Protective Effect of Melatonin on Carbon Tetrachloride - induced Hepatic Fibrogenesis in Rats

Background: Liver cirrhosis is a critical stage of chronic liver diseases that can
produce liver failure, portal hypertension and hepatic carcinoma. Sustained oxidative
stress plays a key role in cell damage and fibrosis induced during liver cirrhosis. Aim
of the work: The aim of the present study was to examine the potential protective
effect of exogenous melatonin co-treatment on liver tissue injury and oxidative stress
processes during induction of early phase of liver fibrosis by carbon tetrachloride
(CCl4) injection in rats. Methods: Hepatic fibrogenesis model was induced in this
study by subcutaneous injection of rats by carbon tetrachloride (CCl4). Eighteen
adult, female albino rats were randomly divided into 3 groups (n = 6): control group
(group I), carbon tetrachloride treated group (group II) and CCl4 + melatonin cotreated
group (group III). Rats in CCl4 treated group were injected subcutaneously
with sterile CCl4 (2 ml/kg of body weight) in a ratio of 1:1 with olive oil twice a week
for 8 weeks. Rats of group III (melatonin co-treated group) were injected with CCl4 in
the same manner as in group II and received intraperitoneal melatonin injection in a
dose of 20 mg/kg twice a week for 8 weeks, starting from the beginning of CCl4
injection. Rats in normal control group were injected subcutaneously with olive oil at
the same dose and frequency as those in CCl4 treated group. At the end of the
experiment, rats were sacrificed, blood samples were collected for biochemical assay.
Liver from each animal was removed for histopathological examination.
Measurement of oxidative stress markers in serum was done by chemical estimation
of serum levels of free radicals: lipid peroxides (LPO) and nitric oxide (NO).
Antioxidant enzymes were estimated by chemical measurement of glutathione
peroxidase (GSH-Px) and superoxide dismutase (SOD) in the serum. Liver injury was
assessed by evaluation of serum levels of liver enzymes (alanine aminotransferase
(ALT), and aspartate aminotransferase (AST)). Determination of development of
early phase of hepatic fibrogenesis was done by chemical measurement of serum level
of hyaluronic acid (HA) using enzyme immunoassay (ELISA), and by
histopathological examinations of hepatic tissues to detect early fibrotic changes as
well as other histological damage of hepatic tissue caused by CCl4 injection with or
without melatonin administration. Results: Results of the present study showed that
CCl4 treatment to rats of group II caused highly significant increase in serum levels
of oxidative stress markers (lipid peroxides and nitric oxide), decrease in serum levels
of antioxidant markers (glutathione peroxidase and superoxide dismutase), increase
of serum levels of hepatic enzymes (ALT and AST) as well as increased serum level of
hyaluronic acid (HA) 8 weeks after CCl4 injections when compared with control
group. Melatonin co-treatment to animals of group III caused significant reduction in 
serum levels of lipid peroxides (LPS) and nitric oxide (NO), significant increase in
plasma levels of glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD),
significant reduction in serum levels of liver enzymes (ALT and AST) as well as
significant decrease in serum level of hyaluronic acid (HA) 8 weeks after CCl4
injections when compared with group II. Histopathological study of liver tissue of
animals of CCl4 treated group showed various manifestation of hepatic cell damage
and early phase of fibrogenesis as necrosis, degeneration, collagen deposition and
few fibrous threads extending into the hepatic lobules. Histopathological study of
hepatic tissue of melatonin co-treated group showed that melatonin caused marked
amelioration of histological manifestations of hepatic cell degeneration and absence
of any sign of fibrogenesis with nearly normalization of the histological appearance
of the hepatic tissue. Compared with CCl4 treated group (group II), histological
appearance of hepatic tissue of rats in melatonin co-treated group (group III) showed
significant improvement. Conclusion: Results of this study suggest that melatonin has
a substantial hepatoprotective effect in a rat hepatic fibrosis model induced by an 8-
weeks’ CCl4 regimen. The protective effect of melatonin may be due to both its direct
radical scavenging properties and indirect effect as a regulator of antioxidant
systems. Therefore, the study proposes that melatonin may be a valuable drug for
inhibition of unwanted fibrosis in patients exposed to different hepatotoxic agents.