FIG 7.

Viral dissemination between macrophages by homotypic cell fusion. (A to C) NLAD8-infected Jurkat cells were cocultured with MDMs for 6 h. After elimination of T cells, MDMs were cultured for 1 or 5 more days and then stained with anti-Gag, phalloidin, and DAPI. (A) Cells were analyzed by confocal microscopy. Bar, 25 μm. (B and C) The number of nuclei per MDM was quantified from images of at least 50 cells. The results are expressed as the percentage of cells with 1, 2, 3, or more than 3 nuclei (B) and as the mean nucleus number per cell (C). Error bars represent 1 SEM. Statistical significance was determined by the Mann-Whitney U-test (****, P < 0.0001). (D) Infected Jurkat cells were cocultured with MDMs for 6 h. After elimination of T cells, autologous MDMs prelabeled with CellTrace were added and cultured for 1 day. MDMs were then stained with anti-Gag, phalloidin, and DAPI. Cells were analyzed by confocal microscopy. Bar, 25 μm. (E) Infected Jurkat cells were cocultured with MDMs for 6 h. After elimination of T cells, MDMs were cultured for 1 day with or without T20 (10 μg/ml) or maraviroc (10 μM) before staining with anti-Gag, phalloidin, and DAPI. Cells were analyzed by confocal microscopy. The number of nuclei was analyzed from images of at least 50 cells. The results are expressed as the percentages of cells with 2, 3, or more than 3 nuclei. (F and G) Infected Jurkat cells were cocultured with MDMs for 6 h. After elimination of T cells, MDMs were cultured for 1, 5, 8, or 12 days with or without T20 (10 μg/ml). (F) The percentage of Gag⁺ MDMs was then evaluated by flow cytometry. (G) In parallel, culture supernatants from MDMs were collected and p24 was quantified. The results shown in panels A to E are representative of those from 4 independent experiments performed with MDMs from 4 donors, while the results shown in panels F and G correspond to the means from 3 independent experiments performed with MDMs from 3 donors. NI, noninfected Jurkat cells cocultured with MDMs.