The EPΔ578ER and EPΔ623ER proteins immortalize progenitors that exhibit strict granulocytic or monocytic differentiation.

Single-cell progenitor's clones were generated that exhibited strict granulocytic or monocytic differentiation upon removal of estrogen, and they were designated ECoM-G (E2a/Pbx1-mediatedConditional Myeloid Differentiation-Granulocytic) and ECoM-M (Monocytic), respectively. (A) Light micrographs and (B) Wright-Giemsa stains were prepared from ECoM-G and ECoM-M cells in the presence (+β-E) and absence (−β-E) of estrogen as indicated above the photographs. (C) NBT reduction and NSE staining. The ECoM-G cells were functionally characterized based on NADPH oxidase activity (NBT reduction), which is up-regulated during myeloid maturation. The ECoM-M cells were analyzed for NSE activity, which is specifically expressed in mature macrophages. (D) Flow cytometric analysis of the cell surface staining with FITC-labeled Ly6G (GR-1), CD11b (Mac-1), and F4/80 on the ECoM-G and ECoM-M cells grown in the presence or absence of estrogen for 7 days. Note that the observed shift in GR-1 staining in the ECoM-M clone is due to increased autofluorescence. (E) Phagocytosis assay. The ability of the ECoM cells to internalize FITC-labeledE coli was evaluated in the presence and absence of estrogen. Hoechst and rhodamine-phalloidin were used to stain the nuclei, and polymerized actin, respectively.