p-辛弗林通过激活AMPK-FoxO1信号通路抑制肝细胞葡萄糖生成

目的：探讨p-辛弗林对肝细胞葡萄糖生成的影响及其作用机制。方法：采用MTS法检测p-辛弗林对HepG2肝细胞株的细胞活力的影响，确定受试物的作用浓度后，比色法测定葡萄糖的生成、葡萄糖-6-磷酸酶（glucose-6-phosphatase，G6Pase）和磷酸烯醇丙酮酸羧基激酶（phosphoenolpyruvate carboxykinase，PEPCK）活力，Western blot蛋白印迹法检测腺苷酸活化蛋白激酶（adenosine 5’-monophosphate-activated protein kinase，AMPK）、磷酸化AMPK（p-AMPK）、乙酰辅酶A羧化酶（acetyl coenzyme A synthetase，ACC）、磷酸化ACC（p-ACC）、叉头框转录因子1（forkhead box class O1，FoxO1）以及磷酸化FoxO1（p-FoxO1）的表达；利用AMPK选择性抑制剂Compound C和AMPK siRNA作用HepG2肝细胞株后，检测p-辛弗林对HepG2肝细胞株葡萄糖的生成、G6Pase和PEPCK活力的影响。结果：p-辛弗林能剂量依赖性地抑制肝细胞葡萄糖的生成，激活AMPK信号通路，促进p-AMPK、p-ACC和p-FoxO1水平增加，极显著抑制G6Pase和PEPCK的活性（P＜0.01）。p-辛弗林的这些影响部分地被Compound C和AMPK siRNA所抑制（P＜0.01）。结论：p-辛弗林能够通过激活AMPK-FoxO1信号通路抑制肝细胞葡萄糖生成。

p-Synephrine Suppresses Hepatic Glucose Production via the AMPK-FoxO1 Signaling Pathway in HepG2 Cells

Objective: This study was designed to determine the effect of p-synephrine, the primary protoalkaloid of bitter orange and other citrus species, on hepatic glucose production in HepG2 cells and to elucidate the underlying mechanisms. Methods: The cell viability was detected by MTS assay. Glucose production and the activity of phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6Pase) were measured by colorimetry. The protein expression levels of adenosine 5’-monophosphate (AMP)-activated protein kinase (AMPK), phosphorylated AMPK (p-AMPK), acetyl coenzyme A synthetase (ACC), phosphorylated ACC (p-ACC), forkhead box class O1 (FoxO1) and phosphorylated FoxO1 (p-FoxO1) were analyzed by Western blot. The cells were incubated in the presence of selective AMPK inhibitor Compound C or AMPK siRNA to examine the impact of p-synephrine on glucose production and G6Pase and PEPCK activity. Results: p-Synephrine significantly inhibited hepatic glucose production in a dose-dependent manner. AMPK, ACC and FoxO1 phosphorylation were stimulated by different concentrations of p-synephrine. In addition, both enzyme activities were significantly suppressed (P < 0.01). These effects were partially reversed by Compound C and AMPK siRNA. Conclusion: p-Synephrine suppresses hepatic glucose production via the AMPK-FoxO1 signaling pathway in HepG2 cells.