Figure 4.

Silent mating‐type information regulation 2 homolog 1 (SIRT1) regulates the level of reactive oxygen species (ROS) in adapted suspension cells (ASC). A, Cells transfected with siControl and siSIRT1 were labeled with dichlorofluorescein diacetate (DCFDA) and analyzed on a fluorescent plate reader. B, Intracellular reduced glutathione (GSH) and oxidized glutathione (GSSG) were measured using a luminescence plate reader in adherent cells (AD) and ASC. Data are given as GSH/GSSG ratio. C, Levels of glutathione‐related enzymes including GPX and GSR in AD and suspension cells (SC) were determined by qRT‐PCR assays. Normalization was carried out using 18S rRNA. D, Cells transfected with siControl and siSIRT1 were treated with N‐acetylcysteine (NAC) at the indicated concentration for 48 h and then the number of cells was counted. E, Cells were transfected with control or SIRT1 siRNA for 24 h and then treated with NAC at the indicated concentrations for 48 h. Level of cleaved poly ADP ribose polymerase (PARP) in SIRT1‐depleted cells was determined by immunoblot assays. *P < .05, **P < .01; n.s., not significant