Figure 2

SOF blocks ZIKV replication in vitro. (a,b) Dose-response curves of SOF against ZIKV in NPCs and in Vero cells, respectively. SOF was tested in 10 points dose-response with two-fold dilution starting at 50 μM, in triplicates. The graphs show the antiviral activity (%) based on reduction of the cytopathic effect by quantification of the cytopathic effects of ZIKV ± SEM on NPCs (a) and Vero cells (b,c). Multi-step growth curve of vehicle or SOF-treated ZIKV-infected (ibH 30656, MOI = 0.1) NPCs at different concentrations over time as measured by viral genome copies of RNA using qRT-PCR. Data represents means ± SD (n = 3 replicates); two-way ANOVA, followed by Tukey’s multiple comparison tests. ****P < 0.0001 compared to DMSO (vehicle) treated NPCs at 120 hours p.i. Note that SD cannot be seen in the log scale. (d) The percentages of apoptotic cells were calculated, averaged, and graphed accordingly (ibH 30656, MOI = 0.1, 96 hours p.i.) in the presence of SOF at different doses (1 nM, 100 nM, 1 μM, 10 μM, 20 μM and 50 μM). One-way ANOVA tests with Tukey multiple comparisons were performed to compare to ZIKV-infected different groups. The presented values are means of TUNEL⁺/DAPI⁺ percentage ± SD (n = 6 images per condition), **P < 0.01, ***P < 0.001, ****P < 0.0001 TUNEL⁺/DAPI⁺ percentage decreases with different doses of SOF. (e) Representative images of mock-infected human neurospheres (left panel), ZIKV-infected vehicle-treated neurospheres (middle panel) and ZIKV-infected SOF-treated (20 μM) (right panel) neurospheres image (ibH30656, MOI = 0.1, 180 hours p.i.), scale bar: 500 μm. (f) Quantification of neurosphere diameter (μm) at 180 hours p.i. One-way ANOVA tests with Tukey multiple comparison were performed to compare different groups. The bars represent the number of spheres counted averaged and plotted for each condition ± SD (n = 6 images captured per condition), *P < 0.05, ***P < 0.001 (n ≥ 100 neurospheres counted per condition). (g) Neurosphere amount was counted over time after infection with ZIKV (ibH30656, MOI = 0.1 at 48, 96 and 120 hours p.i.) treated with vehicle or with SOF (20 μM). Bars represent the number of neurospheres ± SD at a given time (48, 96 and 120 hours p.i.). Two-way ANOVA tests with Tukey multiple comparison were performed to compare different groups, *P < 0.05 (n = 2 independent experiments). (h) Human antiviral response expression signature in human NPCs in monolayer at 120 hours p.i.: mock, ZIKV-infected (ibH30656, MOI 0.1) vehicle or SOF-treated (50 μM) (threshold = two-fold). Bars represent fold changes detected by qPCR from 96 different genes tested, between ZIKV-infected compared to mock-infected NPCs (in white) and between ZIKV + SOF-treated compared to mock-infected NPCs (in black). Note that there are fewer dysregulated genes between ZIKV-infected compared to mock NPCs upon SOF treatment (less black bars) than the ZIKV-infected untreated ones.