Fig. 6
Inhibition of prenylation leads to an enhanced innate immune activation. HepNB2.7 cells seeded in 24-well plates were pre-treated with 50× IC50 concentration of the compounds for 2 h, and then inoculated with 0.5 IU/cell HDV in the presence of the compounds. The medium was replaced with fresh compounds every 2nd/3rd day. The supernatant from day 9 to 12 post infection was collected and one-fifth of the supernatant was used as an inoculum for secondary infection of HuH7-NTCP cells. a Cells of primary (top) and secondary (bottom) infection were fixed, and HDAg was immunostained (scale bar: 100 µm). b HDAg-specific western blot of the cells after primary infection treated with the indicated compounds (top). Western blot quantification of the two HDAg isoforms (bottom). Values represent the mean of three individual blots. c HepNB2.7 cells were fixed after primary infection and co-immunostained with a human serum for the total HDAg (red) and with a rabbit serum specifically for L-HDAg (green) (scale bar: 50 µm). d, e The total RNA was extracted from HepNB2.7 cells after primary infection, reverse transcribed, and the total HDV genomes, interferons, and ISGs were quantified by qPCR (n = 3 replicates). f, g, h Primary human hepatocytes (PHH) were co-infected with 0.5 IU/cell HDV and 150 ge/cell HBV and treated with 25× IC50 concentration of the compounds throughout the infection. The medium was replaced with fresh compounds every 2nd/3rd day. The supernatant from day 10 to 12 post infection was collected and one-fifth of the supernatant was used as an inoculum for secondary infection of HuH7-NTCP cells (f, scale bar: 50 µm). The PHH of the primary infection at d12 were lysed and S- and L-HDAg were analysed by western blot (g). The total RNA was extracted from the PHH at d12, and the total HDV genomes, IFN-lambda, and RSAD2 copies were determined by RT-qPCR (n = 3 replicates) (h). Unpaired two-tailed Student’s t test was used for statistical analyses (*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, n.s. not significant, P > 0.05)
