The Ca-calmodulin kinase kinase β is involved in AMPK activation by capsaicin in HepG2 cells.

A, Cells were incubated with 200μM capsaicin during 1h and levels of phospho-LKB1, and total LKB1 were determined by Western blot. β-tubulin serves as a loading control. The densitometric analysis of bands is shown below. B, Effect of the CaMKK inhibitor STO-609 on AMPK activation. Cells were pre-incubated with 10 μM STO-609 and then treated with 200μM capsaicin during 1h. Levels of phospho-AMPK, phospho-ACC and their corresponding total forms were determined by Western blot. β-tubulin serves as a loading control. The densitometric analysis of bands is shown on the right. C, Effect of CaMKKβ knock down on AMPK activation. Cells were transfected with scrambled siRNA (siControl) or selective CaMKKβ siRNA and incubated with 200μM capsaicin during 1h. Levels of phospho-AMPK, phospho-ACC and their corresponding total forms were determined by Western blot. β-tubulin serves as a loading control. Left, densitometric analysis of CaMKKβ. Right, densitometric analysis of pAMPK and pACC. D, Cell viability determined by MTT of cells transfected with scrambled siRNA (siControl) or selective CaMKKβ siRNA. Data are presented as the mean ± SEM of three different experiments. *, p<0.05 compared with the control group and # p< 0.05 compared with capsaicin by the Student’s t test.