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KMID : 0387820060130020208
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Clinical Pediatric Hematology-Oncology
2006 Volume.13 No. 2 p.208 ~ p.217
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Generation of Functional Dendritic Cells from Human Umbilical Cord Blood CD34+ Cells
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Jung Yun-Jae
Joo Sun-Young
Ryu Kyung-Ha
Woo So-Youn
Seoh Ju-Young
Kang Hyoung-Jin
Shin Hee-Young
Ahn Hyo-Seop
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Abstract
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Purpose: The aim of this study was to generate dendritic cells (DCs) from CD34+ cells of human umbilical cord blood (UCB) and to characterize the requirements for maximizing antigen uptake and the stimulatory function of DCs for effective antigen presentation.
Methods: The CD34+ cells from UCB were cultured at a density of 1.0×105 cells/mL in IMDM supplemented with 10% FBS. Cultures were expanded for 7 days (1st-step culture) with stem cell factor (SCF), flt3-ligand (FL) and thrombopoietin (TPO) and were differentiated into DCs for 5 days (2nd-step culture) with interleukin (IL)-4 and granulocyte macrophage-colony stimulating factor (GM-CSF) with or without tumor necrosis factor (TNF)-α. Immunophenotypical characterization of the cultured cells were analysed with various MoAb on a FACSCalibur flow cytometer. These cells were exposed to stimulation with Lipopolysaccharide (LPS), TNF-α, N-formyl-L-methionyl-L- leucyl-L-phenylalanin (FMLP) and LPS+TNF-α+FMLP for the DCs maturation. These cells were also incubated with fluorescent beads and FITC-conjugated C. albicans for the evaluation of phagocytic activity of generated DCs.
Results: During the 2nd-step culture, the increase of total cell numbers was 2 times higher in the presence of TNF-α than without it. Exposure to stimulants resulted in an up-regulation of costimulatory proteins (CD80, CD86), HLA-DR, CD40 and adhesion molecules (CD11c, CD11b) and the stimulants combination of LPS+TNF+FMLP was superior to LPS, TNF-α, or FMLP on their own. Expansion of CD34+ cells with SCF, FL, TPO, IL-4 and GM-CSF/or TNF-α yielded morphologically and phenotypically mature DCs with phagocytic activity against the beads and C. albicans.
Conclusion: CD34+ cells derived from UCB differentiated into morphologically, phenotypically and functionally mature DCs.
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KEYWORD
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Umbilical cord blood, CD34+ cells, Dendritic cells
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