The Effect of Freezing on the Recovery and Expansion of Umbilical Cord Blood Hematopoietic Stem Cells

Objectives: Cell populations residing in waste tissues (cord blood, umbilical cord, and placenta) may be collected without any medical or ethical contraindications concerning the mother or newborn baby. Cord blood hematopoietic stem cells are routinely used for clinical transplants; however, the low cell dose of the graft limits their therapeutic efficacy as it is associated with increased delayed or failed engraftment. The cell dose can be increased and the efficacy of cord blood transplant potentially improved, by ex vivo expansion before transplant. Materials and Methods: Twelve umbilical cord blood samples were included. The effect of cord blood storage at -80 degrees C on CD34(+) cell count (mean standard deviation [SD]), cell viability (mean SD percent), and cell cycle status (percent quiescent versus dividing) was estimated. Ex vivo culture of cord blood mononuclear cells was done before storage, and after 1 week of freezing, and after 2 weeks of freezing. Ex vivo liquid culture was performed with media supplemented with stem cell factor, interleukin-3 (IL-3), and both. Results: The count of CD34(+) cells in pre-expansion aliquots decreased from 15.00 +/- 9.96 x 10(6) cells before freezing to 7.70 +/- 3.20 x 10(6) cells after 2 weeks of freezing (P = .024). Cell viability in pre-expansion aliquots decreased from 99.5% +/- 1.0% before freezing, to 52.5% +/- 27.5% after 1 week of freezing (P = .013) and to 32.5% +/- 9.5% after 2 weeks of freezing (P = .001). Mean fold of cell expansion and proportion of quiescent versus dividing cells did not change significantly from before to after freezing, and was not significantly different for culture with stem cell factor, IL-3, or both. Conclusion: Although freezing decreased cell count and viability, it did not impair the expansion potential of cord blood hematopoietic cells. Whether IL-3 or stem cell factor should be considered as essential components of expansion media is uncertain.