Effect of Parp-2 deficiency on HSPC proliferation and apoptosis. (A-B) In vivo proliferation of Parp-2^+/+ and Parp-2^−/− LSK and MP cells was determined by intraperitoneal injection of 6-week-old mice with BrdU (1 mg/6 g mouse weight) and provided BrdU in their drinking water for 3 days. BrdU incorporation was analyzed by flow cytometry. Representative histograms (A) from 2 independent experiments including Parp-2^+/+ (n = 3 per experiment) and Parp-2^−/− (n = 3 per experiment) mice are shown. Numbers indicate percent proliferating (BrdU⁺) cells. (B) Bars represent the mean ± SEM values of the percentage of BrdU⁺ cells obtained from 6 mice per genotype. (C) Effect of Parp-2 deficiency on quiescence of LSK and MP cells. Representative staining profiles for LSK and MP cells analyzed for Ki67 and DNA content. The percentage of cells in each quadrant represents the mean from at least 6 mice in each group. (D) Percentage of Parp-2^+/+ and Parp-2^−/− LSK and MP cells that are in G0, G1, and S-G2/M phases of cell cycle. Bars represent the mean ± SEM obtained from 6 mice per genotype. (E) Parp-2^–deficient LSK and MP cells showed increase levels of ROS. Representative histograms showing LSK and MP cells loaded with the ROS detection reagent, 5-(and -6)-carboxy-2′,7′-difluorodihydrofluorescein diacetate (H2DFFDA). (F) The relative ROS level was calculated on the basis of the mean fluorescence intensity of the H2DFFDA and was presented as fold induction compared with the control group. Bars represent the mean ± SEM obtained from 5 mice per genotype. (G) Representative immunofluorescence images of active caspase-3 in LSK and MP cells derived from Parp-2^+/+ and Parp-2^−/− mice at basal and 2 hours after γ-irradiation (IR) (5 Gy). Green represents active caspase-3; blue represents DAPI. (H) Bars represent the percentage of cells positive for active caspase-3. Values represent the mean ± SEM obtained from at least 6 mice per genotype. *Statistically significant difference (P < .05).