The Inhibition Of Anthocyanins From Aronia Melanocarpa On 3T3-L1 Preadipocytes

Aronia melnocarpa,originated in North America,is used in the food industry to produce jam,preserves,fruit tea and dietary supplements.Due to the presence of a large number of polyphenols such as proanthocyanidins,anthocyanins,phenolic acids,flavonols and so on,the taste of them is sour and astringent.The unprocessed Aronia melnocarpa is rarely eaten.The research showed that Aronia melnocarpa has the functions of anti-oxidation,anti-cancer,anti diabetes and so on.Nowadays,with the increasing number of obese and overweight people,the research on anti obesity ingredients from plants has become a hot spot in recent years.Therefore,this study established the differentiation model of 3T3-L1 preadipocyte,at the same time,anthocyanins of Aronia melnocarpa(AAM)was added to intervene the differentiation process,at the end of differentiation,oil red O staining was used to observe the degree of lipid accumulation in the cells,the triglyceride and intake of glucose was measured by spectrophotometry,the content of TNF-α was measured by ELISA,the mRNA and protein expression of differentiation related genes were measured by PCR and Western blot.To explore the inhibition of AAM on 3T3-L1 preadipocytes.(1)Toxicity of AAM to 3T3-L1 preadipocytes was detected by MTT method.After 3T3-L1 preadipocytes were treated with different concentrations of AAM for 48 h,the cell viability gradually decreased with increasing concentration.When the AAM concentration was 10μg/mL,the cell viability was greater than 90%,that is,the effect of AAM on cytotoxicity was negligible.Therefore,2,6,10 μg/mL was selected for subsequent experiments.(2)After AAM interfered with the 3T3-L1 differentiation process,compared with the blank control group,the low,medium,and high dose groups of AAM could inhibit the formation of intracellular lipids to varying degrees,and the inhibition rates reached 16.4%,39.3%,and 60.8%respectively;inhibit the TG release of cells,the inhibition rates reached 7.8%,42.0%and 78.4%respectively;promote the glucose uptake of cells,increase by 7.4%,8.3%and 40.7%respectively;inhibit the release of TNF-α by cells,the inhibition rate 45.5%,72.3%and 81.0%respectively.It shows that AAM intervention can effectively inhibit the differentiation of 3T3-L1 preadipocytes,and there was a certain dose-dependent effect.(3)After AAM intervention in 3T3-L1 differentiation,the effects on fat factors were as follows:in the AAM low dose group,the expression of FABP4 mRNA was significantly down regulated(p<0.05),and the expression of ACC and FAS mRNA was extreme significantly down regulated(p<0.01).In AAM medium dose group,the expression of HSL mRNA was significantly down regulated(p<0.05),and the expression of ACC,FABP4,C/EBP α,Fas,Perilipin and Leptin mRNA were significantly down regulated(p<0.01).In the AAM high dose group,the expression of ACC,FABP4,C/EBP α,FAS,Perilipin,Leptin mRNA was extreme significantly down regulated(p<0.01).The expression of Adiponectin mRNA was significantly increased(p<0.01).The effects of AAM on inflammatory factors was that the expression of IL-6,TLR and MCP-1 mRNA could be significantly decreased in the middle and high dose AAM groups(p<0.01).It shows that AAM can down-regulating ACC,FABP4,C EBP α,FAS,Perilipin,HSL,PPAR γ,Leptin,and SREBP-1c and up-regulate the expression of Adiponectin mRNA,inhibit the differentiation of 3T3-L1 preadipocytes in a dose-dependent manner.(4)AAM intervention in 3T3-L1 differentiation process,AAM medium dose group can significantly improve the level of AMPK and ACC phosphorylation(p<0.05),significantly reduce the level of C/EBP a protein expression(p<0.01).High dose group significantly improve the level of AMPK and ACC phosphorylation,reduce the level of PPAR y protein expression(p<0.01),further indicating that 3T3-L1 preadipocyte differentiation is inhibited by different degrees.The above results show that AAM can regulate the mRNA and protein expression of differentiation and inflammation related genes in 3T3-L1 preadipocytes,inhibit the differentiation process,activate AMPK pathway,alleviate the inflammation caused by differentiation,and maintain the normal morphology of cells.