Effect of ROS scavenging on the Meis1^−/− phenotype. (A) Schematic of NAC administration. We performed daily IP injections for tamoxifen for 14 days followed by daily NAC injections up to 12 days. (B) Flow cytometry profile of LT-HSCs (Lin⁻Sca-1⁺Kit⁺Flk2⁻CD34⁻) of Meis1^+/+ and Meis1^−/− mice after 12 days NAC administration. Note the number of HSCs in Meis1^−/− mice is now similar to Meis1^+/+ values (n = 3). (C) Left panel, FACS plot of Pyronin Y/Hoechst staining of LT-HSCs. Right panel: Quantification of flow cytometric analysis of cell cycle of Meis1^+/+ and Mesi1^−/− LT-HSCs demonstrates restored numbers of G₀ cells in Meis1^−/− cells which indicates restored quiescence of LT-HSCs (n = 3). (D) Quantification of apoptosis in Meis1^+/+ and Meis1^−/− LSK cells (Lin⁻Sca-1⁺Kit⁺) showing persistent trend toward an increase in the number of apoptotic cells, which was not statistically significant (P = .059; n = 3). (E) Quantification of ROS in Meis1^+/+ and Meis1^−/− LT-HSCs (Lin⁻Sca-1⁺Kit⁺Flk2⁻CD34⁻) after NAC treatment showing only a modest increase in ROS in Meis1^−/− HSCs (n = 3). (F) Real-time PCR of HSCs isolated from Meis1^+/+ and Meis1^−/− HSCs after NAC treatment demonstrating no change in p16 and p19 transcripts (n = 3). (G) Schematic of NAC administration and bone marrow transplantations. We performed daily IP injections for tamoxifen and NAC for 14 days followed by bone marrow transplantation. Then, NAC is provided in drinking water for 2 weeks and administrated another 2 weeks. Repopulation was examined at 4 weeks after transplantation. (H) Analysis of repopulation after NAC treatments of BMTs from Meis1^+/+ and Meis1^−/− mice demonstrates restoration of repopulation defect after Meis1 deletion (n = 5). (I) Schematic of proposed model is demonstrating how Meis1 regulates metabolism and maintenance of HSCs through its role on Hif-1α and Hif-2α.