Mechanistic evaluation of the insulin response in H4IIE hepatoma cells: New endpoints for toxicity testing?

DNA microarray analysis, real-time PCR and flow cytometric cell cycle analysis were used to assess the relevance of the insulin response in H4IIE cells. Insulin dose dependently stimulated H4IIE growth. Time dependency of the insulin response was shown with real-time PCR for the known insulin responsive genes: Fasn, Pck1 and Irs2. Based on these results, microarray analysis was performed on H4IIE cells exposed to insulin (100 nM) for 6h and 24h. Genes related to carbohydrate (glycolysis and gluconeogenesis) and lipid metabolism (glycerolipid metabolism, cholesterol synthesis and fatty acid oxidation) were most profoundly afflicted, in accordance with in vivo insulin action in liver. Since changes in carbohydrate and lipid metabolism are pivotal in the pathogenesis of insulin resistance, the presence of a physiological relevant insulin response in H4IIE cells pleads for further testing of its potential use in research on pollutant-driven insulin resistance.本研究采用DNA微阵列分析（DNA microarray analysis）、实时定量PCR（real-time PCR）及流式细胞术细胞周期分析（flow cytometric cell cycle analysis），以评估H4IIE细胞内胰岛素应答的相关性。结果显示，胰岛素可剂量依赖性地促进H4IIE细胞增殖。针对已知胰岛素应答基因Fasn、Pck1及Irs2的实时定量PCR检测证实了胰岛素应答的时间依赖性特征。基于上述发现，本研究对经100 nM胰岛素分别处理6小时与24小时的H4IIE细胞开展了DNA微阵列分析。与肝脏内胰岛素的体内生理作用相符，与碳水化合物代谢（糖酵解与糖异生）及脂质代谢（甘油脂代谢、胆固醇合成及脂肪酸氧化）相关的基因受到最为显著的表达调控。鉴于碳水化合物与脂质代谢的改变在胰岛素抵抗的发病机制中发挥关键作用，而H4IIE细胞具备生理相关性的胰岛素应答能力，因此支持对其在污染物驱动的胰岛素抵抗相关研究中的潜在应用价值开展进一步验证测试。