S-adenosyl-L-methionine modifies antioxidant-enzymes, glutathione-biosynthesis and methionine adenosyltransferases-1/2 in hepatitis C virus-expressing cells

Figure 3 Role of actinomycin D and cycloheximide on the antiviral effect of S-adenosyl-L-methionine. A: SAM decreases replicon RNA levels without affecting its stability. Huh7 HCV replicon cells (1.5 × 10⁵ cells) were incubated with SAM 1 mmol/L, some cells were treated with actinomycin D (4 μg/mL) or CHX (50 μg/mL) for 2 h, then SAM was added. Cells were harvested 16 h later and HCV-RNA levels were quantified by real-time RT-PCR and normalized with GAPDH using ΔΔCt method. Mean results from three experiments are shown; B: CHX partially reverts the downregulated NS5A protein expression induced by SAM. Huh7 HCV replicon cells (1.5 × 10⁵ cells) were incubated with SAM 1 mmol/L, some cells were treated CHX (50 μg/mL) after 4 h of SAM treatment. Cells were harvested at different time points (0-36 h), then total protein was extracted and western blot for NS5A and GAPDH was performed. CHX: Cycloheximide; SAM: S-adenosyl-L-methionine; HCV: Hepatitis C virus.