scholarly journals Hepatitis B Virus RNA-Binding Proteins Associated with Cytokine-Induced Clearance of Viral RNA from the Liver of Transgenic Mice

1999 ◽  
Vol 73 (1) ◽  
pp. 474-481 ◽  
Author(s):  
Tilman Heise ◽  
Luca G. Guidotti ◽  
Victoria J. Cavanaugh ◽  
Francis V. Chisari
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Transgenic Mice ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Viral Rna ◽  
Cytokine Activation ◽  
Tnf Α ◽  
B Virus ◽  
Ifn Γ

ABSTRACT Hepatitis B virus (HBV) gene expression is downregulated in the liver of HBV transgenic mice by a posttranscriptional mechanism that is triggered by the local production of gamma interferon (IFN-γ) and tumor necrosis factor alpha (TNF-α) during intrahepatic inflammation (hepatitis). The molecular basis for this antiviral effect is unknown. In this study, we identified three HBV RNA-binding liver nuclear proteins (p45, p39, and p26) the relative abundance of which correlates with the abundance of HBV RNA in response to the induction of IFN-γ and TNF-α. All three proteins bind to a 91-bp element located at the 5′ end of a previously defined posttranscriptional regulatory element that is thought to mediate the nuclear export of HBV RNA. The presence of p45 correlates directly with the presence of HBV RNA, being detectable under baseline conditions when the viral RNA is abundant and undetectable when the viral RNA disappears in response to IFN-γ and TNF-α. In contrast, p26 is inversely related to HBV RNA, being detectable only when the viral RNA disappears following cytokine activation. Finally, p39 is constitutively expressed, and its abundance and mobility appear to be slightly increased by cytokine activation. These results suggest a model in which hepatocellular HBV RNA content might be controlled by the stabilizing and/or destabilizing influences of these RNA-binding proteins whose activity is regulated by cytokine-induced signaling pathways.

scholarly journals Nitric Oxide Inhibits Hepatitis B Virus Replication in the Livers of Transgenic Mice

2000 ◽  
Vol 191 (7) ◽  
pp. 1247-1252 ◽  
Author(s):  
Luca G. Guidotti ◽  
Heike McClary ◽  
Jacquelyn Moorhead Loudis ◽  
Francis V. Chisari
Keyword(s):  
Nitric Oxide ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Antiviral Activity ◽  
Transgenic Mice ◽  
Hbv Replication ◽  
Polycytidylic Acid ◽  
B Virus ◽  
Ifn Γ ◽  
Deficient Mice

We have previously identified two antiviral cytokines (interferon [IFN]-γ and IFN-α/β) that downregulate hepatitis B virus (HBV) replication in the liver of transgenic mice. The cytokine-inducible downstream events that inhibit HBV replication have not been identified. One possible factor is nitric oxide (NO), a pleiotropic free radical with antiviral activity that is produced in the liver by the inducible NO synthase (iNOS). To examine the role of NO in our model, we crossed transgenic mice that replicate HBV with mice that lack a functional iNOS. Importantly, iNOS-deficient mice were almost completely resistant to the noncytopathic inhibitory effect of HBV-specific cytotoxic T lymphocytes on viral replication, an effect that we have shown previously to depend on the intrahepatic induction of IFN-γ. Conversely, iNOS-deficient mice were not resistant to the antiviral effect of IFN-α/β induced by either polyinosinic-polycytidylic acid complex or by lymphocytic choriomeningitis virus (LCMV) infection. These results indicate that NO mediates the antiviral activity of IFN-γ, whereas the antiviral activity of IFN-α/β is NO independent. We also compared the relative sensitivity of LCMV to control by NO in these animals. Interestingly, LCMV replicated to higher levels in the liver of iNOS-deficient mice than control mice, indicating that NO controls LCMV replication in the liver, as well as HBV.

scholarly journals Cytokine-Sensitive Replication of Hepatitis B Virus in Immortalized Mouse Hepatocyte Cultures

2002 ◽  
Vol 76 (11) ◽  
pp. 5646-5653 ◽  
Author(s):  
Valérie Pasquetto ◽  
Stefan F. Wieland ◽  
Susan L. Uprichard ◽  
Marco Tripodi ◽  
Francis V. Chisari
Keyword(s):  
Gene Expression ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Intracellular Signaling ◽  
Growth Factor Receptor ◽  
Necrosis Factor Alpha ◽  
Tnf Α ◽  
B Virus ◽  
Ifn Γ

ABSTRACT We have previously shown that alpha/beta interferon (IFN-α/β) and gamma interferon (IFN-γ) inhibit hepatitis B virus (HBV) replication by eliminating pregenomic RNA containing viral capsids from the hepatocyte. We have also shown that HBV-specific cytotoxic T lymphocytes that induce IFN-γ and tumor necrosis factor alpha (TNF-α) in the liver can inhibit HBV gene expression by destabilizing preformed viral mRNA. In order to further study the antiviral activity of IFN-α/β, IFN-γ, and TNF-α at the molecular level, we sought to reproduce these observations in an in vitro system. Accordingly, hepatocytes were derived from the livers of HBV-transgenic mice that also expressed the constitutively active cytoplasmic domain of the human hepatocyte growth factor receptor (c-Met). Here, we show that the resultant well-differentiated, continuous hepatocyte cell lines (HBV-Met) replicate HBV and that viral replication in these cells is efficiently controlled by IFN-α/β or IFN-γ, which eliminate pregenomic RNA-containing capsids from the cells as they do in the liver. Furthermore, we demonstrate that IFN-γ, but not IFN-α/β, is capable of inhibiting HBV gene expression in this system, especially when it acts synergistically with TNF-α. These cells should facilitate the analysis of the intracellular signaling pathways and effector mechanisms responsible for these antiviral effects.

scholarly journals Interferon-Regulated Pathways That Control Hepatitis B Virus Replication in Transgenic Mice

2002 ◽  
Vol 76 (6) ◽  
pp. 2617-2621 ◽  
Author(s):  
Luca G. Guidotti ◽  
Amber Morris ◽  
Heike Mendez ◽  
Rick Koch ◽  
Robert H. Silverman ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Antiviral Activity ◽  
Transgenic Mice ◽  
Rnase L ◽  
Hbv Replication ◽  
Antiviral Effects ◽  
B Virus ◽  
Ifn Γ ◽  
Inducible Genes

ABSTRACT We previously showed that the intrahepatic induction of cytokines such as alpha/beta interferon (IFN-α/β) and gamma interferon (IFN-γ) inhibits hepatitis B virus (HBV) replication noncytopathically in the livers of transgenic mice. The intracellular pathway(s) responsible for this effect is still poorly understood. To identify interferon (IFN)-inducible intracellular genes that could play a role in our system, we crossed HBV transgenic mice with mice deficient in IFN regulatory factor 1 (IRF-1), the double-stranded RNA-activated protein kinase (PKR), or RNase L (RNase L) (IRF-1−/−, PKR−/−, or RNase L−/− mice, respectively), three well-characterized IFN-inducible genes that mediate antiviral activity. We showed that unmanipulated IRF-1−/− or PKR−/− transgenic mice replicate HBV in the liver at slightly higher levels than the respective controls, suggesting that both IRF-1 and PKR individually appear to mediate signals that modulate HBV replication under basal conditions. These same animals were responsive to the antiviral effects of the IFN-α/β inducer poly(I-C) or recombinant murine IFN-γ, suggesting that under these conditions, either the IRF-1 or the PKR genes can mediate the antiviral activity of the IFNs or other IFN-inducible genes mediate the antiviral effects. Finally, RNase L−/− transgenic mice were undistinguishable from controls under basal conditions and after poly(I-C) or IFN-γ administration, suggesting that RNase L does not modulate HBV replication in this model.

scholarly journals H-2 Kd-Restricted Hepatitis B Virus-Derived Epitope Whose Specific CD8+ T Lymphocytes Can Produce Gamma Interferon without Cytotoxicity

2005 ◽  
Vol 79 (9) ◽  
pp. 5568-5576 ◽  
Author(s):  
An Chen ◽  
Li Wang ◽  
Jingbo Zhang ◽  
Liyun Zou ◽  
Zhengcai Jia ◽  
...  
Keyword(s):  
T Cells ◽  
Hepatitis B Virus ◽  
T Lymphocytes ◽  
Hepatitis B ◽  
Transgenic Mice ◽  
Cytotoxic Effect ◽  
Cytokine Secretion ◽  
Target Cells ◽  
B Virus ◽  
Ifn Γ

ABSTRACT It is necessary to evaluate the cytokine secretion status of CD8+ T lymphocytes and elucidate the factors influencing cytokine secretion, because the secretion of cytokines is also an important feature of CD8+ T lymphocytes, and the cytokines usually play critical roles in the outcome of diseases. We showed here that peptide AYRPPNAPI, derived from the core antigen of hepatitis B virus (HBV), could bind to H-2 Kd and induce primed splenocytes from HBcAg expression plasmid-immunized mice to produce gamma interferon (IFN-γ) in H-2 Kd- and CD8-dependent manners instead of in a CD4-dependent manner. The induced cells were mainly CD3 and CD8 positive but had no cytotoxic effect on the corresponding target cells. When administered into HBV transgenic mice, these cells can decrease the serum HBV load without causing liver damage. These results suggest that this peptide is a special kind of CD8+ T-cell epitope, for which specific CD8+ T cells can produce IFN-γ when antigenic stimulation is encountered but which have no cytotoxic effect on the corresponding target cells both in vitro and in HBV transgenic mice. This phenomenon indicates initially that the functional mechanisms of CD8+ T cells can be determined by their epitope specificity, which may be associated with the development of epitope-based immunotherapeutic approaches for infectious diseases and tumors.

scholarly journals Hepatitis B virus Core protein nuclear interactome identifies SRSF10 as a host RNA-binding protein restricting HBV RNA production

2020 ◽  
Vol 16 (11) ◽  
pp. e1008593
Author(s):  
Hélène Chabrolles ◽  
Héloïse Auclair ◽  
Serena Vegna ◽  
Thomas Lahlali ◽  
Caroline Pons ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Core Protein ◽  
Restriction Factor ◽  
Dependent Manner ◽  
New Host ◽  
Functional Studies ◽  
B Virus

Despite the existence of a preventive vaccine, chronic infection with Hepatitis B virus (HBV) affects more than 250 million people and represents a major global cause of hepatocellular carcinoma (HCC) worldwide. Current clinical treatments, in most of cases, do not eliminate viral genome that persists as a DNA episome in the nucleus of hepatocytes and constitutes a stable template for the continuous expression of viral genes. Several studies suggest that, among viral factors, the HBV core protein (HBc), well-known for its structural role in the cytoplasm, could have critical regulatory functions in the nucleus of infected hepatocytes. To elucidate these functions, we performed a proteomic analysis of HBc-interacting host-factors in the nucleus of differentiated HepaRG, a surrogate model of human hepatocytes. The HBc interactome was found to consist primarily of RNA-binding proteins (RBPs), which are involved in various aspects of mRNA metabolism. Among them, we focused our studies on SRSF10, a RBP that was previously shown to regulate alternative splicing (AS) in a phosphorylation-dependent manner and to control stress and DNA damage responses, as well as viral replication. Functional studies combining SRSF10 knockdown and a pharmacological inhibitor of SRSF10 phosphorylation (1C8) showed that SRSF10 behaves as a restriction factor that regulates HBV RNAs levels and that its dephosphorylated form is likely responsible for the anti-viral effect. Surprisingly, neither SRSF10 knock-down nor 1C8 treatment modified the splicing of HBV RNAs but rather modulated the level of nascent HBV RNA. Altogether, our work suggests that in the nucleus of infected cells HBc interacts with multiple RBPs that regulate viral RNA metabolism. Our identification of SRSF10 as a new anti-HBV restriction factor offers new perspectives for the development of new host-targeted antiviral strategies.

scholarly journals Interleukin-18 Inhibits Hepatitis B Virus Replication in the Livers of Transgenic Mice

2002 ◽  
Vol 76 (21) ◽  
pp. 10702-10707 ◽  
Author(s):  
Kiminori Kimura ◽  
Kazuhiro Kakimi ◽  
Stefan Wieland ◽  
Luca G. Guidotti ◽  
Francis V. Chisari
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Transgenic Mice ◽  
Nk Cells ◽  
Natural Killer ◽  
Nkt Cells ◽  
Interleukin 18 ◽  
Hbv Replication ◽  
B Virus ◽  
Ifn Γ

ABSTRACT Interleukin-18 (IL-18) produced by activated antigen-presenting cells stimulates natural killer (NK) cells, natural killer T (NKT) cells, and T cells to secrete gamma interferon (IFN-γ). In this study, injection of a single 10-μg dose of recombinant murine IL-18 rapidly, reversibly, and noncytopathically inhibited hepatitis B virus (HBV) replication in the livers of HBV transgenic mice. Furthermore, HBV replication was inhibited by as little as 1 μg of IL-18 injected repetitively, and also by a single 0.1-μg dose of IL-18 injected together with 1 ng of IL-12, neither of which inhibited HBV replication individually, demonstrating synergy between these cytokines in this system. The antiviral effect of IL-18 was mediated by its ability to activate resident intrahepatic NK cells and NKT cells to produce IFN-γ and by its ability to induce IFN-α/β production in the liver. These results suggest that IL-18 has the potential to contribute to the control of HBV replication during self-limited infection and that it may have therapeutic value for the treatment of patients with chronic hepatitis.

scholarly journals Inhibition of Hepatitis B Virus Replication during Schistosoma mansoni Infection in Transgenic Mice

2000 ◽  
Vol 192 (2) ◽  
pp. 289-294 ◽  
Author(s):  
Heike McClary ◽  
Rick Koch ◽  
Francis V. Chisari ◽  
Luca G. Guidotti
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Transgenic Mice ◽  
Schistosoma Mansoni ◽  
Antiviral Effect ◽  
Hbv Replication ◽  
Frequent Event ◽  
B Virus ◽  
Helper Cell Type ◽  
Ifn Γ

Although coinfection of hepatitis B virus (HBV) and Schistosoma mansoni is a frequent event in humans, little is known about the interactions between these two pathogens. S. mansoni infection induces T helper cell type 2 (Th2)–type cytokines in the liver of humans and mice. The intrahepatic induction of nitric oxide (NO) and Th1-type cytokines, such as interferon (IFN)-γ and IFN-α/β, inhibits HBV replication noncytopathically in the liver of transgenic mice. To examine whether S. mansoni infection and the accompanying induction of Th2-type cytokines could interfere with HBV replication in the liver, HBV transgenic mice were infected with S. mansoni. By 5 wk after infection, HBV replication disappeared concomitant with the intrahepatic induction of NO and Th1-type cytokines, and in the absence of Th2-type cytokines. By 6–8 wk after infection, HBV replication remained undetectable and this was associated with further induction of NO and Th1-type cytokines together with the appearance of Th2-type cytokines. The S. mansoni–dependent antiviral effect was partially blocked by genetically deleting IFN-γ, although it was unaffected by deletion of IFN-α/β. These results indicate that IFN-γ (probably via NO) mediates most of this antiviral activity and that Th2-type cytokines do not counteract the antiviral effect of IFN-γ. Similar events may suppress HBV replication during human S. mansoni infection.

scholarly journals Natural Killer T Cell Activation Inhibits Hepatitis B Virus Replication in Vivo

2000 ◽  
Vol 192 (7) ◽  
pp. 921-930 ◽  
Author(s):  
Kazuhiro Kakimi ◽  
Luca G. Guidotti ◽  
Yasuhiko Koezuka ◽  
Francis V. Chisari
Keyword(s):  
Immune Response ◽  
T Cells ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Transgenic Mice ◽  
Nkt Cells ◽  
Innate Immune ◽  
Hbv Replication ◽  
B Virus ◽  
Ifn Γ

We have previously reported that hepatitis B virus (HBV)–specific CD8+ cytotoxic T lymphocytes and CD4+ helper T lymphocytes can inhibit HBV replication in the liver of HBV transgenic mice by secreting interferon (IFN)-γ when they recognize viral antigen. To determine whether an activated innate immune system can also inhibit HBV replication, in this study we activated natural killer T (NKT) cells in the liver of HBV transgenic mice by a single injection of α-galactosylceramide (α-GalCer), a glycolipid antigen presented to Vα14+NK1.1+ T cells by the nonclassical major histocompatibility complex class I–like molecule CD1d. Within 24 h of α-GalCer injection, IFN-γ and IFN-α/β were detected in the liver of HBV transgenic mice and HBV replication was abolished. Both of these events were temporally associated with the rapid disappearance of NKT cells from the liver, presumably reflecting activation-induced cell death, and by the recruitment of activated NK cells into the organ. In addition, prior antibody-mediated depletion of CD4+ and CD8+ T cells from the mice did not diminish the ability of α-GalCer to trigger the disappearance of HBV from the liver, indicating that conventional T cells were not downstream mediators of this effect. Finally, the antiviral effect of α-GalCer was inhibited in mice that are genetically deficient for either IFN-γ or the IFN-α/β receptor, indicating that most of the antiviral activity of α-GalCer is mediated by these cytokines. Based on these results, we conclude that α-GalCer inhibits HBV replication by directly activating NKT cells and by secondarily activating NK cells to secrete antiviral cytokines in the liver. In view of these findings, we suggest that, if activated, the innate immune response, like the adaptive immune response, has the potential to control viral replication during natural HBV infection. In addition, the data suggest that therapeutic activation of NKT cells may represent a new strategy for the treatment of chronic HBV infection.

scholarly journals Indispensable Role for TNF-α and IFN-γ at the Effector Phase of Liver Injury Mediated by Th1 Cells Specific to Hepatitis B Virus Surface Antigen

2000 ◽  
Vol 165 (2) ◽  
pp. 956-961 ◽  
Author(s):  
Akio Ohta ◽  
Masashi Sekimoto ◽  
Marimo Sato ◽  
Toshiaki Koda ◽  
Shin-ichiro Nishimura ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Liver Injury ◽  
Hepatitis B ◽  
Surface Antigen ◽  
Th1 Cells ◽  
Virus Surface ◽  
Effector Phase ◽  
Tnf Α ◽  
B Virus ◽  
Ifn Γ

scholarly journals Concentrations of Cytokines, Soluble Interleukin-2 Receptor, and Soluble CD30 in Sera of Patients with Hepatitis B Virus Infection during Acute and Convalescent Phases

2002 ◽  
Vol 9 (6) ◽  
pp. 1372-1375 ◽  
Author(s):  
Francisca Monsalve-de Castillo ◽  
Tania A. Romero ◽  
Jesús Estévez ◽  
Luciana L. Costa ◽  
Ricardo Atencio ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Acute Phase ◽  
Surface Antigen ◽  
Interleukin 2 ◽  
Soluble Form ◽  
Convalescent Phase ◽  
Tnf Α ◽  
B Virus ◽  
Ifn Γ

ABSTRACT The immunoregulatory roles of interleukin-2 (IL-2), IL-4, IL-10, gamma interferon (IFN-γ), tumor necrosis factor alpha (TNF-α), the soluble form of the IL-2 receptor (sIL-2R), and the soluble form of CD30 (sCD30) were evaluated in patients with hepatitis B virus (HBV) infection. Two groups of subjects were studied: 15 healthy individuals without hepatitis antecedents and 15 patients with HBV infection. Blood samples were taken during the acute and convalescent phases. The analysis of the samples was done by the enzyme-linked immunosorbent assay technique. IFN-γ and TNF-α levels decreased in the convalescent phase. IL-10, IL-2, and sIL-2R levels increased in the acute and convalescent phases, while sCD30 levels increased during the acute phase. The IL-4 concentrations decreased in both phases. During the acute phase, IFN-γ and TNF-α induced increases in IL-2, sIL-2R, IL-10, and sCD30 levels in serum, which allowed the development of immunity characterized by the nonreactivity of the HBV surface antigen, the onset of antibodies to the HBV surface antigen (anti-HBs), and normal alanine aminotransferase levels during the convalescent phase. Increased IL-2 levels during the acute phase would stimulate the activities of NK cells and CD8+ lymphocytes, which are responsible for viral clearing. The raised sIL-2R levels reveal activation of T lymphocytes and control of the IL-2-dependent immune response. The sCD30 increment during the acute phase reflects the greater activation of the Th2 cellular phenotype. Its decrease in the convalescent phase points out the decrease in the level of HBV replication. The increase in IL-10 levels could result in a decrease in IL-4 levels and modulate IFN-γ and TNF-α levels during both phases of disease, allowing the maintenance of anti-HBs concentrations.

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