No abstract

Hepatitis E is the most common type of acute hepatitis prevalent worldwide. The open reading frame 3 protein of HEV (HEV ORF3) is proposed to create a favorable environment for viral replication and pathogenesis. However, the mechanisms by which HEV overcomes the effects of host immunity, particularly the role of ORF3, remain to be established. Expression of IFNα and IFNβ in supernatant and cell samples was examined via ELISA and quantitative RT-PCR. The protein levels of specific signaling factors in cells overexpressing HEV ORF3 were examined via western blot. Analyses of cells transfected with vectors expressing ORF3 demonstrated that HEV ORF3 significantly impairs the generation of endogenous type I interferon through downregulating TLR3 and TLR7 as well as their corresponding downstream signaling pathways. Moreover, inhibition of NFκB, JAK/STAT and JNK/MAPK signaling pathways contributed significantly to suppression of increased levels of TLR7. Levels of p-P65, p-STAT1 and p-JNK were markedly impaired in ORF3-expressing cells, even upon treatment with the respective agonists. HEV ORF3 inhibits the production of endogenous type I interferon through downregulation of TLR3 and TLR7. Furthermore, suppression of TLR7 is achieved through impairment of multiple signaling pathways, including NFκB, JAK/STAT and JNK/MAPK.

Hepatitis B virus (HBV)has four open reading frames (ORFs) of which ORF C is consists of the pre Core and Core genes encodes the Hepatitis B core antigen (HBcAg) and Hepatitis B e antigen (HBeAg). Studies have shown that HBeAg significantly inhibits the NLRP3 inflammasome activation and interleukin-1β (IL-1β) production.However, the role of HBcAg and ORF C proteins (in this paper, ORF C proteins = HBcAg + HBeAg) were remain unclear. Our study aims to assess whether HBcAg and ORF C proteins can affect the NLRP3 inflammasome pathway. Vectors expressing ORF C proteins and HBcAg were designed and transfected into HepG2 cells. And then, cells were stimulated with lipopolysaccharide (LPS). Activation of the NLRP3 inflammasome and the levels of IL-1β and IL-18 were evaluated by Western blot analysis, quantitative reverse-transcription polymerase chain reaction, enzyme-linked immunosorbent assay, and immunofluorescence. The expression of NLRP3 and IL-1β peaked when HepG2 cells were stimulated with 1000 ng/mL LPS for 18 to 24 hours.HBcAg, but not ORF C proteins, promoted LPS-induced NLRP3 inflammasome activation and IL-1β production. These findings provide a novel mechanism on how the HBV causes liver inflammation and may provide insights into the search for new therapeutic strategies. K E Y W O R D Shepatitis B core antigen, interleukin-18, interleukin-1β, NLRP3, ORF C proteins

Background & Aims: Hepatitis B virus (HBV) infection is the most critical factor underlying liver cirrhosis and hepatocellular carcinoma worldwide. IL-1β and IL-18, generated by activation of the inflammasome/caspase-1 signaling pathway, play important roles in the control and clearance of HBV. However, the specific relationship between the inflammasome response and IFN-α resistance or viral persistence is yet to be established. Methods: Blood samples of patients and supernatant fractions of HBV cell lines were collected for analysis and the effects on inflammasome activation and IL-1β production evaluated via enzyme-linked immunosorbent assay (ELISA), western blot, quantitative RT-PCR and immunofluorescence. Results: IL-1β and IL-18 levels produced in sera of IFN-α non-responders were significantly lower than those of responders and normal donors. Additionally, expression of IL-1β and inflammasome components was decreased in peripheral blood mononuclear cells (PBMC) of non-responders, compared with those of responders. In vitro experiments on HepG2, HepG2.2.15 and HepAD38 cell lines showed that HBV induces a significant decrease in IL-1β production through inhibiting activation of the NF-κB signaling and inflammasome/caspase-1 pathways. And hepatitis B virus polymerase (HBV-Pol) appeared crucial for these inhibitory effects of HBV.Conclusion: IL-1β production is suppressed in HBV carriers and IFN-α non-responders. HBV induces a significant decrease in IL-1β production through inhibiting the NF-κB signaling and inflammasome pathways, for which HBV-Pol is a crucial requirement.