Defects in Gdf15^−/−stroma contribute to impaired expansion of SEPs in vitro. (A-E) Gdf15^−/− BM cells were cultured in SEEM in vitro with or without the addition of WT CD11b⁺Ly6C⁺ monocyte (WT MO). After coculturing for 7 days, total cell numbers are presented in panel A. (B) Representative flow cytometry analysis of erythroid progenitor surface markers. (C) The percentages of Kit⁺Sca1⁺ SEPs (left) and absolute numbers of Kit⁺Sca1⁺ SEPs (right). (D) The percentages (left) and absolute numbers (right) of CD34⁺CD133⁺Kit⁺Sca1⁺ SEPs. Addition of WT monocytes promoted the expansion of Gdf15^−/− SEPs. (E) Stress BFU-E colony production in cultures ± WT monocytes. (F-I) WT and Gdf15^−/− BM cells were cultured in SEEM for 3 days. Nonadherent progenitor cells were collected with culture media and plated on the stromal layer of the indicated genotype. Progenitors were cultured for another 4 days before analysis. (F) Flow cytometry analysis of living vs dead progenitor cells. (G) Flow cytometry analysis of Kit⁺Sca1⁺ stress erythroid progenitors of the indicated genotype grown on the indicated stromal genotype. (H) Statistical analyses on percentages (left) and absolute numbers (right) of Kit⁺Sca1⁺ SEPs. (I) Statistical analyses on percentages (left) and absolute numbers (right) of CD34⁺CD133⁺Kit⁺Sca1⁺ SEPs. Data are shown as individual subject and the mean ± SEM. *P < .05, **P < .01, and ***P < .001.