Departments of 1Medicine, Indiana University School of Medicine, Indianapolis, Indiana.
Am J Physiol Gastrointest Liver Physiol. 2010 Jun;298(6):G1004-12. doi: 10.1152/ajpgi.00482.2009. Epub 2010 Mar 11.
Ethanol treatment of cultured hepatoma cells and of mice inhibited the activity of AMP-activated protein kinase (AMPK). This study shows that the inhibitory effect of ethanol on AMPK phosphorylation is exerted through the inhibition of the phosphorylation of upstream kinases and the activation of protein phosphatase 2A (PP2A).Inhibition of AMPK phosphorylation by palmitate was attributed to ceramide-dependent PP2A activation. We hypothesized that the inhibitory effect of ethanol on AMPK phosphorylation was mediated partly through the generation of ceramide. The effect of ethanol and inhibitors of ceramide synthesis on AMPK phosphorylation, ceramide levels, and PP2A activity were assessed in rat hepatoma cells (H4IIEC3). The effect of ethanol on hepatic ceramide levels was also studied in C57BL/6J mice fed the Lieber-DeCarli diet. In H4IIEC3 cells, ceramide reduced AMPK phosphorylation when they were treated for between 4 and 12 h. The basal level of AMPK phosphorylation in hepatoma cells was increased with the treatment of ceramide synthase inhibitor, fumonisin B1. Ethanol treatment significantly increased cellular ceramide content and PP2A activity by approximately 18-23%, when the cells were treated with ethanol for between 4 and 12 h. These changes in intracellular ceramide concentrations and PP2A activity correlated with the time course over which ethanol inhibited AMPK phosphorylation. The activation of PP2A and inhibition of AMPK phosphorylation caused by ethanol was attenuated by fumonisin B1 and imipramine, an acid sphingomyelinase (SMase) inhibitor. There was a significant increase in the levels of ceramide and acid SMase mRNA in the livers of ethanol-fed mice compared with controls. We concluded that the effect of ethanol on AMPK appears to be mediated in part through increased cellular levels of ceramide and activation of PP2A.
乙醇处理培养的肝癌细胞和小鼠可抑制 AMP 激活的蛋白激酶（AMPK）的活性。本研究表明，乙醇对 AMPK 磷酸化的抑制作用是通过抑制上游激酶的磷酸化和蛋白磷酸酶 2A（PP2A）的激活来实现的。棕榈酸对 AMPK 磷酸化的抑制归因于神经酰胺依赖性 PP2A 激活。我们假设，乙醇对 AMPK 磷酸化的抑制作用部分是通过神经酰胺的产生来介导的。在大鼠肝癌细胞（H4IIEC3）中评估了乙醇和神经酰胺合成抑制剂对 AMPK 磷酸化、神经酰胺水平和 PP2A 活性的影响。还研究了乙醇对 Lieber-DeCarli 饮食喂养的 C57BL/6J 小鼠肝神经酰胺水平的影响。在 H4IIEC3 细胞中，当用神经酰胺处理 4 至 12 小时时，神经酰胺降低了 AMPK 磷酸化。在用神经酰胺合酶抑制剂福莫丁 B1 处理时，肝癌细胞中的 AMPK 磷酸化基础水平增加。乙醇处理可使细胞内神经酰胺含量和 PP2A 活性分别增加约 18-23%，当细胞用乙醇处理 4 至 12 小时时。这些细胞内神经酰胺浓度和 PP2A 活性的变化与乙醇抑制 AMPK 磷酸化的时间过程相关。福莫丁 B1 和酰基鞘氨醇酶（SMase）抑制剂丙咪嗪可减轻乙醇引起的 PP2A 激活和 AMPK 磷酸化抑制。与对照组相比，乙醇喂养的小鼠肝脏中神经酰胺和酸性 SMase mRNA 的水平显著增加。我们得出结论，乙醇对 AMPK 的作用部分是通过增加细胞内神经酰胺水平和激活 PP2A 来介导的。
