SDF-1 increases the number of megakaryocytic progenitor-derived colonies from PB Inc− and Inc+ CD34+ cells.
PB CD34+ cells were purified (A) without incubation on a plastic support (Inc−) or (B) after overnight incubation (Inc+) and plated in duplicate at a density of 1 × 104 cells/mL in a serum-free Easymega medium containing IL3, IL6, and TPO in the presence or absence of SDF-1 (0.05 ng/mL) or anti–SDF-1 antibody (5 ng/mL) or isotype control (5 ng/mL). Colonies were scored after 10 to 14 days of culture. Two differentiation stages were distinguished on the basis of clone size on day 14. Colonies containing more than 10 cells were classed as BFU-MK, whereas colonies containing fewer than 10 cells were classed as CFU-MK. Results are expressed as mean percentage of SDF-1 untreated control cells (CT) ± SD. Control colony numbers for CFU-MK and BFU-MK from PB Inc− cells were 118.7 ± 43 and 35.6 ± 12.1, and for PB Inc+ cells, 149 ± 27.7; 46.4 ± 15.3, respectively. Control plating efficiency calculated on the total number of MK colonies derived from PB Inc− and PB Inc+ CD34+ cells was 1.5% ± 0.6% and 2% ± 0.75%, respectively (n = 3 independent experiments). * indicates significant difference from control values, P < .05.
