Warfarin compromises hematopoiesis and HSC function. (A-B) Total number of leukocytes (P = .009 and P = .003; ANOVA, Tukey test, n = 10-16) (A) and monocytes (P = .008 and P = .001; ANOVA, Tukey test, n = 10) (B) in peripheral blood of control mice (red circles) or mice treated with 0.5 mg/kg (blue squares) or 0.05 mg/kg (green triangles) warfarin 14 days after the beginning of treatment. The data were generated in 3 independent experiments. (C) Percentage of CD11b⁺ myeloid cells in peripheral blood of control mice (red) or mice treated with 0.5 mg/kg (blue) warfarin 14 days after begin of treatment (P = .008; t test, n = 8). The data were generated in 3 independent experiments. (D) Number of colonies in methylcellulose derived from BM cells from controls (red circles) or warfarin (blue)-treated mice (P = .02; t test) on day 10 after plating. Ten thousand Lin⁻ cells were plated. Five donor mice were used. The experiment was performed in triplicate for each donor mouse. The data were generated in 2 independent experiments. (E-F) Total number of leukocytes (P = .004; t test) 7 days after a 1-time treatment with 200 mg/kg 5-FU (n = 10) (E) and Kaplan-Meier-style survival curve (F) of control mice (red line) (n = 8) or mice treated with 0.5 mg/kg (blue dashed line) (P = .0004; Log-rank test, n = 8) or 0.05 mg/kg warfarin (green dotted line; = .016; Log-rank test, n = 5) cotreated with 75 mg/kg 5-FU on days 0, 7, 14, 21 (starting on day 14 of warfarin treatment). The schematic details the treatment schedule. (G-I) Competitive (P values as indicated; t test, n = 5) (G), serial (P values as indicated; t test, n = 5) (H), and limiting dilution (P = .0006) (I) transplantation assays using 1 × 10⁶ CD45.1⁺ total donor BM cells from control (red line) mice or warfarin-treated mice (blue line) for the competitive and serial transplantation assay. In the competitive transplantation assay 10⁶ CD45.2⁺ competitor cells were cotransplanted. The limiting dilution assay (I), in which 5 × 10⁵, 12.5 × 10⁴, 6 × 10⁴, and 1.5 × 10⁴ pooled CD45.1⁺ BM cells were transplanted into 5 CD45.2⁺ mice per group, shows the percentage of nonengrafted mice (y-axis) in relation to the dose of transplanted BM. The lines fitted by regression analysis (L-Calc) allow estimation of functional HSC frequency, which was 7.6-fold lower in warfarin-treated donors. (J) Number of colonies arising from 50 000 plated Lin⁻ BM cells pretreated with vehicle (red) or 2 μM warfarin (blue) for 7 days in methylcellulose in the absence of drugs. Colonies were scored on day 10 after plating (n = 6). The data were generated in 3 independent experiments. (K) Relative leukocyte count 7 days after a 1-time treatment with 200 mg/kg 5-FU in control mice treated without (red circles) or with vitamin K (blue squares; 15 mg/kg) or mice treated with 0.5 mg/kg warfarin (green triangles) or warfarin plus vitamin K (purple inverted triangles; P = .02; Kruskal-Wallis test, n = 4-10). Warfarin and vitamin K had been administered concomitantly for 14 days before the dose of 5-FU.