Expression of APRIL and its receptors during MK differentiation. (A) Flow cytometry analysis of cord blood CD34⁺ cells undergoing unilineage MK differentiation (from day 3 to day 14) labeled with recombinant Flag-APRIL (open histograms) as compared with staining controls (black histograms). Jurkat and 293 cells were used as positive and negative controls, respectively. (B-C) Semiquantitative RT-PCR (B) and real-time (C) analyses of CD34⁺ and differentiating megakaryocytic cells. Peripheral blood mononuclear cells (PBMCs) and hematopoietic progenitors transduced with empty vector (PINCO) or with APRIL cDNA (APRIL) were used as controls. Error bars indicate ± standard deviation (SD). (D) Immunoblot analysis of APRIL expression in CD34⁺ and differentiating megakaryocytic cells. 293 cells transfected with empty vector (P) or with APRIL cDNA (APRIL) were used as controls.