| Background:Nonalcoholic fatty liver disease(NAFLD)is a kind of disease characterized by diffuse hepatocyte bullous steatosis and inflammatory cell infiltration.The prevalence of NAFLD in adults is not less than 30%,but in obesity and diabetes is even as high as 80%.The rise in global prevalence of NAFLD is positively correlated with the incidence rate of obesity,diabetes and metabolic syndrome.Finally,it may develop into cirrhosis and liver cancer,which brings great burden to individuals and society.The liver is an important organ for fat synthesis and storage in the body,and also plays an important role in the transportation and metabolism of fat tissue.Any obstacle will cause excessive accumulation of fat in liver cells and form fatty liver.The excessive increase of adipose tissue promoted the activation of Toll like receptor 4(TLR-4)by saturated fatty acids,Tumor necrosis factor-?(TNF-?)mediates the development of chronic inflammation and participates in a variety of pathological processes through the death receptor signaling pathway.Traf3,a downstream signaling pathway of TNF-?,can cause the formation of autophosphorylation after the ubiquitination of TAK1 and activate NF-?B inflammatory pathway,which plays an important role in the pathogenesis of inflammatory lesions in nonalcoholic fatty liver disease.Lingguizhugan decoction is a commonly used dehumidifier in traditional Chinese medicine,which has the effects of Warming Yang,removing dampness,resolving phlegm and strengthening spleen.Good at treating phlegm syndrome caused by deficiency of Spleen Yang.It is often used to treat those who stop drinking water due to various causes and pathogenesis.On the basis of previous studies,the target of this study is TLR4/TNF-a/NF-?B signaling pathway.We intend to explore the mechanism of Modified Lingguizhugan Decoction regulating the occurrence and development of inflammatory reaction through animal experiments.Objective:The effect and mechanism of Modified Lingguizhugan Decoction on lipid metabolism and inflammatory response of liver cells in rats.Methods:The experimental rats were purchased by Hubei experimental animal research center.The male WINSTAR rats were 8-10 weeks old and weighing 300-350g.Experimental group:the experimental rats were randomly divided into five groups,10 rats in each group,respectively:? control group:normal diet,perfusion of normal saline;?model group:NAFLD induced by high fat diet,perfusion of normal saline;? low dose group:NAFLD induced by high fat diet,perfusion of low dose modified Lingguizhugan Decoction;? middle dose group:NAFLD induced by high fat diet,perfusion of middle dose modified Lingguizhugan Decoction treatment;? high dose group:NAFLD induced by high-fat diet,infusion of high-dose modified Lingguizhugan Decoction treatment,experimental model establishment:to adopt high-fat diet(the energy provided by fat accounts for 60%of the total energy).Modeling and intervention treatment:rats were fed with high-fat diet for 12 weeks to establish obesity model induced by high-fat diet;rats were treated with modified Lingguizhugan Decoction for 8 weeks.After 8 weeks of modeling,blood was collected from carotid artery,tissue samples were taken and tested.Single factor analysis of variance and t-test were used.P<0.05 showed that the difference was statistically significant.Results:(1)The weight of the model group was significantly higher than that of the normal group,with poor spirit,slow response,large amount of drinking water,small amount of food intake,disordered body hair,yellow hair color and mushy stool.The liver is large and round,rough in surface,with a sense of granule when stroked,tight in membrane,hard in texture,yellow brown in color,and yellow spots,which are obviously different from those in the normal group.The body weight,activity and liver texture of each dose group were better than those of the model group.(2)The liver index and serum biochemical index showed that the liver index,serum TC,TG and LDL levels in each dose group were significantly lower than those in the model group(P<0.05).(3)HE staining results of liver tissue:the hepatocytes in model group had obvious pathological characteristics:the structure of liver lobule was destroyed;the arrangement of cells was disordered and the radial arrangement disappeared;a large number of fat drop vacuoles with different sizes and clear boundaries were scattered in the field of vision,necrotic non nuclear hepatocytes were scattered,and at the same time,uneven inflammatory infiltration was seen.Compared with the model group,the treatment group(low,medium and high dose)had different degrees of improvement in the changes of hepatocyte tissue structure,steatosis and inflammatory infiltration.(4)Westen blot:Compared with the normal group,the expression of TRAF-3 and NF-?B(p65/P-p65)in the model group increased in varying degrees,with statistical differences between them(P<0.05);compared with the model group,the above indexes in the treatment group(low,medium and high doses)were significantly reduced,with statistical differences between them(P<0.01).Compared with the normal group,the expression of TLR4 in the model group was significantly increased(P<0.05);in the treatment group,compared with the model group,the expression of TLR4 in the high dose group was significantly reduced,with statistical differences between them(P<0.01);compared with the model group,the expression of TLR4 in the low and middle dose groups was not statistically different(P>0.05).(5)RT-qPCR:In rat liver tissues,the expression levels of lipid metabolism factors(SREBP,Leptin)and inflammatory factors(TNF-?,IL-1?,IL-10 and IL-12)were compared between the model group and the normal group,and the expression levels of the model group were significantly higher than that of the normal group(P<0.05).Treatment group(low,middle and high dose)lipid metabolism in rat liver tissue factor(SREBP,Leptin),inflammatory factors(TNF-?,IL-1?,IL-10,IL-12)expression level with the model group respectively to carry on the contrast between two groups,each dose group were lower than that of model group,in addition to the low dose group of TNF alpha mRNA relative expression of no statistical significance,contrast has significant differences between groups(P<0.05).Conclusion:(1)Modified Lingguizhugan decoction can improve the general condition of rats with nonalcoholic fatty liver disease,regulate the disorder state of lipid metabolism,reduce the state of liver cell injury,fatty degeneration and inflammatory infiltration.It can improve lipid metabolism and protect liver.(2)Modified Lingguizhugan decoction can significantly reduce the expression level of TRAF3,NF-?B(p65/P-p65),TLR4 pathway and related proteins,as well as the expression of genes such as lipid metabolism and inflammatory factor(IL).(3)Modified Lingguizhugan decoction can reduce the fat deposition in the liver of rats,improve the degree of fatty change and inflammatory infiltration of liver cells,reduce fat,anti-inflammatory and protect the liver,showing a good therapeutic effect on non-alcoholic fatty liver disease. |
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