Early-stage Lyn^-/-erythroid progenitor cells accumulate at a Kit^posCD71^high stage of development and are deficient in Epo and SCF proliferative responsiveness. (A) At the indicated time points, Kit^posCD71^low, Kit^posCD71^high, and Kit^negCD71^high erythroblast formation was assayed. At 48 hours, 2 subpopulations of Kit^pos cells reproducibly were detected (and are designated cohorts I and II). For Lyn^-/- erythroblasts, note accumulations at a Kit^posCD71^high stage (especially at 72 and 120 hours) and corresponding deficits in Kit^negCD71^high cell formation. Data illustrated are representative of 3 independent experiments (and additional representative experiments are illustrated in Figure S1, available at the Blood website; see the Supplemental Figures link at the top of the online article). (B) Also graphed are ratios of Kit^pos/Kit^neg erythroblast formation for wild-type versus Lyn^-/- progenitors at 48, 72, and 120 hours of culture. (C) Over an extended period of culture, expansion capacities of Lyn^-/- and control Lyn^+/+ cultures also were assayed by direct cell counts. Note the deficient expansion of Lyn^-/- erythroblasts (observed in 3 independent experiments). (D) Levels of Epo- and SCF-induced ³HdT incorporation in expanded Lyn^-/- and control erythroblasts. Expanded Kit^posCD71^high erythroblasts were isolated by lineage depletion and MACS, and cultured for 24 hours in SP34-EX medium in the presence of Epo and/or SCF as indicated. Cultures then were pulsed with ³HdT, and mean incorporation rates (± SE) were determined.