Expansion of primitive human hematopoietic progenitors in a perfusion bioreactor system with IL-3, IL-6, and stem cell factor.
Present methods for long-term hematopoietic culture (LTHC) employ a static culture environment which is not well-characterized. Primitive long-term culture-initiating cell (LTC-IC) numbers have been shown to decline in conventional static human LTHC, even with exogenous cytokine combinations. We have expanded human hematopoietic cells from umbilical cord blood on a preformed marrow stroma with synergistic cytokine combinations in a novel perfusion bioreactor system, which continuously maintained culture conditions within desired ranges. Interleukin-3 (IL-3) and interleukin-6 (IL-6) in perfusion culture resulted in rapid 7-day expansion of granulocyte-macrophage colony forming units (CFU-GM, 11-fold), erythroid burst-forming units (BFU-E, 2.5-fold), and granulocyte-erythroid-macrophage colony forming units (CFU-Mix, 2.4-fold), compared to 6-fold, 1.4-fold, and no expansion, respectively, in static cultures. Addition of stem cell factor (SCF) to IL-3/IL-6 in static culture increased the extent of CFU-GM expansion (to 9-fold), but did not result in BFU-E or CFU-Mix expansion. In perfusion cultures with IL-3/IL-6/SCF, much greater expansions of CFU-GM (18-fold) and CFU-Mix (5.3-fold) were obtained. More importantly, expansion of LTC-IC (nearly 3-fold in two of three experiments) was only obtained with IL-3/IL-6/SCF and perfusion. The ability to expand hematopoietic cells while maintaining or expanding primitive progenitors has potential clinical applications in bone marrow transplantation and gene therapy.
Koller
M R
MR
Northwestern University, Department of Chemical Engineering, Evanston, IL 60208-3120.
Bender
J G
JG
Miller
W M
WM
Papoutsakis
E T
ET
D002452:Cell Count; D002478:Cells, Cultured; D016207:Cytokines / Q000378:metabolism; D015672:Erythroid Precursor Cells / Q000166:cytology; D005312:Fetal Blood / Q000166:cytology; D005947:Glucose / Q000378:metabolism; D006098:Granulocytes / Q000166:cytology; D016298:Hematopoietic Cell Growth Factors / Q000494:pharmacology*; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology*; D015850:Interleukin-6 / Q000494:pharmacology*; D007773:Lactates / Q000378:metabolism; D019344:Lactic Acid; D008264:Macrophages / Q000166:cytology; D010477:Perfusion; D019089:Stem Cell Factor
D016207:Cytokines; D016298:Hematopoietic Cell Growth Factors; D007377:Interleukin-3; D015850:Interleukin-6; D007773:Lactates; D019089:Stem Cell Factor; D019344:Lactic Acid; D005947:Glucose