Constitutive mitochondrial release of cytochromecin Kostmann progenitor cells. (A) Mitochondria of CD34⁺ and CD33⁺ cells were stained with MitoTracker Red, and cytochrome c localization was revealed by indirect immunofluorescence. Representative images of cells from patient 3 and from a healthy control are displayed. The punctate yellow pattern denotes mitochondrial localization of cytochrome c, and the diffuse green pattern indicates cytochrome c that has been released into the cytosol (see insets for typical examples). Original magnification × 40. (B) Release of cytochrome c from mitochondria of CD34⁺ progenitors of Kostmann patients and healthy controls, as determined by confocal microscopy. Black bars indicate freshly isolated cells; white bars, 4 hours in vitro incubation (no G-CSF); and gray bars, 4 hours in vitro incubation (with G-CSF; 100 ng/mL). All Kostmann patients were maintained off therapy for 60 hours prior to bone marrow aspiration. (C) Release of cytochrome c from mitochondria of CD33⁺ progenitors of Kostmann patients and controls, determined as in panel B.