小檗碱的降糖作用独立于AMPK的信号通路

Berberine enhances glucose metabolism in absence of AMP-activated protein kinase activation

目的：研究小檗碱的降糖作用是否依赖于单磷酸腺苷激活蛋白激酶（AMPK）信号途径。方法培养HepG2细胞和C2C12细胞，给予不同浓度小檗碱处理。葡萄糖消耗实验和乳酸生成实验用于检测小檗碱的降糖以及刺激糖酵解的作用。AMPK抑制剂化合物C（Compound C，CC）和显性失活突变型AMPK，即腺病毒负显性AMPK（Ad-DN-AMPK）腺病毒用于抑制AMPK的表达和活性。Western印迹法用于检测AMPK以及乙酰辅酶A羧化酶（ACC）磷酸化水平，以评估AMPK通路的活性。结果小檗碱显著刺激了HepG2细胞和C2C12细胞的葡萄糖消耗和乳酸生成，并表现出剂量依赖性的药物作用。5和10μmol/L的小檗碱显著增加AMPK及其下游蛋白ACC的磷酸化水平。CC和Ad-DN-AMPK腺病毒转染能明显抑制细胞内AMPK信号通路的活性。然而，在AMPK活性被抑制的条件下，小檗碱依然能够显著增加细胞的葡萄糖消耗和乳酸生成。结论小檗碱通过刺激糖酵解而上调细胞的糖代谢，该作用无需AMPK信号通路的参与。即使在AMPK的表达或者活性被抑制的情况下，小檗碱依然能够发挥显著的降糖作用。

Objective To investigate whether berberine exerts hypoglycemic effect in dependence of adenosine 5￠-monophosphate-activated protein kinase （AMPK） signal pathway. Methods After human liver hepatocellular cell line HepG2 and mouse myoblast cell line C2C12 were treated with berberine at different concentrations, glucose consumption and lactate production were used to evaluate the glucose-lowering and anaerobic glycolysis effects of berberine. Compound C （CC）, an AMPK inhibitor, and recombinant adenovirus Ad-DN-AMPK expressing dominant negative mutant of AMPK, were used to inhibit the expression and block the activity of AMPK in the cells. AMPK phosphorylation and acetyl coA carboxylase （ACC） phosphorylation were measured with Western blotting to assess the activity of AMPK pathway. Results Berberine significantly increased glucose consumption and lactate production in HepG2 and C2C12 cells in a dose-dependent manner. AMPK and ACC phosphorylation were significantly stimulated by 5 and 10μmol/L berberine. CC and adenovirus-dominant negative-AMPK （Ad-DN-AMPK） adenovirus obviously inhibited AMPK activity, but failed to diminish the promotion of glucose utilization and lactate production induced by berberine. Conclusion Berberine significantly enhances glucose consumption by stimulating glucose metabolism, in absence of AMPK pathway activation. Therefore, even in the case of the expression or activity of AMPK is inhibited, berberine is still able to exert a significant hypoglycemic effect.