scholarly journals Apoptosis of Hepatitis B Virus-Infected Hepatocytes Prevents Release of Infectious Virus

2010 ◽  
Vol 84 (22) ◽  
pp. 11994-12001 ◽  
Author(s):  
Silke Arzberger ◽  
Marianna Hösel ◽  
Ulrike Protzer
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Inverse Correlation ◽  
Infection Model ◽  
Virus Particles ◽  
Hbv Replication ◽  
Culture Model ◽  
B Virus ◽  
A Cell ◽  
Infected Cells

ABSTRACT Apoptosis of infected cells is critically involved in antiviral defense. Apoptosis, however, may also support the release and spread of viruses. Although the elimination of infected hepatocytes is required to combat hepatitis B virus (HBV) infection, it is still unknown which consequences hepatocyte apoptosis has for the virus and whether or not it is advantageous to the virus. To study this, we designed a cell culture model consisting of both HBV-producing cell lines and primary human hepatocytes serving as an infection model. We showed that the release of mature, enveloped virions was 80% to 90% reduced 24 h after the induction of apoptosis in HBV-replicating hepatoma cells or HBV-infected hepatocytes. Importantly, HBV particles released from apoptotic hepatocytes were immature and nonenveloped and proved not to be infectious. We found an inverse correlation between the strength of an apoptotic stimulus and the infectivity of the virus particles released: the more potent the apoptotic stimulus, the higher the ratio of nonenveloped capsids to virions and the lower their infectivity. Furthermore, we demonstrated that HBV replication and, particularly, the expression of the HBx protein transcribed from the viral genome during replication do not sensitize cells to apoptosis. Our data clearly reject the hypothesis that the apoptosis of infected hepatocytes facilitates the propagation of HBV. Rather, these data indicate that HBV needs to prevent the apoptosis of its host hepatocyte to ensure the release of infectious progeny and, thus, virus spread in the liver.

scholarly journals 3-Hydroxypyrimidine-2,4-Diones as Novel Hepatitis B Virus Antivirals Targeting the Viral Ribonuclease H

2017 ◽  
Vol 61 (6) ◽  
Author(s):  
Andrew D. Huber ◽  
Eleftherios Michailidis ◽  
Jing Tang ◽  
Maritza N. Puray-Chavez ◽  
Maria Boftsi ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Antiviral Activity ◽  
Essential Role ◽  
Rnase H ◽  
Ribonuclease H ◽  
Hbv Replication ◽  
B Virus ◽  
Infected Cells ◽  
Micromolar Range

ABSTRACT Hepatitis B virus (HBV) RNase H (RNH) is an appealing therapeutic target due to its essential role in viral replication. RNH inhibitors (RNHIs) could help to more effectively control HBV infections. Here, we report 3-hydroxypyrimidine-2,4-diones as novel HBV RNHIs with antiviral activity. We synthesized and tested 52 analogs and found 4 that inhibit HBV RNH activity in infected cells. Importantly, 2 of these compounds inhibited HBV replication in the low micromolar range.

scholarly journals Replenishment of Hepatitis B Virus cccDNA Pool Is Restricted by Baseline Expression of Host Restriction Factors In Vitro

2019 ◽  
Vol 7 (11) ◽  
pp. 533 ◽  
Author(s):  
Sergey Brezgin ◽  
Anastasiia Kostyusheva ◽  
Ekaterina Bayurova ◽  
Ilya Gordeychuk ◽  
Maria Isaguliants ◽  
...  
Keyword(s):  
Dna Damage ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Real Time ◽  
Real Time Pcr ◽  
Hbv Replication ◽  
Hbv Cccdna ◽  
B Virus ◽  
Infected Cells ◽  
Cell Cycling

Background: Covalently closed circular DNA (cccDNA) of hepatitis B virus (HBV) is the major cause of viral persistence in patients with chronic HBV infection. Understanding the mechanisms underlying stability and persistence of HBV cccDNA in hepatocytes is critical for developing novel therapeutics and managing chronic hepatitis B. In this study, we observed an unexpected increase in HBV cccDNA levels upon suppression of transcription by de novo DNA methyltransferase DNMT3A and uncovered additional mechanisms potentially involved in HBV cccDNA maintenance. Methods: HBV-expressing cell lines were transfected with a DNMT3A-expressing plasmid. Real-time PCR and HBsAg assays were used to assess the HBV replication rate. Cell cycling was analyzed by fluorescent cell sorting. CRISPR/Cas9 was utilized to abrogate expression of APOBEC3A and APOBEC3B. Alterations in the expression of target genes were measured by real-time PCR. Results: Similar to previous studies, HBV replication induced DNMT3A expression, which in turn, led to reduced HBV transcription but elevated HBV cccDNA levels (4- to 6-fold increase). Increased levels of HBV cccDNA were not related to cell cycling, as DNMT3A accelerated proliferation of infected cells and could not contribute to HBV cccDNA expansion by arresting cells in a quiescent state. At the same time, DNMT3A suppressed transcription of innate immunity factors including cytidine deaminases APOBEC3A and APOBEC3B. CRISPR/Cas9-mediated silencing of APOBEC3A and APOBEC3B transcription had minor effects on HBV transcription, but significantly increased HBV cccDNA levels, similar to DNMT3A. In an attempt to further analyze the detrimental effects of HBV and DNMT3A on infected cells, we visualized γ-H2AX foci and demonstrated that HBV inflicts and DNMT3A aggravates DNA damage, possibly by downregulating DNA damage response factors. Additionally, suppression of HBV replication by DNMT3A may be related to reduced ATM/ATR expression. Conclusion: Formation and maintenance of HBV cccDNA pools may be partially suppressed by the baseline expression of host inhibitory factors including APOBEC3A and APOBEC3B. HBV inflicts DNA damage both directly and by inducing DNMT3A expression.

scholarly journals Hepatitis B Virus-Mediated Changes of Apolipoprotein mRNA Abundance in Cultured Hepatoma Cells

2003 ◽  
Vol 77 (9) ◽  
pp. 5503-5506 ◽  
Author(s):  
Pamela A. Norton ◽  
Qiaoke Gong ◽  
Anand S. Mehta ◽  
Xuanyong Lu ◽  
Timothy M. Block
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hepatoma Cell ◽  
Inverse Correlation ◽  
Apolipoprotein Ai ◽  
Hepatoma Cell Lines ◽  
B Virus ◽  
Infected Cells ◽  
Steady State Levels ◽  
Third Line

ABSTRACT An inverse correlation between hepatitis B virus (HBV) and steady-state levels of apolipoprotein AI and CIII mRNAs was observed in two hepatoma cell lines. Analysis of a third line containing an inducible viral genome implicated viral pregenomic RNA in apolipoprotein mRNA reduction. We conclude that HBV alters infected cells despite the absence of overt cytopathogenicity.

scholarly journals CRISPR/Cas9 delivery by NIR-responsive biomimetic nanoparticles for targeted HBV therapy

2022 ◽  
Vol 20 (1) ◽  
Author(s):  
Dan Wang ◽  
Ling Chen ◽  
Chengbi Li ◽  
Quanxin Long ◽  
Qing Yang ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
High Efficiency ◽  
Efficient Treatment ◽  
Hbv Replication ◽  
Hbv Cccdna ◽  
Potential System ◽  
B Virus ◽  
Biomimetic Nanoparticles ◽  
Infected Cells

Abstract Background Currently, there are no curative drugs for hepatitis B virus (HBV). Complete elimination of HBV covalently closed circular DNA (cccDNA) is key to the complete cure of hepatitis B virus infection. The CRISPR/Cas9 system can directly destroy HBV cccDNA. However, a CRISPR/Cas9 delivery system with low immunogenicity and high efficiency has not yet been established. Moreover, effective implementation of precise remote spatiotemporal operations in CRISPR/Cas9 is a major limitation. Results In this work, we designed NIR-responsive biomimetic nanoparticles (UCNPs-Cas9@CM), which could effectively deliver Cas9 RNP to achieve effective genome editing for HBV therapy. HBsAg, HBeAg, HBV pgRNA and HBV DNA along with cccDNA in HBV-infected cells were found to be inhibited. These findings were confirmed in HBV-Tg mice, which did not exhibit significant cytotoxicity and minimal off-target DNA damage. Conclusions The UCNPs-based biomimetic nanoplatforms achieved the inhibition of HBV replication via CRISPR therapy and it is a potential system for efficient treatment of human HBV diseases. Graphical Abstract

scholarly journals Experimental transfection of Macaca sylvanus with cloned human hepatitis B virus

2002 ◽  
Vol 83 (7) ◽  
pp. 1645-1649 ◽  
Author(s):  
Tarik Gheit ◽  
Souad Sekkat ◽  
Lucyna Cova ◽  
Michèle Chevallier ◽  
Marie Anne Petit ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Hbv Dna ◽  
Macaca Sylvanus ◽  
Primate Model ◽  
Virus Particles ◽  
Hbv Replication ◽  
Post Transfection ◽  
B Virus ◽  
Dane Particles

Due to the absence of easily accessible animal models for the study of hepatitis B virus (HBV), the possibility of using Macaca sylvanus, a monkey originating from Morocco, North Africa, was investigated. Three monkeys were intrahepatically inoculated with a replication-competent head-to-tail HBV DNA plasmid dimer construct. The HBV surface antigen and HBV DNA were detected prior to alanine aminotransferase elevation in the serum of two of three HBV-inoculated monkeys at day 2 post-transfection and persisted for several weeks. This indicates that transfected animals developed markers of HBV infection. In addition, electron microscopy of the serum 3 weeks post-transfection showed the presence of virus particles whose shape and size were similar to complete 42 nm HBV Dane particles. Histological examination of liver tissues also revealed pathological changes not observed in uninfected controls, which strongly suggested acute hepatitis. HBV DNA was also detected by PCR in these monkey livers. Taken together, these results indicate that HBV can successfully replicate in this model and that M. sylvanus could be a potentially useful new primate model for the study of HBV replication.

scholarly journals A cell surface protein that binds avian hepatitis B virus particles.

1994 ◽  
Vol 68 (4) ◽  
pp. 2091-2096 ◽  
Author(s):  
K Kuroki ◽  
R Cheung ◽  
P L Marion ◽  
D Ganem
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Cell Surface ◽  
Surface Protein ◽  
Cell Surface Protein ◽  
Virus Particles ◽  
B Virus ◽  
A Cell

In vitro Anti-Human Immunodeficiency Virus and Anti-Hepatitis B Virus Activities and Pharmacokinetic Properties of Heterodinucleoside Phosphates Containing AZT or ddC

1998 ◽  
Vol 9 (2) ◽  
pp. 117-126 ◽  
Author(s):  
PA Peghini ◽  
R Zahner ◽  
H Kuster ◽  
H Schott ◽  
RA Schwendener
Keyword(s):  
Human Immunodeficiency Virus ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Blood Levels ◽  
Hbv Replication ◽  
B Virus ◽  
Immunodeficiency Virus ◽  
Pharmacokinetic Properties ◽  
Infected Cells

In vitro activities, against human immunodeficiency virus (HIV)- and hepatitis B virus (HBV)-infected cells, of four amphiphilic heterodinucleoside phosphates containing 3′-azido-2′,3′-dideoxythymidine (AZT) or 2′,3′-dideoxycytidine (ddC) as antiviral monomers were evaluated. The four compounds were N4-hexadecyl-2′-deoxyribocytidylyl-(3′→5′)-3′-azido-2′,3′-deoxythymidine (N4-hxddC-AZT), N4-palmitoyl-2′-deoxyribocytidylyl-(3′→5′)-3′-azido-2′,3′-deoxythymidine (N4-pamdC-AZT), N4-hexadecyl-2′-deoxycytidylyl-(3′→5′)-2′,3′-dideoxycytidine (N4-hxddC-ddC) and 2′-deoxythymidylyl-(3′→5′)-N4-palmitoyl-2′,3′-dideoxycytidine (dT-N4-pamddC). All four dimers were active against HIV, dT-N4-pamddC being the most active and least toxic. dT-N4-pamddC also exhibited strong antiviral effects against a panel of eight AZT-resistant HIV strains. The ddC-containing heterodimers incorporated in liposomes additionally inhibited HBV replication by 50–80% in HepG2 2.2.15 cells. AZT and the AZT-containing dimers were ineffective. Differences in pharmacokinetic properties between the antiviral monomers and the heterodimers were evaluated using liposomal formulations of3H-labelled AZT heterodimers as model compounds. The cellular distribution of AZT in H9 cells was predominantly cytoplasmic, whereas the amphiphilic dimers were distributed more evenly throughout the cytoplasm, nuclear membranes and microsomes. Blood levels of the heterodimers decreased at a rate two- to threefold slower than AZT and the areas-under-the-curves were five- to sevenfold higher for N4-pamdC-AZT and N4-hxddC-AZT, respectively. Compared to AZT, the peak levels of the dimers were three to four times higher in blood and five to six times higher in the liver. Analysis of blood samples showed that 34% of N4-pamdC-AZT was metabolized to AZT, whereas only 9% of N4-hxddC-AZT released AZT. Considering the antiviral potency and the favourable pharmacokinetic properties of the heterodimers, these compounds merit further exploration as antiviral drugs.

scholarly journals A cell-penetrating whole molecule antibody targeting intracellular HBx suppresses hepatitis B virus via TRIM21-dependent pathway

Theranostics ◽  
2018 ◽  
Vol 8 (2) ◽  
pp. 549-562 ◽  
Author(s):  
Jun-Fang Zhang ◽  
Hua-Long Xiong ◽  
Jia-Li Cao ◽  
Shao-Juan Wang ◽  
Xue-Ran Guo ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
B Virus ◽  
Cell Penetrating ◽  
A Cell ◽  
Antibody Targeting ◽  
Dependent Pathway

scholarly journals Hepatitis B Virus Replication Is Associated with an HBx-Dependent Mitochondrion-Regulated Increase in Cytosolic Calcium Levels

2007 ◽  
Vol 81 (21) ◽  
pp. 12061-12065 ◽  
Author(s):  
Stephanie L. McClain ◽  
Amy J. Clippinger ◽  
Rebecca Lizzano ◽  
Michael J. Bouchard
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Molecular Mechanisms ◽  
Mitochondrial Permeability Transition Pore ◽  
Mitochondrial Permeability Transition ◽  
Permeability Transition Pore ◽  
Permeability Transition ◽  
Cytosolic Calcium ◽  
Hbv Replication ◽  
B Virus

ABSTRACT The nonstructural hepatitis B virus (HBV) protein HBx has an important role in HBV replication and in HBV-associated liver disease. Many activities have been linked to HBx expression; however, the molecular mechanisms underlying many of these activities are unknown. One proposed HBx function is the regulation of cytosolic calcium. We analyzed calcium levels in HepG2 cells that expressed HBx or replicating HBV, and we demonstrated that HBx, expressed in the absence of other HBV proteins or in the context of HBV replication, elevates cytosolic calcium. We linked this elevation of cytosolic calcium to the association of HBx with the mitochondrial permeability transition pore.

scholarly journals Phosphorothioate Di- and Trinucleotides as a Novel Class of Anti-Hepatitis B Virus Agents

2004 ◽  
Vol 48 (6) ◽  
pp. 2199-2205 ◽  
Author(s):  
Radhakrishnan P. Iyer ◽  
Yi Jin ◽  
Arlene Roland ◽  
John D. Morrey ◽  
Samir Mounir ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Nucleoside Analogs ◽  
Hbv Dna ◽  
Parallel Synthesis ◽  
Hbv Replication ◽  
Long Term Treatment ◽  
B Virus ◽  
Dna Strand

ABSTRACT Several nucleoside analogs are under clinical development for use against hepatitis B virus (HBV). Lamivudine (3TC), a nucleoside analog, and adefovir dipivoxil (ADV), an acyclonucleotide analog, are clinically approved. However, long-term treatment can induce viral resistance, and following the cessation of therapy, viral rebound is frequently observed. There continues to be a need for new antiviral agents with novel mechanisms of action. A library of more than 600 di- and trinucleotide compounds synthesized by parallel synthesis using a combinatorial strategy was screened for potential inhibitors of HBV replication using the chronically HBV-producing cell line 2.2.15. Through an iterative process of synthesis, lead optimization, and screening, three analogs were identified as potent inhibitors of HBV replication: dinucleotides ORI-7246 (drug concentration at which a 10-fold reduction of HBV DNA was observed [EC90], 1.4 μM) and ORI-9020 (EC90, 1.2 μM) and trinucleotide ORI-7170 (EC90, 7.2 μM). These analogs inhibited the replication of both strands of HBV DNA. No suppression of HBV protein synthesis or intracellular core particle formation by these analogs was observed. No inhibition of HBV DNA strand elongation by the analogs or their 5′-triphosphate versions was apparent in in vitro polymerase assays. Although the exact mechanism of action is not yet identified, present data are consistent with an inhibition of the HBV reverse transcriptase-directed priming step prior to elongation of the first viral DNA strand. In transient-transfection assays, these analogs inhibited the replication of 3TC-resistant HBV. Synergistic interactions in combination treatments between the analogs and either 3TC or ADV were observed. These compounds represent a novel class of anti-HBV molecules and warrant further investigation as potential therapeutic agents.

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