Establishment of stable cell lines in which the HBV genome replicates episomally for evaluation of antivirals

Sun, Suofeng; Li, Yuan; Liu, Bowei; Zhang, Bingyong; Han, Sun‐Young; Li, Xiuling

doi:10.5114/aoms.2018.79712

Cited by 3 publications

(1 citation statement)

References 28 publications

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“…A part of the nucleocapsid is recycled into the nucleus and the rcDNA is converted to cccDNA. Although the viral episomal genomic cccDNA serves as a template for transcription, it cannot be integrated into its formation [8,9]. HBV cccDNA is the template for transcription of all viral mRNAs and accumulates as a stable episome in the nucleus of infected cells.…”

Section: Introductionmentioning

confidence: 99%

HBx Regulation on HBV Pregenome Promoter in the Episomal Form Versus the Integrated Form

Hu¹,

Zhu²,

Liu³

et al. 2023

Discovery Medicine

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Background: Hepatitis B virus (HBV) genome structure is an incomplete closed double stranded circular DNA and it uses covalently closed circular DNA (cccDNA) as template for replication. To study the antiviral effect on different HBV replication forms, a stable cell line expressing HBV using Huh7 cells with shuttle plasmid to imitate the real HBV replication form was stablished. Unlike the HepG2.2.15 cells, the replication of HBV-expressing Huh7 cells present significant decrease after 9 days of interferon-α (IFN-α) treatment. This study aimed to verify whether hepatitis B virus X (HBx) epigenetic regulation by HBV promoter is affected by the DNA form and discuss the differences between the episomal form and the integrated form. Material and Methods: Huh7 cells were used with two different plasmids containing HBV genome to imitate HBV-expressing cells with the episomal form and the integrated form. Luciferase reporting system was used to determine the activation of the promoter after treatment with IFN-α with different concentrations and promoter regulation factor HBx. HBx-expressing plasmid was transfected to evaluate its effect on HBV replication in the episomal form. HBV DNA and pregenomic RNA (pgRNA) in HBx knockdown cell line was determined and HBx-expressing plasmid was transfected to evaluate its effect on HBx in the episomal form. Results: The two cell lines were established successfully and used for further experiments after selection. IFN-α showed significant inhibition effect on HBV pregenome promoter in the episomal form DNA while was not observed in the integrated form. After HBx-expressing plasmid was transfected, HBV pregenome promoter activity was higher in the episomal form rather than the integrated form. HBx showed a concentration-dependant activation on HBV replication in the episomal form. HBx knockdown reduced HBV production and HBV concentration significantly increased after transfection by HBx-expressing plasmid. Conclusions: HBx regulation effect on HBV pregenome promoter is influenced by the HBV genome form. The epigenetic regulation effect on HBV pregenome promoter is more active in the episomal form rather than the integrated form.

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Section: Introductionmentioning

confidence: 99%

HBx Regulation on HBV Pregenome Promoter in the Episomal Form Versus the Integrated Form

Hu¹,

Zhu²,

Liu³

et al. 2023

Discovery Medicine

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Novel protein kinase C phosphorylated kinase inhibitor-matrine suppresses replication of hepatitis B virus via modulating the mitogen-activated protein kinase signal

Zhou

Gao

et al. 2022

Bioengineered

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Solamargine acts as an antiviral by interacting to MZF1 and targeting the core promoter of the hepatitis B virus gene

Chen,

Zhao,

Huang

et al. 2024

Aging

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Background: Hepatitis B virus (HBV) infection is still a serious threat to global health and can lead to a variety of liver diseases, including acute and chronic hepatitis, liver cirrhosis, liver failure, hepatocellular carcinoma (HCC), and so on. At present, there are mainly two kinds of drugs for the treatment of hepatitis B at home and abroad: interferon (IFN) and nucleoside/nucleotide analogs (NAs). In recent years, natural compounds have been considered an important source for the development of new anti-HBV drugs due to their complex structure, diverse components, high efficiency, and low toxicity. Many studies have demonstrated that Solamargine has significant anticancer activity, but the antiviral effect is rarely studied. This study aimed to verify the anti-HBV effect of Solamargine and to explore the specific mechanism. Method: The relative expression of HBV pregenomic RNA (pgRNA) was detected by reverse transcription real-time fluorescence quantitative PCR (RT-qPCR). Northern blot and western blot were used to detect the relative expression of HBV pgRNA and target protein. PCR was used in the construction of HBV pg-promoter, ENII/BCP, and a series of gene deletion mutant fluorescent reporter vectors. The fluorescence relative expression of each mutant was detected by Renilla luciferase assay. Results: By binding to MZF1 (Myeloid zinc finger protein 1, MZF1), Solamargine inhibits HBV core promoter activity, reduces pregenomic RNA level, and inhibits HBV, achieving antiviral effects.

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Establishment of stable cell lines in which the HBV genome replicates episomally for evaluation of antivirals

Cited by 3 publications

References 28 publications

HBx Regulation on HBV Pregenome Promoter in the Episomal Form Versus the Integrated Form

HBx Regulation on HBV Pregenome Promoter in the Episomal Form Versus the Integrated Form

Novel protein kinase C phosphorylated kinase inhibitor-matrine suppresses replication of hepatitis B virus via modulating the mitogen-activated protein kinase signal

Solamargine acts as an antiviral by interacting to MZF1 and targeting the core promoter of the hepatitis B virus gene

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