Abstract 615: LKB1 is Required for Statin-induced Activation of the AMP-activated Protein Kinase
Statins, lipid-lowering drugs, exert many beneficial effects on the cardiovascular system. Recent studies have demonstrated that clinically relevant concentrations of statins might exert their therapeutic effects by activating the AMP-activated kinase (AMPK) in cultured endothelial cells, but the mechanism remains elusive. The aim of the study was to determine the molecular mechanism by which statins activate AMPK. Confluent bovine aortic endothelial cells (BAEC) were exposed to simvastatins for 1 to 24 h. Exposure of BAEC to simvastatins caused a time-dependent and dose-dependent increase of AMPK-Thr172 phosphorylation. In parallel, simvastatins significantly increased the phosphorylation of both LKB1 at serine 428 and protein kinase C (PKC)-ζ at Thr410/403. Further, adenoviral overexpression of LKB1 kinase-dead (LKB1 D194A) mutants, but not adenoviruses encoding green fluorescence proteins (GFP), ablated simvastatin-induced AMPK-Thr172 phosphorylation, suggesting LKB1 is required for statin-enhanced AMPK activation. Conversely, neither inhibition of calcium calmodulin-dependent kinase kinase (CaMKK) with STO609 (1 μM), a CaMKK-specific inhibitor, nor transfection of CaMKK-specific siRNA altered simvastatin-enhanced AMPK Thr172 phosphorylation, suggesting that CaMKK is not required for statin-enhanced AMPK activation. Moreover, overexpression of PKC-ζ dominant negative mutants or PKC-ζ-specific siRNA abolished simvastatin-enhanced phosphorylation of both LKB1-Ser428 and AMPK-Thr172, implying that PKC-ζ is the upstream enzyme of LKB1. Finally, compared to lean mice, the phosphorylation of both AMPK-Thr172 and LKB1-Ser428 was significantly suppressed in the hearts and livers of diabetic DB/DB mice (n=4 or 6, p<0.05). Treatment of diabetic DB/DB mice with fluvastatin (22.5 mg/kg/day in drinking water) for 4 months significantly increased the levels of total AMPK as well as AMPK-Thr172 phosphorylation in the livers and hearts of DB/DB mice. In addition, fluvastatin significantly increased detection of LKB1-Ser428 phosphorylation. We conclude that statins increase AMPK activation by increasing PKC-ζ-dependent LKB1 phosphorylation at Ser428 in vivo.