Table_1_Extracellular ATP Increases Glucose Metabolism in Skeletal Muscle Cells in a P2 Receptor Dependent Manner but Does Not Contribute to Palmitate-Induced Insulin Resistance.docx

Saturated fatty acids such as palmitate contribute to the development of Type 2 Diabetes by reducing insulin sensitivity, increasing inflammation and potentially contributing to anabolic resistance. We hypothesized that palmitate-induced ATP release from skeletal muscle cells may increase inflammatory cytokine production and contribute to insulin/anabolic resistance in an autocrine/paracrine manner. In C2C12 myotubes differentiated at physiological glucose concentrations (5.5 mM), palmitate treatment (16 h) at concentrations greater than 250 μM increased release of ATP and inflammatory cytokines IL-6 and MIF, significantly blunted insulin and amino acid-induced signaling and reduced mitochondrial function. In contrast to our hypothesis, degradation of extracellular ATP using apyrase, did not alter palmitate-induced insulin resistance nor alter release of cytokines. Moreover, treatment with ATPγS (16 h), a non-hydrolysable ATP analog, in the absence of palmitate, did not diminish insulin sensitivity. Acute treatment with ATPγS produced insulin mimetic roles; increased phosphorylation of PKB (aka AKT), S6K1 and ERK and enhanced GLUT4-mediated glucose uptake in the absence of exogenous insulin. The increases in PKB and S6K1 phosphorylation were completely prevented by pre-incubation with broad spectrum purinergic receptor (P2R) blockers PPADs and suramin but not by P2 × 4 or P2 × 7 blockers 5-BDBD or A-438079, respectively. Moreover, ATPγS increased IL-6 yet decreased MIF release, similar to the cytokine profile produced by exercise. Acute and chronic treatment with ATPγS increased glycolytic rate in a manner that was differentially inhibited by PPADs and suramin, suggesting heterogeneous P2R activation in the control of cellular metabolism. In summary, our data suggest that the palmitate-induced increase in ATP does not contribute to insulin/anabolic resistance in a cell autonomous manner.

饱和脂肪酸如棕榈酸通过降低胰岛素敏感性、加剧炎症反应以及可能促进合成代谢抵抗而促成2型糖尿病的发展。本研究假设，由棕榈酸诱导的骨骼肌细胞ATP释放可能增加炎症细胞因子（如IL-6和MIF）的产生，并通过自分泌/旁分泌途径导致胰岛素/合成代谢抵抗。在C2C12肌管分化于生理葡萄糖浓度（5.5 mM）时，棕榈酸处理（16小时）在浓度高于250 μM的情况下，显著增加了ATP和炎症细胞因子IL-6及MIF的释放，显著减弱了胰岛素和氨基酸诱导的信号传导，并降低了线粒体功能。与我们的假设相反，使用apyrase降解细胞外ATP并未改变棕榈酸诱导的胰岛素抵抗，也未改变细胞因子的释放。此外，在无棕榈酸存在的情况下，使用非水解性ATP类似物ATPγS（16小时）处理并未降低胰岛素敏感性。急性ATPγS处理产生了胰岛素模拟作用；在没有外源性胰岛素的情况下，增加了PKB（又称AKT）、S6K1和ERK的磷酸化，并增强了GLUT4介导的葡萄糖摄取。PKB和S6K1磷酸化的增加可被广泛谱系的嘌呤能受体（P2R）阻断剂PPADs和suramin完全抑制，但不能被P2×4或P2×7阻断剂5-BDBD或A-438079抑制。此外，ATPγS增加了IL-6的释放，但降低了MIF的释放，这与运动产生的细胞因子谱相似。急性及慢性ATPγS处理以不同的方式增加了糖酵解速率，这可被PPADs和suramin不同地抑制，表明在细胞代谢控制中P2R激活的异质性。总之，我们的数据表明，棕榈酸诱导的ATP增加并不以细胞自主的方式导致胰岛素/合成代谢抵抗。