肿瘤坏死因子α和吡格列酮对3T3-L1脂肪细胞葡萄糖转运子4表达的影响

Influences of tumor necrosis factor alpha and pioglitazone on glucose transporter 4 expression in 3T3-L1 adipocytes

目的:观察肿瘤坏死因子α和噻唑烷二酮类药物吡格列酮对3T3-L1脂肪细胞中葡萄糖转运子4(glucose transporter4,GLUT4)mRNA和蛋白表达的影响,并了解其是否有时间依赖性。方法:实验于2005-09/2006-05在安徽医科大学病原微生物分子生物学教研室进行。对3T3-L1细胞进行培养并诱导分化为成熟的3T3-L1脂肪细胞后随机分为对照组、肿瘤坏死因子α组和吡格列酮组3组。肿瘤坏死因子α组和吡格列酮组分别加含20μg/L肿瘤坏死因子α或10-4mol/L吡格列酮培养基培养,提取干预后1,3,6d细胞总RNA和总蛋白做RT-PCR和Western-blot,测定GLUT4mRNA和蛋白的表达,以未加任何药物干预组做对照。结果:①GLUT4mRNA表达:对照组细胞为0.506±0.049;肿瘤坏死因子α组干预后1,3,6d表达低于对照组(0.465±0.039,0.410±0.010,0.320±0.019,F=17.8,P=0.001),并随干预时间呈递减关系;吡格列酮组干预后1,3,6d表达高于对照组(0.544±0.064,0.616±0.065,0.664±0.070,F=4.87,P=0.043),且与干预时间呈正比。②GLUT4蛋白的相对含量:对照组细胞为0.624±0.093;肿瘤坏死因子α组干预后1,3,6d低于对照组(0.549±0.112,0.460±0.111,0.286±0.117,F=7.39,P=0.011),且随干预时间递减;而吡格列酮组GLUT4干预后1,3,6d高于对照组(0.693±0.098,0.750±0.106,0.866±0.074,F=4.0,P=0.048),随干预时间呈递增关系。结论:①肿瘤坏死因子α可降低3T3-L1脂肪细胞中GLUT4mRNA和蛋白表达量,吡格列酮的作用与肿瘤坏死因子α相反,两者作用均呈时间依赖性。②在3T3-L1脂肪细胞中,吡格列酮可能通过增加GLUT4的表达来提高胰岛素敏感性,并可能部分拮抗肿瘤坏死因子α诱导的胰岛素抵抗。

AIM： To investigate the effects of tumor necrosis factor-α （TNFα） and pioglitazone on glucose transporter 4 （GLUT4） mRNA and protein expression in 3T3-L1 adipocytes, and explore the time dependence. METHODS： The experiment was completed in the Research Room of Pathogenic Microorganism and Molecular Biology of Anhui Medical University from September 2005 to May 2006. The differentiated 3T3-L1 adipocyte cells were divided randomly into three groups： control group, TNFα group and pioglitazone group. 20 μg/L TNFα and 10^-4 mol/L pioglitazone were adopted in the TNFα group and pioglitazone group, respectively. After intervening for 1, 3 and 6 days, total RNA and total protein received RT-PCR and Western-blot to determine the expressions of GLUT4mRNA and protein, and then compared with the control group. RESULTS：（1）GLUT4mRNA expression： It was 0.506±0.049 in the control group. It was lower in the TNFα group after intervening for 1, 3 and 6 days （0.465±0.039,0.410±0.010,0.320±0.019,F =17.8,P =0.001 ）, and decreased gradually with time dependence. It was higher in the pioglitazone group after intervening for 1, 3 and 6 days than the control group （0.544±0.064,0.616±0.065,0.664±0.070, F =4.87, P =0.043）, and showed direct proportion with the time of intervention. （2）Relative amount of GLUT4： It was 0.624±0.093 in the control group. It was lower in the TNFα group after intervening for 1, 3 and 6 days （0.549±0.112,0.460±0.111,0.286±0.117,F =7.39,P =0.011 ）, and decreased gradually with time dependence. It was higher in the pioglitazone group after intervening for 1, 3 and 6 days than the control group （0.693±0.098,0.750±0.106,0.866±0.074, F =4.0,P =0.048 ）, and showed increase with the time of intervention. CONCLUSION： （1）TNFα can decrease the contents of GLUT4mRNA and protein expression in 3T3-L1 adipocytes. TNFα and pioglitazone have opposite effects that have time dependence.（2）Pioglitazone appeares to improve insulin sensitivity by increasing the GLUT4 expression and partly inhibits the effect of TNFα on insulin resistance in 3T3-L1 adipocytes.