PCNA impaired pro–caspase-9 activation in vitro. (A) Kinetics of immunodetection of cleaved caspase-9 after an in vitro procaspase-9 activation assay performed using neutrophil cytosol, 50 µM cytochrome c, and 1 mM ATP, without and with 50 µM recombinant PCNA. (B) Kinetics of PCNA immunodetection using PCNA pAb performed in the same in vitro assay as in A in the absence of exogenous recombinant PCNA to detect only endogenous PCNA. (C) Kinetics of cleaved caspase-9 immunodetection as in A in cytosol of neutrophils isolated from G-CSF–treated donors or controls (CT). PCNA Western blot analysis confirmed higher PCNA levels in the G-CSF–treated donors (right). A–C show data from representative experiments that were performed at least four times, yielding identical results.