Secondary bile acids expand myeloid cells via direct interaction with bone marrow. (A) Whole bone marrow was placed in either (1) colony-forming media or (2) liquid coculture with AFT024 cells. (B) Myelopoiesis schematic representation; CFU-GEMM, CFU-GM, MEP, and BFU-E. (C-E) Whole bone marrow was cultured in colony-forming media with DCA, CA, or LCA. Colonies were analyzed as (C) percentage of total colonies, (D) number of individual colony types, and (E) total colony numbers. (F) Wright-Giemsa staining of FAC-sorted GMPs from colony-forming assays; bar represents 50 μM size. (G-J) Flow cytometric analysis of frequencies of (G) CMPs, (H) CMPs, (I) neutrophils, and (J) macrophages from liquid coculture experiments. All data are shown and represent 3 (colony-forming assay) or 2 (liquid coculture) experimental replicates and were analyzed using one-way analysis of variance (ANOVA) with Tukey post hoc test, or two-way ANOVA with Dunnett post hoc test. ∗P < .05; ∗∗P < .01; ∗∗∗P < .001; ∗∗∗∗P < .0001. MEPs, megakaryocyte erythrocyte progenitors.