Cytokine-mediated expansion does not deplete cord blood cells with stem cell characteristics

Cord blood (CB) has been successfully used to regenerate the hematopoietic system after myeloablative therapy. We investigated whether cytokine mediated expansion depletes CB of cells with stem cell characteristics. CB mononuclear cells (MNC) were enriched for quiescent (primitive) stem cells by incubation with 25 μg/ml 5-Fluorouracil (5-FU) and control CB MNC were incubated with media alone. Cells were then incubated for 7 days with Interleukin-1 (IL1)+IL3+ Stem Cell Factor (SCF) and progenitor content, cell cycle status, nucleated cell count, immunophenotype and resistance to 25 μg/ml 5-FU (primitive stem cells) were evaluated before and after cytokine exposure. Incubation with IL1+IL3+SCF caused an increase (fold expansion) in committed (28.6 ± 8.1), immature (5.8 ± 1.8), and primitive progenitors (4.1 ± 0.8) among control CB MNC compared to a decrease in committed progenitors (0 ± 0) but an increase in both immature (8.4 ± 4.8) and primitive progenitors (7 ± 2.9) among 5-FU resistant CB MNC. An increase in the proportion of CD34⁺ cells occurred in both fractions. Expanded control CB MNC showed a significant increase in numbers of 5-FU resistant committed (p = 0.024), immature (p = 0.014) and primitive progenitors (p = 0.01) as compared with fresh CB MNC. Re-exposure of 5-FU resistant expanded CB MNC to 5-FU shows growth of some immature and primitive progenitors. Cytokine-mediated expansion of untreated and quiescent CB cells is possible and cytokine-mediated expansion does not deplete CB cells with stem cell characteristics.