{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,11,27]],"date-time":"2025-11-27T13:55:29Z","timestamp":1764251729210,"version":"build-2065373602"},"reference-count":51,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2021,11,13]],"date-time":"2021-11-13T00:00:00Z","timestamp":1636761600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001659","name":"Deutsche Forschungsgemeinschaft","doi-asserted-by":"publisher","award":["Projektnummer 272983813 \u2013 TRR179"],"award-info":[{"award-number":["Projektnummer 272983813 \u2013 TRR179"]}],"id":[{"id":"10.13039\/501100001659","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Viruses"],"abstract":"<jats:p>Immunity against hepatitis B virus (HBV) infection is complex and not entirely understood so far, including the decisive factors leading to the development of chronic hepatitis B. This lack of a mechanistic understanding of HBV-specific immunity is also caused by a limited number of suitable animal models. Here, we describe the generation of a recombinant adenovirus expressing an HBV 1.3-overlength genome linked to luciferase (Ad-HBV-Luc) allowing for precise analysis of the quantity of infected hepatocytes. This enables sensitive and close-meshed monitoring of HBV-specific CD8 T cells and the onset of anti-viral immunity in mice. A high dose of Ad-HBV-Luc developed into chronic hepatitis B accompanied by dysfunctional CD8 T cells characterized by high expression of PD1 and TOX and low expression of KLRG1 and GzmB. In contrast, a low dose of Ad-HBV-Luc infection resulted in acute hepatitis with CD8 T cell-mediated elimination of HBV-replicating hepatocytes associated with elevated sALT levels and increased numbers of cytotoxic HBV-specific CD8 T cells. Thus, the infectious dose was a critical factor to induce either acute self-limited or chronic HBV infection in mice. Taken together, the new Ad-HBV-Luc vector will allow for highly sensitive and time-resolved analysis of HBV-specific immune responses during acute and chronic infection.<\/jats:p>","DOI":"10.3390\/v13112273","type":"journal-article","created":{"date-parts":[[2021,11,14]],"date-time":"2021-11-14T20:52:28Z","timestamp":1636923148000},"page":"2273","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["In Vivo Bioluminescence Imaging of HBV Replicating Hepatocytes Allows for the Monitoring of Anti-Viral Immunity"],"prefix":"10.3390","volume":"13","author":[{"given":"Katrin","family":"Manske","sequence":"first","affiliation":[{"name":"Institute of Molecular Immunology, Klinikum Rechts der Isar, TUM School of Medicine, Technical University of Munich, 80333 Munich, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Annika","family":"Schneider","sequence":"additional","affiliation":[{"name":"Institute of Molecular Immunology, Klinikum Rechts der Isar, TUM School of Medicine, Technical University of Munich, 80333 Munich, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6512-1602","authenticated-orcid":false,"given":"Chunkyu","family":"Ko","sequence":"additional","affiliation":[{"name":"Institute of Virology, Technical University of Munich, 81675 Munich, Germany"},{"name":"Infectious Diseases Therapeutic Research Center, Therapeutics & Biotechnology Division, Korea Research Institute of Chemical Technology (KRICT), Daejeon 34114, Korea"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Percy A.","family":"Knolle","sequence":"additional","affiliation":[{"name":"Institute of Molecular Immunology, Klinikum Rechts der Isar, TUM School of Medicine, Technical University of Munich, 80333 Munich, Germany"},{"name":"German Center for Infection Research (DZIF), Munich Partner Site, 81675 Munich, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7269-5433","authenticated-orcid":false,"given":"Katja","family":"Steiger","sequence":"additional","affiliation":[{"name":"Institute of Pathology, Technical University of Munich, 81675 Munich, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9421-1911","authenticated-orcid":false,"given":"Ulrike","family":"Protzer","sequence":"additional","affiliation":[{"name":"Institute of Virology, Technical University of Munich, 81675 Munich, Germany"},{"name":"German Center for Infection Research (DZIF), Munich Partner Site, 81675 Munich, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3861-5150","authenticated-orcid":false,"given":"Dirk","family":"Wohlleber","sequence":"additional","affiliation":[{"name":"Institute of Molecular Immunology, Klinikum Rechts der Isar, TUM School of Medicine, Technical University of Munich, 80333 Munich, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,11,13]]},"reference":[{"key":"ref_1","unstructured":"WHO (2021, June 26). Hepatitis B Fact Sheet. Available online: https:\/\/www.who.int\/news-room\/fact-sheets\/detail\/hepatitis-b."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"e559","DOI":"10.1016\/S2214-109X(16)30130-9","article-title":"Acceptability and feasibility of a screen-and-treat programme for hepatitis B virus infection in The Gambia: The prevention of liver fibrosis and cancer in africa (prolifica) study","volume":"4","author":"Lemoine","year":"2016","journal-title":"Lancet Glob. Health"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"S289","DOI":"10.1093\/cid\/ciw090","article-title":"A novel collaborative community-based hepatitis B screening and linkage to care program for African immigrants","volume":"62","author":"Shankar","year":"2016","journal-title":"Clin. Infect. Dis."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"258","DOI":"10.1016\/j.patbio.2009.11.001","article-title":"Pathogenesis of hepatitis B virus infection","volume":"58","author":"Chisari","year":"2010","journal-title":"Pathol. Biol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"S54","DOI":"10.1002\/hep.21060","article-title":"The liver as an immunological organ","volume":"43","author":"Racanelli","year":"2006","journal-title":"Hepatology"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"201","DOI":"10.1038\/nri3169","article-title":"Living in the liver: Hepatic infections","volume":"12","author":"Protzer","year":"2012","journal-title":"Nat. Reviews. Immunol."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"447","DOI":"10.1172\/JCI118055","article-title":"Cell-specific expression of transforming growth factor-beta in rat liver. Evidence for autocrine regulation of hepatocyte proliferation","volume":"96","author":"Bissell","year":"1995","journal-title":"J. Clin. Investig."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"226","DOI":"10.1016\/0168-8278(95)80433-1","article-title":"Human Kupffer cells secrete IL-10 in response to lipopolysaccharide (LPS) challenge","volume":"22","author":"Knolle","year":"1995","journal-title":"J. Hepatol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1193","DOI":"10.1053\/j.gastro.2013.12.036","article-title":"Hepatic immune regulation and its involvement in viral hepatitis infection","volume":"146","author":"Knolle","year":"2014","journal-title":"Gastroenterology"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"729","DOI":"10.1002\/hep.22034","article-title":"Liver: An organ with predominant innate immunity","volume":"47","author":"Gao","year":"2008","journal-title":"Hepatology"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"4215","DOI":"10.1128\/JVI.02844-06","article-title":"Characterization of hepatitis B virus (HBV)-specific T-cell dysfunction in chronic HBV infection","volume":"81","author":"Boni","year":"2007","journal-title":"J. Virol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1835","DOI":"10.1172\/JCI33402","article-title":"Bim-mediated deletion of antigen-specific CD8 T cells in patients unable to control HBV infection","volume":"118","author":"Lopes","year":"2008","journal-title":"J. Clin. Investig."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"859","DOI":"10.1038\/nm.3251","article-title":"Pathogenesis of chronic viral hepatitis: Differential roles of T cells and NK cells","volume":"19","author":"Rehermann","year":"2013","journal-title":"Nat. Med."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1494","DOI":"10.1002\/hep.24249","article-title":"Role of the coinhibitory receptor cytotoxic T lymphocyte antigen-4 on apoptosis-Prone CD8 T cells in persistent hepatitis B virus infection","volume":"53","author":"Schurich","year":"2011","journal-title":"Hepatology"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"325","DOI":"10.1053\/j.gastro.2018.10.032","article-title":"Targeting innate and adaptive immune responses to cure chronic HBV infection","volume":"156","author":"Gehring","year":"2019","journal-title":"Gastroenterology"},{"key":"ref_16","unstructured":"WHO (2015). Guidelines for the Prevention, Care and Treatment of Persons with Chronic Hepatitis B Infection, WHO Library Cataloguing-in-Publication Data."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"45","DOI":"10.1111\/liv.13291","article-title":"Hepatitis B virus long-term impact of antiviral therapy nucleot(s)ide analogues (NUCs)","volume":"37","author":"Grossi","year":"2017","journal-title":"Liver Int."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"S41","DOI":"10.1016\/j.jhep.2016.02.009","article-title":"Attacking hepatitis B virus cccDNA\u2014The holy grail to hepatitis B cure","volume":"64","author":"Lucifora","year":"2016","journal-title":"J. Hepatol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"183","DOI":"10.1006\/jmbi.2000.4481","article-title":"Structural organization of the hepatitis B virus minichromosome","volume":"307","author":"Bock","year":"2001","journal-title":"J. Mol. Biol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1386","DOI":"10.1016\/S0016-5085(99)70289-1","article-title":"Direct ex vivo analysis of hepatitis B virus-specific CD8(+) T cells associated with the control of infection","volume":"117","author":"Maini","year":"1999","journal-title":"Gastroenterology"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"68","DOI":"10.1128\/JVI.77.1.68-76.2003","article-title":"CD8(+) T cells mediate viral clearance and disease pathogenesis during acute hepatitis B virus infection","volume":"77","author":"Thimme","year":"2003","journal-title":"J. Virol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"682","DOI":"10.1053\/j.gastro.2009.09.052","article-title":"Antiviral intrahepatic T-cell responses can be restored by blocking programmed death-1 pathway in chronic hepatitis B","volume":"138","author":"Fisicaro","year":"2010","journal-title":"Gastroenterology"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"373","DOI":"10.1002\/hep.24707","article-title":"Hepatocyte polarization is essential for the productive entry of the hepatitis B virus","volume":"55","author":"Schulze","year":"2012","journal-title":"Hepatology"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"S281","DOI":"10.1093\/cid\/ciw023","article-title":"Mechanism of hepatitis B virus persistence in hepatocytes and its carcinogenic potential","volume":"62","author":"Dandri","year":"2016","journal-title":"Clin. Infect. Dis."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"327","DOI":"10.1038\/nrgastro.2017.44","article-title":"Viral hepatitis: The bumpy road to animal models for HBV infection","volume":"14","author":"Protzer","year":"2017","journal-title":"Nat. Rev. Gastroenterol. Hepatol."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"13825","DOI":"10.1073\/pnas.202398599","article-title":"Hydrodynamic injection of viral DNA: A mouse model of acute hepatitis B virus infection","volume":"99","author":"Yang","year":"2002","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1762","DOI":"10.1053\/j.gastro.2020.01.032","article-title":"Knockdown of virus antigen expression increases therapeutic vaccine efficacy in high-titer hepatitis B virus carrier mice","volume":"158","author":"Michler","year":"2020","journal-title":"Gastroenterology"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"5108","DOI":"10.1128\/JVI.75.11.5108-5118.2001","article-title":"Transfer of hepatitis B virus genome by adenovirus vectors into cultured cells and mice: Crossing the species barrier","volume":"75","author":"Sprinzl","year":"2001","journal-title":"J. Virol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"2089","DOI":"10.1002\/hep.30080","article-title":"Outcome of anti-viral immunity in the liver is shaped by the level of antigen expressed in infected hepatocytes","volume":"68","author":"Manske","year":"2018","journal-title":"Hepatology"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1231","DOI":"10.1016\/j.jhep.2018.08.012","article-title":"Hepatitis B virus genome recycling and de novo secondary infection events maintain stable cccdna levels","volume":"69","author":"Ko","year":"2018","journal-title":"J. Hepatol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"105140","DOI":"10.1016\/j.antiviral.2021.105140","article-title":"Intramolecular recombination enables the formation of hepatitis B virus (HBV) cccdna in mice after HBV genome transfer using recombinant AAV vectors","volume":"194","author":"Ko","year":"2021","journal-title":"Antivir. Res."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"444","DOI":"10.1038\/s41586-021-03233-8","article-title":"Auto-aggressive CXCR6(+) CD8 T cells cause liver immune pathology in NASH","volume":"592","author":"Dudek","year":"2021","journal-title":"Nature"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"a021477","DOI":"10.1101\/cshperspect.a021477","article-title":"Mouse models of hepatitis B virus pathogenesis","volume":"5","author":"Iannacone","year":"2015","journal-title":"Cold Spring Harb. Perspect. Med."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Liu, Y., Maya, S., and Ploss, A. (2021). Animal models of hepatitis B virus infection\u2013success, challenges, and future directions. Viruses, 13.","DOI":"10.3390\/v13050777"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"6158","DOI":"10.1128\/jvi.69.10.6158-6169.1995","article-title":"High-level hepatitis B virus replication in transgenic mice","volume":"69","author":"Guidotti","year":"1995","journal-title":"J. Virol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"5554","DOI":"10.1128\/JVI.03134-12","article-title":"Adeno-associated virus-mediated gene transfer leads to persistent hepatitis B virus replication in mice expressing HLA-A2 and HLA-DR1 molecules","volume":"87","author":"Dion","year":"2013","journal-title":"J. Virol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"1447","DOI":"10.1053\/j.gastro.2012.03.006","article-title":"Transfer of HBV genomes using low doses of adenovirus vectors leads to persistent infection in immune competent mice","volume":"142","author":"Huang","year":"2012","journal-title":"Gastroenterology"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"6669","DOI":"10.1073\/pnas.0401771101","article-title":"Genomic analysis of the host response to hepatitis B virus infection","volume":"101","author":"Wieland","year":"2004","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"1347","DOI":"10.1016\/j.jhep.2020.06.026","article-title":"Reduced mitochondrial resilience enables non-canonical induction of apoptosis after TNF receptor signaling in virus-infected hepatocytes","volume":"73","author":"Lampl","year":"2020","journal-title":"J. Hepatol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"9652","DOI":"10.1128\/JVI.00867-09","article-title":"The size of the viral inoculum contributes to the outcome of hepatitis B virus infection","volume":"83","author":"Asabe","year":"2009","journal-title":"J. Virol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"703","DOI":"10.1002\/hep.29112","article-title":"Sodium taurocholate cotransporting polypeptide is the limiting host factor of hepatitis B virus infection in macaque and pig hepatocytes","volume":"66","author":"Lempp","year":"2017","journal-title":"Hepatology"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"1647","DOI":"10.1053\/j.gastro.2017.08.034","article-title":"Viral load affects the immune response to HBV in mice with humanized immune system and liver","volume":"153","author":"Dusseaux","year":"2017","journal-title":"Gastroenterology"},{"key":"ref_43","doi-asserted-by":"crossref","unstructured":"Bility, M.T., Cheng, L., Zhang, Z., Luan, Y., Li, F., Chi, L., Zhang, L., Tu, Z., Gao, Y., and Fu, Y. (2014). Hepatitis B virus infection and immunopathogenesis in a humanized mouse model: Induction of human-specific liver fibrosis and m2-like macrophages. PLoS Pathog., 10.","DOI":"10.1371\/journal.ppat.1004032"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"2044","DOI":"10.1136\/gutjnl-2018-316091","article-title":"Hbv infection-induced liver cirrhosis development in dual-humanised mice with human bone mesenchymal stem cell transplantation","volume":"68","author":"Yuan","year":"2019","journal-title":"Gut"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"36","DOI":"10.1016\/j.bbrc.2012.08.138","article-title":"Amp-activated protein kinase stimulates myostatin expression in C2C12 cells","volume":"427","author":"Das","year":"2012","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"955","DOI":"10.1089\/hum.2014.048","article-title":"Cellular immune response against firefly luciferase after sleeping beauty-mediated gene transfer in vivo","volume":"25","author":"Vezys","year":"2014","journal-title":"Hum. Gene Ther."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"8623","DOI":"10.1073\/pnas.0809818106","article-title":"High antigen levels are the cause of T cell exhaustion during chronic viral infection","volume":"106","author":"Mueller","year":"2009","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1038\/s41586-019-1326-9","article-title":"Tox reinforces the phenotype and longevity of exhausted T cells in chronic viral infection","volume":"571","author":"Alfei","year":"2019","journal-title":"Nature"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"1819","DOI":"10.1084\/jem.20150598","article-title":"High antigen levels induce an exhausted phenotype in a chronic infection without impairing T cell expansion and survival","volume":"213","author":"Utzschneider","year":"2016","journal-title":"J. Exp. Med."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"805","DOI":"10.1038\/cmi.2015.80","article-title":"Effective intrahepatic CD8+ T-cell immune responses are induced by low but not high numbers of antigen-expressing hepatocytes","volume":"13","author":"Ochel","year":"2016","journal-title":"Cell Mol. Immunol."},{"key":"ref_51","doi-asserted-by":"crossref","unstructured":"Isogawa, M., Chung, J., Murata, Y., Kakimi, K., and Chisari, F.V. (2013). Cd40 activation rescues antiviral CD8(+) T cells from PD-1-mediated exhaustion. PLoS Pathog., 9.","DOI":"10.1371\/journal.ppat.1003490"}],"container-title":["Viruses"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1999-4915\/13\/11\/2273\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:29:46Z","timestamp":1760167786000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1999-4915\/13\/11\/2273"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,11,13]]},"references-count":51,"journal-issue":{"issue":"11","published-online":{"date-parts":[[2021,11]]}},"alternative-id":["v13112273"],"URL":"https:\/\/doi.org\/10.3390\/v13112273","relation":{},"ISSN":["1999-4915"],"issn-type":[{"type":"electronic","value":"1999-4915"}],"subject":[],"published":{"date-parts":[[2021,11,13]]}}}