Melatonin Hepatoprotective Effects During Ischemia/Reperfusion Proceed In Hepatic Cirrhosis Mice

Objective:The aim of the study was to verify the hepatoprotective effects of melatonin (MT) during the ischemia/reperfusion (I/R) procedure in hepatic cirrhosis mice, to evaluate the effects of MT to survival rate, hepatic function, apoptosis rate, oxidative stress and inflammatory cytokines expression, to probe the potential mechanism of these effects.Background:Study shows that more than50%primary hepatic carcinoma patients were complicated with cirrhosis. Liver function reserve with cirrhosis was not enough for liver resection. And cirrhosis patients were more sensitive than normal liver in ischemia/reperfusion (I/R) injury. The liver failure rate and death rate after hepatic operation was higher than uncirrhosis tumor patients. In the ischemia reperfusion procedure, reactive oxygen species (ROS)-the metabolite of Association Tennis Professional (ATP)-stimulate the expression of nuclear factor-KB (NF-KB) to induce the apoptosis of hepatic cell directly and indirectly. We hypothesised that it could alleviated the hepatic injury if the ROS were scavenged. MT was isolated from pineal and also secreted from other organs such as liver. MT could regulate the body’s rhythmic physiological activities. In recent years, the antioxidant capacity of MT was concerned and studied. Melatonin was a powerful antioxidant with some advantage such as smaller molecular weight, higher lipid solubility, wider spreading, smaller toxicity and higher safety. Melatonin (MT) could scavenge ROS directly and had hepatoprotective potential in hepatic ischemia/reperfusion injury in uncirrhosis mouse. But it existed certain inflammatory molecules expression in liver cirrhosis. The blood distribution and hepatocellular metabolism in liver cirrhosis were also different from normal liver. And the effect of MT in cirrhosis mice was still unclear. The subject of this study was to investigate the effect of MT during the procedure of I/R in cirrhosis mouse, and study the specific mechanism of the effect.Methods:100BAL/BC mice were randomly divided into2groups, one group was gavaged with20%carbon tetrachloride (CCL4) for4weeks (0.5ml/kg,2/w) to induced liver cirrhosis. The surgical procedure:the vascular clamp was taken off after selective hepatic vascular exclusion for45min with clamp to reperfusion. Mice were sacrificed by taken blood from apex at different time30min,1h,3h,6h to collect hepatic tissue specimen to test oxidative stress indicators:Methane Dicarboxylic Aldehyde(MDA),Myeloperoxidase(MPO), Superoxide dismutase (SOD),and inflammatory indicators:NF-KB, Tumor Necrosis Factor a (TNF-a), interleukin-6(IL-6);apoptosis indicators:Anoxin v Prodium Iodide(PI) and fluorescein isothiocyanate(FITC), and collect serum sample to test liver function indicaters:glutamic-pyruvic transaminase(ALT), glutamic oxalacetic transaminase(AST), lactate dehydrogenase(LDH) to evaluate the liver function after reperfusion. Immunohistochemical(IHC) method to test the pathological change.Results:Paraffin section sirius red stain shows that collagenous fiber increase obviously, and more pseudolobule in the cirrhosis liver group. Hydroxyproline content was significant higher in CCL4gavaged group. The survival rate of normal group, cirrhosis MT pretreatment group and cirrhosis control group was100%,87.5%and62.5%. Paraffin section HE stain result shows that hepatic tissue necrosis and degeneration phenomenon was alleviated in MT pretreatment group. The liver function indexs ALT, AST,LDH and inflamation indexs NF-KB, TNF-a, IL-6were all lower in MT pretreatment groups. The content of mRNA of NF-KB, TNF-a, IL-6was lower significantly in MT pretreatment group than control group. Hepatocyte apoptosis rate was lower in MT pretreatment group ether. But IHC result of proliferating cell nuclear antigen(PCNA)shows nonsignificant hepatocyte proliferation in MT pretreatment group.Conclusions:The study suggests that MT has hepatoprotection in procedure of ischemia reperfusion. The potential mechanism of alleviated I/R injury in cirrhosis mouse were to scavenging reactive oxygen species (ROS) and down-regulating the synthesis of NF-KB,TNF-a and IL-6.But MT could not promote the proliferation of hepatocyte post I/R obviously.