyl radicals, was increased five-fold in rats treated with thioacetamide+dimethylsulfoxide (p<0.001), suggesting a role for hydroxyl radical scavenging in the protection from fulminant hepatic failure in this model. In the group of thioacetamide-treated rats that were pretreated with L-NAME, liver enzymes, blood ammonia levels and the mortality rate were higher than in the control group, treated with thioacetamide only. CONCLUSIONS In thioacetamide-induced fulminant hepatic failure, the hydroxyl radical scavengers dimethylsulfoxide and dimethylthiourea prevent liver injury. Neither N-acetylcysteine nor antagonists of tumor necrosis factor-alpha are protective in this rat model. Inhibition of nitric oxide formation aggravates liver damage and reduces the survival of rats with thioacetamide-induced liver damage.

The Ras antagonist, farnesylthiosalicylic acid (FTS), inhibits experimentally-induced liver cirrhosis in rats

BACKGROUND/AIMS Protooncogenes may play an important role, not only in carcinogenesis, but also in the regulation of normal cellular proliferation and differentiation. Several studies have indicated increased expression of the Ras protooncogenes in the liver in animal models and in patients with liver cirrhosis. The aim of the present study was to examine whether a synthetic Ras antagonist, S-farnesylthiosalicylic acid (FTS), which specifically dislodges Ras from the membrane of Ras-transformed fibroblasts (EJ cells), can prevent experimentally-induced liver cirrhosis in rats. METHODS Cirrhosis was induced in male Wistar rats by intraperitoneal administration of thioacetamide (200 mg/kg twice weekly for 12 weeks). The Ras antagonist, farnesylthiosalicylic acid (FTS, 5 mg/kg), was administered during the study period 3 times a week. Ras expression in the liver was determined by Western blot analysis with pan anti-Ras antibodies and by immunohistochemistry. RESULTS Rats treated with thioacetamide and the Ras antagonist, farnesylthiosalicylic acid (FTS), for 12 weeks had lower histopathologic scores of fibrosis and inflammation (p-values of 0.003 and 0.008, respectively) than those treated with thioacetamide only. There were no differences between the histopathologic scores in vehicle (control) and in Ras-antagonist (FTS) only treatments. Analysis of hepatic hydroxyproline levels from the two thioacetamide-treated groups and controls confirmed the histopathologic scores (7.7+/-0.9 mg/g protein in the TAA-treated vs. 3.8+/-0.5 mg/g protein in the TAA+FTS treated group, p = 0.007). Ras levels, determined by Western blot analysis, were markedly increased in the livers treated with TAA (17-fold over control) and significantly decreased (by about 70%) in the livers of rats treated with TAA and FTS. Studies in isolated human hepatic stellate cells demonstrated that FTS inhibited both DNA synthesis and migration of those cells (p<0.05). CONCLUSION These results indicate that inhibition of Ras expression in the liver during fibrogenesis, prevents the development of experimentally-induced hepatic cirrhosis.

Pentoxifylline prevents concanavalin A-induced hepatitis by reducing tumor necrosis factor α levels and inhibiting adhesion of T lymphocytes to extracellular matrix

BACKGROUND/AIMS Concanavalin A activates T lymphocytes and causes T cell-mediated hepatic injury in mice. Tumor necrosis factor alpha is a critical mediator in this experimental model. T-cell-mediated liver injury involves the migration of immune cells, notably CD4+ T lymphocytes, into liver tissue. Pentoxifylline is a strong suppressor of tumor necrosis factor alpha release and prevents leukocyte adherence to vascular endothelium and down-regulates the expression of intercellular adhesion molecule-1 in monocytes. In this study, we examined the efficacy of pentoxifylline as a potential therapeutic compound for the treatment of concanavalin A hepatitis. METHODS Balb/c mice were injected with 12 mg/kg concanavalin A with or without a single injection of pentoxifylline (5-300 mg/kg), 2 h prior to concanavalin A administration. Liver damage was evaluated by determining serum levels of liver enzymes and tumor necrosis factor alpha, and hepatic histopathology compared to mice treated with concanavalin A only. We also assessed the effects of pentoxifylline on the adhesive properties of T lymphocytes to fibronectin, as a paradigm for immune cell-extracellular matrix interactions required for migration. Pretreatment with pentoxifylline significantly reduced serum levels of liver enzymes (3800+/-650 vs 150+/-28 IU/l) and tumor necrosis factor alpha (710+/-105 vs 113+/-15 pg/ml) with no evidence of inflammation in histopathologic examination compared to control mice treated with concanavalin A. Pentoxifylline also inhibited the binding of murine T cells to fibronectin. All the effects of pentoxifylline were dose-dependent. CONCLUSIONS These results indicate that high doses of pentoxifylline can prevent concanavalin A hepatitis by suppression of tumor necrosis factor alpha release and inhibition of T cells adhesion to extracellular matrix.

Hypothyroidism Protects Rat Liver from Acetaminophen Hepatotoxicity

Recent data from animal studies suggest thatinduced hypothyroidism inhibits the development of liverinjury in several animal models, including livercirrhosis and fulminant hepatic failure in rats, and immune-mediated acute liver injury in mice. Theaim of the present study was to determine whetherhypothyroidism would likewise preventacetaminophen-induced hepatic damage in rats. Liverdamage was induced by acetaminophen (2 g/kg) administered bygavage to fasting rats as a single dose. Hypothyroidismwas induced by methimazole, propylthiouracil, orsurgical thyroidectomy and confirmed by elevated serum levels of TSH. Hypothyroidism significantlyinhibited acetaminophen-induced liver damage asmanifested by the decreased serum levels of liverenzymes, malondialdehyde and blood ammonia, as well asby the higher hepatic glutathione content, in allthree groups of hypothyroid rats compared to euthyroidcontrols (P < 0.01). Histopathologic analysis showedsignificantly less liver necrosis and inflammation in the acetaminophen-treated hypothyroid rats.Oxygen extraction, measured in isolated perfused ratliver preparation, was also reduced in the hypothyroidlivers to 42 ± 8% compared to 81 ± 14% ofcontrols (P < 0.01). However, the expression ofCYP2E1 in the livers of hypothyroid rats, as measured bywestern blot analysis, was not decreased compared tocontrol rats. These results suggest that inducedhypothyroidism, regardless of the mode of induction, protectsrat liver from acetaminophen hepatotoxicity. This effectmay be related to hypometabolism of liver cells, but theexact mechanism needs further clarification.