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. 2020 Apr 3;11:1677. doi: 10.1038/s41467-020-15337-2

Fig. 4. Modeling polarized infection of hepatitis A and E viruses.

Fig. 4

a Newly secreted focus-forming infectious particles (FFU) from HEV infected pol-HLCs released in either apical or basolateral compartment were titered on hepatoma cells. Pol-HLCs were treated with DMSO or 10 μm of the HEV replication inhibitor Sofosbuvir (n = biological replicates). b HEV particles released from HEV infected pol-HLCs 7 d post infection were also measured by qRT-PCR quantification of viral RNA copies or ORF2 capsid ELISA (n = biological replicates). c Newly secreted infectious HEV particles from infected pol-HLCs were treated with 1:100 anti-HEV capsid ORF2 antibody and titered on hepatoma cells (n = biological replicates). d Relative infectivity of HEV particles recovered from the lysate or released in the supernatant of hepatoma cells transfected with HEV RNA from strain Kernow-C1 P6, 7 d post-transfection. Harvested HEV particles were treated with 1:100 anti-ORF2 antibody, IgG-control antibody and/or 0.1% sodium deoxycholate (DOC) for 30 min at RT. HEV infectivity was determined by titration on hepatoma cells (n = biological replicates). e Intra- and extracellular HEV particles from HEV P6 RNA-transfected hepatoma cells were mixed with either apical or basal supernatant from pol-HLCs and/or 1:100 anti-ORF2 antibody prior to titration on hepatoma cells (n = biological replicates). f Western blot analysis of cell lysates from shCYP8B1-inducible, H9-derived pol-HLCs (H9/shCYB8B1). Cells were treated or untreated for 48 h with 3 μg/ml doxycycline (DOX) to induce shRNA expression. Shown are representative images of n = 2. g Apical total bile acid release from H9/shCYP8B1-derived pol-HLCs treated with 3 μg/ml DOX as indicated (n = biological replicates). h Extracellular HEV particles from HEV P6 RNA-transfected S10-3 cells were mixed with apical supernatant from H9/shCYP8B1-derived pol-HLCs treated with or without 3 μg/ml DOX (n = biological replicates). i HEV secretion model from non-polarized and polarized HLCs. j Pol-HLCs were infected with HAV strain HM175/18 f. 48 h post infection, newly released particles were harvested and titered on hepatoma cells (n = biological replicates). Statistical analysis was performed using a two-tailed unpaired t test with Bonferroni adjustment for multiple comparisons.