The Experimental Study Of The Treatment Of Non-alcoholic Fatty Liver Disease Through Thioredoxin Reductase Up-regulated

Research background:Non-alcoholic fatty liver is a group of fatty degeneration of liver cells,necrosis,inflammatory cell infiltration,such as the main features of the clinical pathological syndrome,the liver lesions with histology similar to alcoholic liver disease,but no history of excessive alcohol consumption.According to pathological changes can be divided into simple fatty liver,fatty hepatitis,fatty liver fibrosis and may gradually progress to cirrhosis,liver cancer,such as end-stage liver disease,has become second only to chronic viral hepatitis and alcoholic key pre-cirrhotic liver disease.Patients with NAFLD often accompanied by obesity, dyslipidemia,hypertension and abnormal glucose metabolism,such as metabolic syndrome(MS) of the main components,the metabolic syndrome be considered the performance of the liver. Epidemiology shows that,NAFLD prevalence in all parts of the world,there is concern that, with the prevalence of obesity,obese children and adolescents gradually increase,or even a significant portion of patients had neither diabetes nor obesity.This indicates that the non-alcoholic fatty liver disease(NAFLD) to the prevalence rate increased significantly,thereby seriously endangering the health of the people,which,was concidered as an important public health and social problems,but has yet to find an effective treatment.Therefore,explore its pathogenesis,and the search for effective treatment with a very positive social significance.Objective:Non-alcoholic fatty liver disease(NAFLD),Recent study found that the incidence rate of its rise and the gradual onset of age,However,its pathogenesis is not yet clear,is still on the lack of an effective clinical treatment.Lipodystrophy environment so that an imbalance of fatty liver are the basis of hepatic steatosis,and oxidative stress be considered variable development of fatty liver steatohepatitis catalyst.Oxidative stress so that the reaction of the body of a large amount of oxides(ROS),generated more than when the body's antioxidant system to remove the ability or cells function to protect the system,it will cause macromolecular damage,including apoptosis,including pathological changes.Antioxidant substances reduced or lack of inactivation of reactive oxygen species barriers,and enhanced oxidative stress.Oxidant / antioxidant imbalance in the development of NAFLD is an important reason,thioredoxin reductase(TrxR) is a selenium enzyme with an important role in anti-oxidative damage.It has been reported,hepatitis,liver cirrhosis in patients with TrxR activity were lower than normal people.TrxR of high-fat-induced non-alcoholic fatty liver disease(NAFLD) reported a rare study of efficacy.This experiment using high-fat cholesterol diet set up non-alcoholic fatty liver rat model to observe the rat liver TrxR mRNA expression of TrxR in the pathogenesis of NAFLD in the preliminary study of the role,while observing the Jiangzhi Yigan granule and sodium selenite on liver tissue expression of TrxR mRNA explore its possible mechanism of molecular therapy,in order to provide experimental basis for clinical application.Methods:1.Animal Model and Specimen Collection:60 male Wistar rats fed a normal diet after a week,were randomly divided into normal control group(N group):normal diet;the model group(M group),drug treatment group(D group),selenium group(X group):given high fat diet(85%normal diet,13%lard and 2%cholesterol),N group and M group 20 rats in each group,D group and X group 10 rats in each group.The experimental set up 8,13 two weeks time point,N group and M group 10 rats for each time point,groups corresponding to the N1,N2, M1,M2.No.8 in the experimental group over the weekend will be N1 and M1 rats were killed, while D and X group were separately given Jiangzhi Yigan granule 1ml/100g,to 0.8g / ml concentration and the concentration of sodium selenite 4μg/ml dissolved gavage,twice daily,N2 and M2 rats were given drinking water equivalent gavage,twice daily,were continued 5W.After 13 weeks of the experiment all the rats were killed,since the abdominal aortic blood,quickly remove the liver,according to conventional preparation of serum,liver homogenate and liver biopsy,and to retain a small amount of liver tissue cryopreservation in -70℃.2.Detect indicators:measured body weight,body length,wet weight of liver and spleen,and calculate the liver index(liver wet weight / body weight×100%),Lee's index(0.33 weight / body length×1000) and the ratio of liver and spleen(liver wet weight / spleen wet weight) and other physical indicators.Automatic biochemical analyzer determination of alanine aminotransferase(ALT),aspartate aminotransferase(AST),blood lipids and liver tissue lipid content,glucose oxidase enzymatic determination fasting blood glucose(FBG),copper staining method for measuring serum and liver tissue levels of FFA,RIA of serum fasting insulin(FINS) and TNF-αlevels,calculated insulin sensitivity index(ISI=In〔1 /(FINS×FBG)〕);xanthine oxidase enzymatic Determination of liver tissue SOD activity;thiobarbituric acid method for measuring liver tissue MDA content;Determination of NADPH-coupled liver tissue GSH-PX activity,DTNB Determination of TrxR activity;light microscope to assess steatosis and inflammatory activity;reverse transcription polymerase chain reaction(RT-PCR) determination of liver tissue thioredoxin reductase mRNA expression.Results:1.The lobuli hepatis of normal group rats structural integrity,liver cord around the central venous actinomorphous distribution,cells were polygonal,more or less uniform size; various biochemical indices were within the normal range;thioredoxin reductase mRNA expression expression more.2.After 8 weeks,control with the N1 group,model group M1 rats,there is a clear lipid metabolism disorders,accompanied by hyperinsulinemia and insulin resistance;liver tissue had mild to moderate fatty change,and no inflammatory cell infiltration;serum transaminase levels were significantly higher than normal,TNF-αlevels;liver tissue TC,TG,MDA increased,SOD, GSH-PX,TrxR activity decreased;liver TrxR mRNA expression than the control group decreased significantly.(P<0.05).3.After 13 weeks,the degree of insulin resistance of M2 rats than the M1 group no significant progress;Compared to normal group,there are scattered inflammatory cell infiltration and necrosis at the point within the lobuli hepatis,inflammatory activity scoring significantly higher than normal group(P <0.01);M2 Group on the extent of oxidative stress more clearly the progress of M1 Group,serum and liver lipid content,serum TNF-αcontent and transaminase levels were significantly increased;HDL-C were significantly reduced;liver tissue of TC,TG,MDA increased,SOD,GSH-PX,TrxR activity decreased;TrxR mRNA in liver tissue a significant reduction in expression,and compared with the pre-significant difference(P<0.01). The target in rats serum and liver tissue of drug treatment group and selenium group were significantly improved;lipid changes in liver tissue and the degree of inflammatory activity reduced significantly(P<0.05,P<0.01);TrxRmRNA expression in liver tissue was significantly higher.(P<0.01).Conclusion:1.High-fat and high-cholesterol diet can successfully copy the NAFLD rat model;insulin resistance,lipid metabolism disorders,oxidative stress and inflammatory cytokines such as TNF-αdisorders are closely related to the occurrence of NAFLD.2.Thioredoxin reductase can be expressed in normal rat liver,NAFLD rat liver thioredoxin reductase gene expression is lower,and with the model time longer,a significant reduction in expression levels.First demonstration TrxR mRNA expression of NAFLD in animal models are reduced.3.Jiangzhi Yigan granule has a good anti-therapeutic effects for combat NAFLD.(1) can significantly reduce the NAFLD rat serum and liver tissue TC,TG,LDL-C,FFA content, significantly increased HDL-C content,to improve high blood lipids,high liver fat,and thus a good adjust lipid metabolism to reduce liver lipid fatty degeneration of liver cells to alleviate.(2) can increase the expression of TrxR mRNA in liver tissue.(3) be able to intervene the occurrence and development of NAFLD,parts of its role may be associated with raised liver thioredoxin reductase mRNA expression.4 Selenite can effectively control rat nonalcoholic steatohepatitis,may be associated with raised liver thioredoxin reductase mRNA expression of lipid metabolism by regulating the liver against oxidative stress and lipid peroxidation injury,to reduce lipid-related liver,which can provide a new train of thought for early clinical prevention and treatment of NAFLD.