Myeloid differentiation of human cord blood CD34⁺ cells during ex vivo expansion using thrombopoietin, flt3-ligand and/or granulocyte-colony stimulating factor

We investigated the phenotypic changes of human umbilical cord blood (CB) CD34⁺ cells during ex vivo expansion using thrombopoietin (TPO), flt3- ligand (FL), and/or granulocyte-colony stimulating factor (G-CSF). During ex vivo expansion of CD34⁺ cells isolated from human CB for up to 5 weeks, surface expression of molecules on the cultured cells including CD64 (FcγRI), CD32 (FcγRII). CD16 (FcγRIII), CD11b (MAC-1) and CD18 (β₂- integrin) was analysed by flow cytometry along with simultaneous measurement of apoptosis by 7-aminoactinomycin D staining method. CD64, CD32 and/or CD18 expressing cells appeared in the cultures both with and without the addition of G-CSF until file tenth day. However, without G-CSF, CD16⁺ fractions did not appear and CD11b⁺ fractions were not maintained. With G-CSF, the CD16⁺ or CD11b⁺ fractions appeared only from the second week. These results suggest that G-CSF is necessary for the late stage of myeloid maturation during which CD16 and CD11b are expressed.