Chemerin过表达致3T3-L1脂肪细胞葡萄糖消耗减少  

Chemerin reduces glucose consumption in 3T3-L1 adipocytes

作　　者：李雪梅[1] 翟丽东[2] 王鹏华[1] 于佩[1] 于德民[1] 

机构地区：[1]天津医科大学代谢病医院内分泌研究所卫生部激素与发育重点实验室,300070 [2]天津医科大学基础医学院人体解剖和组织胚胎系

出　　处：《中华糖尿病杂志》2014年第12期881-886,共6页CHINESE JOURNAL OF DIABETES MELLITUS

摘　　要：目的 应用重组慢病毒构建3T3-L1脂肪细胞chemerin过表达模型并进一步探讨其对糖代谢的影响及可能机制.方法 构建鼠chemerin过表达重组慢病毒,并设对照慢病毒,感染3T3-L1细胞,实时定量聚合酶链反应（RT-PCR）法检测转染后chemerin表达水平;应用胰岛素、3-异丁基1-甲基黄嘌呤、地塞米松诱导3T3-L1前脂肪细胞分化为成熟脂肪细胞,油红O染色鉴定;诱导分化第8天加入慢病毒重组体,继续培养5d,葡萄糖氧化酶法检测各组葡萄糖消耗;RT-PCR法检测各组胰岛素受体底物1（IRS1）、胰岛素受体底物2（IRS2）、蛋白激酶B1 （Akt1）、叉头状转录因子O1 （FoxO1）基因表达水平;Western-blotting检测chemerin、丝氨酸/苏氨酸蛋白激酶（Akt）、磷酸化丝氨酸/苏氨酸蛋白激酶（pAkt）、FoxO1、磷酸化叉头状转录因子O1 （pFoxO1）蛋白水平.两组数据比较应用t检验.结果 Chemerin过表达慢病毒感染3T3-L1细胞72 h后细胞中可见红色荧光,RT-PCR结果显示：过表达组与空载对照组相比chemerin基因表达明显增加（分别为3.04±0.19比1.01±0.11,t=15.65,P＜0.05）;chemerin过表达组葡萄糖消耗减少[分别为（3.30± 1.44）比（6.07±1.15） mmol/L,t=-0.35,P＜0.05];RT-PCR结果显示：IRS1、IRS2基因水平无明显变化（均P＞0.05）,Akt1基因表达下降（分别为0.76±0.08比1.07±0.15,t=-3.11,P＜0.05）,FoxO1基因表达上调（分别为1.53±0.30与1.03±0.21,t=2.34,P＜0.05）.Western-blotting结果显示：Chemerin过表达后chemerin蛋白水平增加（相对表达量分别为1.08±0.06比0.72±0.03,t=-10.12;P＜0.05）;Akt、pAkt蛋白水平均降低（分别为0.74±0.21比1.23±0.20,0.58±0.17比0.92±0.07;t=2.81、3.17,均P＜0.05）,FoxO1蛋白水平升高（分别为1.04±0.09比0.76±0.14,t=-2.91,P＜0.05）、pFoxO1蛋白水平降低（分别为0.61±0.13比0.89±0.10,t=2.93,P＜0.05）.结论 Chemerin可能通过下调Akt1 mRNA使3T3-L1脂肪细胞葡萄糖消耗减少.Objective To investigate the influence of overexpression of Chemerin on 3T3-L1 adipocyte constructed by recombinant lentiviral on glycometabolism and the potential mechanism.Methods Rat Chemerin expression of recombinant lentivirus was constructed.This recombinant lentivirus and control lentivirus transfected 3T3-L1 cells respectively.The expression of chemerin in 3T3-L1 cells was detected by real-time reverse transcriptase-polymerase chain reaction （RT-PCR）.3T3-L1 preadipocytes were induced into mature adipocyte by insulin,IBMX and dexamethasone.Oil red O was used to identify the mature 3T3-L1 adipocytes.Recombination lentiviruses were added in culture medium at the eighth day of differentiation.The cells were continuously cultured for five days and then were harvested for series of determinations.Glucose oxidase method was used to test the glucose consumption.The mRNA expression of IRS1,IRS2,Akt1 and FoxO1 were detected by RT-PCR.The protein expression of chemerin,Akt,pAkt,FoxO1 and pFoxO1 were determined by western blotting.The measurement data were measured by mean ± standard deviation （（x） ± s）.Student＇ s t-test was used for comparison of two groups.Results Red fluorescence can be observed in 3T3-L1 cells after being infected with recombination lentivirus for 72 h.The expression of chemerin mRNA significantly increased （relative expression：（3.04±0.19）vs（1.01 ± 0.11）,t=15.65,P〈0.05） and glucose consumption decreased （（3.30± 1.44）vs（6.07±1.15） mmol/L,t=-0.35,P〈0.05） in over expression chemerin group comparing with empty-vector control group.There were no difference of IRS1and IRS2 mRNA between the two groups （all P〉0.05）.Compared to the empty-vector control group,the over expression chemerin group have higher FoxO1 mRNA and protein expression （relative expression：（1.53±0.30） vs （1.03±0.21）,t=2.34,P〈0.05 ; （1.04±0.09）vs（0.76±0.14）,t=-2.91,P〈0.05）,and lower Akt1 mRNA expression （relative expression：0.76±0.08 vs 1.07±0.15,

关 键 词：慢病毒 CHEMERIN 过表达 3T3-L1脂肪细胞 葡萄糖消耗 

分 类 号：R587.1[医药卫生—内分泌]