FGF signaling regulates HSPC quiescence versus cycling states and HSPC intracellular ROS levels. C57BL/6 mice were treated with either PBS or FGF-2 (n > 7). (A) Percentage of G₀ quiescent (Ki67⁻/7-amino-actinomycin D-low [7AAD^low]) SKL cells determined by flow cytometry. (B) Representative flow cytometric analysis of cell-cycle status of SKL cells. Numbers indicate the mean percentage ± SEM of G₀ quiescent (Ki67⁻/7AAD^low) SKL cells. (C) Percentage of G₀ quiescent (Ki67⁻) SLAM cells determined by flow cytometry. (D) Representative flow cytometric analysis of Ki67 expression inside SLAM cells. Numbers indicate mean percentage ± SEM of G₀ quiescent (Ki67⁻) SLAM cells. (E) Percentage of ROS^low-expressing SKL cells as determined by flow cytometry. (f) Representative flow cytometric analysis of ROS expression inside SKL cells. SKL were further divided into subpopulations of ROS^low (ROS-L)–, ROS^intermediate (ROS-I)–, and ROS^high (ROS-H)–expressing cells. Lin⁻ progenitors were enriched from C57BL/6 mice total BM cells and cultured for 5 days with a proliferation-inducing cytokine mixture (erythropoietin, SCF, GM-CSF, and IL-3) supplemented with FGF-2 or FGFR inhibitor (SU5402; n = 4 experimental repeats). (G) Frequency of cultured SKL cells as measured by flow cytometry. (H) Percentage of ROS^low-expressing cultured SKL cells as determined by flow cytometry. (I) Representative flow cytometric analysis of ROS expression inside cultured SKL cells. (J) B6.SJL recipient mice were sublethally irradiated (n = 15) and transplanted with C57BL/6 3 × 10⁴ Lin⁻ cultured cells from DMSO-, FGF-2-, or SU5402-treated cultures (n = 5). Mice were killed and levels of engraftment were determined by examining the percentage of CD45.2 donor–derived chimerism. *P < .05; **P < .01. Data shown are means ± SEM.