Engelman J A, Berg A H, Lewis R Y, Lisanti M P, Scherer P E
Department of Molecular Pharmacology and Diabetes Research and Training Center, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Mol Endocrinol. 2000 Oct;14(10):1557-69. doi: 10.1210/mend.14.10.0542.
Tumor necrosis factor-alpha (TNFalpha) has been implicated as a contributing mediator of insulin resistance observed in pathophysiological conditions such as obesity, cancer-induced cachexia, and bacterial infections. Previous studies have demonstrated that TNFalpha confers insulin resistance by promoting phosphorylation of serine residues on insulin receptor substrate 1 (IRS-1), thereby diminishing subsequent insulin-induced tyrosine phosphorylation of IRS-1. However, little is known about which signaling molecules are involved in this process in adipocytes and about the temporal sequence of events that ultimately leads to TNFalpha-stimulated IRS-1 serine phosphorylation. In this study, we demonstrate that specific inhibitors of the MAP kinase kinase (MEK)1/2-p42/44 mitogen-activated protein (MAP) kinase pathway restore insulin signaling to normal levels despite the presence of TNFalpha. Additional experiments show that MEK1/2 activity is required for TNFalpha-induced IRS-1 serine phosphorylation, thereby suggesting a mechanism by which these inhibitors restore insulin signaling. We observe that TNFalpha requires 2.5-4 h to markedly reduce insulin-triggered tyrosine phosphorylation of IRS-1 in 3T3-L1 adipocytes. Although TNFalpha activates p42/44 MAP kinase, maximal stimulation is observed within 10-30 min. To our surprise, p42/44 activity returns to basal levels well before IRS-1 serine phosphorylation and insulin resistance are observed. These activation kinetics suggest a mechanism of p42/44 action more complicated than a direct phosphorylation of IRS-1 triggered by the early spike of TNFalpha-induced p42/44 activity. Chronic TNFalpha treatment (>> 72 h) causes adipocyte dedifferentiation, as evidenced by the loss of triglycerides and down-regulation of adipocyte-specific markers. We observe that this longer term TNFalpha-mediated dedifferentiation effect utilizes alternative, p42/44 MAP kinase-independent intracellular pathways. This study suggests that TNFalpha-mediated insulin resistance, but not adipocyte dedifferentiation, is mediated by the MEK1/2-p42/44 MAP kinase pathway.
肿瘤坏死因子-α（TNFα）被认为是在肥胖、癌症引起的恶病质和细菌感染等病理生理状况下所观察到的胰岛素抵抗的促成介质。先前的研究表明，TNFα通过促进胰岛素受体底物1（IRS-1）上丝氨酸残基的磷酸化来赋予胰岛素抵抗，从而减少随后胰岛素诱导的IRS-1酪氨酸磷酸化。然而，关于脂肪细胞中此过程涉及哪些信号分子以及最终导致TNFα刺激的IRS-1丝氨酸磷酸化的事件的时间顺序，人们知之甚少。在本研究中，我们证明尽管存在TNFα，丝裂原活化蛋白激酶激酶（MEK）1/2-p42/44丝裂原活化蛋白（MAP）激酶途径的特异性抑制剂可将胰岛素信号恢复至正常水平。额外的实验表明，TNFα诱导的IRS-1丝氨酸磷酸化需要MEK1/2活性，从而提示了这些抑制剂恢复胰岛素信号的一种机制。我们观察到，在3T3-L1脂肪细胞中，TNFα需要2.5至4小时才能显著降低胰岛素触发的IRS-1酪氨酸磷酸化。尽管TNFα激活p42/44 MAP激酶，但在10至30分钟内观察到最大刺激。令我们惊讶的是，在观察到IRS-1丝氨酸磷酸化和胰岛素抵抗之前，p42/44活性就已恢复到基础水平。这些激活动力学表明，p42/44作用的机制比由TNFα诱导的p42/44活性早期峰值触发的IRS-1直接磷酸化更为复杂。长期TNFα处理（>> 72小时）会导致脂肪细胞去分化，甘油三酯的丧失和脂肪细胞特异性标志物的下调证明了这一点。我们观察到，这种长期的TNFα介导的去分化效应利用了替代的、不依赖p42/44 MAP激酶的细胞内途径。本研究表明，TNFα介导的胰岛素抵抗而非脂肪细胞去分化是由MEK1/2-p42/44 MAP激酶途径介导的。
