To Explore The Effect Of Zhige Oral Solution On Insulin Resistance In Rats With Alcoholic Liver Disease Based On IRS-1/PI3K/AKT Pathway

Objective:The rat model of alcoholic liver disease was successfully established by alcohol induction,which confirmed the presence of insulin resistance in alcohol-induced alcoholic liver disease rats,and to explore the effect of Zhige oral liquid on insulin resistance in alcoholic liver disease rats and its possible mechanism.Methods:One hundred healthy male SD rats were randomly divided into five groups,each group of 20 rats.Normal group was given distilled water gavage 1.0 m L/100g/d,the rats model group of alcoholic liver disease was given 52% Luzhou Laobaigan 1.0 m L/100g/d,and Zhige oral solution low-dose,medium-dose and high-dose groups were given different doses of Zhige oral solution 0.25 m L/100g/d,0.5 m L/100g/d and 1.0 m L/100g/d on the basis of model group after 6-8 hours.After 12 weeks of Fasting Fasting Blood Glucose(FBG)and Fasting Insulin(FIns)levels were measured by Fasting Insulin ELISA,Fasting Insulin Resistance Index(HOMA-IR)and Insulin sensitivity Index(HOMA-IS)could be calculated according to that.Random execution model group line 3 rats HE dyeing tests confirmed some alcoholic liver damage,continue the treatment for 4 weeks and continual after 16 weeks,the last for 12 hours after fasting drink,take on an empty stomach tail venous blood(ELisa method for detecting FBG,FINS)after lavage 50% Glucose,line of Oral Glucose Tolerance Test sugar(Oral Glucose how Test,OGTT)and calculate the Area under the curve(Area under the curve,AUC)and put to death in the rat,abdominal aorta take blood,detecting FBG,FINS at the end of the time,The insulin resistance of alcoholic liver injury rat model was confirmed.Hepatic tissues were collected for HE staining and immunohistochemistry to observe the hihiopathological changes of rats in each group.Total protein was extracted from liver to detect IRS-1,PI3 K,AKT,GLUT-2 and other IRS-1/PI3K/AKT pathway related protein expression levels by Western blotting.Results:1.Comparison of histopathological changes in the liver of rats in each group(HE staining ×400): The morphology and structure of liver cells in the normal group were orderly and regular,with distinct boundary and radial distribution.The structure of liver cells in the model group was disordered,with obvious enlargement of liver cells and blurred boundaries,and a large number of vesicular fat vacuoles of different sizes.The structure of hepatocytes in the high-dose group was orderly and regular,and most of which in the high-dose group were similar to those in the normal group,with radial arrangement and a few tiny fat vacuoles visible.Compared with the model group,the morphological structure of liver cells in the medium-dose group was improved,but worse than that in the high-dose group.The liver cells were slightly swollen,with scattered fat vacuoles and slightly blurred boundaries.The low-dose group showed obvious hepatocyte enlargement,irregular structure and scattered fat vacuoles of different sizes,which showed no significant improvement compared with the model group..2.Groups of rats with insulin resistance related indicators in comparison: compared with normal group,model group in different stages of the insulin resistance index(FBG,FINS,HOMA-IR)increased significantly,HOMA-IS significantly reduced(P<0.01 or P<0.05),compared with model group,definite ge oral liquid of high,middle dose group of insulin resistance index(FBG,FINS,HOMA-IR)significantly decreased(P<0.05),HOMA-IS significantly higher(P<0.05);According to the calculation of OGTT AUC,compared with the normal group,the OGTT AUC of the model group was significantly increased(P<0.01).Compared with model group,the OGTT AUC of Zhige oral liquid dosage groups were significantly decreased(P<0.01 or P<0.05).3.Groups of rats with protein expression levels related to IRS-1/PI3K/AKT signaling pathway in liver in comparison: Western blotting semi-quantitative analysis of average gray value showed that compared with the normal group,the protein expression levels of IRS-1,PI3 K,P-PI3 K,AKT,P-AKT and GLUT-2 in model group were significantly decreased(P<0.01).Compared with model group,the expression levels of all detected proteins in Zhige oral solution high-dose and medium-dose group were both obviously increased(P<0.01 or P<0.05),but the medium-dose group was slightly worse than that in the high--dose group.Compared with the model group,PI3 K,P-PI3 K and AKT of he low-dose group were improved(P<0.01 or P<0.05),but GLUT-2,IRS-1 and P-AKT were not significantly improved(P>0.05).Immunohistochemical method was used to semi-quantitatively analyze the mean optical density value.Compared with the normal group,the protein expression levels of IRS-1,PI3 K,P-PI3 K,AKT,P-AKT and GLUT-2 in liver tissues with model group were conspicuously decreased(P<0.01).Compared with model group,the protein expression levels of IRS-1,PI3 K,P-PI3 K,AKT,P-AKT and GLUT-2 in liver tissues of Zhige oral solution high-dose,medium-dose and low-dose groups were increased,and the expression levels of IRS-1,PI3 K,P-AKT and GLUT-2 in liver tissues were the most significant in high-dose and medium-dose groups(P<0.01),followed by low-dose group(P<0.05).Conclusion:1.Alcohol can successfully induce alcoholic liver injury in rats,insulin resistance and changes in IRS-1/PI3K/AKT signaling pathway in alcohol-induced alcoholic liver disease rat model;2.Alcohol inhibits the expression of proteins related to IRS-1/PI3K/AKT signaling pathway,Zhige oral liquid can improve insulin resistance in rats with alcoholic liver disease,and the mechanism of action may be closely related to the activation of IRS-1/PI3K/AKT signaling pathway in the regulation of glucose metabolism.