Fig. 3. HCV infection triggers NF-κB activation with delayed kinetics in 7.5-TLR3 cells.

(A) PRDII promoter activities in Huh7.5 and 7.5-TLR3 cells at various time points post-infection with JFH1 virus (MOI=1). “*” and “**” indicate statistical differences exist between mock- and HCV-infected 7.5-TLR3 cells with a p-value of <0.05 and <0.01, respectively. (B) through (D) 7.5-TLR3 cells were infected with JFH1 virus (MOI=0.5) for the indicated time periods. An uninfected culture treated with TNF (15 ng/ml, 2 h) was included as a positive control for NF-κB activation. (B) Immunoblot analysis was conducted to determine the time-course IκBα degradation, which was quantified by densitometry (lower graph). (C) Immunoblotting of p65 accumulation in the nuclear extracts at various time points post-infection. Immunoblotting of Lamin A/C was shown as a loading control. (D) EMSA analysis of time-course nuclear p65/p50 binding to an NF-κB probe derived from the immunoglobulin gene promoter. Both p65 and p50 antibodies supershifted the NF-κB complex in nuclear extracts of 7.5-TLR3 cells infected with HCV for 48 h (lanes 6 and 7). TNFα treatment (15 ng/ml, 2 h) was included as a positive control for NF-κB activation (lane 8)