RESUMO
BACKGROUND & AIMS: Conflicting results have been reported regarding the impact of hepatitis B virus X protein (HBx) expression on liver regeneration triggered by partial hepatectomy (PH). In the present report we investigated the mechanisms by which HBx protein alters hepatocyte proliferation after PH. METHODS: PH was performed on a transgenic mouse model in which HBx expression is under the control of viral regulatory elements and liver regeneration was monitored. LPS, IL-6 neutralizing antibody, and SB203580 were injected after PH to evaluate IL-6 participation during liver regeneration. RESULTS: Cell cycle progression of hepatocytes was delayed in HBx transgenic mice compared to WT animals. Moreover, HBx induced higher secretion of IL-6 soon after PH. Upregulation of IL-6 was associated with an elevation of STAT3 phosphorylation, SOCS3 transcript accumulation and a decrease in ERK1/2 phosphorylation in the livers of HBx transgenic mice. The involvement of IL-6 overexpression in cell cycle deregulation was confirmed by the inhibition of liver regeneration in control mice after the upregulation of IL-6 expression using LPS. In addition, IL-6 neutralization with antibodies was able to restore liver regeneration in HBx mice. Finally, the direct role of p38 in IL-6 secretion after PH was demonstrated using SB203580, a pharmacological inhibitor. CONCLUSIONS: HBx is able to induce delayed hepatocyte proliferation after PH, and HBx-induced IL-6 overexpression is involved in delayed liver regeneration. By modulating IL-6 expression during liver proliferation induced by stimulation of the cellular microenvironment, HBx may participate in cell cycle deregulation and progression of liver disease.
Assuntos
Interleucina-6/fisiologia , Regeneração Hepática/fisiologia , Transativadores/fisiologia , Animais , Anticorpos Neutralizantes/administração & dosagem , Ciclo Celular , Proliferação de Células , Elementos Facilitadores Genéticos , Hepatectomia , Vírus da Hepatite B/genética , Vírus da Hepatite B/patogenicidade , Vírus da Hepatite B/fisiologia , Hepatite B Crônica/imunologia , Hepatite B Crônica/patologia , Hepatite B Crônica/virologia , Hepatócitos/imunologia , Hepatócitos/patologia , Hepatócitos/virologia , Interações Hospedeiro-Patógeno , Humanos , Imidazóis/administração & dosagem , Interleucina-6/antagonistas & inibidores , Regeneração Hepática/genética , Regeneração Hepática/imunologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Modelos Animais , Regiões Promotoras Genéticas , Piridinas/administração & dosagem , Transativadores/genética , Proteínas Virais Reguladoras e AcessóriasRESUMO
Current treatments for HBV chronic carriers using interferon alpha or nucleoside analogues are not effective in all patients and may induce the emergence of HBV resistant strains. Bay 41-4109, a member of the heteroaryldihydropyrimidine family, inhibits HBV replication by destabilizing capsid assembly. The aim of this study was to determine the antiviral effect of Bay 41-4109 in a mouse model with humanized liver and the spread of active HBV. Antiviral assays of Bay 41-4109 on HepG2.2.15 cells constitutively expressing HBV, displayed an IC(50) of about 202 nM with no cell toxicity. Alb-uPA/SCID mice were transplanted with human hepatocytes and infected with HBV. Ten days post-infection, the mice were treated with Bay 41-4109 for five days. During the 30 days of follow-up, the HBV load was evaluated by quantitative PCR. At the end of treatment, decreased HBV viremia of about 1 log(10) copies/ml was observed. By contrast, increased HBV viremia of about 0.5 log(10) copies/ml was measured in the control group. Five days after the end of treatment, a rebound of HBV viremia occurred in the treated group. Furthermore, 15 days after treatment discontinuation, a similar expression of the viral capsid was evidenced in liver biopsies. Our findings demonstrate that Bay 41-4109 displayed antiviral properties against HBV in humanized Alb-uPA/SCID mice and confirm the usefulness of Alb-uPA/SCID mice for the evaluation of pharmaceutical compounds. The administration of Bay 41-4109 may constitute a new strategy for the treatment of patients in escape from standard antiviral therapy.
Assuntos
Albuminas/metabolismo , Antivirais/farmacologia , Vírus da Hepatite B/metabolismo , Hepatite B/tratamento farmacológico , Piridinas/farmacologia , Pirimidinas/farmacologia , Animais , Biópsia/métodos , DNA Viral/metabolismo , Hepatócitos/citologia , Humanos , Imuno-Histoquímica/métodos , Cinética , Fígado/metabolismo , Fígado/virologia , Camundongos , Camundongos SCID , Carga ViralRESUMO
BACKGROUND: Treatment of HBV chronic carriers using interferon (IFN)-α or nucleoside/nucleotide analogues fails to suppress viral infection. Type-II IFN-γ has been shown to inhibit HBV replication. The goal of the present work was to evaluate the antiviral efficacy against HBV of a thermostable IFN-γ variant isolated using Massive Mutagenesis and thermoresistant selection (THR) technologies. METHODS: The thermostability of wild-type (wt) and S63C IFN-γ was determined in vitro and in vivo. Activation of the IFN-γ responsive element by wt and S63C IFN-γ was tested using a luciferase assay. HepG2.2.15 cells constitutively expressing HBV were used to analyse the antiviral activity of wt and S63C IFN-γ against HBV replication. Intracellular HBV DNA was detected by Southern blot and quantified by real-time PCR analyses. RESULTS: S63C IFN-γ was shown to be more thermostable and had a longer half-life than wt IFN-γ. Both wt and S63C IFN-γ displayed a similar capacity to activate the IFN pathway. The treatment of HepG2.2.15 cells with wt or S63C IFN-γ induced the inhibition of HBV viral replication. After heating, S63C IFN-γ displayed better conservation of its antiviral activity against HBV when compared with wt IFN-γ. CONCLUSIONS: These results confirm that the THR method can be used to isolate mutants with enhanced thermostability and demonstrate that a thermostable IFN-γ variant presents antiviral properties against HBV replication. This molecule could provide a new strategy to treat patients who do not respond to antiviral therapy.