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534
The e Antigen Is an Interferon Resistance
Protein That Allows the Hepatitis B Virus to
Evade the Early Immune Response.
Zina Valaydon1, Gregor Ebert2, Liana Mackiewicz3, Simon
Preston4, Paul V. Desmond5, Vitina Sozzi6, Peter Revill7,
Alexander J. V. Thompson8 and Marc Pellegrini2, (1)
Gastroenterology, St Vincent’s Hospital, Melbourne, (2)Walter
and Eliza Hall Institute, (3)Walter an Eliza Hall Institute, (4)
Wehi, (5)Gastroenterology, St Vincent’s Hospital Melbourne,
(6)Victorian Infectious Disease Reference Laboratory, (7)The
University of Melbourne, (8)Department of Gastroenterology,
St. Vincent’s Hospital
Background: The precore antigen, HBeAg, is thought
to be a tolerogen in the natural history of Hepatitis B virus
(HBV) infection. However, the exact underlying immune
mechanisms are unknown. The basal core promoter (BCP)
and precore (PC) variants of the hepatitis B reduce and
abolish HBeAg production, respectively, and are associated
with increased risk of cirrhosis and hepatocellular carcinoma.
We used a novel immunocompetent mouse model of HBV
infection to study viral kinetics and immunological differences
between wild type (WT) virus and HBV variants with little
or no HBeAg.Methods: An HBV 1.2 overlength genome
was flanked by adeno-associated virus inverted terminal
repeats within a plasmid vector. The genome was mutated
using site-directed mutagenesis to introduce dual A1762T/
G1764A BCP and G1896A PC mutations. WT, PC and
BCP plasmids were hydrodynamically injected in the tail
vein of C57/BL6 mice. Interferon-alpha receptor knockout
(IFNaR-KO) and interferon-gamma knockout mice were
used to investigate the role of Type 1 interferon (IFN) and
IFN-gamma respectively. To investigate the role of tumour
necrosis factor-alpha (TNF-a), mice were treated with TNF-a
neutralizing antibodies. Viral DNA was extracted from serum
and quantitated by qPCR and serology was performed.
Transcriptomic and proteomic studies were carried out on
liver tissue to determine differences in expression profile
between mutants and WT. Results: Significant differences
in viral kinetics were seen with the BCP and PC mutants
vs. WT (Fig1a). There was a rapid drop in viral load (VL) in
the mutants in the early phase of infection in weeks 1-7 (p
<0.03 vs. WT at week 2-7). ALT levels were also significantly
increased in mutants vs WT (Mean ALT 128 IU/ml vs 40 IU/
ml, p<0.02). To further interrogate the phenomenon a series
of immune experiments were performed. The experiment was
then performed in IFNaR-KO mice. There was a significant
increase in VL in both PC and BCP mutants (p <0.05) and the
early difference in VL between mutant and WT strains was
abolished when the influence of IFN-a was removed (Fig 1b).
No effect was seen with the neutralization of The IFN-gamma
or TNF pathways. Transcriptomic study of liver tissue showed
that there is a significant upregulation of the Type 1 interferon
pathway and gene signatures in HBeAg negative mutants vs.
WT (P=0.04)- Fig 2. Proteomic studies show that interferon
related proteins are significantly overexpressed in HBeAg
negative mutants (Fig 3). Conclusion: These studies shows
that in Type 1 IFN is a key mediator of the early innate immune
response and that in absence of HBeAg, HBV is much more
susceptible to Type 1 IFN. This is supported by viral kinetic
differences, immune studies as well as transcriptomic and
proteomics data. Collectively our studies show that HBeAg is
an IFN resistance protein.
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发表于 2018-10-24 19:45
