The Mechanism Of Dai Medicine Pabengban Protects Against Chronic Alcoholic Liver Disease

Background: Chronic excessive alcohol consumption leads to a board spectrum of alcoholic liver diease(ALD),including steatosis,hepatitis,fibrosis,cirhosis,and even hepatocellular carcinoma.Although multiple attempts have been made worldwide to improve patient outcome,we have yet to find a reliable treatment.Pabengban(Gynura Procumbens),a traditional Dai medicine,exhibited protective effects on the acute ethanol-induced liver injury in our previous study.Thus,Pabengban might have positive effect on severe chronic alcoholic steatosis.Aim: To explore the effect of Pabengban on chronic ALD and its molcular mechanism;and to provide new therapeutic target for drug development and enrich the experience of Dai medicine treatment in ALD.Methods: 1.The study of Dai medicine Pabengban on chronic alcoholic liver disease(1)Mouse model of alcoholic fatty liver was established through feeding mice with Lieber-De Carli liquid for 8 weeks.For the last 2 weeks,those mice of treatment groups were treated with Pabengban ethanol extract,60% ethanol eluted fraction(the active fraction of Pabengban),and chlorogenic acid(one of main ingredients of 60% ethanol eluted fraction),respectively.Silymarin,a hepatoprotective drug,was used as a positive control.(2)Histological evaluation of the effects of Pabengban against ethanol-induced lipid accumulation.The body weight and liver index were tested.Mouse livers were subjected to standard H&E and Oil Red O staining.These two staining above were quantified by fat score and multispectral imaging ananlyses,respectively.(3)Serological evaluation of Pabengban against ethanol-induced abnormal liver function.The serum concentration of AST,ALT,TCHO,TG,DLC and HDLC levels were measured with commercially available kits.Mouse serum endotoxin level was measured using an ELISA kit,and intestinal permeability was tested by the concentration of FITC-D4 in serum.Mouse serum ethanol concentration was measured by headspace gas chromatography.(4)Molecular evaluation of Pabengban treatment in chronic alcoholic fatty liver.The m RNA expression of lipid metabolism genes invovled in fatty acid sythensis(SREBP-1c,FASN,ACC and SCD-1),TG-secretion(Apo B and MTTP),fatty acid ?-oxidation(PPAR-?,PGC-1,m Fabp-1,PPAR-?,CPT1 A and CYP2E1A),and inflammation markers(TNF-? and IL-1?)were tested by RT-PCR assay.Moreover,the protein expression of SREBP-1c and its upstream regulators(MAPK and AMPK)and downstream effector(ACC)were detected by Western blot assay.2.The intestine absorption characteristics of chrologenic acid in Gynura procumbens was improved by the other component AFG and chrologenic acid absorption characteristics in ileum at p H 7.8,6.9,and 5.5 was detected by the gravimetric method in rat single-pass intestinal perfusion model,The absorption rate of AFG was the largest at p H 7.8.In addition,AFG absorption characteristics were also detected in duodenum,jejunum,and colon at p H 7.8.Results: 1.Pabengban ethanol extract,60% ethanol eluted fraction,and chrologenic acid reversed ethanol-induced serum abnormal liver fuction and lipid accumulation.Ethanol significantly increased m RNA expression of fatty acid synthesis and oxidation genes and upregulated the activity of SREBP-1c and ACC,and deceased the phosphorylation of MAPK(p44/42 and p38).Pabengban ethanol extract and 60% ethanol eluted fraction protected against ethanol-induced liver steatosis,possibly through ameliorating hepatic lipid accumulation by modulating lipid metabolism-related genes through MAPK/SREBP-1c-dependent pathway.As one of the major active ingredients of 60% ethanol eluted fraction,chlorogenic acid canceled the negative effect of ethanol on ACC expression and activation.However,unlike 60% ethanol eluted fraction,chlorogenic acid did not change the MAPK or SREBP-1c activity in the current study.This result indicated that chlorogenic acid inhibited ACC in a MAPK/SREBP-1c-independent manner.Futhermore,the liver index and the m RNA expression of TNF-? and IL-1? have no significatly change.2.The absorption rate of chrologenic acid was the largest at p H 5.5 in ileum,while the absorption rate of AFG and chlorogenic acid in AFG was the largest at p H 7.8 in ileum.Conclusion: 1.The study of Dai medicine Pabengban on chronic alcoholic liver disease Pabengban protected against ethanol-induced liver steatosis,possibly through ameliorating hepatic lipid accumulation by modulating lipid metabolism-related genes through MAPK/SREBP-1c-dependent and-independent pathways.2.The intestine absorption characteristics of chrologenic acid in Gynura procumbens was improved by the other component Other component in the AFG changed the absorption characteristics of chlorogenic acid in AFG,which conduct a better absorption characteristics in physiological p H in small intestine.This may explain why AFG showed better potential than chlorodenic acid to prevent ethanol-induced liver steatosis.