Dépôt numérique

An in Vitro Culture Model for The Induction of HDV Replication in a HBV Expressing Cell Line

Blanchet, Matthieu; Angelo, Léna; Tetreault, Yasmine; Khabir, Marwa; Sureau, Camille; Vaillant, Andrew et Labonté, Patrick ORCID logoORCID: https://orcid.org/0000-0001-7262-3125 (2023). An in Vitro Culture Model for The Induction of HDV Replication in a HBV Expressing Cell Line In: Meeting of the American-Association-for-the-Study-of-Liver-Diseases (AASLD), November 10-14, 2023, Boston, MA.

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Background: Individual’s chronically infected with both Hepatitis B virus (HBV) and Hepatitis D virus (HDV) present an increased risk to develop cirrhosis and hepatocellular carcinoma in comparison to HBV mono-infected individual’s. The number of HDV infected individual’s worldwide is estimated to be between 20 and 40 million, a strong incentive to further engage in the search for new antivirals using innovative and practical in vitro models. Here we present the HepG2BD cell line as a novel in vitro culture system for inducible HDV replication in HBV replicating cells. Methods: Engineering: The HepG2BD cell line derives from HepG2.2.15 cells in which a HDV cDNA sequence under the control of a Tet-Off promoter is inserted into the adeno-associated virus integration site 1 (AAVS1) safe harbor site. The insertion at this precise location was conducted using the CRISPR-Cas9 technology for reliable initiation/ repression of HDV RNA replication, and production of infectious HDV virions in a cell line that constitutively expresses HBV. Characterization: HDV RNA in cells and supernatants was quantified by RT-qPCR and Northern blot, HBV RNA in cells was monitored by RTqPCR. Secreted HBV particle concentrations were evaluated by qPCR after anti-PreS1 immunoprecipitation. Secreted HBsAg was quantified by ELISA. Cellular presence and localization of HDAg in various cell culture conditions was monitored by confocal immunofluorescence. Infectivity of secreted HDV virions was measured by inoculation of NTCPHuh7 cells followed by RT-qPCR. Results: The presented cell line is, to our knowledge, the first one to allow full replication of both HBV and HDV. This cell line shows proper induction of the HDV lifecycle. The interplay between HBV and HDV was detected in many instances. Differentiation into more classic hepatocyte morphology was associated with an increased concentration of both HDV and HBV in the supernatant. Conclusion: This new and unique model will be instrumental for the screening/characterization of new antivirals targeting both viruses (such as REP 2139). The HepG2BD cell line is also expected to help in our understanding of the dynamic interplay between the HBV and HDV lifecycles, and in identifying cellular factors involved.

Type de document: Document issu d'une conférence ou d'un atelier
Informations complémentaires: Présentation orale 1200-C Hepatology 78 (Suppl.1): S367 http://dx.doi.org/ 10.1097/HEP.0000000000000580
Mots-clés libres: -
Centre: Centre INRS-Institut Armand Frappier
Date de dépôt: 06 janv. 2024 22:31
Dernière modification: 06 janv. 2024 22:31
URI: https://espace.inrs.ca/id/eprint/13965

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