| As obesity results from the imbalance between energy consumption and energy intake,it is effective to treat obesity by increasing energy expenditure or reducing energy intake.White adipose tissue(WAT)stores energy in the form of triglycerides,while brown adipose tissue(BAT)distributes energy in the form of heat.Stimulated with cold and other conditions,there will be another cell type similar to brown adipocytes called the beige adipocytes.The accumulation of beige adipocytes in WAT is often referred to as ‘browning’ of WAT.Browning of WAT can increase energy consumption through adipocyte thermogenesis,which is generally accepted as a promising strategy to mitigate obesity and related metabolic diseases.Various ways can promote the browning of WAT,such as cold treatment,exercise,hormone and metabolic regulation.However,few effective and safe agents are available clinically that promote browning of WAT.Both brown and beige adipocytes is characteristic with high expression levels of Uncoupling protein-1(Ucp1),which is located in the mitochondrial membrane.Ucp1 can convert the proton power potential into heat to increase energy consumption.In this study,20 traditional Chinese herbal medicines were screened by Ucp1-2A-luciferase to examine whether they induce browning of WAT.We found that Astragalus membranaceus extract significantly enhanced the expression of Ucp1,promoting the browning of WAT.To identify the constituents involved in adipocyte thermogenesis,each of these constituents was used to treat the adipocytes.Formononetin was found the critical component of Astragalus membranaceus to promote the browning of WAT.Additionally,real-time quantitative PCR(qPCR)showed that formononetin elevated the expression of other thermogenic genes.Thus formononetin appears to be the critical constituent that mediates the effect of Astragalus membranaceus extract on adipocyte thermogenesis.Immun-ofluorescence staining further showed that formononetin can increase the UCP1 protein level.The effect of formononetin on adipocyte thermogenesis was further verified by Seahorse bioanalyzer,which indicated that treatment of formononetin is capable of increasing uncoupled mitochondrial respiration.After 8 weeks of HFD feeded,C57BL6/J mice were treated with formononetin or wehicle for 8 weeks.Res μlts show that formononetin-treated mice showed lower bodyweight than the control mice.Consistent with this finding,weights of the white adipose tissue(eWAT)and the white adipose tissue(eWAT)in the experimental mice were significantly lower than that in the control group,and the weight of brown adipose tissue(BAT)was not significantly changed.All of these results showed that formononetin reduced only fat mass but not lean mass in DIO mice.Consistently,ins μlin sensitivity was improved in the formononetin treated mice,as the glucose tolerance test(GTT)showed.H&E staining analysis also confirmed that the size of the adipocytes in the adipose tissue was smaller in formononetin-treated mice than in the control mice.The expression levels of the brown adipocyte gene Prdm16,Dio2,pgc-1,and Cidea were significantly increased in the white adipose tissue(eWAT)、 the white adipose tissue(eWAT)and brown adipose tissue(BAT)of mice after the treatment of formononetin.Furthermore,these results suggest that formononetin have the same molecular target as rosiglitazone.Both formononetin and rosiglitazone dose-dependently promoted luciferase activity.Furthermore,siRNA knockdown of PPARγ dramatically reduced adipocyte thermogenesis induction by formononetin.To further verify that formononetin can bind directly with PPARγ,AlphaScreen binding assay was performed.Formononetin binds with PPARγ to form a heterodimer with RXR to drive expression of Ucp1 in adipocytes.Taken together,our study demonstrates that the Chinese herbal medicine Astragalus membranaceus and its constituent formononetin have the potential to reduce obesity and associated metabolic disorders. |
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