FIG 7.

Langerin-mediated IAV infection can occur via multiple entry pathways. (A) Lec2-Lg cells were incubated at 37°C for 30 min in serum-free medium alone (mock) or serum-free medium containing either 5 μM Pistop2-100 or 50 μg/ml genistein. After the addition of 15 μg/ml of Alexa Fluor 594-conjugated transferrin (Tf; clathrin dependent) or 50 μg/ml of Alexa Fluor 488-conjugated cholera toxin B (CTB; caveolin dependent), cells were incubated at 37°C for a further 15 min and then fixed, stained for double-stranded nucleic acid (DAPI), and observed by confocal microscopy. Representative images are shown. (B) Effects of chemical inhibitors on IAV infection of CHO-ctrl and Lec2-Lg cells. Monolayers of CHO-ctrl (white bars) and Lec2-Lg (black bars) cells were incubated in serum-free medium alone (0) or in serum-free medium containing increasing concentrations of Pitstop2-100 or genistein at 37°C for 30 min. Next, 10⁷ PFU BJx109 (left panels) or 10⁵ FFU RSV (right panels) was added (while maintaining the concentration of chemical inhibitors) and incubated at 37°C for 1 h. Cells were then washed, 10 mM NH₄Cl was added to prevent further infection, and the cells were cultured for a further 7 h (BJx109) or 18 h (RSV) and then fixed and stained for virus-infected cells. Data show the mean percent infection (±1 SD). Statistical significance was assessed using one-way ANOVA. *, P < 0.05; **, P < 0.01; ***, P < 0.001; n.s., not significant.