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Virology Journal
Published in: Virology Journal 1/2013

Open Access 01-12-2013 | Research

Inhibition of hepatitis B virus (HBV) gene expression and replication by HBx gene silencing in a hydrodynamic injection mouse model with a new clone of HBV genotype B

Authors: Lei Li, Hong Shen, Anyi Li, Zhenhua Zhang, Baoju Wang, Junzhong Wang, Xin Zheng, Jun Wu, Dongliang Yang, Mengji Lu, Jingjiao Song

Published in: Virology Journal | Issue 1/2013

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Abstract

Background

It has been suggested that different hepatitis B virus (HBV) genotypes may have distinct virological characteristics that correlate with clinical outcomes during antiviral therapy and the natural course of infection. Hydrodynamic injection (HI) of HBV in the mouse model is a useful tool for study of HBV replication in vivo. However, only HBV genotype A has been used for studies with HI.

Methods

We constructed 3 replication-competent clones containing 1.1, 1.2 and 1.3 fold overlength of a HBV genotype B genome and tested them both in vitro and in vivo. Moreover, A HBV genotype B clone based on the pAAV-MCS vector was constructed with the 1.3 fold HBV genome, resulting in the plasmid pAAV-HBV1.3B and tested by HI in C57BL/6 mice. Application of siRNA against HBx gene was tested in HBV genotype B HI mouse model.

Results

The 1.3 fold HBV clone showed higher replication and gene expression than the 1.1 and 1.2 fold HBV clones. Compared with pAAV-HBV1.2 (genotype A), the mice HI with pAAV-HBV1.3B showed higher HBsAg and HBeAg expression as well as HBV DNA replication level but a higher clearance rate. Application of two plasmids pSB-HBxi285 and pSR-HBxi285 expressing a small/short interfering RNA (siRNA) to the HBx gene in HBV genotype B HI mouse model, leading to an inhibition of HBV gene expression and replication. However, HBV gene expression may resume in some mice despite an initial delay, suggesting that transient suppression of HBV replication by siRNA may be insufficient to prevent viral spread, particularly if the gene silencing is not highly effective.

Conclusions

Taken together, the HI mouse model with a HBV genotype B genome was successfully established and showed different characteristics in vivo compared with the genotype A genome. The effectiveness of gene silencing against HBx gene determines whether HBV replication may be sustainably inhibited by siRNA in vivo.
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Literature
1.
Lai CL, Ratziu V, Yuen MF, Poynard T: Viral hepatitis B. Lancet. 2003, 362: 2089-2094. 10.1016/S0140-6736(03)15108-2.PubMedCrossRef
2.
Ganem D, Prince AM: Hepatitis B virus infection—natural history and clinical consequences. N Engl J Med. 2004, 350: 1118-1129. 10.1056/NEJMra031087.PubMedCrossRef
3.
Liaw YF, Chu CM: Hepatitis B virus infection. Lancet. 2009, 373: 582-592. 10.1016/S0140-6736(09)60207-5.PubMedCrossRef
4.
5.
Guidotti LG, Matzke B, Schaller H, Chisari FV: High level hepatitis B virus replication in transgenic mice. J Virol. 1995, 69: 6158-6169.PubMedPubMedCentral
6.
Araki K, Miyazaki J, Hino O, Tomita N, Chisaka O, et al: Expression and replication of hepatitis B virus genome in transgenic mice. Proc Natl Acad Sci USA. 1989, 186: 207-211.CrossRef
7.
Moriyama T, Guilhot S, Klopchin K, Moss B, Pinkert CA, et al: Immunobiology and pathogenesis of hepatocellular injury in hepatitis B virus transgenic mice. Science. 1990, 248: 361-364. 10.1126/science.1691527.PubMedCrossRef
8.
Larkin J, Clayton M, Sun B, Perchonock CE, Morgan JL, et al: Hepatitis B virus transgenic mouse model of chronic liver disease. Nat Med. 1999, 5: 907-912. 10.1038/11347.PubMedCrossRef
9.
Böcher WO, Marcus H, Shakarchy R, Dekel B, Shouval D, et al: Antigen-specific B and T cells in human/mouse radiation chimera following immunization in vivo. Immunology. 1999, 96: 634-641. 10.1046/j.1365-2567.1999.00704.x.PubMedPubMedCentralCrossRef
10.
Ilan E, Burakova T, Dagan S, Nussbaum O, Lubin I, et al: The hepatitis B virus-trimera mouse: a model for human HBV infection and evaluation of anti-HBV therapeutic agents. Hepatology. 1999, 29: 553-562. 10.1002/hep.510290228.PubMedCrossRef
11.
Dandri M, Burda MR, Török E, Pollok JM, Iwanska A, et al: Repopulation of the mouse liver with human hepatocytes and in vivo infection with hepatitis B virus. Hepatology. 2001, 33: 981-988. 10.1053/jhep.2001.23314.PubMedCrossRef
12.
Huang LR, Wu HL, Chen PJ, Chen DS: An immunocompetent mouse model for the tolerance of human chronic hepatitis B virus infection. Proc Natl Acad Sci USA. 2006, 103: 17862-17867. 10.1073/pnas.0608578103.PubMedPubMedCentralCrossRef
13.
Takehara T, Suzuki T, Ohkawa K, Hosui A, Jinushi M, et al: Viral covalently closed circular DNA in a non-transgenic mouse model for chronic hepatitis B virus replication. J Hepat. 2006, 44: 267-274. 10.1016/j.jhep.2005.07.030.CrossRef
14.
Cobleigh MA, Buonocore L, Uprichard SL, Rose JK, Robek MD: A vesicular stomatitis virus-based hepatitis B virus vaccine vector provides protection against challenge in a single dose. J Virol. 2010, 84: 7513-7522. 10.1128/JVI.00200-10.PubMedPubMedCentralCrossRef
15.
Lin YJ, Huang LR, Yang HC, Tzeng HT, Hsu PN, et al: Hepatitis B virus core antigen determines viral persistence in a C57BL/6 mouse model. Proc Natl Acad Sci USA. 2010, 107: 9340-9345. 10.1073/pnas.1004762107.PubMedPubMedCentralCrossRef
16.
Yang PL, Althage A, Chung J, Maier H, Wieland S, et al: Immune effectors required for hepatitis B virus clearance. Proc Natl Acad Sci USA. 2010, 107: 798-802. 10.1073/pnas.0913498107.PubMedPubMedCentralCrossRef
17.
Yin Y, Wu C, Song J, Wang J, Zhang E, et al: DNA Immunization with Fusion of CTLA-4 to Hepatitis B Virus (HBV) Core Protein Enhanced Th2 Type Responsesand Cleared HBV with an Accelerated Kinetic. PLoS One. 2011, 6: 1-11.CrossRef
18.
Wu C, Deng W, Deng L, Cao L, Qin B, et al: Amino acid substitutions at the positions 122 and 145 of hepatitis B surface antigen (HBsAg) determine the antigenicity and immunogenicity of HBsAg and influence in vivo HBsAg clearance. J Virol. 2012, 86: 4658-4669. 10.1128/JVI.06353-11.PubMedPubMedCentralCrossRef
19.
20.
Zoulim F: Infection Hepatitis B virus resistance to antiviral drugs: where are we going?. Liver Int. 2011, Suppl 1: 111-116.CrossRef
21.
Haasnoot PC, Cupac D, Berkhout B: Inhibition of virus replication by RNA interference. J Biomed Sci. 2003, 10: 607-616.PubMedCrossRef
22.
Randall G, Rice CM: Interfering with hepatitis C virus RNA replication. Virus Res. 2004, 102: 19-25. 10.1016/j.virusres.2004.01.011.PubMedCrossRef
23.
Wu J, Nandamuri KM: Inhibition of hepatitis viral replication by siRNA. Expert Opin Biol Ther. 2004, 4: 1649-1659. 10.1517/14712598.4.10.1649.PubMedCrossRef
24.
Li BJ, Tang Q, Cheng D, Qin C, Xie FY, et al: Using siRNA in prophylactic and therapeutic regimens against SARS coronavirus in Rhesus macaque. Nat Med. 2005, 11: 944-951.PubMed
25.
Tang H, Oishi N, Kaneko S, Murakami S: Molecular function and biological roles of hepatitis B virus x protein. Cancer Sci. 2006, 97: 977-983. 10.1111/j.1349-7006.2006.00299.x.PubMedCrossRef
26.
Wei Y, Neuveut C, Tiollais P, Buendia MA: Molecular biology of the hepatitis B virus and role of the X gene. Pathol Biol. 2010, 58: 267-272. 10.1016/j.patbio.2010.03.005.PubMedCrossRef
27.
28.
Meng Z, Qiu S, Zhang X, Wu J, Schreiter T, et al: Inhibition of woodchuck hepatitis virus gene expression in primary hepatocytes by siRNA enhances the cellular gene expression. Virology. 2009, 384: 88-96. 10.1016/j.virol.2008.11.012.PubMedCrossRef
29.
Tseng TC, Kao JH: HBV genotype and clinical outcome of chronic hepatitis B facts and puzzles. Gastroenterology. 2008, 134: 1272-1273. 10.1053/j.gastro.2007.12.046.PubMedCrossRef
30.
Kao JH, Chen PJ, Lai MY, Chen DS: Hepatitis B genotypes correlate with clinical outcomes in patients with chronic hepatitis B. Gastroenterology. 2000, 118: 554-559. 10.1016/S0016-5085(00)70261-7.PubMedCrossRef
31.
Yuan J, Zhou B, Tanaka Y, Kurbanov F, Orito E, et al: Hepatitis B virus (HBV) genotypes/subgenotypes in china: mutations in core promoter and precore/core and their clinical implications. J Clin Virol. 2007, 39: 87-93. 10.1016/j.jcv.2007.03.005.PubMedCrossRef
32.
Durantel D, Carrouée-Durantel S, Werle-Lapostolle B, Brunelle MN, Pichoud C, et al: A new strategy for studying in vitro the drug susceptibility of clinical isolates of human hepatitis B virus. Hepatology. 2004, 40: 855-864.PubMedCrossRef
33.
Yang H, Westland C, Xiong S, Delaney WE: In vitro antiviral susceptibility of full-length clinical hepatitis B virus isolates cloned with a novel expression vector. Antiviral Res. 2004, 61: 27-36. 10.1016/j.antiviral.2003.07.003.PubMedCrossRef
34.
Lu M, Isogawa M, Xu Y, Hilken G: Immunization with the gene expressing woodchuck hepatitis virus nucleocapsid protein fused to cytotoxic-T-lymphocyte-associated antigen 4 leads to enhanced specific immune responses in mice and woodchucks. J Virol. 2005, 79: 6368-6376. 10.1128/JVI.79.10.6368-6376.2005.PubMedPubMedCentralCrossRef
35.
Kuhns MC, Kleinman SH, McNamara AL, Rawal B, Glynn S, et al: Lack of correlation between HBsAg and HBV DNA levels in blood donors who test positive for HBsAg and anti-HBc: implications for future HBV screening policy. Transfusion. 2004, 44: 1332-1339. 10.1111/j.1537-2995.2004.04055.x.PubMedCrossRef
36.
Manesis EK, Hadziyannis E, Angelopoulou OP, Hadziyannis SJ: Prediction of treatment-related HBsAg loss in HBeAg-negative chronic hepatitis B: a clue from serum HBsAg levels. Antivir Ther. 2007, 12: 73-82.PubMed
37.
Wiegand J, Wedemeyer H, Finger A, Heidrich B, Rosenau J, et al: A decline in hepatitis B virus surface antigen (hbsag) predicts clearance, but does not correlate with quantitative hbeag or HBV DNA levels. Antivir Ther. 2008, 13: 547-554.PubMed
38.
Simonetti J, Bulkow L, McMahon BJ, Homan C, Snowball M, et al: Clearance of hepatitis B surface antigen and risk of hepatocellular carcinoma in acohort chronically infected with hepatitis B virus. Hepatology. 2010, 51: 1531-1537. 10.1002/hep.23464.PubMedCrossRef
39.
Chen CC, Ko TM, Ma HI, Wu HL, Xiao X, et al: Long-term inhibition of hepatitis B virus in transgenic mice by double-stranded adeno-associated virus 8-delivered short hairpin RNA. Gene Ther. 2007, 14: 11-19. 10.1038/sj.gt.3302846.PubMedCrossRef
40.
Yang Z, Zhang J, Cong H: A retrovirus-based system to stably silence GDF-8 expression and enhance myogenic differentiation in human rhabdomyosarcoma cells. J Gene Med. 2008, 10: 825-833. 10.1002/jgm.1216.PubMedCrossRef
41.
Sebestyen MG, Budker VG, Budker T, Subbotin VM, Zhang G, Monahan SD, et al: Mechanism of plasmid delivery by hydrodynamic tail vein injection. I. Hepatocyte uptake of various molecules. J Gene Med. 2006, 8: 852-873. 10.1002/jgm.921.PubMedCrossRef
42.
Crespo A, Peydro A, Dasi F, Benet M, Calvete JJ, Revert F, et al: Hydrodynamic liver gene transfer mechanism involves transient sinusoidal blood stasis and massive hepatocyte endocytic vesicles. Gene Ther. 2005, 12: 927-935. 10.1038/sj.gt.3302469.PubMedCrossRef
43.
Liu F, Lei J, Vollmer R, Huang L: Mechanism of liver gene transfer by mechanical massage. Mol Ther. 2004, 9: 452-457. 10.1016/j.ymthe.2003.12.003.PubMedCrossRef
44.
Kobayashi N, Nishikawa M, Hirata K, Takakura Y: Hydrodynamics-based procedure involves transient hyperpermeability in the hepatic cellular membrane: implication of a nonspecific process in efficient intracellular gene delivery. J Gene Med. 2004, 6: 584-592. 10.1002/jgm.541.PubMedCrossRef
45.
Zamore PD, Tuschl T, Sharp PA: RNAi: double-stranded RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell. 2000, 101: 25-33. 10.1016/S0092-8674(00)80620-0.PubMedCrossRef
46.
Baer M, Nilsen TW, Costigan C, Altman S: Structure and transcription of a human gene for H1 RNA, the RNA component of human RNase P. Nucleic Acids Res. 1990, 18: 97-103. 10.1093/nar/18.1.97.PubMedPubMedCentralCrossRef
47.
Brummelkamp TR, Bernards R, Agami R: A System for Stable Expression of Short Interfering RNAs in Mammalian Cells. Science. 2002, 296: 550-553. 10.1126/science.1068999.PubMedCrossRef
48.
Cougot D, Neuveut C, Buendia MA: HBV induced carcinogenesis. J Clin Virol. 2005, 34 (suppl 1): S75-S78.PubMedCrossRef
49.
Hung L, Kumar V: Specific inhibition of gene expression and transactivation functions of hepatitis B virus X protein and c-myc by small interfering RNAs. FEBS Lett. 2004, 560: 210-214. 10.1016/S0014-5793(04)00113-9.PubMedCrossRef
50.
Han Q, Zhang C, Zhang J, Tian Z: Reversal of hepatitis B virus-induced immune tolerance by an immunostimulatory 3p-HBx-siRNAs in a retinoic acid inducible gene I-dependent manner. Hepatology. 2011, 54: 1179-1189. 10.1002/hep.24505.PubMedCrossRef
51.
Reynolds A, Leake D, Boese Q, Scaringe S, Marshall WS: Rational siRNA design for RNA interference. Nat Biotechnol. 2004, 22: 326-330. 10.1038/nbt936.PubMedCrossRef
52.
Morrissey DV, Blanchard K, Shaw L, Jensen K, Lockridge JA, et al: Activity of stabilized short interfering RNA in a mouse model of hepatitis B virus replication. Hepatology. 2005, 41: 1349-1356. 10.1002/hep.20702.PubMedCrossRef
53.
Morrissey DV, Lockridge JA, Shaw L, Blanchard K, Jensen K, et al: Potent and persistent in vivo anti-HBV activity of chemically modified siRNAs. Nat Biotechnol. 2005, 23: 1002-1007. 10.1038/nbt1122.PubMedCrossRef
54.
Xuan B, Qian Z, Hong J, Huang W: EsiRNAs inhibit hepatitis B virus replication in mice model more efficiently than synthesized siRNAs. Virus Res. 2006, 118: 150-155. 10.1016/j.virusres.2005.12.005.PubMedCrossRef
55.
Aigner A: Gene silencing through RNA interference (RNAi) in vivo: strategies based on the direct application of siRNAs. J Biotechnol. 2006, 124: 12-25. 10.1016/j.jbiotec.2005.12.003.PubMedCrossRef
56.
Kim SI, Shin D, Choi TH, Lee JC, Cheon GJ, et al: Systemic and specific delivery of small interfering RNAs to the liver mediated by apolipoprotein A-I. Mol Ther. 2007, 15: 1145-1152.PubMed
57.
Garcia T, Li J, Sureau C, Ito K, Qin Y, Wands J, Tong S: Drastic reduction in the production of subviral particles does not impair hepatitis B virus virion secretion. J Virol. 2009, 83: 11152-11165. 10.1128/JVI.00905-09.PubMedPubMedCentralCrossRef
58.
Kuhns MC, Kleinman SH, McNamara AL, Rawal B, Glynn S, Busch MP, REDS Study Group: Lack of correlation between HBsAg and HBV DNA levels in blood donors who test positive for HBsAg and anti-HBc: implications for future HBV screening policy. Transfusion. 2004, 44: 1332-1339. 10.1111/j.1537-2995.2004.04055.x.PubMedCrossRef
59.
Tian YJ, Xu Y, Zhang ZH, Meng ZJ, Tan L, et al: A substitution at the amino acid position 122 of hepatitis B surface antigen (HBsAg) strongly impairs the antigenicity of HBsAg. J Clin Microbiol. 2007, 45: 2971-2978. 10.1128/JCM.00508-07.PubMedPubMedCentralCrossRef
60.
Mendy ME, Kaye S, van der Sande M, Rayco-Solon P, Waight PA, et al: Application of real-time PCR to quantify hepatitis B virus DNA in chronic carriers in The Gambia. Virology. 2006, 3: 23-10.1186/1743-422X-3-23.CrossRef
61.
Lu M, Yao X, Xu Y, Lorenz H, Dahmen U, et al: Combination of an antiviral drug and immunomodulation against hepadnaviral infection in the woodchuck model. J Virol. 2008, 82: 2598-2603. 10.1128/JVI.01613-07.PubMedPubMedCentralCrossRef
62.
Meng Z, Xu Y, Wu J, Tian Y, Kemper T, et al: Inhibition of hepatitis B virus gene expression and replication by endoribonuclease-prepared siRNA. J Virol Methods. 2008, 150: 27-33. 10.1016/j.jviromet.2008.02.008.PubMedCrossRef
63.
von Freyend MJ, Untergasser A, Arzberger S, Oberwinkler H, Drebber U, et al: Sequential control of hepatitis B virus in a mouse model of acute, self-resolving hepatitis B. J Viral Hepat. 2011, 18: 216-226. 10.1111/j.1365-2893.2010.01302.x.PubMedCrossRef
64.
Zhang X, Zhang E, Ma Z, Pei R, Jiang M, et al: Modulation of hepatitis B virus replication and hepatocyte differentiation by MicroRNA-1. Hepatology. 2011, 53: 1476-1485. 10.1002/hep.24195.PubMedCrossRef
Metadata
Title
Inhibition of hepatitis B virus (HBV) gene expression and replication by HBx gene silencing in a hydrodynamic injection mouse model with a new clone of HBV genotype B
Authors
Lei Li
Hong Shen
Anyi Li
Zhenhua Zhang
Baoju Wang
Junzhong Wang
Xin Zheng
Jun Wu
Dongliang Yang
Mengji Lu
Jingjiao Song
Publication date
01-12-2013
Publisher
BioMed Central
Published in
Virology Journal / Issue 1/2013
Electronic ISSN: 1743-422X
DOI
https://doi.org/10.1186/1743-422X-10-214

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