Background The non-structural (NS) 3/4A protease/helicase of the hepatitis C virus is known to modulate signalling pathways in the infected hepatocyte by cleaving CARD adaptor inducing IFNβ (Cardif), T-cell protein tyrosine phosphatase (TC-PTP) and TIR domain-containing adaptor inducing IFNβ (TRIF), but the effects of NS3/4A in vivo still remain unclear.
Aim To investigate the influence of NS3/4A on intracellular and intercellular signalling in vivo by analysing the intrahepatic inflammatory response of naïve, lipopolysaccharide (LPS)/d-galactosamine (d-galN) or tumour necrosis factor-α (TNFα)/d-galN-treated NS3/4A-transgenic (Tg) mice.
Methods The intrahepatic immunity of naïve and LPS/d-galN- or TNFα/d-galN-treated NS3/4A-Tg mice was determined using western blot, ELISA, real-time PCR, flow cytometry and survival monitoring. The injection of cytokines or antibodies against signalling components was performed to analyse the relevance of the respective pathways for the investigated issues. A Tg mouse lineage expressing an inactivated NS3/4A protease (NS3/4AIle1073Ala-Tgs) was generated to examine if protective effects were NS3/4A protease dependent.
Results The activation of hepatic signal transducer and activator of transcription 1 and 2 was impaired in NS3/4A-Tg mice after treatment with LPS/d-galN or TNFα/d-galN. This was paralleled by a reduction in hepatic interferon-γ (IFNγ). Reconstitution of IFNγ reverted the resistance to LPS/TNFα in NS3/4A-Tg mice. Subsequently, blocking IFNγ in vivo rendered wild-type mice resistant against treatment with LPS/TNFα. A new Tg mouse expressing an inactivated NS3/4A protease had the same phenotype as wild-type mice with respect to hepatic IFNγ levels and sensitivity to LPS/d-galN. Finally, the chemokine profile was altered in the NS3/4A-Tg mice towards an anti-inflammatory state, which helps to explain the altered immune cell subsets and reduction in hepatic IFNγ production.
Conclusions Our data demonstrate that the NS3/4A protease reduces the intrahepatic production of IFNγ and alters TNFα-mediated effects, thereby impairing the hepatic inflammatory response. This may contribute to viral persistence.
- Viral hepatitis
- liver injury
- hepatitis C
- liver immunology
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Funding This work was supported by the Swedish Cancer Society (MS), the Swedish Research Council (MS) and the Stockholm County Council (MS). EDB was supported by grants from the Ruth and Richard Juhlin Foundation, the Goljes Memorial Fund, the Lars Hierta Memorial Foundation, the Swedish Royal Academy of Sciences, the Professor Nanna Svartz Fund and Karolinska Institutet. JS was supported by grants from the Erik and Edith Fernström's Foundation, Foundation Blanceflor Boncompagni-Ludovisi and the Filip Lundberg's Foundation. LF was supported by grants from the Swedish Society of Medical Research, the Swedish Society of Medicine, the Goljes Memorial Fund, the Åke Wiberg Foundation, the Royal Swedish Academy of Sciences, Foundation Längmanska Kulturfonden and Karolinska Institutet. JGB was supported by grants from the Deutsche Forschungsgemeinschaft through FOR 729, SFB 575, Heisenberg-Program and GK 1045.
Competing interests None to declare.
Ethics approval This study was conducted with the approval from the local committee on human research ethics and the Swedish Medical Products Agency (Läkemedelsverket; approved on January 20, 2006, with the approval number 2005-1311-31/3).
Provenance and peer review Not commissioned; externally peer reviewed.