Transgenic mice expressing hepatitis B virus X protein are more susceptible to carcinogen induced hepatocarcinogenesis

ABSTRACT Chronic infection with hepatitis B virus (HBV) is one of the major etiological factors in the development of human hepatocellular carcinoma. Transgenic mice that express the HBV X protein (HBx) have previously been shown to be more sensitive to the effects of hepatocarcinogens, although the mechanism for this cofactor role remains unknown. The ability of HBx to inhibit DNA repair in transiently transfected cell lines suggests one possible pathway. In the present study, primary hepatocytes isolated from transgenic mice that possess the HBV X gene under the control of the human α-1-antitrypsin regulatory region (ATX mice) were found to be deficient in their ability to conduct unscheduled DNA synthesis in response to UV-induced DNA damage. In order to measure the impact of HBx expression on DNA repair in vivo, double-transgenic mice that express HBx and possess a bacteriophage lambda transgene were sacrificed at 30, 90, and 240 days of age. Mutation frequency was determined for high-molecular-weight liver DNA of ATX and control mice by functional analysis of the lambda transgene. Expression of HBx did not significantly increase the accumulation of spontaneous mutations. These results are consistent with previous studies of HBx transgenic mice in which no effect of HBx on liver histology was apparent. This new animal model provides a powerful system in which to investigate the in vivo cooperation between HBx expression and environmental carcinogens.

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Expression of Hepatitis B Virus X Protein Does Not Alter the Accumulation of Spontaneous Mutations in Transgenic Mice

Journal of Virology ◽

10.1128/.74.11.5266-5272.2000 ◽

2000 ◽

Vol 74 (11) ◽

pp. 5266-5272 ◽

Cited By ~ 1

Author(s):

Charles R. Madden ◽

Milton J. Finegold ◽

Betty L. Slagle

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Transgenic Mice ◽

Spontaneous Mutations ◽

X Protein ◽

B Virus

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Enhancement of Hepatitis B Virus Replication by Its X Protein in Transgenic Mice

Journal of Virology ◽

10.1128/jvi.76.5.2579-2584.2002 ◽

2002 ◽

Vol 76 (5) ◽

pp. 2579-2584 ◽

Cited By ~ 93

Author(s):

Zhenming Xu ◽

T. S. Benedict Yen ◽

Lanying Wu ◽

Charles R. Madden ◽

Wenjie Tan ◽

...

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Transgenic Mice ◽

Viral Gene Expression ◽

Viral Gene ◽

X Protein ◽

Multifunctional Protein ◽

Hbv Replication ◽

B Virus

ABSTRACT Hepatitis B virus (HBV) X gene encodes a multifunctional protein that can regulate cellular signaling pathways, interact with cellular transcription factors, and induce hepatocellular oncogenesis. In spite of its diverse activities, the precise role of the X protein in the viral life cycle of HBV remains unclear. To investigate this question, we have produced transgenic mice that carry either the wild-type HBV genome or a mutated HBV genome incapable of expressing the 16.5-kDa X protein. Our results indicate that while the X protein is not absolutely essential for HBV replication or its maturation in transgenic mice, it can enhance viral replication, apparently by activating viral gene expression. These results demonstrate a transactivation role of the X protein in HBV replication in transgenic mice.

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Hepatitis B virus X protein enhances the development of liver fibrosis and the expression of genes associated with epithelial-mesenchymal transitions and tumor progenitor cells

Carcinogenesis ◽

10.1093/carcin/bgz109 ◽

2019 ◽

Vol 41 (3) ◽

pp. 358-367 ◽

Cited By ~ 3

Author(s):

James Ahodantin ◽

Bouchra Lekbaby ◽

Myriam Bou Nader ◽

Patrick Soussan ◽

Dina Kremsdorf

Keyword(s):

Hepatitis B Virus ◽

Liver Fibrosis ◽

Hepatitis B ◽

Transgenic Mice ◽

Epithelial Mesenchymal Transition ◽

Biological Effects ◽

Matrix Remodeling ◽

X Protein ◽

Mesenchymal Transition ◽

B Virus

Abstract The hepatitis B virus X protein (HBx) has pleiotropic biological effects, which underlies its potential role in cell transformation. However, its involvement in hepatic fibrosis remains unclear. In this study, we wanted to clarify, in vivo, the role of HBx protein in the development of liver fibrosis. Mice transgenic for the full-length HBx (FL-HBx) were used. To create liver fibrosis, FL-HBx transgenic and control mice were chronically exposed to carbon tetrachloride (CCl4). Modulation of the expression of proteins involved in matrix remodeling, hepatic metabolism and epithelial-mesenchymal transition (EMT) were investigated. In transgenic mice, FL-HBx expression potentiates CCl4-induced liver fibrosis with increased expression of proteins involved in matrix remodeling (Collagen1a, α-Sma, PdgfR-β, MMP-13). In FL-HBx transgenic mice, an increase in EMT was observed with a higher transcription of two inflammatory cytokines (TNF-α and TGF-β) and a decrease of glutamine synthetase expression level. This was associated with a sustained cell cycle and hepatocyte polyploidy alteration consistent with p38 and ERK1/2 overactivation, increase of PLK1 transcription, accumulation of SQSTM1/p62 protein and increase expression of Beclin-1. This correlates with a higher expression of tumor progenitor cell markers (AFP, Ly6D and EpCam), indicating a higher risk of progression from fibrosis to hepatocellular carcinoma (HCC) in the presence of FL-HBx protein. In conclusion, our results show that FL-HBx protein enhances the development of liver fibrosis and contributes to the progression of liver disease from chronic hepatitis to HCC.

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Blocking of G1/S transition and cell death in the regenerating liver of Hepatitis B virus X protein transgenic mice

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2005.12.089 ◽

2006 ◽

Vol 340 (3) ◽

pp. 916-928 ◽

Cited By ~ 81

Author(s):

Bo-Kuan Wu ◽

Chao-Chin Li ◽

Huei-Jane Chen ◽

Junn-Liang Chang ◽

King-Song Jeng ◽

...

Keyword(s):

Cell Death ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Transgenic Mice ◽

Regenerating Liver ◽

X Protein ◽

B Virus

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Altered DNA Mutation Spectrum in Aflatoxin B1-Treated Transgenic Mice That Express the Hepatitis B Virus X Protein

Journal of Virology ◽

10.1128/jvi.76.22.11770-11774.2002 ◽

2002 ◽

Vol 76 (22) ◽

pp. 11770-11774 ◽

Cited By ~ 27

Author(s):

Charles R. Madden ◽

Milton J. Finegold ◽

Betty L. Slagle

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Transgenic Mice ◽

Aflatoxin B1 ◽

Environmental Carcinogens ◽

X Protein ◽

Dna Mutation ◽

Dna Mutations ◽

B Virus

ABSTRACT Humans chronically infected with hepatitis B virus (HBV) are at further risk of liver cancer upon exposure to dietary aflatoxin B1 (AFB1), a carcinogenic product of the mold Aspergillus flavus. For the present study, we utilized double-transgenic mice (ATX mice) that express the HBV X protein (HBx) and possess a bacteriophage lambda transgene to evaluate the in vivo effect of HBx expression on AFB1-induced DNA mutations. The expression of HBx correlated with a 24% increase in mutation frequency overall and an approximately twofold increase in the incidence of G/C-to-T/A transversion mutations following AFB1 exposure. These results are consistent with a model in which expression of HBx during chronic HBV infection may contribute to the development of hepatocellular carcinoma following exposure to environmental carcinogens.

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Hepatitis B Virus X Protein Increases 8-Oxo-7,8-Dihydro-2ʹ-Deoxyguanosine (8-Oxodg) Level via Repressing MTH1/ MTH2 Expression in Hepatocytes

Cellular Physiology and Biochemistry ◽

10.1159/000495166 ◽

2018 ◽

Vol 51 (1) ◽

pp. 80-96 ◽

Cited By ~ 5

Author(s):

Yan-Ting Lin ◽

Wei Liu ◽

Yun He ◽

Yun-Li Wu ◽

Wan-Nan Chen ◽

...

Keyword(s):

Oxidative Stress ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Transgenic Mice ◽

Hbv Infection ◽

Luciferase Reporter ◽

Sequencing Analysis ◽

X Protein ◽

Chronic Hepatitis B Virus ◽

B Virus

Background/Aims: Chronic hepatitis B virus (HBV) infection markedly increases the risk of development of hepatocellular carcinoma (HCC). Among the seven viral proteins that HBV encodes, HBV X protein (HBx) appears to have the most oncogenic potential. The mitochondria-associated HBx can induce oxidative stress in hepatocytes, leading to the production of abundant reactive oxygen species (ROS). High levels of ROS usually induce oxidative DNA damage and 8-hydroxy-2-deoxyguanosine (8-OHdG), also known as 8-oxo-7,8-dihydro-2-deoxyguanosine (8-oxodG), which is one of the major products of DNA oxidation and an important biomarker for oxidative stress and carcinogenesis. Cells have evolved a mechanism to prevent oxidized nucleotides from their incorporation into DNA through nucleotide pool sanitization enzymes of MTH1 (NUDT1), MTH2 (NUDT15), MTH3 (NUDT18) and NUDT5. However, little is known as to whether HBx can regulate the expression of those enzymes and modulate the formation and accumulation of 8-oxodG in hepatocytes. Methods: The level of 8-oxodG was assessed by ELISA in stable HBV-producing hepatoma cell lines, an HBV infectious mouse model, HBV and HBx transgenic mice and HBV-infected patients versus their respective controls. Expression of MTH1, MTH2, MTH3 and NUDT5 was determined by a real-time quantitative PCR and western blot analysis. Transcriptional regulation of MTH1 and MTH2 expression by HBx and the effect of HBx on MTH1 and MTH2 promoter hypermethylation were examined using a luciferase reporter assay and bisulfite sequencing analysis. Results: In comparison with controls, significantly higher levels of 8-oxodG were detected in the genome and culture supernatant of stable HBV-producing HepG2.2.15 cells, in the sera and liver tissues of HBV infectious mice and HBV or HBx transgenic mice, and in the sera of HBV-infected patients. Expression of HBx in hepatocytes significantly increased 8-oxodG level and reduced the expression of MTH1 and MTH2 at both mRNA and protein levels. It was also demonstrated that HBx markedly attenuated the MTH1 or MTH2 promoter activities through hypermethylation. Furthermore, enhancement of 8-oxodG production by HBx was reversible by overexpression of MTH1 and MTH2. Conclusion: Our data show that HBx expression results in the accumulation of 8-oxodG in hepatocytes through inhibiting the expression of MTH1 and MTH2. This may implicate that HBx may act as a tumor promoter through facilitating the mutational potential of 8-oxodG thus connecting a possible link between HBV infection and liver carcinogenesis.

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