Fig. 8

Inhibition of TGFβ signaling regulates HSPC in miR-143/145^−/− mouse marrow and in human del(5q) MDS cells. a Colony-forming unit (CFU) analysis of marrow from WT and miR-143/145^−/− mice transduced with shCtl or shDab2 (mean ± SEM, n = 3). b Primary CFU marrow was replated in equal proportions per condition, normalized to the number of input cells, to generate secondary CFUs (mean ± SEM, n = 3). c Primary and (d) secondary CFU analysis of marrow from WT and miR-143/145^−/− mice transduced with shCtl or shSmad3 (mean ± SEM, n = 3). e CFU assay of marrow cells from WT and miR-143/145^−/− mice treated with the indicated concentrations of galunisertib (mean ± SEM, n = 3). f Primary CFU cells were replated in equal proportions per condition, normalized to the number of input cells, to generate secondary CFUs (mean ± SEM, n = 3). g Marrow was harvested from mice transplanted with miR-143/145^−/− marrow transduced with shCtl or shDab2, and a long-term culture-initiating cell (LTC-IC) assay was performed. Shown is a log-fraction plot of the limiting dilution model. The slope of the line is the log-active cell fraction. The dotted lines give the 95% CI. h Western blot of DAB2 to verify protein knockdown in MDS-L cells using shRNA constructs. i Flow cytometry analysis of marrow from moribund mice transplanted with del(5q) MDS-L cells transduced with shCtl or shDAB2. The proportion of CD38⁻ or CD38⁺ cells within the CD34⁺ compartment is shown (mean ± SEM, n = 4). j Peripheral blood from mice transplanted with del(5q) MDS-L cells transduced with shCtl (n = 13) or shDAB2 (n = 9) was analyzed by flow cytometry 5 weeks post transplant. The percentage of GFP⁺CD33⁺ cells in the live cell population is shown (mean ± SEM)