The Signaling Mechanism Of Resveratrol In Intervening Inflammatory Injury In Nonalcoholic Fatty Liver Of Mice

Non-alcoholic fatty liver disease(NAFLD)is a disease that prominent characterized by fatty degeneration of hepatocytes and liver excessive fat deposition caused by nonalcoholic and other secondary hepatic steatosis factors.In recent years,the incidence of obesity,non insulin dependent diabetes,hyperlipidemia,hypertension and their related metabolic syndromes has increased as people living standard improving and eating habits changing which make quite a number of people in danger of suffering from NAFLD.The main reason is that the number of the cardiovascular events increased significantly in nonalcoholic fatty liver disease patients.So the NAFLD,which does serious threat to human health and social development,is becoming a new a major health problem in China,has become a new challenge in the field of contemporary liver disease.NAFLD is characterized by high levels of non-esterified fatty acids(NEFA),inflammation and hepatic steatosis.Inflammation plays a crucial role in the development of fatty liver.Existing research shows that Resveratrol(RSV)supplement,a stilbene polyphenols phytoalexin,could intervene inflammatory response and hepatic steatosis,whereas,the mechanism by which how to play is not well understand.Inflammation plays an important role in the process of genesis and development in NAFLD.Nuclear transcription factors-kappa B(NF-?B)is closely related to the occurrence of inflammation.The activated NF-?B pathway enhances the transcription of tumor necrosis factor-?(TNF-?),interleukin-6(IL-6)and interleukin-1beta(IL-1?).TNF-?,IL-6 and IL-1? can promote peripheral fat decomposition,hepatic steatosis,inflammation,necrosis and apoptosis,which lead to the occurrence of NAFLD.RSV can activate AMP-activated protein kinase(AMPK)/Sirtuins(SIRT)pathway to reduce the expression of fat synthesis-related genes,reduce lipid production,and improve hepatic steatosis.However,whether RSV activates the AMPK/SIRT1 pathway,thereby inhibiting NF-?B inflammatory pathway,to intervene NAFLD is not clear.In view of the therapeutical effect of RSV on the NAFLD,clearing the inflammatory signal mechanism of RSV intervene NAFLD has important theoretical and practical significance to explore the original drug for the treatment of NAFLD.This research exposited the above problems from three aspects.The first,body level.Mice were fed high fat diets(HFD)to establish NAFLD animal models and added 30 mg/kg/d RSV.The pathological changes of the liver were observed,the levels of NEFA,aspartate aminotransferase(AST),alanine aminotransferase(ALT),alkaline phosphatase(ALP),?-glutamyl transpeptidase(GGT),lactate dehydrogenase(LDH),TNF-?,IL-6 and IL-1? in serum were detected.In addition,we also detected the expressions of key molecules of AMPK/SIRT1 pathway and the key factor of NF-?B inflammatory pathway,and the transcription of inflammatory cytokines such as TNF-?,IL-6 and IL-1? in liver.The results indicated that mice fed with HFD showed severe liver injury,lipid deposition and high concentrations of NEFA,GGT,AST,ALT,ALP and LDH.In addition,feeding HFD can inhibit the activation of AMPK/SIRT1 pathway in the liver,induce NF-?B inflammatory pathway activation,and promote the expression of TNF-?,IL-6 and IL-1?.And feeding with HFD and RSV can activate AMPK/SIRT1 pathway,inhibit the expression of proteins in NF-?B signaling pathway and inflammatory cytokines TNF-?,IL-6 and IL-1?,significantly intervene lipid deposition in liver of mice,improve the liver inflammatory injury.The second,cellular level.NEFA(0,0.6,1.2 and 2.4 m M)were added into primary hepatocytes of mice cultured in vitro to establish cell models.The results showed that NEFA treatment significantly increased the protein expression of p-I?B?/I?B? and p-NF-?B p65/NF-?B p65 and the m RNA transcription of TNF-?,IL-6 and IL-1? in hepatocytes,indicating that NEFA could activate NF-?B inflammatory pathway and promote the transcription of proinflammatory cytokines in primary hepatocytes of mice,and then induce hepatocellular inflammatory injury.The third,gene signal transduction level.Based on different concentrations of NEFA induced hepatocytes inflammatory injury models,RSV(50?M and 100?M)and respective inhibitors of AMPK? and SIRT1(Nicotinamide,Nic and Compound C,Com C)were added into primary hepatocytes of mice cultured in vitro.The results showed that the phosphorylation of AMPK? and the protein expression of SIRT1 in RSV+NEFA group were significantly higher than those in NEFA group,which indicated that RSV could intervene the inhibitory effect of NEFA on AMPK? and SIRT1 expression.In addition,the activation of NF-?B and the m RNA transcriptions of proinflammatory cytokines TNF-?,IL-6 and IL-1? were significantly lower in the RSV+NEFA group than in the NEFA group.And also,the expression of NF-?B and the m RNA transcriptions of proinflammatory cytokines TNF-?,IL-6 and IL-1? were significantly higher in NEFA+RSV+Com C group and NEFA+RSV+Nic group than in the NEFA+RSV group,indicating that inhibition of AMPK? and SIRT1 further reduced the effect of RSV on the overactivation of NF-?B inflammatory pathway induced by NEFA.These data suggested that RSV attenuates NEFA-induced activation of NF-?B inflammatory pathway and proinflammatory cytokines by activating AMPK? / SIRT1 pathway in hepatocytes,thereby intervening the inflammatory damage of hepatocytes induced by high concentrations of NEFA.This research confirmed from three levels that RSV inhibited the activation of NF-?B inflammatory pathway and decreased the expression of proinflammatory cytokines TNF-?,IL-6 and IL-1? by activating AMPK?/SIRT1 pathway,thereby intervening the inflammatory response of the important parts in the occurrence and development in NAFLD.This research laid a theoretical foundation for the exploitation of the clinical application of RSV.