Elevated levels of ROS and oxidative DNA damage in primitive CML-CP CD34⁺ subsets. (A) DHE, MSR, CC1, and DCFDA were used to detect various ROS with different DNA damaging capabilities (left). ROS was measured in CD34⁺CD38⁻, quiescent CD34⁺CFSE^max (CFSE^max), and total CD34⁺ cells from healthy donors (black bars) and CML-CP patients (gray bars) in 17% and 1% O₂, as indicated. (B) Diagram showing oxidative DNA lesions: 8-oxoG and DSBs (stained by γ-H2AX; left). 8-oxoG and γ-H2AX were detected by specific fluorescence in CD34⁺CD38⁻, quiescent CD34⁺CFSE^max (CFSE^max), and total CD34⁺ cells from healthy donors (black bars) and CML-CP patients (gray bars) maintained in 17% and 1% O₂, as indicated (CD34⁺ ± NAC). CD34⁺ cells from healthy donors (black bars) and those from CML-CP patients were incubated (white bars) or not (gray bars) with 50μM N-acetyl-L-cysteine for 48 hours. ROS (DCFDA fluorescence), 8-oxoG, and γ-H2AX foci were detected. (C-D) ROS (DHE, MSR, CC1, and DCFDA fluorescence) and/or 8-oxoG was detected in CD34⁺CD38⁻ and CD34⁺CD38⁺ CML-CP cells (C) and in CD34⁺(CFSE^max or CTV^max) quiescent and CD34⁺(CFSE^low or CTV^low) proliferating CML-CP cells (D). Results represent means of 3-20 samples/group ± SD. *P < .05 compared with healthy counterparts (A-B), these treated with NAC (B), and CFSE^low or CTV^low (D).