Hypoxia suppresses growth and imatinib-induced apoptosis in CML progenitors. (A) CP CML CD34⁺CD38⁻ or CD34⁺CD38⁺ cells (1 × 10⁵ cells each, n = 4) were treated with imatinib at 21% or 0.5% O₂ for 96 hours, and the total number of viable cells was determined by trypan blue exclusion and flow cytometry. Cell numbers are indicated on the top of each column. (B) Cells were treated as in panel A, and the percentage of apoptotic cells was measured by Annexin V staining using flow cytometry (n = 4). Representative flow cytometry dot plots are shown. (C) Flow cytometric evaluation of pCrkL in CML cells. CD34⁺ cells were cultured for 24 or 96 hours and subjected to flow cytometric analysis. Viable cells were identified with the fixable viability dye (FVD; eFluor), and pCrkL levels were measured by intracellular staining in both CD34⁺CD38⁻ and CD34⁺CD38⁻ subpopulations. Flow cytometry dot plots and histograms show the results after 24 hours treatment. The background signals were determined in the same populations stained with the appropriate isotype controls. (D) Mean fluorescence intensity of pCrkL in multiple samples is shown (n = 5 and 3). *P < .05, **P < .01.