HIV-1 replication in osteoclasts. (A) The induced osteoclasts (circles) and macrophages (triangles) were incubated for 6 hours with a JR-FL or NL4-3 strain at the indicated doses and washed to remove free virus. Viral p24 concentration in the cell culture supernatants collected on day 3, 6, and 9 was monitored by ELISA. (*p < 0.05, **p < 0.01 by t test between day 3 and each day in each cell type) (B) The induced osteoclasts (black bars) and macropharges (white bars) were infected with JR-FL (2.5 × 105 IFU/ml) or NL4-3 (2.5 × 105 IFU/ml) in the presence of TFV at the indicated concentrations. Viral p24 concentrations in the cell culture supernatants on day 6 (JR-FL) or day 9 (NL4-3) were measured by ELISA. (*p < 0.05, **p < 0.01 by t test between None and each TFV sample) ND means not detected. (C) The supernatants from CD14-derived macrophages and osteoclasts infected with JR-FL (2.5 × 105 IFU/ml) or NL4-3 (2.5 × 105 IFU/ml) in the absence or presence of 4 μg/ml TFV were collected on day 9 after infection (shown in Figure 3C). MAGIC5 cells, susceptible to both R5 and X4 HIV-1, were incubated for 6 hours with each supernatant. Three days after infection, the cells were fixed and stained with anti-p24 antibody and Hoechst 33258. The supernatant from the uninfected macrophages was used as a control (Cont.). Scale bar, 100 μm.