Research On The Effects Of Main Liposoluble Organic Sulfides In Garlic Oil Against Alcoholic Liver Injury Of Mice

ObjectiveAlcoholic liver disease is a series of liver disease caused by large amounts of alcohol intake for a long time,which ranges from alcoholic fatty liver,alcoholic hepatitis,alcoholic fibrosis and alcoholic cirrhosis.ALD has become a worldwide public health problem due to the gradual increase of morbidity and mortality of ALD.Although there has been great progress in the exploration of the mechanisms of ALD,it is still lack of effective preventive and therapeutic measures.Previous studies have clearly demonstrated that the critical roles of oxidative stress in the pathogenesis of ALD.NF-E2 related factors 2(Nrf-2)is most important nuclear transcription factor responsible for the activation of a variety of antioxidant enzymes and phase Ⅱenzymes.Therefore,some natural or synthetic compounds which could effectively active Nfr-2 may be useful for the treatment of ALDGarlic has long been used as a folk medicine.Accumulating evidences from studies in past decades have revealed that garlic-derived organosulfur compounds(OSCs)possess various biological activities.Previous studies have provided some evidences that the OSCs in garlic could suppress the progression of ALD.This study compared the protective effects of 5 major lipid-soluble OSCs in garlic agaist ALD,and to explore the changes of Nrf-2 for the potential mechanisms explorationMethods1.Animal and treatment SPF male ICR mice were divided into 10 groups,i.e.control group,ethanol group,DAS(50mg/kg)/ethanol group,DAS(100m-g/kg)/ethanol group,DADS(50mg/kg)/ethanol group,DADS(100mg/kg)/etha-nol group,DATS(50mg/kg)/ethanol group,DATS(100mg/kg)/ethanol group,AMDS(100mg/kg)/ethanol group,and AMTS(100mg/kg)/ethanol group.Mice in control group were fed with control liquid diet,while the mice in other groups received ethanol(5%,w/v)containing liquid diet.The OSCs were d-issolved in corn oil and were delivered to mice by garvage.The mice i n all groups except the control were gavaged with 5g/kg.bw ethanol at the end of 4weeks of liqiud diet feeding.The mice were sacrificed 4h after e-thanol garvage,and the blood and liver tissues were collected for analysi s.2.Measurement of the serum aminotransferase activities The blood was centrifuged at 3000rpm/min for 15 min for the separation of serum,and the activities of serum ALT and AST were determined using automatic biochemistry analyzer.3.Examination of hepatic triglyceride(TG)level The hepatic TG levels were determined by using the commercial assay kits provided by Beijing applygen Co.and normalized with protein concentrations in the homogenates.4.Histological examination Frozen section were prepared for oil red o staining,while formalin-fixed liver tissues were embedded in paraffin,sectioned,and then stained with hematoxylin and eosin(H&E).5.Examination of hepatic glutathione(GSH)level The hepatic GSH level were determined by the commercial assay kits provided by Beyotime biotechnology Co.and normalized with protein concentrations in the homogenates.6.Determination of the hepatic Nrf-2,peroxisome proliferator-activated receptor a(PPAR-a),and sterol regulatory element binding protein lc(SREBP-lc)The total protein samples were prepared using RIPA lysis buffer,cytoplasmic and nuclear proteins was extracted by using the cytoplasmic and nuclear proteins extraction kits,and western blotting was performed to determine the protein expression of related factors.Results1.Effects of OSCs on the liver weight,bodyweight,and liver index Compared with control group,the body weight of mice in model group did not significantly altered(P>0.05),while the liver weight and liver index were significantly increased(P<0.05);compared with mice in model group,the body weight of OSCs-treated mice was slightly decreased(P<0.05),while the liver weight and liver index were significantly decreased(P<0.05).2.Effects of OSCs on the activities of serum ALT and AST.Compared with the mice in control group,the serum ALT and AST activities were all significantly increased(P<0.05).Compared with the model group,all OSCs-treatment led to a significant decline of the AST activities(P<0.05),while DAS(100mg/kg)and DADS(50mg/kg and 100mg/kg)groups significantly suppressed the elevation of ALT induced by ethanol exposure(P<0.05).3.Effects of OSCs on the hepatic TG levels.Compared with control group,the hepatic TG level in mice of model group was significantly increased(P<0.05).DADS(100mg/kg)and DATS(100mg/kg)significantly suppressed ethanol induced uptake of hepatic TG level(P<0.05).4.Effects of OSCs on hepatic GSH levels.Comparing to control group,the GSH content in liver tissue of mouses in model group reduced obviously(P<0.05).Comparing to model group,the GSH significantly increased in DADS(50mg/kg and 100mg/kg)groups and DATS(50mg/kg and 100mg/kg)groups(P<0.05).5.Effects of DATS and DADS on the protein expression of Nrf-2 Comparing to control group,the level of nuclear Nrf-2 in liver tissue in model group was decreased significantly(P<0.05),and the level of y-GCLc,NQO-1,HO-1 and SOD were significantly decreased(P<0.05).while the protein level of Keap-1 did not change(P>0.05).Comparing to model group,the protein levels of nuclear Nrf-2 in DADS and DATS groups increased significantly(P<0.05),the protein level of Keap-1 in DADS and DATS groups did not change(P>0.05).While protein levels of HO-1,NQO-1,y-GCLC were all increased obviously(P<0.05).6.Effects of DADS on the protein expression of PPAR-a and SREBP-lc in mice liver.Compared with control group,the protein level of PPAR-a and SREBP-lc in model group were significantly decreased(P<0.05),while they were increased in DADS and DATS grouos(P<0.05).Compared to control group,SCD protein level was increased in model group(P<0.05),whicn was not affected by DADS treatment(P>0.05).Comparing to control group,there is no obvious difference of FAS protein level among control group,model group and intervention groups(P>0.05).Compared with control group,ACC protein expression in model group was significantly increased(P<0.05),which was not affected by DADS treatment(P>0.05).Comparing with control group,the protein level of FABP in model group was decreased significantly(P<0.05),which was not affected by DADS intervention(P>0.05).Comparing to control group,the protein level of ACOX-1,CPT-1,P-ACC in model group did not change significantly(P>0.05).Comparing to model group,the protein level of these factors in DADS groups did not change significantly(P>0.05).7.Effects of DADS on the protein level of LC3 and p62 in mice liver.Compared with control group,the protein level of LC3 and p62 were significantly increased in model group(P<0.05).Compared with model group,the protein level of LC3 and p62 in DADS groups were significantly decreased(P<0.05).