Melatonin attenuates hepatic ischemia through mitogen-activated protein kinase signaling

Melatonin exerts a protective effect during hepatic ischemia-reperfusion (I/R) injury through modulation of the apoptotic cell death program. Mitogen-activated protein kinases mediate various intracellular processes such as cell differentiation, survival, and death. This study investigated whether melatonin exerts a protective effect through the activation of Raf-MEK-ERK and its downstream targets, including 90 ribosomal S6 kinase (p90RSK) and Bad, during hepatic I/R damage.

Hepatic ischemia was induced in mice by occlusion of the hepatic artery, portal vein, and bile duct. Adult mice were subjected to 1 h of hepatic ischemia and 3 h of reperfusion. Vehicle or melatonin (10 mg/kg, intraperitoneal) was injected 15 min before ischemia and just before reperfusion. Serum aspartate aminotransferase and alanine aminotransferase levels were measured, and terminal deoxynucleotidyl transferase dUTP nick-end labeling histochemistry was performed. Moreover, Western blot and immunoprecipitation analyses were performed.

Melatonin treatment attenuated hepatic I/R-induced increases in alanine aminotransferase and aspartate aminotransferase levels and also ameliorated hepatic injury-induced pathologic lesions and increases of positive terminal deoxynucleotidyl transferase dUTP nick-end labeling staining in hepatic tissues. Hepatic I/R injury induced decreases in the phosphorylation of Raf-1, MEK1/2, and extracellular-regulated kinase (ERK)1/2, whereas melatonin attenuated decreases in these phosphorylation levels. Moreover, melatonin prevented the injury-induced decreases in phosphorylation of downstream targets, p90RSK and Bad. Immunoprecipitation analysis showed that the interaction between phospho-Bad and 14-3-3 was decreased in vehicle-treated animals, while melatonin prevented this decrease. Melatonin also attenuated the injury-induced increase in cleaved caspase-3. In cultured hepatocytes, melatonin treatment prevented the hydrogen peroxide-induced cell death and decrease in phosphorylation of ERK1/2. Moreover, blocking MEK by PD98059 attenuated the effect of melatonin.

These data suggest that melatonin protects hepatic cells against hepatic I/R damage through the activation of the Raf-MEK-ERK cascade and phosphorylation of its downstream targets.