Txnip过表达抑制猪前体脂肪细胞分化并削弱葡萄糖对分化的诱导  

Inhibition of Porcine Preadipocytes Differentiation and Reduction of Glucose Induction by Txnip-overexpression

作　　者：于淇 岳小婧 张国华[1] 戴洪伟[1] 陈妍[1] 李建江[1] 卢建雄[1] YU Qi;YUE Xiao-Jing;ZHANG Guo-Hua;DAI Hong-Wei;CHEN Yan;LI Jian-Jiang;LU Jian-Xiong(College of Life Science and Engineering,Northwest Minzu University,Lanzhou 730030,China)

机构地区：[1]西北民族大学生命科学与工程学院,兰州730030

出　　处：《农业生物技术学报》2020年第8期1400-1409,共10页Journal of Agricultural Biotechnology

基　　金：国家自然科学基金(31560639,31860633);西北民族大学中央高校基本科研业务费专项资金(31920190001);西北民族大学研究生科研创新项目(Yxm2019139)。

摘　　要：硫氧还蛋白互作蛋白(thioredoxin interacting protein,Txnip)是硫氧还蛋白氧化还原系统的组成成分,通过抑制硫氧还蛋白的活性,调节细胞氧化还原平衡,影响细胞多种生理过程,然而在猪(Sus scrofa)脂肪细胞分化营养调控中的作用尚不明确.本研究合成猪Txnip基因cDNA序列,构建Txnip过表达慢病毒(lentivirus,LV)载体LV5-Txnip,转染原代培养猪前体脂肪细胞,其转染率约90%,Txnip mRNA表达水平显著上调约80倍.转染LV5-Txnip的猪前体脂肪细胞用成脂分化培养液诱导后,分别以5和15 mmol/L葡萄糖处理,每隔1d检测细胞成脂分化及相关基因表达.结果表明,在检测的各时间点,其分化比转染阴性对照慢病毒和未转染细胞显著减弱(P<0.05),过氧化物酶体增殖物激活受体γ(peroxisome proliferator-activated receptorγ,PPARγ)和脂肪酸合酶(fatty acid synthase,FAS)基因mRNA表达水平显著降低(P<0.05).与低糖培养相比,高浓度葡萄糖显著诱导对照细胞分化(P<0.05)及PPARγ、碳水化合物反应元件结合蛋白基因(carbohydrate response element binding protein,ChREBP)和葡萄糖转运子4基因(glucose transporter 4,Glut4)表达(P<0.01),但是葡萄糖对细胞分化和基因表达的诱导作用被Txnip过表达显著降低(P<0.05).本研究提示,Txnip过表达通过降低PPARγ表达抑制猪前体脂肪细胞分化,Txnip可能是其分化的抑制因子;Txnip通过降低Glut4的表达减少细胞对葡萄糖的摄取,下调ChREBP及PPARγ表达水平,从而削弱葡萄糖对分化的影响.本研究结果将为深入探讨猪脂肪形成的营养调控机制提供参考依据.Thioredoxin interacting protein(Txnip),a redox regulatory protein,binds to thioredoxin(Trx)and inhibits its activity,regulating the redox state in various physiological processes of cells,but its role in differentiation of porcine fat cells regulated by nutrient is unclear.In this study,the cDNA sequence of Sus scrofa Txnip gene was synthesized and the recombinant lentivirus(LV)overexpressing Txnip(LV5-Txnip)was constructed.The isolated porcine preadipocytes were cultured and transfected with LV5-Txnip and the transfection efficiency was around 90%.The Txnip mRNA level was raised by about 80 folds.The porcine preadipocytes transfected with LV5-Txnip were induced by adipogenic differentiation medium and treated with 5 or 15 mmol/L concentration of glucose,respectively.The differentiation and mRNA expressions of adipogenic-related genes for the cells were detected every other day using oil red O staining extraction assay and qRT-PCR,respectively.The results showed that differentiation of the cells transfected with LV5-Txnip was significantly inhibited(P<0.05),compared to the cells transfected with negative control virus and to the non-transfected cells,the expression level of peroxisome proliferator-activated receptorγ(PPARγ)and fatty acid synthase(FAS)was also reduced significantly(P<0.05).The differentiation and mRNA expression of PPARγ,carbohydrate response element binding protein(ChREBP)and glucose transporter 4(Glut4)of control preadipocytes were induced significantly by high concentration of glucose,but this promotion was reduced by Txnip-overexpression(P<0.05).Txnip-overexpression suppressed differentiation of porcine preadipocytes through down-regulating PPARγexpression,which indicating Txnip could be an inhibitor of porcine adipocyte differentiation.Txnip impaired the promotion of glucose on the cell differentiation via decreased Glut4 expression leading to lower glucose transport and expression of ChREBP and PPARγ.The present study provides a reference for the investigation of the adipogenesis regula

关 键 词：硫氧还蛋白互作蛋白(Txnip) 过表达 猪 前体脂肪细胞 分化 葡萄糖 

分 类 号：S811.3[农业科学—畜牧学]