Fig. 3.

A) Percentages of infected cells using a 3D model with HeLa spheroids pretreated with 40 μM RSV were obtained by quantification of CFSE-positive cells by cytometry analysis. As controls, spheroids were left untreated or pre-incubated with the drug vehicle (1% DMSO for 18 h). All infections were performed for 4 h with the addition of the drug vehicle (−) or RSV at final concentrations of 4 μM (+) or 40 μM (++) (indicated in the figure as “added during infection” above the line). B) Fluorescence images of in vitro infections performed on HeLa spheroids pre-incubated with RSV or DMSO, showing HeLaR2 cells imaged at 530 nm (red) merged with CFSE-labeled parasites imaged with a 488 nm (green). Images were acquired with a 40 × objective, and the spheroid was scanned at 10, 20, 30, 40 and 50 μm in depth from the surface. Graphics below show the quantification of parasites within the spheroids in each stack scanned. Quantification was obtained using the particles analysis tool from ImageJ software of five different spheroids from each treatment per experiment. All assays are expressed as the mean ± SD of three independent experiments (∗p < 0,05; ∗∗p < 0,005). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)