Umbilical cord blood progeny cells that retaina CD34+ phenotype after ex vivo expansion have less engraftment potential than unexpanded CD34+ cells

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Abstract
BACKGROUND: Because of the limitation of cell numbers associated with cord blood harvests, there is a need to determine the efficacy of using ex vivo‐expanded cord blood cells in a transplantation setting. In this study, limiting‐dilution analysis was used in nonobese diabetic mice with severe combined immunodeficiency (NOD/SCID) to compare the engraftment potential of progeny cells expressing the CD34+ phenotype after expansion with that of uncultured CD34+ cells.STUDY DESIGN AND METHODS: Cord blood CD34+ cells were cultured in Iscove's modified Dulbecco medium supplemented with 10‐percent fetal calf serum (FCS) and IL‐6, SCF, megakaryocyte growth and development factor, and Flt3 ligand. The resulting ex vivo‐expanded products were assessed for total numbers of nucleated cells, CD34+ cells, and CFUs and long‐term culture‐initiating cell activity. The engraftment potentials of cultured progeny CD34+ cells and uncultured CD34+ cells were determined by using NOD/SCID mice.RESULTS: After 14 days of culture, total nucleated cell counts increased over input values by 180 ± 59‐fold, CD34+ cell numbers by 44 ± 13‐fold, CFU activity by 23 ± 5‐fold, and long‐term culture‐initiating cell activity by 20 ± 6‐fold (mean ± SD; n = 6). The frequency of SCID‐repopulating cells (SRC) in mice transplanted with uncultured products was 1 per 20,000 CD34+ cells (95% CI, 1:10,000‐1:38,000) and that in mice receiving ex vivo‐expanded products was 1 per 418,000 progeny CD34+ cells (95% CI, 1:158,000‐1:1,100,000). Taken together, these data indicated that, after 2 weeks of culture, there was a modest twofold increase in the total number of SRCs. However, the levels of human CD45 cell engraftment in NOD/SCID recipients of progeny CD34+ cells were significantly lower than those in mice receiving equivalent numbers of uncultured CD34+ cells (pCONCLUSION: Umbilical cord blood progeny cells retaining a CD34+ phenotype after ex vivo expansion have less engraftment potential than do unexpanded CD34+ cells.