The Regulatory Mechanism Of Insulin/AKT/S6K1 Signaling Pathway In FLHS By Resveratrol

The present experiments in vivo and in vitro were carried out to respectively observe the regulation and protection mechanism of resveratrol to Insulin/AKT/S6K1 signaling pathway in liver of Fatty liver hemorrhagic syndrome layers induced by high-energy-low-protein diet.1)Animal experiment.A total of 288 Hyline brown laying hens were randomly divided into 4 groups with six repeats in each group and 12 birds in each repeat,group ? was control group with birds feeding basic diets(metabolic energy 2678.99 kcal/kg,crude protein 15.86%);group ?was pathologic model group with birds feeding high-energy-low-protein diet(metabolic energy 3100.00 kcal/kg,crude protein 12.00%);group ?and group ? were treatment groups with birds feeding a basic diets and high-energy-low-protein diet containing a resveratrol dosage of 400 mg/kg,respectively.The whole trial period lasted 120 days,and the samples were randomly collected for relevant experiments on 40 days,60 days,80 days,100 days and 120 days.Results showed as follows:(1)Compared with the control group,the concentration of TG and T-Ch in group ?were significantly elevated,whereas the content of HDL-Ch was significantly increased in the early stages and then decreased(P<0.05);Activities of ALT,AST were markedly increased(P<0.05);the liver index,liver fat rate and abdominal fat rate were significantly elevated(P<0.05);HE and oil red stained found that liver cells presented fatty pathological change,with abundant lipid droplet accumulating in cytoplasm and nucleus being extruded to cell edge;the addition of 400 mg/kg resveratrol in groups ? and ?significantly improved the serum lipid metabolism level,reliving liver injury of FLHS caused by high-energy-low-protein diet and easing liver lipid deposition.(2)The concentrations of blood glucose,insulin,HOMA-IR index,IL-6 and TNF-? in group ? were significantly increased compared with birds in control ?(P<0.05);through oral glucose tolerance test(OGTT)and insulin sensitive experiment(IST),it was found that the blood glucose area under curve(AUC)in group ? was significantly elevated compared with other three groups(P<0.05);the addition of 400 mg/kg resveratrol in groups ? and ? diets could significantly decreased the concentrations of blood glucose,insulin,IL-6 and TNF-?,and OGTT and IST found glucose AUC remarkably decreased(P<0.05);(3)P-Tyr of IRS-1 and AKT phosphorylation levels of layers in group ? declined,whereas the S6K1 protein phosphorylation level went up(P<0.05);gene expression levels of insulin receptor(InsR),4EBP-1,Glut-1 and Glut-3 were notably decreased(P<0.05);gene protein expression of TOR?S6K1?FOXO1 were also significantly elevated(P<0.05);addition of 400 mg/kg resveratrol in groups ? and ? could remarkably improve the Insulin/AKT/S6K1 signaling pathway and its downstream gene expression(P<0.05).(4)Comparing laying hens liver metabolites in group ? to group ?,it was found 46 different metabolites,of which 14 kinds of metabolism molecules including fumaric acid,linoleic acid and alanine in group II were increased and other 32 kinds of small metabolism molecules including threonine,methionine and other essential amino acids were decreased;results demonstrated that high-energy-low-protein diet caused liver Krebs cycle and amino acid metabolic blocking in layer;throught electron microscopy observation,jjjjjjjj mitochondrial cristae structure got blurred and swollen,mitochondrial abundance declined,autophagosome in cytoplasm went up,which hints that mitochondrial function of FLHS layers got obstacles further leading to an increased level of autophagy.2)Experiment in vitro.This present experiment used perfusion separation method to extract liver cells in order to respectively investigate the protection mechanism of resveratrol to layer liver cell oxidative stress and fatty degeneration model caused by high glucose and high fatty acid.And results showed as follow:(1)30 mM glucose processing the primary cells of layers significantly decreased T-SOD,CAT and GSH-Px enzymes activity in cell,which caused MDA content remarkably increased(P<0.05);mRNA expression of Nrf-2 was notably decreased,while NF-kB,IL-1? and TNF-? genes expression were significantly elevated(P<0.05).Resveratrol could remarkably upregulate mRNA expression of Nrf-2 and downregulate mRNA expression of NF-kB,IL-1? and TNF-?,further improving liver cell ant oxidative level.(2)1 mM palmitic acid processing layer's primary cells significantly decreased T-SOD enzyme activity,which caused MDA and TG contents remarkably increased in cell.Resveratrol could notably improve antioxidant condition,significantly upregulated mRNA expression of InsR and PPAR-?,and suppress mRNA expression of TOR,S6K1 and Srebp-1c compared with group ?and group ?,respectively.In conclusion,resveratrol could remarkably improve the lipid metabolism disorders,insulin and its signaling transduction obstacles of FLHS layers caused by high-energy-low-protein diet,further reducing the liver damage of FLHS and lipidosis levels.