Figure 1
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+CFSEmax (CFSEmax), 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+CFSEmax (CFSEmax), 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+(CFSEmax or CTVmax) quiescent and CD34+(CFSElow or CTVlow) 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 CFSElow or CTVlow (D).

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+CFSEmax (CFSEmax), 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+CFSEmax (CFSEmax), 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+(CFSEmax or CTVmax) quiescent and CD34+(CFSElow or CTVlow) 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 CFSElow or CTVlow (D).

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