Hepatitis C Virus NS5A Protein Triggers Oxidative Stress by Inducing NADPH Oxidases 1 and 4 and Cytochrome P450 2E1

Figure 2

Domain I of NS5A protein triggers ROS production via activation of the TGFβ1 NOX1 COX-2 NOX4 cascade. (a, b) Huh7 cells were transfected with NS5A-expression constructs and subjected to gene expression analysis by real-time RT-PCR (a) or Western blotting (b). (c, d) NS5A-expressing cells were treated with anti-TGFβ1/2 antibodies (c) or anti-HCV core antibodies as negative control (d), and after an additional 30 hrs gene expression was quantified using RT-qPCR. (d–f) Huh7 cells expressing full-length NS5A were transfected with siRNA and 48 hrs later subjected to gene expression analysis by RT-qPCR (e) or measurement of ROS levels using the DHE (f) or H₂DCFDA (g) assays. Results are mean ± SD from six replicates; or versus DMSO-treated cells transfected with pVax1 (ANOVA with Tukey-Kramer post hoc analysis); versus DMSO-treated cells transfected with the respective NS5A form (ANOVA with Tukey-Kramer post hoc analysis); (-test).

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