基于AMPK通路研究葛根素对猪前体脂肪细胞成脂分化的影响  

Effect of Puerarin on Adipogenic Differentiation of Porcine Preadipocytes Based on AMPK Pathway

作　　者：吕香州 张琪[1] 李欣 于永生[1] LYU Xiangzhou;ZHANG Qi;LI Xin;YU Yongsheng(Institute of Animal Biotechnology,Jilin Academy of Agricultural Sciences,Gongzhuling 136100,China)

机构地区：[1]吉林省农业科学院动物生物技术研究所,公主岭136100

出　　处：《中国畜牧兽医》2022年第6期2088-2097,共10页China Animal Husbandry & Veterinary Medicine

基　　金：中央引导地方科技发展资金(202002066JC);吉林省农业科技创新工程(C12044104)。

摘　　要：【目的】探究葛根素是否通过AMPK通路促进松辽黑猪前体脂肪细胞分化,以期为在饲粮中添加葛根素改善猪肉肉质提供参考。【方法】待松辽黑猪前体脂肪细胞汇合度达90%时进行为期4 d的诱导分化。诱导分化时,将细胞分为对照组(Con)、葛根素组(Pue)、葛根素+AMPK激活剂AICAR组(AICAR)及葛根素+AMPK抑制剂CompoundC组(CompoundC),对照组诱导液中不添加葛根素,Pue组添加40μmol/L葛根素,AICAR组添加40μmol/L葛根素+500μmol/L AICAR,CompoundC组添加40μmol/L葛根素+20μmol/L CompoundC。诱导分化结束,收集各组细胞,用甘油三酯(TG)酶法检测细胞中甘油三酯浓度;实时荧光定量PCR检测AMPK各亚基以及成脂分化相关基因的表达水平;用Western blotting检测过氧化物酶体增殖物激活受体γ(PPARγ)、AMP依赖的蛋白激酶(AMPK)、p-AMPK(Thr-172)、乙酰辅酶A羧化酶(ACC)的蛋白表达水平;通过Autodock Vina 1.20对葛根素、AMPKα亚基做分子对接预测。【结果】甘油三酯测定结果显示,与Con组相比,Pue组甘油三酯浓度显著增加(P<0.05);与Pue组相比,AICAR组甘油三酯浓度显著降低(P<0.05)。实时荧光定量PCR结果显示,与Con组相比,Pue组PPKAG2、PPKAB1、PPKAB2、PPKAA1、PGC-1α表达量均显著降低(P<0.05),Pue组C/EBPα、PPARγ、ACC表达量均显著增加(P<0.05);与Pue组相比,AICAR组PPKAG1、PPKAG2、PPKAB1、PPKAB2、PPKAA1表达量显著增加(P<0.05),AICAR组C/EBPα、PPARγ、ACC表达量均显著降低(P<0.05),CompoundC组PPKAG1、PPKAG2、PPKAB2表达量均显著降低(P<0.05),CompoundC组C/EBPα、ACC表达量均显著增加(P<0.05)。Western blotting结果显示,与Con组相比,Pue组PPARγ、ACC蛋白表达量显著增加(P<0.05),AMPK、p-AMPK蛋白表达量显著下降(P<0.05),且p-AMPK/AMPK显著下降(P<0.05);与Pue组相比,AICAR组PPARγ、ACC蛋白表达量显著下降(P<0.05),AMPK、p-AMPK蛋白表达量显著增加(P<0.05),CompoundC组PPARγ、ACC蛋白表达量显著增加(P<0.05),AMPK�【Objective】This study was aimed to explore whether puerarin could promote the differentiation of Songliao Black pig precursor adipocytes through the AMPK pathway,in order to provide a reference for adding puerarin to diet to improve pork meat quality.【Method】When the confluence degree of precursor adipocytes of Songliao Black pig reached 90%,the differentiation was induced for 4 days.When inducing differentiation,the cells were divided into control group(Con),puerarin group(Pue),puerarin+AMPK activator AICAR group(AICAR)and puerarin+AMPK inhibitor CompoundC group(CompoundC).Puerarin was not added to the induction solution in the control group,while 40μmol/L puerarin was added in the Pue group,40μmol/L puerarin+500μmol/L AICAR were added in AICAR group,40μmol/L puerarin+20μmol/L CompoundC were added in CompoundC group.After induction and differentiation,the cells of each group were collected and the triglyceride(TG)concentration was detected by triglyceride enzyme method,the expression levels of AMPK subunits and adipogenic differentiation related genes were detected by Real-time quantitative PCR,peroxisome proliferator activated receptorγ(PPARγ)AMP-dependent protein kinase(AMPK),p-AMPK(Thr-172),acetyl-CoA carboxylase(ACC)were detected by Western blotting.Autodock Vina 1.20 was used to predict the molecular docking of puerarin and AMPKαsubunits.【Result】The results of triglyceride determination showed that compared with Con group,the triglyceride concentration of Pue group was significantly up-regulated(P<0.05);Compared with Pue group,the triglyceride concentration of AICAR group was significantly decreased(P<0.05).The results of Real-time quantitative PCR showed that compared with Con group,the expressions of PPKAG2,PPKAB1,PPKAB2,PPKAA1 and PGC-1αin Pue group were significantly decreased(P<0.05),the expressions of C/EBPα,PPARγand ACC in Pue group were significantly increased(P<0.05);Compared with Pue group,the expressions of PPKAG1,PPKAG2,PPKAB1,PPKAB2 and PPKAA1 in AICAR group were signific

关 键 词：葛根素 松辽黑猪 AMPK通路 成脂分化 脂肪细胞 

分 类 号：S828[农业科学—畜牧学]