AMPK Activates UCP1 To Inhibit Inflammation Of White Adipose Tissue And Attenuates Atherosclerosis

Objective:To explore the role and mechanism of AMP-activated protein kinase activation of Uncoupling Protein 1 to regulate the beige coloration of white adipose tissue aroundblood vessels,thereby inhibiting inflammation of white adipose tissue around the tube and reducing Atherosclerosis.Methods:Two experimental methods,in vivo and in vitro,were used to comprehensively study the mechanism of AMPK activated UCP-1 to inhibitinflammation of PVAT and reduce atherosclerosis.（1）Apo E knockout(Apo E^-/-)mice were fed with atherosclerosis model in vivo,and different AMPK agonists（A769662,Metformin）were given for 12 weeks.Body weight was recorded weekly,and food was recorded every3 days.The amount of blood was taken at the tail tip for an oral glucose tolerance（OGTT）test at 12 weeks.The blood of the orbit was used to determine the blood lipid level ofApo E^-/-mice;Paraffin sections and HE staining were used for athological analysis;Western Blot,RT-q PCR were used to detect the expression of white fat beige genes and proteins;in vitro vascular tone measurement system was used to detect vascular endothelial function and smooth muscle function;The number and size of arterial plaques were observed by oil red O staining.（2）In vitro induction of 3T3-L1 preadipocytes with different AMPKagonists（A769662,Metformin）.After stimulation with AMPK inhibitors,Western Blot was used to detect beige protein,and adipocytes and macrophages after inductionadministration were used.The cells were co-cultured to detect the levels of inflammation and related factors.Results:（1）The weight of Apo E^-/-mice changed with time.Compared with control group,the weight of model group mice increased significantly（P<0.01）,compared with model group,the weight of AMPK group mice decreased significantly（P<0.01）.There was no difference in food intake between groups;（2）The weight of fat in Apo E^-/-mice was changed.Compared with the control group,the weight of liver（P<0.05）,epididymal fat（white fat）（P<0.01）and scapular fat（brown fat）in the model group were increased,while the weight of liver,epididymal fat and scapular fat in the AMPK agonist group were decreased（P<0.05）and increased compared with the model group In addition（P<0.05）;（3）Improve the glucose tolerance of Apo E^-/-mice,the fasting blood glucose level of model mice was significantly higher than that of control mice（P<0.01）.OGTTexperimental results showed that the high-fat feeding of Apo E^-/-mice caused the damage of islet function and the abnormality of glucose tolerance.After the administration ofA769662 and Metformin,the phenomenon of glucose tolerance and insulin resistance could be improved;（4）Improve the abnormality of blood lipid of Apo E^-/-mice.Compared with the control group,the level of triglyceride in the model group increased（P<0.05）,the level of total cholesterol increased（P<0.01）,the level of high-density lipoprotein cholesteroldecreased（P<0.01）,compared with the model group,the level of triglyceride in theA769662 group decreased significantly（P<0.01）,the level of triglyceride（P<0.01）and low density lipoprotein（P<0.05）in the Metformin group decreased significantly and high density lipoprotein increased significantly（P<0.01）;（5）Morphological changes ofadipocytes in Apo E^-/-mice,and the results of white fat HE staining showed that,compared with control group,the volume of white fat cells and subcutaneous white fat cells inepididymis of Apo E^-/-mice model group increased significantly（P<0.01）,and the volume of fat cells and subcutaneous fat cells in epididymis of mice decreased significantly（P<0.01）after AMPK agonist;（6）Morphological changes of fat cells aroundApo E^-/-mice,compared with control group,PVAT adipocytes in abdominal aorta of model group increased significantly（P<0.01）,and PVAT adipocytes decreased significantly（P<0.01）after AMPK agonist;（7）Compared with the control group,the m RNA expression of PGC-1α（P<0.01）,UCP-1（P<0.01）,and CIDEA（P<0.05）in the model group decreased significantly.Compared with the model group,the m RNA expression of PRDM-16（P<0.05）,Tmem26（P<0.05）,PGC-1α（P<0.01）,UCP-1（P<0.01）,CIDEA（P<0.05）increased significantly,inflammatory factor MCP-1 and IL-6 decreased significantly in A769662 group（P<0.01）,the m RNA expression of PGC-1αand UCP-1 in the Metformin group were increased significantly（P<0.01）,and the expression of inflammatory factors IL-6 decrease significantly（P<0.01）.Western Blot indicated compared with the control group,the expression of PGC-1α,UCP-1 and p-AMPK in the model group decreased（P<0.05）,and the expression of MCP-1 and NF-κB in the model group increased（P<0.01）.Compared with the model group,p-AMPK（P<0.05）and UCP-1（P<0.05）,PGC-1α（P<0.01）,PRDM-16（P<0.01）in the A769662 group increased,and the expression of inflammatory factors MCP-1,NF-κB（P<0.01）decreased,After administration ofMetformin,expression of UCP-1,PRDM-16 increased（P<0.01）,and expression of inflammatory factors NF-κB decrease（P<0.05）;（8）Changes aortal plaque ofApo E^-/-mice,the results of full-length oil red O staining showed that the aortic plaque in model group was significantly higher than that in control group（P<0.01）.Compared with model group,the arterial plaque was significantly reduced after AMPK agonist（P<0.01）;（9）The expression of beige gene and inflammatory factor in 3T3-L1 adipocytes waschanged.Compared with control group,the concentration of A769662 and Metformin indifferent concentrations were different.The expression of UCP-1,PRDM-16 and PGC-1αin adipocytes increased（P<0.05）,and the expression of MCP-1 and NF-κB in adipocytesdecreased（P<0.01）;（10）After the administration of AMPK,the expression of MCP-1 and NF-κB in adipocytes supernatant co cultured with macrophages decreased significantly（P<0.01）.Conclusion:AMPK agonists can inhibit the weight,improve glucose tolerance,reduce triglyceride and cholesterol levels,reduce the volume and fat content of fat cells,increase the expression of white fat beige gene and reduce inflammatory factors.Therefore,AMPK agonists can reduce atherosclerosis by activating UCP-1 to inhibit inflammation of PVAT,it can resist atherosclerosis through adventitia.