Stem cell factor and FLT3-ligand are strictly required to sustain the long-term expansion of primitive CD34+DR- dendritic cell precursors.
We studied cytokine-driven differentiation of primitive human CD34(+)HLA-DR(-) cells to myeloid dendritic cells (DC). Hemopoietic cells were grown in long-term cultures in the presence of various combinations of early acting cytokines such as FLT3-ligand (FLT3-L) and stem cell factor (SCF) and the differentiating growth factors GM-CSF and TNF-alpha. Two weeks of incubation with GM-CSF and TNF-alpha generated fully functional DC. However, clonogenic assays demonstrated that CFU-DC did not survive beyond 1 wk in liquid culture regardless of whether FLT3-L and/or SCF were added. FLT3-L or SCF alone did not support DC maturation. However, the combination of the two early acting cytokines allowed a 100-fold expansion of CFU-DC for >1 month. Phenotypic analysis demonstrated the differentiation of CD34(+)DR(-) cells into CD34(-)CD33(+)DR(+)CD14(+) cells, which were intermediate progenitors capable of differentiating into functionally active DC upon further incubation with GM-CSF and TNF-alpha. As expected, GM-CSF and TNF-alpha generated DC from committed CD34(+)DR(+) cells. However, only SCF, with or without FLT3-L, induced the expansion of DC precursors for >4 wk, as documented by secondary clonogenic assays. This demonstrates that although GM-CSF and TNF-alpha do not require additional cytokines to generate DC from primitive human CD34(+)DR(-) progenitor cells, they do force terminal differentiation of DC precursors. Conversely, FLT3-L and SCF do not directly affect DC differentiation, but instead sustain the long-term expansion of CFU-DC, which can be induced to produce mature DC by GM-CSF and TNF-alpha.
Curti
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Institute of Hematology and Medical Oncology "Ludovico e Ariosto Seràgnoli," University of Bologna, Bologna, Italy.
Fogli
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Ratta
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Tura
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Lemoli
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RM
D000276:Adjuvants, Immunologic / Q000502:physiology; D018952:Antigens, CD34 / Q000096:biosynthesis*; D001854:Bone Marrow Cells / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000276:immunology; D002455:Cell Division / Q000276:immunology; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D003713:Dendritic Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D004338:Drug Combinations; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006684:HLA-DR Antigens / Q000096:biosynthesis*; D006410:Hematopoiesis / Q000276:immunology*; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D008024:Ligands; D008565:Membrane Proteins / Q000502:physiology*; D019089:Stem Cell Factor / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D000276:Adjuvants, Immunologic; D018952:Antigens, CD34; D004338:Drug Combinations; D006684:HLA-DR Antigens; D008024:Ligands; D008565:Membrane Proteins; D019089:Stem Cell Factor; D014409:Tumor Necrosis Factor-alpha; C084881:flt3 ligand protein; D016178:Granulocyte-Macrophage Colony-Stimulating Factor