SRP54 mutations induce a great impairment of granulocytic differentiation. CD34⁺ cells from healthy donors (controls) or patients were cultured for 14-21 days in serum free-medium with SCF, IL-3, and G-CSF. (A) The fold increase in proliferation in log scale was assessed individually by Trypan blue exclusion, using controls (= 4) and patients (n = 5). (B) Fold increase in proliferation in patients and controls at days (D) 7, 10, and 14. Mean ± SEM; ***P < .001. (C) Apoptotic granulocytic cells were analyzed by flow cytometry using Annexin V⁺ assay in controls (n = 2) and patients (n = 2). The MDM2 antagonist, Nutlin 3a (20 µM for 24 hours), was used as positive control at D9. (D) The results represent the mean of 3 independent experiments at days 7, 9, and 13 with controls (n = 2) and patients (n = 2). Mean ± SEM; *P < .05; ***P < .001. (E) P53 target genes (P21, BAX, NOXA) expression levels were checked by qRT-PCR related to PPIA and HPRT at days (D) 7, 10, and 14 with controls (n = 3-4) and patients (n = 2-4). Mean ± SEM; *P < .05. (F) Granulocytic cells were sorted as CD15⁺CD11b⁺CD36⁻ cells between days 10 and 14, using CD36 to specifically eliminate monocytes. P53 target genes (P21, BAX, NOXA) expression levels were checked by qRT-PCR related to PPIA and HPRT with controls (n = 4) and patients (n = 3). Mean ± SEM; *P < .05; **P < .01.