Generation of dendritic cells from bone marrow progenitors using GM-CSF, TNF-alpha, and additional cytokines: antagonistic effects of IL-4 and IFN-gamma and selective involvement of TNF-alpha receptor-1.
We report the generation of dendritic cells (DC) starting from CD34+ bone marrow (BM) progenitor cells, using a two-stage culture system in which, besides granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumour necrosis factor-alpha (TNF-alpha), stem-cell factor (SCF) was added during the first 5 days, while interleukin-4 (IL-4) and/or interferon-gamma (IFN-gamma) were added during the secondary culture period of 9 days. Addition of IL-4 favoured the outgrowth of CD1a+, HLA-DR+, CD4+, CD40+, CD80+ but CD14- cells with dendritic morphology and strong antigen-presenting capacity. Addition of IFN-gamma selectively induced HLA-DR and CD86 but did not up-regulate CD1a expression or antigen-presenting capacity of the differentiated cells. An antagonism between IL-4 and IFN-gamma could further be confirmed in that, as compared with IL-4 alone, the simultaneous addition of IL-4 and IFN-gamma to GM-CSF plus TNF-alpha during maturation reduced both the phenotypical (CD1a, CD4, CD40) and functional characteristics of DC. Using receptor-specific TNF-alpha mutants, we investigated the relative involvement of TNF-alpha receptors R1 and R2 in the generation of DC. The induction of CD1a and HLA-DR, as well as the increase in allostimulatory capacity were dependent on TNF-R1 triggering, whereas triggering through TNF-R2 had no measurable effect. We conclude first, that the expansion of DC from BM progenitors could most effectively be enhanced in a two-stage culture assay using SCF, GM-CSF, TNF-alpha and IL-4; second, that the effect of TNF-alpha in DC generation involves signalling via the TNF-R1 receptor; and third, that IFN-gamma counteracts some of the effects of IL-4 in DC generation.
Lardon
F
F
Laboratory of Experimental Haematology, University of Antwerp, Belgium.
Snoeck
H W
HW
Berneman
Z N
ZN
Van Tendeloo
V F
VF
Nijs
G
G
Lenjou
M
M
Henckaerts
E
E
Boeckxtaens
C J
CJ
Vandenabeele
P
P
Kestens
L L
LL
Van Bockstaele
D R
DR
Vanham
G L
GL
D017951:Antigen Presentation; D018952:Antigens, CD34 / Q000032:analysis; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000276:immunology; D016207:Cytokines / Q000276:immunology*; D003713:Dendritic Cells / Q000276:immunology*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000276:immunology; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000276:immunology*; D006801:Humans; D016130:Immunophenotyping; D015847:Interleukin-4 / Q000276:immunology; D009154:Mutation; D018124:Receptors, Tumor Necrosis Factor / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000276:immunology
D018952:Antigens, CD34; D016207:Cytokines; D018124:Receptors, Tumor Necrosis Factor; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor