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. 2016 Feb 24;7(11):12075–12088. doi: 10.18632/oncotarget.7679

Figure 7. Blocking of sirt1 activity using siRNA and sirtinol inhibited the activation of autophagy flux by melatonin.

Figure 7

HaCaT keratinocytes were incubated with melatonin in the presence of sirtinol for 24 hr. Total keratinocyte extracts were prepared and analyzed by Western blot for LC3-II and p62 protein levels A. Sirt1 small interfering RNA (siSirt1) or negative control siRNA (NC siRNA) transfected HaCaT keratinocytes were incubated with 10 μM of melatonin. Western blot for LC3-II and p62 proteins was analyzed from HaCaT keratinocytes B. HaCaT keratinocyte was transfected by overexpressing adenovirus (Ad-Sirt1) or lacZ-bearing adenovirus (Ad-lacZ). Western blot for LC3-II and p62 proteins were analyzed from HaCaT keratinocytes C. β-actin was used as loading control. The cells were immunostained with p62 antibody (green) and observed in the fluorescent view Scale bar: 10 μm D.