In Vitro Mass Production of Human Erythroid Cells from the Blood of Normal Donors and of Thalassemic Patients

We describe a new two-step culture method for mass production in vitro of erythroid cells from either CD34+ (105 cells/mL) or light-density (106 cells/mL) cells purified from the blood of normal donors and thalassemic patients. The method includes (i) culture of the cells in the presence of dexamethasone and estradiol (10−6 M each) and (ii) the growth factors SCF (50 ng/mL), IL-3 (1 ng/mL), and EPO (1 U/mL). In their proliferative phase, these cultures generated 1–2 × 107 erythroblasts for each milliliter of blood collected from normal donors or thalassemic patients. They were composed mostly (90%) of CD45low/glycophorin (GPA)neg/CD711ow cells at day 7, 50–60% of which became CD45neg/GPA+/CD71high by days 15–20. However, when cells from days 7 to 12 of the proliferative phase were transferred in differentiation medium containing EPO and insulin, they progressed to mature erythroblasts (>90% benzidinepos and CD45neg/GPA+/CD71medium) in 4 days. Because of the high number of erythroid cells that are generated from modest volumes of blood, this method will prove useful in donor-specific studies of erythroid differentiation.