A24 inhibits Tf uptake and impairs TfR expression and recycling. (A) A24 inhibits Tf uptake. PBMCs were incubated with PHA and IL-2 for 24 hours before being plated in culture. To assay Tf uptake, activated cells were incubated for different time intervals with A24 (□) or 30.9 isotype control (▦) in the presence of 2.5 μM [⁵⁵Fe]-Tf. Data are means ± SD of one experiment of 3 separate experiments. (B-C) A24 impairs TfR expression and recycling. (B) Expression of TfR and CD25 was followed on PBMCs from healthy donor (i) and patients with chronic (ii) and acute (iii) ATL. Cells were activated with PHA and IL-2 for 72 hours, in the presence of A24 or a control antibody (30.9). The inhibition percentage of the mean fluorescence intensity linked to TfR and CD25 expression is indicated in histograms located at the upper right corner of right panels. (C) CIB cells were stained at 4°C with A24 and plated under culture conditions at 37°C for different times (0-180 minutes) to determine the effect of A24 binding on TfR trafficking. TfR revealed with an antimouse-Cy-5 (blue) colocalized with the WGA-Alexa-488 membrane staining (green) at the initial stage (0 minutes). Colocalization disappeared progressively thereafter. Original magnification, × 400.