Department of Immunology, Key Laboratory of Immuno Microenvironment and Disease of the Educational Ministry of China, Tianjin Medical University, Tianjin 300070, China.
Eur J Cell Biol. 2011 Dec;90(12):1000-15. doi: 10.1016/j.ejcb.2011.06.004. Epub 2011 Oct 1.
Adipose tissue hypoxia is an early phenotype in obesity, associated with macrophage infiltration and local inflammation. Here we test the hypothesis that adipocytes in culture respond to a hypoxic environment with the release of pro-inflammatory factors that stimulate macrophage migration and cause muscle insulin resistance. 3T3-L1 adipocytes cultured in a 1% O2 atmosphere responded with a classic hypoxia response by elevating protein expression of HIF-1α. This was associated with elevated mRNA expression and peptide release of cytokines TNFα, IL-6 and the chemokine monocyte chemoattractant protein-1 (MCP-1). The mRNA and protein expression of the anti-inflammatory adipokine adiponectin was reduced. Conditioned medium from hypoxia-treated adipocytes (CM-H), inhibited insulin-stimulated and raised basal cell surface levels of GLUT4myc stably expressed in C2C12 myotubes. Insulin stimulation of Akt and AS160 phosphorylation, key regulators of GLUT4myc exocytosis, was markedly impaired. CM-H also caused activation of JNK and S6K, and elevated serine phosphorylation of IRS1 in the C2C12 myotubes. These effects were implicated in reducing propagation of insulin signaling to Akt and AS160. Heat inactivation of CM-H reversed its dual effects on GLUT4myc traffic in muscle cells. Interestingly, antibody-mediated neutralization of IL-6 in CM-H lowered its effect on both the basal and insulin-stimulated cell surface GLUT4myc compared to unmodified CM-H. IL-6 may have regulated GLUT4myc traffic through its action on AMPK. Additionally, antibody-mediated neutralization of MCP-1 partly reversed the inhibition of insulin-stimulated GLUT4myc exocytosis caused by unmodified CM-H. In Transwell co-culture, hypoxia-challenged adipocytes attracted RAW 264.7 macrophages, consistent with elevated release of MCP-1 from adipocytes during hypoxia. Neutralization of MCP-1 in adipocyte CM-H prevented macrophage migration towards it and partly reversed the effect of CM-H on insulin response in muscle cells. We conclude that adipose tissue hypoxia may be an important trigger of its inflammatory response observed in obesity, and the elevated chemokine MCP-1 may contribute to increased macrophage migration towards adipose tissue and subsequent decreased insulin responsiveness of glucose uptake in muscle.
脂肪组织缺氧是肥胖的早期表型，与巨噬细胞浸润和局部炎症有关。在这里，我们验证了这样一个假设，即在低氧环境下，培养的脂肪细胞会释放促炎因子，刺激巨噬细胞迁移，并导致肌肉胰岛素抵抗。在 1%氧气环境中培养的 3T3-L1 脂肪细胞通过提高 HIF-1α 的蛋白表达，对缺氧环境产生了典型的缺氧反应。这与细胞因子 TNFα、IL-6 和趋化因子单核细胞趋化蛋白-1（MCP-1）的 mRNA 表达和肽释放升高有关。抗炎脂肪因子脂联素的 mRNA 和蛋白表达减少。缺氧处理脂肪细胞的条件培养基（CM-H）抑制了胰岛素刺激和基础细胞表面水平的稳定表达在 C2C12 肌管中的 GLUT4myc。胰岛素刺激 Akt 和 AS160 磷酸化，GLUT4myc 胞吐的关键调节因子，明显受损。CM-H 还导致 C2C12 肌管中 JNK 和 S6K 的激活，以及 IRS1 的丝氨酸磷酸化升高。这些作用与降低胰岛素信号向 Akt 和 AS160 的传递有关。CM-H 的热失活逆转了其对肌肉细胞中 GLUT4myc 转运的双重影响。有趣的是，抗体中和 CM-H 中的 IL-6 降低了其对基础和胰岛素刺激的细胞表面 GLUT4myc 的作用，与未修饰的 CM-H 相比。IL-6 可能通过其对 AMPK 的作用来调节 GLUT4myc 转运。此外，抗体中和未修饰 CM-H 引起的胰岛素刺激的 GLUT4myc 胞吐作用部分逆转。在 Transwell 共培养中，缺氧挑战的脂肪细胞吸引 RAW 264.7 巨噬细胞，与脂肪细胞在缺氧期间释放的 MCP-1 升高一致。在脂肪细胞 CM-H 中中和 MCP-1 阻止了巨噬细胞向其迁移，并部分逆转了 CM-H 对肌肉细胞胰岛素反应的影响。我们得出结论，脂肪组织缺氧可能是肥胖症中观察到的炎症反应的一个重要触发因素，升高的趋化因子 MCP-1 可能导致更多的巨噬细胞向脂肪组织迁移，并随后降低肌肉对葡萄糖摄取的胰岛素反应。
