Serum supplement, inoculum cell density, and accessory cell effects are dependent on the cytokine combination selected to expand human HPCs ex vivo

BACKGROUND: The prolonged periods of pancytopenia associated with cord bloo d transplants suggest that in some cases cell numbers may be limiting. The possibility that limiting cell numbers may be overcome and prolonged period s of pancytopenia abrogated by the transplantation of human umbilical cord blood cells expanded ex vivo has led to efforts to define optimal culture c onditions for these cells. STUDY DESIGN AND METHODS: Cord blood CD34+ cells were cultured with three c ytokine combinations: SCF+G-CSF+GM-CSF+MGDF (SGGM); IL-6+ SCF+MGDF+Flt3-lig and (6SMF); and IL-1+IL-3+IL-6+G-CSF+GM-CSF+SCF+Epo (GFmix). Serum effects, inoculum concentration (cells/mL) seeding density (cell/ cm(2)) and access ory cell effects on the expansion of CD34+ cells were determined. RESULTS: Cellular outputs were significantly higher with fetal calf serum ( FCS) than with cord blood serum (CBS) or adult group AB serum (ABS) in the presence of 6SMF, however, CBS was as effective as FCS. The best seeding co ncentrations varied for each of the cytokine combinations, and inoculum den sities exceeding 1000 cells per cm(2) proved detrimental for cultures conta ining GFmix and SGGM. Accessory cell studies indicated that populations exp ressing the CD33 antigen inhibited the expansion of purified CD34+ cells in the presence of GFmix or SGGM, but not in the presence of 6SMF. CONCLUSION: Serum supplement, inoculum cell concentration, seeding densitie s, and accessory cell effects are dependent upon the cytokine combination s elected to expand cord blood HPCs ex vivo. Thus, each of these measures sho uld be assessed to establish reproducible and reliable conditions for the s election of different cytokine combinations to culture cord blood HPCs.