Long-term culture of human CD34(+) progenitors with FLT3-ligand, thrombopoietin, and stem cell factor induces extensive amplification of a CD34(-)CD14(-) and a CD34(-)CD14(+) dendritic cell precursor.
Current in vitro culture systems allow the generation of human dendritic cells (DCs), but the output of mature cells remains modest. This contrasts with the extensive amplification of hematopoietic progenitors achieved when culturing CD34(+) cells with FLT3-ligand and thrombopoietin. To test whether such cultures contained DC precursors, CD34(+) cord blood cells were incubated with the above cytokines, inducing on the mean a 250-fold and a 16,600-fold increase in total cell number after 4 and 8 weeks, respectively. The addition of stem cell factor induced a further fivefold increase in proliferation. The majority of the cells produced were CD34(-)CD1a- CD14(+) (p14(+)) and CD34(-)CD1a-CD14(-) (p14(-)) and did not display the morphology, surface markers, or allostimulatory capacity of DC. When cultured with granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4), both subsets differentiated without further proliferation into immature (CD1a+, CD14(-), CD83(-)) macropinocytic DC. Mature (CD1a+, CD14(-), CD83(+)) DCs with high allostimulatory activity were generated if such cultures were supplemented with tumor necrosis factor-alpha (TNF). In addition, p14(-) cells generated CD14(+) cells with GM-CSF and TNF, which in turn, differentiated into DC when exposed to GM-CSF and IL-4. Similar results were obtained with frozen DC precursors and also when using pooled human serum AB+ instead of bovine serum, emphasizing that this system using CD34(+) cells may improve future prospects for immunotherapy.
Arrighi
J F
JF
Department of Dermatology and Allergy Unit, Geneva University Hospital, Geneva, Switzerland.
Hauser
C
C
Chapuis
B
B
Zubler
R H
RH
Kindler
V
V
D000818:Animals; D018949:Antigens, CD1 / Q000032:analysis; D018952:Antigens, CD34 / Q000032:analysis*; D001790:Blood Physiological Phenomena; D002417:Cattle; D002452:Cell Count; D002454:Cell Differentiation / Q000187:drug effects; D003470:Culture Media / Q000494:pharmacology; D003713:Dendritic Cells / Q000166:cytology*; D005312:Fetal Blood / Q000166:cytology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects*; D006801:Humans; D015847:Interleukin-4 / Q000494:pharmacology; D018950:Lipopolysaccharide Receptors / Q000032:analysis*; D007959:Lymphocyte Culture Test, Mixed; D008565:Membrane Proteins / Q000494:pharmacology*; D019089:Stem Cell Factor / Q000494:pharmacology*; D013926:Thrombopoietin / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D018949:Antigens, CD1; D018952:Antigens, CD34; D003470:Culture Media; D018950:Lipopolysaccharide Receptors; D008565:Membrane Proteins; D019089:Stem Cell Factor; D014409:Tumor Necrosis Factor-alpha; C084881:flt3 ligand protein; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D013926:Thrombopoietin