FIG 4.

The wing and β-ladder domains of flavivirus NS1 are necessary for inducing endothelial cell hyperpermeability. (A) HPMEC monolayer was seeded in the apical chambers of 24-well Transwell inserts and treated with either WT or chimeric NS1 proteins at 2.5 μg/mL. The transendothelial electrical resistance (TEER) between the apical and basolateral chamber was measured over time and normalized to the untreated controls at each respective time point. (B) Quantification of the area between the curve and Y = 1.0 in (A) (“area under curve”), correlating to a decrease in electrical resistance and an increase in permeability. (C, E) Endothelial hyperpermeability (TEER) assay as in (A), but with the indicated NS1 protein at (C) 5 and (E) 10 μg/mL, respectively. (D, F) Area of the curve of (C) and (E), respectively. AUC, area under the curve. a.u., arbitrary units. Data represent at least n = 3 biological replicates, plotted as mean ± standard error of the mean (SEM). *, P < 0.05; **, P < 0.01; ***, P < 0.005; ****, P < 0.001 by one-way ANOVA with multiple comparisons. Star colors indicate the respective control to which each construct was compared.