STAT5-induced long-term growth is enhanced in the absence of GATA1. (A) CD34⁺ CB cells were transduced with STAT5A-ER and the indicated lentiviral shRNA constructs. After transduction, cells were cultured on MS5 stromal cells and either left untreated or stimulated with 4-OHT. Each week, half of the medium was removed and the cells were counted and analyzed by flow cytometry to calculate the number of double-transduced cells. The cumulative expansion of double-transduced cells was calculated relative to the starting amount of double-transduced cells, which was set at 1. Data are mean ± SD of 2 experiments, which is representative of 4 experiments performed. (B) The same cells as in panel A were stained for GPA, CD11b, CD14, and CD15. The cumulative fold expansion of the cells, separated by the respective phenotype, is shown. (C) Double-transduced CD34⁺ CB cells as in panel A were sorted after 1, 2, 3, and 5 weeks, and progenitor frequencies per 10 000 plated cells were determined by culture in CFC-mix methylcellulose medium. (D) Cumulative expansion of progenitors was calculated based on the progenitor frequencies depicted in panel C and the cumulative expansion as shown in panel A. (E) The absolute amount of colonies at week 1, divided by BFU-E, and CFU-GM was calculated according to the expansion shown in panel A. (F) Early (day 10) cobblestone frequency of CB CD34⁺ cells, transduced as in panel A and cultured in limiting dilution on MS5 stromal cells. Significant differences (2-tailed t test): *P < .05, **P < .01.