Lycium Ruthenicum Murray Anthocyanin Extracts Promotes Adaptive Thermogenesis Induced By Browning Of White Fat In C57BL/6 Mice Through AMPK/ACC1 Pathway

The main function of brown adipose tissue is to convert its chemical energy into heat energy to keep the body temperature constant.The main function of white adipose tissue is to store energy in the form of triglycerides.While the main difference between brown adipose tissue and white adipose tissue is that brown adipose tissue can consume energy through heat production.Therefore,it is necessary to explore how to promote the browning of white adipose tissue.Objective: To study the effects of lyceum barbarum anthocyanin(AC),cold stimulation and AC+cold stimulation on the adaptive thermogenesis induced by browning of white adipose tissue in mice through AMPK/ACC1 signaling pathway.Besides,we explored the role of AC and cold exposure in promoting the transformation of white fat into brown adipose tissue to provide a theoretical basis for obesity and related diseases.Methods: C57BL/6 mice(n=40)were randomly divided into 4 groups: the normal room temperature group(Ⅰ group: 23 ℃),the normal cold stimulation group(Ⅱ group,4 ℃),the normal room temperature + anthocyanins groups(Ⅲgroup,23 ℃,40 mg/kg)and normal cold stimulation + anthocyanins(Ⅳ group,4 ℃,40 mg/kg),the corresponding indicators were detected:1.The changes of mouse body weight,food intake,percentage of fat and liver weight,glucose and insulin resistance,and blood lipid and cell morphology were measured;2.The adipose-related factor expression of UCP1,PGC-1α,PRDM16,PPARγ,Di O2,Leptin and Cidea at m RNA level were explored by RT-RCR;3.Western Blot was used to detect the expression of UCP1,PGC-1α,FABP4 and Leptin at protein level;4.UCP1,PGC-1α,FABP4 and Leptin intensity of immunochromogenic were searched by immunohistochemical detection;5.The protein expression of the key factors of AMPK/ACC1 signal pathway(p-AMPKα、AMPKα、p-ACC and ACC)were studied by western blot.Results:1.AC and cold stimulation and their combined effect can inhibit the growth of body weight in mice.Cold stimulation can increase the food intake of mice,AC can inhibit the food intake of mice.In the experiment of the percentage of fat and liver weight,AC,cold stimulation and AC+cold stimulation can reduce the weight of white fat,increase the proportion of brown fat,and reduce the formation of white fat(P<0.05).In glucose tolerance and insulin resistance experiments,AC,cold stimulation and AC+cold stimulation both can improve the glucose tolerance and insulin sensitivity of mice.In the detection of blood lipid four experiments,it was found that AC and cold stimulation and AC+cold stimulation extremely reduced the blood content of TG and LDL-C(P<0.01),while AC and cold stimulation significantly reduced T-CHO and increased HDL-C(P <0.05).It was found that the characteristics of brown fat appeared in the combination of AC and cold stimulus by HE staining.Scanning electron microscope was used to observe the brown adipose tissue,and after treatment with AC and cold stimulus and combination,the fiber bundles gradually appeared together phenomenon.2.By RT-PCR,In i WAT,AC extremely up-regulated Cidea,prdm16,pparγand UCP1 expression at RNA level(P <0.01),while significantly up-regulated PGC-1α and down-regulated Leptin expression at RNA level(P <0.05).Cold stimulation significantly up-regulated PGC-1α and UCP1 expression at RNA level(P<0.01),while significantly up-regulate Cidea and down-regulate Leptin expression at RNA level(P <0.05).The combined effect significantly up-regulated Cidea,prdm16,pparγ,PGC-1α,Di O2 and UCP1 and extremely down-regulated Leptin expression at RNA level(P<0.01).In e WAT,AC extremely reduced Leptin(P<0.01)and significantly increased the expression of prdm16,Di O2 and pparγ(P<0.05)expression at RNA level.Cold stimulation extremely reduced Leptin(P<0.01)and extremely increased Cidea expression at RNA level(P<0.01).The binding effect extremely up-regulated Cidea,prdm16,Di O2,ppar γ and UCP1 and extremely reduced the expression of Leptin expression at RNA level(P<0.01).In BAT,AC and cold stimulation and the binding effect significantly increased Prdm16 and AC significantly PGC-1α was up-regulated(P<0.05)and the combination extremely increased the expression of PGC-1α and UCP1 expression at RNA level(P <0.01).3.Protein expression of UCP1,PGC-1α,FABP4 and Leptin were detected by WB.AC,cold stimulation and AC+cold stimulation can up-regulate the expression of UCP1 and PGC-1α and inhibit the protein expression intensity of FABP4 and Leptin.4.The immunochromatographic intensity of UCP1,PGC-1α,FABP4 and Leptin was detected by immunohistochemistry.The adipose factor expression of UCP1,PGC-1α,FABP4 and Leptin induced by AC+ cold stimulation appeared significant positive reaction;.5.It was found that p-AMPKα and p-ACC were significantly higher in mice treated with AC compared to the blank group while p-AMPKα and p-ACC were extremely upregulated in Compound C+AC group through AMPK / ACC1 signaling pathway(P<0.01).Compared with AC group,the protein expression of p-AMPKα was extremely extremely promoted in Compound C and Compound C+AC group(P<0.01).Conclusion: In summary,cold stimulation + AC had a certain effect on browning of white fat,reducing the accumulation of white fat and promoting browning of white fat.In mice,the AMPK / ACC1 signaling pathway can be activated by black wolfberry anthocyanins,and its efficacy can be blocked by Compound C.