FIGURE 2.

NF-κB activation triggered by IL-1 and TNFα was critical for anti-HBV activity. A–D, F, and I, HepaRG cells were left untreated (control) or treated with varying concentrations of IL-1β (1, 10, 30, and 100 ng/ml) or 25 units/ml heparin (A), with 30 ng/ml IL-1β together with or without a neutralizing anti-IL-1RI antibody at 20 μg/ml (B), with 10 ng/ml IL-1β or varying concentrations of IL-1Ra (10, 30, and 100 ng/ml) (C), with 3 ng/ml IL-1β together with or without PD98059, SP600125, SB203580, or Bay11-7082 (D), or QNZ or BMS-345541 (F), or with TNFα (10, 100, and 300 ng/ml) (I) according to the treatment schedule shown in Fig. 1A. HBV infection was monitored by HBs protein secretion into the medium in A, C, D, F, and I and with HBc protein in the cells in B. E, G, and H, NF-κB (E and G) and ISRE activity (H) were measured by reporter assay in the cells transfected with the reporter plasmid expressing luciferase driven from five tandem repeats of NF-κB elements (E, upper graph, and G) or ISRE (H, upper graph) or by RT-PCR in the cells (E and H, lower panels) upon signaling inhibitors used in D and F together with or without IL-1β (E and G), or upon IL-1β (10, 30, and 100 ng/ml) or IFNα 100 IU/ml as a positive control (H) for 6 h. The white and black bars in the upper graph of E and G show the data in the absence or presence of IL-1β, respectively. Bands for mRNA for cIAP, XIAP, and GAPDH (E) or ISG56, PKR, or GAPDH (H) are presented in the lower panels. All of the data are based on averages of three independent experiments.