Figure 2.

Bmi-1 Is Sufficient to Induce the Extensive Ex Vivo Self-Renewal of Adult Erythroblasts

(A) Lentiviral transduction of mouse Bmi-1 led to prolonged proliferation of bone-marrow-derived SREs grown in erythroid expansion media. Erythroid cells transduced with an empty vector proliferated for 2 weeks, while erythroid cells transduced with a Bmi-1 overexpression vector stably proliferated for more than a month (representative date from one of ten independent experiments). The dotted line represents expected cell proliferation if cells divide daily.

(B) Bmi-1-induced ESRE (iESRE) was maintained for 100 days, exhibiting more than 10³⁰-fold total erythroblast expansion.

(C) The percentage of erythroblasts transduced with Bmi-1 (GFP⁺ cells) when analyzed 3 days and 18 days after infection. Representative data from one of two independent experiments are shown.

(D) BMI-1 protein expression is increased in erythroblasts transduced with FUGW-Bmi1. Actin served as an internal control. Representative data from one of three for ESREs and one of four for iESREs independent experiments are shown.

(E) Bmi-1-induced iESREs exhibited a high nuclear to cytoplasmic ratio and basophilic cytoplasm resembling proerythroblasts.

(F) iESREs express both KIT (CD117) and transferrin receptor (CD71) on their cell surface. Representative data from one of three independent experiments are shown.

(G) iESREs remain dependent on the continued presence of EPO, SCF, and DEX for ex vivo self-renewal. Withdrawal of individual factors resulted in the rapid loss of cell proliferation. One representative culture of five independent experiments is shown.

See also Figure S2.