| Background and objective:Propolis has been widely used as a traditional natural medicine for a long time because of its antibacterial,antioxidant,anti-inflammatory and metabolic regulating functions.Research evidences have shown that propolis can effectively alleviate obesity,insulin resistance,type 2 diabetes and other metabolic diseases,but the exact mechanism is still unclear.In recent years,a large number of evidences have shown that gut microbiota plays an important role in the pathogenesis of metabolic diseases,and diet is the key factor to change gut microbiota.It has been shown that propolis can regulate the gut microbiota of colitis animal model induced by dextran sulfate and high fat diet fed animal model.We speculated that propolis may play a role in the prevention and treatment of metabolic diseases by regulating gut microbiota.Therefore,this study was to study the effects of ethanol extract of propolis(EEP)on insulin resistance,glycolipid metabolism and gut microbiota of obese mice induced by high fat diet(HFD),and to analyze the relationship between the structural changes of gut microbiota and insulin resistance,obesity,glycolipid metabolism and inflammation in HFD-fed mice.Methods:Eight-week-old male wild-type mice of the C57BL/6J genetic lineage were randomly distributed into four groups with 10 individuals in each group and fed once a day by intragastric gavage:1)Chow diet group(Chow group):mice were fed with chow diet and water,2)High-fat diet group(HFD group):mice were fed with high-fat diet and polyethylene glycol,3)High-fat diet and 1%EEP group(HFD+1%EEP group):mice were fed with high-fat diet and 1%EEP,and 4)High-fat diet and 2%EEP group(HFD+2%EEP group):mice were fed with high-fat diet and 2%EEP.After 12 weeks of treatment,the samples of organs,tissues,feces,and serum were collected from each mouse for subsequent analyses.The main components and contents of phenolic compounds in EEP were detected by high performance liquid chromatography(HPLC).Glucose tolerance and insulin resistance of mice were assessed by oral glucose tolerance test(OGTT)and insulin tolerance test(ITT).The expression of tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),interleukin-6(IL-6),and interleukin-10(IL-10)in liver and adipose tissue of mice were detected by fluorescence quantitative PCR,and the serum levels of these inflammatory cytokines and lipopolysaccharide binding proteins(LBP)were detected by enzyme-linked immunosorbent assay(ELISA).The fat accumulation of liver and epididymis was detected by immunohistochemistry and oil red O staining.Liquid chromatography-mass spectrometry(LC-MS)was used to analyze the lipid of liver tissue in mice;Miseq sequencing of 16SrRNA gene of intestinal bacteria in mice was used to analyze the bacterial structure and metabolism function.Results:(1)A total of 12 phenolic compounds(Protocatechuic acid,caffeic acid,p-coumaric acid,ferulic acid,daidzein,cinnamic acid,quercetin,naringenin,genistein,keampferol,hesperetin,and biochanin A were detected in the EEP using HPLC.(2)EEP significantly improved insulin resistance and glucose tolerance in HFD-fed mice:EEP decreased fasting insulin,insulin resistance index and area under ITT curve in a dose-dependent manner,and decreased fasting blood glucose and area under OGTT curve in a dose-dependent manner in HFD fed mice.(3)EEP significantly reduced the obesity of HFD-fed mice:EEP decreased the weight gain,fat accumulation of liver and epididymis in HFD-fed mice in a dose-dependent manner,and these effects were not related to the food intake of mice.(4)EEP significantly reduced the expression of inflammatory cytokines and endotoxemia induced by HFD in mice:EEP decreased the expression of TNF-α,IL-1βand IL-6 in HFD-fed mice in a dose-dependent manner,increased the expression of IL-10,and decreased the serum level of LBP.The serum levels of inflammatory markers were significantly correlated with the metabolic markers of obesity,insulin resistance,blood glucose and lipid in HFD-fed mice.(5)EEP significantly improved the blood lipid level and liver lipid metabolism level in HFD-fed mice:EEP significantly reduced the serum levels of TG and TC in HFD-fed mice.LC-MS showed that EEP significantly changed the contents of liver lipid in HFD fed mice,a total of 122 lipid components were significantly changed,including 74 glycerides(GL),37 Glycerophospholipids(GP)and 11 sphingolipids(SP).(6)EEP can significantly regulate the structure and composition of gut microbiota in HFD-fed mice:it can increase the abundance of anti-inflammatory bacteria in the intestinal flora of mice,such as Roseburia and Intellimonas,Paracharacteroidesgoldsteinii and Paracharacteroidesdistasonis,and reduce the abundance of pro-inflammatory bacteria such as Faecalibaculum and Prevotella,and Bacteroides vulgatus.(7)The dominant gut bacterial genera and species altered by EEP supplementation were closely associated with the parameters of insulin resistance,glucose and lipid metabolism and obesity in HFD-fed mice.Moreover,EEP can partially reverse the function of intestinal microbiota of HFD-fed mice to a similar level to that in chow diet mice.Conclusion:EEP supplementation reduced HFD-induced weight gain and liver fat accumulation,proinflammatory cytokines and insulin resistance,and improved glucose tolerance and lipid profile.Meanwhile,EEP supplementation in HFD-fed mice increased anti-obesity and anti-inflammatory bacteria,and reduced pro-inflammatory bacteria.Moreover,these dominant bacterial taxa altered by EEP were significantly associated with the metabolic parameters of insulin resistance and obesity in HFD-fed mice.These results in this study indicated that EEP reduced HFD-induced obesity and insulin resistance and improved glucose tolerance and lipid profile,which may be mediated by modulating the composition and function of gut microbiota in mice.However,further studies,especially clinical studies,are still needed to provide more evidence to support these conclusions. |
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