ROS levels regulate proplatelet-forming MKs. Mature MKs were obtained from cord blood CD34⁺ progenitors after differentiation for 12 days. They were treated for 5 hours (A-D) under normoxia (20% O₂) with the indicated drugs (NAC, 1 mg/mL; NAM, 5 µM; PEITC, 5 µM) or the appropriate solvent (PBS; A-C; dimethyl sulfoxide; D) as the control. Alternatively, they were incubated for 24 hours (E-F) in a 20% (normoxia) or 95% (hyperoxia) O₂ incubator chamber. (A,E) Representative images of the observed mature MKs upon a 5-hour (A) or 24-hour (E) incubation are presented. (A: *nondeformed MKs; arrowheads, slightly deformed MKs; arrows, proplatelet-forming MKs). Bar represents 10 µm. (B,D,E) Percentage proplatelet-forming MKs relative to total cells observed by phase-contrast microscopy; dot plots show individual data from 3 to 7 independent experiments; means ± SEM (ratio paired Student t test; **P ≤ .01; ***P ≤ .001). (C,F) Cells were incubated with MitoSOX fluorescent dye before being processed by flow cytometry. Mean fluorescence intensity is expressed as fold change over control cells. Data are means ± SEM; ratio paired Student t test; *P ≤ .05.