Differentiation of monocyte-derived dendritic cells under the influence of platelets.
BACKGROUND
Monocytapheresis has been established to collect a sufficient number of monocytes (MO) for differentiation to dendritic cells (DC) as a cancer vaccine. Platelets (Plt) are invariably found as a contaminant in the final monocytapheresis product. The aim of this study was to investigate DC differentiation under the influence of Plt with regard to their function and phenotype.


METHODS
MO were isolated and co-cultured with autologous Plt at different MO:Plt ratios (1:1.7, 1:5, 1:15, 1:45 and 1:135) in the presence of interleukin-4 (IL-4) and granulocyte-macrophage colony-stimulating factor (GM-CSF). IL-12p70 release after ligation of CD40L was determined in the supernatant by enzyme-linked immunosorbent assay (ELISA). For T-cell stimulation, tetanus toxoid was added to immature DC and maturation was induced by adding cytokines (IL-1beta, IL-6, tumor necrosis factor-alpha and prostaglandin E(2)). Stimulated T cells were analyzed for activation and proliferation as well as for intracellular cytokines by flow cytometry.


RESULTS
All DC cultures were strongly positive for CD83. At a contaminating concentration of 5 Plt/MO, matured DC showed the highest expression of HLA-DR, CD80 and CD86, inducing a strong T-cell proliferation with high production of IL-4 and interferon-gamma. The highest level of IL-12p70 production was observed by the same DC group.


CONCLUSIONS
Plt did not negatively influence DC maturation but enhanced the expression of co-stimulatory molecules and the release of IL-12. Functionally this was reflected by a strong T-cell response that involved T-helper 1 (Th1)- as well as Th2-biased T cells. Our findings show that controlling the Plt concentration may provide important advantages for the generation of DC for use in immunotherapy.
Nguyen
X D
XD
Institute of Transfusion Medicine and Immunology, Medical Faculty Mannheim, Heidelberg University, Red-Cross Blood Donation Service of Baden-Wurttemberg-Hessen, Germany. x.nguyen@blutspende.de
Müller-Berghaus
J
J
Kälsch
T
T
Schadendorf
D
D
Borggrefe
M
M
Klüter
H
H
D001792:Blood Platelets / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000276:immunology; D016238:Cytapheresis; D016207:Cytokines / Q000276:immunology / Q000494:pharmacology; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology* / Q000502:physiology; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000276:immunology / Q000494:pharmacology; D006801:Humans; D016130:Immunophenotyping; D018664:Interleukin-12 / Q000096:biosynthesis / Q000276:immunology; D015847:Interleukin-4 / Q000276:immunology / Q000494:pharmacology; D009000:Monocytes / Q000187:drug effects / Q000276:immunology* / Q000502:physiology; D006377:T-Lymphocytes, Helper-Inducer / Q000276:immunology* / Q000378:metabolism
D016207:Cytokines; D018664:Interleukin-12; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor