Evaluation Of The Impact Of Advanced Glycation End Products(AGEs) On Inflammation And Glucolipid Metabolism Disorders

AGEs are a group of heterogeneous compounds,produced by the reaction of reducing sugar with proteins,nucleic acids and fats.Food thermal processing contributes to the formation of food flavor and color,but it is also the main way for AGEs formation.Dietary AGEs and their precursors in food can be absorbed by intestine and accumulate in various cells and tissues.Their excessive accumulation may have adverse effects on human health.Many experiments have shown that dietary AGEs intake is positively correlated with inflammatory markers in the circulatory system,suggesting that dietary AGEs can promote inflammation.However,the specific molecular mechanism is still unclear.Dysregulation of metabolism induced by inflammation is considered to be an important factor for metabolic diseases.Therefore,we intended to study the effects of high dietary AGEs intake on metabolism by means of molecular biology and cell biology.And we would analyze its possible molecular mechanism from the perspective of inflammation using adipose and liver tissue as target organs.Methods: 1.Study on inflammation induced by AGEs in RAW264.7 cells: After 24 hours of treatment with AGEs,the toxicity of exogenous AGEs on cells was detected firstly;Then,the effects of exogenous AGEs on oxidative stress and m RNA expression of inflammatory markers were measured by testing kits and q PCR;Finally,the proteins of RAGE?MAPK/NF-?B ? NLRP3 inflammasome were detected by Western blot.2.The effects of pterostilbene on AGEs-induced inflammation in RAW264.7 cells: Firstly,we determined the effects of pterostilbene on oxidative stress and gene expression of inflammatory markers,such as MCP-1 and TNF-?;Then,we determined the activation of RAGE/MAPK/NF-?B signaling pathway after co-treatment with pterostilbene;3.Ten C57BL/6 mice were randomly divided into two groups: normal chow group(Ctrl group)and high dietary AGEs group(Heated group).After 12 weeks feeding,fasting weight,organ index,blood glucose,insulin and triglyceride levels were measured firstly.Then,the distribution of CML and oxidative stress levels in liver,serum,subcutaneous fat,perirenal fat and gland fat of mice were measured.Next,the m RNA expressions of M1/M2 macrophage markers,lipolysis-related enzymes and glucose transporter 4 gene in adipose tissue were detected by q PCR;Furthermore,the proteins of insulin signaling pathway in liver were detected by Western blot and hepatic gene expressions of key enzymes involved in glycometabolism were detected by q PCR,which were verified using Hep G2 cells in vitro;Finally,hepatic transcription in mice was analyzed by RNA-sequence.Results: 1.The contents of ROS and oxidative protein products such as AOPP and protein carbonyls were significantly increased by AGEs treatment compared with BSA treatment.Additionally,the gene expression of inflammatory markers such as IL-1?TNF-??COX-2 and MCP-1,as well as the secretion of inflammatory mediator NO and chemokine MCP-1,were also increased in AGEs-treated cell.2.The proteins of RAGE,NLRP3 and c-caspase-1,as well as the phosphorylation levels of p38,ERK1/2,JNK and p65 proteins were increased in AGEs-treated cells.3.Pterostilbene exerted a beneficial inhibition on oxidative stress and pro-inflammatory gene expression,mainly through blocking RAGE-mediated signaling pathway and interacting with MAPKs and NF-?B 4.The fasting blood glucose,insulin and insulin resistance index of mice fed with high dietary AGEs were significantly higher than those of mice fed with normal chow.And the accumulations of CML in serum,liver and adipose tissue were also significantly increased in high dietary AGEs group.5.High dietary AGEs intake induced an increase in pro-inflammatory genes of M1 macrophage marker(CD80,Nos2 and MCP-1)and a decrease in anti-inflammatory genes of M2 macrophage marker(Arg1,YM1 and Fizz1)in adipose tissue.Furthmore,high dietary AGEs group saw an increase in gene expression of LPL and leptin,and a decrease in Glut4 gene in adipose tissue.6.High dietary AGEs intake induced the decrease of IRS1 protein content and the gene expression of Gc K,while it up-regulated the gene expression of G6 PC in mice hepatic tissues.In vitro,CML treatment increased insulin-induced phosphorylation of IRS1(Ser)and decreased p-AKT protein in Hep G2 cells.Additionally,the gene expression of PEPCK and G6 PC was also up-regulated.7.RNA-sequence analysis showed that the high dietary AGEs intake induced the activation of inflammatory pathway and immune responses in mice liver.Conclusion: 1.Exogenous AGEs treatment can induce the oxidative stress and inflammation in macrophages via RAGE-mediated NF-?B and NLRP3 inflammasome pathway.2.Pterostilbene alleviated AGEs-induced oxidative stress and inflammation via RAGE-mediated MAPK/NF-?B signaling pathway.3.High dietary AGEs can accumulate in mice serum,adipose and hepatic tissues,leading to inflammation and disorders of glucolipid metabolism,and ultimately inducing insulin resistance.