Smad5-deficient progenitors possess unperturbed differentiation and proliferation and show normal sensitivity to TGF-β1 inhibition in vitro. (A) Single-cell culture evaluating the proliferation capacity of hematopoietic progenitors. Primitive LSK cells from induced mice were purified by FACS and cultured in supplemented serum-free medium containing SCF, TPO, and FL for 12 days (n = 4). Colony assays were performed to evaluate the ability of committed progenitors to form (B) myeloid and (C) erythroid colonies (n = 3). (D-E) Proliferation and colony assay examining the sensitivity of primitive hematopoietic cells and myeloid progenitors to TGF-β1 inhibition, respectively. (D) LSK cells were grown as described for panel A with 0, 0.1, or 1 ng/mL TGF-β1 (n = 3). (E) CFU-GMs were plated with 0, 0.1, 1, or 10 ng/mL TGF-β1 (n = 12, 5, 8, and 4 for the different concentrations, respectively). Data represent mean values ± SD. ▦ indicates wt cells; ▪, Smad5-deficient cells.