The Effects Of Shenlingjianpiwei Granule On TNF-? & Cytb In Non-alcoholic Fatty Liver Mouse Model

Objective:To initiate nonalcoholic fatty liver disease in mice model by high fat diet.Characteristics of serum liver function,blood lipids,inflammatory factors and cytochrome b expression was observed dynamically in NAFLD model mice liver tissue.That NAFLD model mice was treated by Shenlingjianpiwei granule was observed.At same time,inflammatory factors and liver cytochrome b was observed.Methods:1.Animals grouped:Male kunming mouse?150?were divided randomLy into 2 groups,the control group?50?and high fat diet model group?100?.The high fat diet model group was randomLy divided into 4 groups that include high fat diet model group,Slinglingjianpiwei granule low-dose group,middle-dose group and high-dose group on 6^thweekend when mouse model of NAFLD was replicated successfully.2.Observed dynamically points set:Observed dynamically points is 4^thweekend,6^thweekend,10^thweekend and12^thh weekend.Mouse in the control group&the high fat diet group was executed in order to be observed on 4^thweekend,6^thweekend,8^thweekend,10^thweekend&12^thweekend.Mouse in the Shenlingjianpiwei granule low-dose group,middle-dose group&high-dose group was executed in order to be observed on 8^thweekend,10^thweekend&12^thweekend.3.Animal model preparation:The high fat diet consist of lard?10%?,sucrose?15%?,egg yolk powder?15%?,cholesterol?2.5%?,sodium cholate?2.5%?,propylthiouracil?0.2%?,the basal feed?54.8%?.The model of NAFLD mouse model was built by high fat diet.4.Intragastric:After NAFLD model built,give corresponding treatment?7-12 weeks?,with the control group and high fat diet model group was given saline intragastrically(0.1mL·10g^-1·d^-1),and mouse in Shenlingjianpiwei granule low-dose group,middle-dose group&high-dose group were given different concentration of Slingjianpiweigranuleaqueoussolutionbygastric(0.1mL·10g^-1·d^-1).Shenlingjianpiwei granule low-dose group contains Shenlingjianpiwei granule0.13g·mL^-1,Shenlingjianpiwei granule middle-dose group contains Shenlingjianpiwei granule 0.26 g·mL^-1,and Shenlingjianpiwei granule high-dose group contains Shenlingjianpiwei granule 0.52 g·mL^-1.5.Pathological examination:The histopathological changes of liver tissues were observed by hematoxylin-eosin staining.6.Serum biochemical and liver tissue index detection:With automatic biochemical analyzer test in mice serum AST,ALT,TC,TG,LDL-C,HDL-C,ELISA method to detect serum and liver of mice TNF-alpha level,Western blotting test mice liver Cytb relative expression level,fluorescence quantitative PCR method to detect the mice liver Cytb mRNA expression quantity relatively.7.Serum and liver TNF-?levels were detected by ELISA.Western bloting was used to detect Cytb levels in mice.Fluorescence quantitative PCR was used to detect the relative expression of Cytb mRNA in mouse liver.8.Analysis of statistical data by one-way ANOVA.Results:1.Mouse model of NAFLD established and change dynamically:1.1 Mice liver tissue morphology showed that 4^thh weekend,high fat diet model group appeared a few small fat vacuoles,liver cells slight edema,mild disorders,liver can identify,no inflammatory factor.On 6^thh weekend,in high fat diet model group appear a large number of small fat vacuoles,fat cavitation area accounted for a third view area,liver cell cytoplasm slightly lighter color,change a balloon samples,disordered arrangement of cells,distinguish between hepatic cord,nuclei are deepened,appear a small number of inflammatory factors,shows that NAFLD mouse model to replicate success.In 8-12 weekends,fat cavitation increased,liver cell cytoplasm color becomes shallow,disordered arrangement of cells,distinguish between hepatic cord,nuclei are deepened,inflammatory factor increases gradually in high fat diet model group.1.2 Serum biochemical index in mice showed that 4^th,6^th,8^th,10^th&12^thh weekends,compared to cycle,high in fat model group compared with the normal group,the serum AST,ALT,TC,TG,LDL-C,HDL-C rise,statistically significant?P<0.05?.1.3 The results of TNF-alpha in mice showed that compared with the normal group,the serum and liver TNF-alpha were all higher than those in the normal group at4^th,6^th,8^th,10^th&12^thh weekends,with statistically significant differences?P<0.05?.1.4 Mice liver Cytb,according to the results of 4^th,6^th,8^th,10^th&12^thh weekends,compared to cycle,high in fat model group compared with normal group,the liver Cytb and Cytb mRNA relative expression are elevated,statistically significant?P<0.05?.2.The curative effect and part mechanism of shenlingjianpiwei granule:2.1 Rat liver tissue morphology showed that 8^thh at the weekend,and three spleen and stomach granule low-dose group fat cavitation compared with high-fat model group,the number and area of less,occasional inflammatory factor,disordered arrangement of liver cells,distinguish between hepatic cord.The low dose of fat vacuoles in the low-dose group of shenlingjianpiwei granule is small and the area is small,the liver cells are arranged in disorder,and the hepatic cord can not be distinguished.The liver cells were clear,the liver was clear and the inflammatory factors were observed.At the end of 10^thh weekends,the amount and area of the low dose group fat vacuolar and the high-fat model group were reduced,the inflammatory factors were infiltrated,the liver cells were in disorder,and the liver was indistinct.The fat vacuoles in the low-dose group were larger,the inflammatory factors were infiltrated,the liver cells were in disorder,and the liver was indistinct.The fat vacuoles in the high-dose group were observed,the liver cells were clear,the hepatic cord was clear,and a small number of inflammatory factors were infiltrated.At the end of the 12th week,the amount and area of the low-dose group fat vacuolar and the high-fat model group were reduced,the inflammatory factors were infiltrated,the liver cells were in disorder,and the liver was indistinct.The fat vacuoles in the low-dose group were larger,the inflammatory factors were infiltrated,the liver cells were in disorder,and the liver was indistinct.The fat vacuoles in the high-dose group of shenlingjianpiwei granule were observed,the liver cells were loose,the liver was not clear,and the inflammatory factor infiltration was observed.2.2 Serum biochemical index in mice showed that 8^th,10^th,12^thh weekends,compared to cycle,and three spleen and stomach granular particles of low,medium and high dose group compared with high-fat model group,the serum AST,ALT,TC,TG,LDL-C,HDL-C are reduced,statistically significant?P<0.05?2.3 TNF-alpha in mice,according to the results of 8^th,10^th,12^thh weekends,compared to cycle,and three spleen and stomach granule low,medium and high dose group compared with high-fat model group,the serum and liver TNF alpha being reduced,statistically significant?P<0.05?.2.4 Mice liver Cytb,according to the results of the 8^thh weekend,and three spleen and stomach granule low dose group compared with high fat diet model,liver Cytb no statistical difference?P<0.05?,8 at the weekend,and three grains of spleen and stomach,high dose and high fat compared with model group,liver Cytb being reduced,statistically significant?P<0.05?.On the 8th weekend,compared with the high-fat model group,the relative expression of Cytb mRNA was decreased in the low,medium and high-dose group compared with the high fat diet model group,and the statistical difference was statistically significant?P<0.05?.10^thh and 12^thh weekends,compared to cycle,and three spleen and stomach granule low,medium and high dose group compared with high-fat model group,liver Cytb and Cytb m RNA expression relatively amount reduced,the statistically significant?P<0.05?.Conclusion:1.This experimental method replicates the mouse NAFLD model in the 6th weekend,and it was developed from simple fatty liver to fatty liver disease during the 10^th-12^thh weekend.2.TNF-alpha and hepatic Cytb play a role in the formation and development of NAFLD.With the progress of NAFLD,serum and liver TNF-alpha and hepatic Cytb levels rise.3.Slingjianpiwei granules can reduce the inflammatory response of NAFLD and inhibit the development of simple non-alcoholic fatty liver to NASH.4.Slingjianpiwei granules can remove aggregation of lipid,reduce NAFLD inflammatory reaction,suppression and may reverse simple to NASH development of nonalcoholic fatty liver disease,adjust NAFLD mitochondrial oxidative respiration environment,protect the liver cells.