Hepatitis B virus efficiently infects non-adherent hepatoma cells via human sodium taurocholate cotransporting polypeptide

Abstract

Sodium taurocholate cotransporting polypeptide (NTCP) has been reported as a functional receptor for hepatitis B virus (HBV) infection. However, HBV could not efficiently infect HepG2 cells expressing NTCP (NTCP-HepG2 cells) under adherent monolayer-cell conditions. In this study, NTCP was mainly detected in the basolateral membrane region, but not the apical site, of monolayer NTCP-HepG2 cells. We hypothesized that non-adherent cell conditions of infection would enhance HBV infectivity. Non-adherent NTCP-HepG2 cells were prepared by treatment with trypsin and EDTA, which did not degrade NTCP in the membrane fraction. HBV successfully infected NTCP-HepG2 cells at a viral dose 10 times lower in non-adherent phase than in adherent phase. Efficient infection of non-adherent NTCP-HepG2 cells with blood-borne or cell-culture-derived HBV was observed and was remarkably impaired in the presence of the myristoylated preS1 peptide. HBV could also efficiently infect HepaRG cells under non-adherent cell conditions. We screened several compounds using our culture system and identified proscillaridin A as a potent anti-HBV agent with an IC50 value of 7.2 nM. In conclusion, non-adherent host cell conditions of infection augmented HBV infectivity in an NTCP-dependent manner, thus providing a novel strategy to identify anti-HBV drugs and investigate the mechanism of HBV infection.

Drug repurposing for new, efficient, broad spectrum antivirals

“…Although the precise mechanism of action is not fully clear, these drugs altered viral mRNA processing, blocking replication before viral DNA synthesis. Some cardiac glycosides like procillaridin A, bufallin, covallatoxin and digitoxin are able to inhibit hepatitis B virus (HBV) in cell cultures (Okuyama-Dobashi et al, 2015). In this study, however, digoxin did not show an anti-HBV effect.…”

Synthetically derived bat influenza A-like viruses reveal a cell type- but not species-specific tropism

“…This characteristic probably reflects the basolateral localization of the putative cellular receptor of HL17 and HL18 in MDCK II cells. Several other viruses are also known to use basolateral receptors for entry into epithelial cells, including VSV (19), hepatitis B virus (23,24), hepatitis C virus (25), adenovirus type 2 and 5 (26), vaccinia virus (27), and measles virus (28). Interestingly, many of these viruses exploit junctional and adhesion proteins of the host cell for attachment and entry (29).…”

Hepatitis C virus infection suppresses hepatitis B virus replication via the RIG-I-like helicase pathway

“…HBc antigen monoclonal antibodies were generated by immunizing of BALB/c mice with recombinant HBc protein synthesized in a wheat cell-free protein production system, as previously described 29 . HBc antigen was probed by using a mouse monoclonal antibody against HBc antigen (clone 7B2, culture supernatant of the hybridoma) 30 diluted 100-fold in PBS (1 hour at room temperature). HCV NS5A was stained by using a rabbit monoclonal antibody against NS5A diluted 100-fold in PBS (1 hour at room temperature).…”