Figure 6

Functional features of Mo-DC from patients receiving pegfilgrastim. Monocytes were purified from the peripheral blood of patients given pegfilgrastim and were cultured in the presence of either pre-G-CSF or post-G-CSF serum (20% v/v) for 6 days, as detailed in Materials and Methods. Control cultures consisted of immunogenic DC preparations that were differentiated with GM-CSF and IL-4 without the provision of additional maturation stimuli (^GM4DC). Panel A: Uptake of FITC-conjugated dextran by monocytes cultured in vitro in the presence of pre-pegfilgrastim serum (day 0) or post-pegfilgrastim serum (days +7 and +11). Median values and interquartile range are shown. *p < 0.05 compared with Mo-DC differentiated with GM-CSF and IL-4; §p < 0.05 compared with cells cultured with pre-G-CSF serum. Panel B: Representative experiment; red histograms depict the uptake of FITC-conjugated dextran by monocytes kept at 4°C (negative control) and empty histograms depict the uptake of FITC-conjugated dextran by the monocyte preparations kept at 37°C. Panel C: CD4⁺CD25^- T cells and monocytes were purified from the peripheral blood of healthy donors as detailed in the main text. After irradiation, monocytes were cultured with CFDA-SE loaded, allogeneic T cells at a fixed monocyte-to-T cell ratio (1:27) for 7 days, either in the absence or presence of patient serum (20% v/v). The proliferation index of T-cell cultures established in the presence of patient serum collected before and after G-CSF administration is shown. The bars depict median and interquartile range recorded in 3 independent experiments performed in duplicate. Panel D: Results of a representative experiment out of 3 with similar results. The percentage of parental, undivided cells (U; depicted in blue) is indicated. The analysis of CFDA-SE fluorescence was performed with the proliferation wizard of the ModFit software package, as previously detailed [12].