TBI induced hematopoietic suppression. (A) TBI induces a transient decrease in the number of BM-MNCs. Mice were exposed to 6.5 Gy TBI. At 1, 3, 7, 14, 28, and 56 days after TBI, BM-MNCs were isolated and enumerated. The results are expressed as the mean of BM-MNCs/hind leg ± SE (n = 3 mice/group). A group of 6 unirradiated mice was used as control (day 0). (B) Representative phenotypic analysis of BM-MNCs by flow cytometry. BM-MNCs (1 × 10⁶) from control or irradiated mice were incubated with biotin-conjugated antibodies against CD3e, CD45R/B220, Gr-1, Mac-1, and Ter-119 and then with streptavidin-FITC. After incubation with anti-CD16/CD32 antibody, they were stained with anti-Sca-1-PE and anti-c-kit-APC antibodies. Cells incubated with appropriate conjugated rat antimouse isotypes were included as controls. For each sample, a minimum of 200 000 cells was analyzed on a FACS Caliber (Becton Dickinson) and the resultant data were analyzed using CellQuest software (Becton Dickinson) after gating on viable cells. (C) TBI induces a sustained reduction in the frequencies of LKS^- and LKS⁺ cells in BM-MNCs. The frequencies of LKS^- and LKS⁺ cells in BM-MNCs from control and irradiated mice were determined by flow cytometry as illustrated in panel B. Top panel: Representative flow cytometric analyses of LKS^- and LKS⁺ cells in BM-MNCs from control or irradiated mice. The frequencies of LKS^- and LKS⁺ cells are indicated in the LKS^- and LKS⁺ cell gates as a percentage of total BM-MNCs. Bottom panel: Kinetics of LKS^- and LKS⁺-cell recovery after TBI. The frequencies of LKS^- and LKS⁺ cells are expressed as a mean percentage of BM-MNCs ± SE (n = 4 to 6). ANOVA analysis reveals that TBI induced a time-dependent reduction in the number of BM-MNCs and frequencies of LKS^- and LKS⁺ cells (P < .001).