Figure 2.

Synergic antiviral effect of S-adenosyl-L-methionine plus standard interferon treatment. A: Effect of SAM and PEG-IFN + RBV on HCV-RNA levels. Huh7 HCV replicon cells (2 × 10⁵ cells) were incubated with 1 mmol/L SAM alone or SAM plus PEG-IFN (1000 UI/mL) and RBV (50 μmol/L). Cells were harvested at 24, 48 and 72 h and HCV-RNA levels were quantified by real-time RT-PCR and normalized with GAPDH and RPS18 using delta delta Ct method. Mean results from three independent experiments are shown. ^bP ≤ 0.01 comparison of SAM vs untreated cells; ^dP ≤ 0.01 comparison of SAM + PEG-IFN + RBV vs untreated cells; B: HCV NS5A protein levels in Huh7 HCV replicon cells treated with SAM PEG-IFN + RBV. Huh7 HCV replicon cells (2 × 10⁵) cells were incubated with 1 mmol/L SAM alone or SAM plus PEG-IFN (1000 UI/mL) and RBV (50 μmol/L). Cell lysates were prepared, and equal amounts of protein extracts (40 μg) were subjected to immunoblot analysis to detect NS5A and actin levels. The ratios of NS5A and actin were quantified with ImageJ 1.46r 2011 software, means of three blots are represented in the graphic. ^bP ≤ 0.01 comparison of SAM vs untreated cells; ^dP ≤ 0.01 comparison of SAM + PEG-IFN + RBV vs untreated cells; C: Huh7 HCV replicon cells (1.5 × 10⁴ cells) were incubated with 1 mmol/L SAM alone, or SAM plus PEG-IFN (1000 UI/mL) and RBV (50 μmol/L). Cells were harvested at 24, 48 and 72 h and total proteins were obtained. Cell lysates were prepared, and equal amounts of protein extracts (40 μg) were subjected to immunoblot analysis to detect STAT1, PKR and actin levels. SAM: S-adenosyl-L-methionine; HCV: Hepatitis C virus; IFN: Interferon; RBV: Ribavirin.