Cell density-dependent proliferation in frequently-fed peripheral blood mononuclear cell cultures

Background Our goal was to prentice granulocyte progenitor (CFU-G) and post-progenitor (CD15(+)CD11b(+/-)) cells for subsequent transplantation. We hypothesized that increasing the feeding frequency and maintaining constant densities ma y overcome inhibitory growth conditions tie. low pH) in high-density cultur es Methods To study the effect of cell density on total cell expansion, differentiatio n and lactate production, 50% daily medium exchanges were used in cultures of peripheral blood mononuclear cells (PB MNC) maintained at constant densi ties (ranging from 5 x 10(4) cells/mL, to 2.5 x 10(6) cells/mL). Results We observed a significant increase in total cell expansion when the density was increased from 5 x 10(4) cells/mL to 1 x 10(6) cells/mL, but a further increase to 2.5 x 10(6) cells/mL resulted in a decline in cell expansion. Increasing feeding to 90% daily exchange in cultures with2.5 x 10(6) cells/ mL did not enhance cell expansion; nor did reducing the extent of feeding i n cultures with 5 x 10(4) cells/mL to 10% daily exchange. We did nor observ e a relationship between cell density and the percentage of granulocyte pro genitor and post-progenitor (CD15(+)CD11b(-/+) cells. While specific lactat e production (q(lac)) in cultures with 2.5 x 10(6) cells/mL was approximate ly 60% of those observed in lower density cultures by Day 13, this differen ce was largely eliminated by increasing the extent of feeding in cultures w ith 2.5 x 10(6) cells/mL. Discussion Our results suggest that feeding rates must be adjusted according to cell d ensity to maximize culture performance. They also suggest that cellular cro wding on the culture surface can limit expansion in suspension (nonadherent ) cultures.