FIGURE 3.

Lentivirus-mediated knockdown of SIRT1 diminishes the basal and polyphenol-induced AMPK activation in HepG2 cells. HepG2 cells were infected without or with lentivirus expressing either an shRNA control or SIRT1 shRNA and then selected in 0.6 μg/ml puromycin. Cells were allowed to recover from the selection for 1 week prior to the experiments. A, co-expression of GFP in both control shRNA cells and SIRT1 shRNA cells was observed under fluorescence microscopy. B, endogenous SIRT1 expression was largely suppressed by lentivirus expressing SIRT1 shRNA. Representative immunoblots for the expression of SIRT1 and β-actin are shown in duplicates under the identical condition. C–E, knockdown of SIRT1 by lentivirus-mediated SIRT1 shRNA down-regulates the basal and polyphenol-stimulated AMPK and ACC phosphorylation. HepG2 cells expressing either control or SIRT1 shRNA were quiesced in serum-free medium overnight and treated with S17834 (10 μm, 1 h). *, p < 0.05 versus untreatment in cells expressing control shRNA; #, p < 0.05 versus S17834 treatment in cells expressing control shRNA (mean ± S.E., n = 3).