A novel approach for the generation of human dendritic cells from blood monocytes in the absence of exogenous factors.
Human dendritic cells (DCs) for research and clinical applications are typically derived from purified blood monocytes that are cultured in a cocktail of cytokines for a week or more. Because it has been suggested that these cytokine-derived DCs may be deficient in some important immunological functions and might not accurately represent antigen presenting cell (APC) populations found under normal conditions in vivo, there is an interest in developing methods that permit the derivation of DCs in a more physiologically relevant manner in vitro. Here, we describe a simple and reliable technique for generating large numbers of highly purified DCs that is based on a one-way migration of blood monocytes through a layer of human umbilical vein endothelial cells (HUVECs) that are cultured to confluency in the upper chamber of a Transwell device. The resultant APCs, harvested from the lower Transwell chamber, resemble other cultured DC populations in their expression of major histocompatibility (MHC) and costimulatory molecules, ability to phagocytose protein antigens and capacity to trigger primary antigen-specific T cell responses. This technique offers several advantages over the standard method of in vitro cytokine-driven DC development, including: (1) the rapidity of this approach, as DC differentiation occurs in only 2 days, (2) the differentiation process itself, which is more akin to the development of DCs under physiologic conditions and (3) the cost-effectiveness of the system, since no monocyte pre-selection is required and DC development occurs in the absence of expensive recombinant cytokines.
Schanen
Brian C
BC
VaxDesign Corporation, Orlando, FL 32826, USA.
Drake
Donald R
DR
D018929:Cell Culture Techniques; D002465:Cell Movement; D002478:Cells, Cultured; D018920:Coculture Techniques; D016207:Cytokines / Q000378:metabolism; D003713:Dendritic Cells / Q000276:immunology*; D042783:Endothelial Cells / Q000276:immunology*; D017629:Gap Junctions / Q000276:immunology; D006680:HLA Antigens / Q000378:metabolism; D006801:Humans; D007156:Immunologic Memory; D008213:Lymphocyte Activation; D008567:Membranes, Artificial; D009000:Monocytes / Q000276:immunology*; D010587:Phagocytosis; D010641:Phenotype; D011076:Polycarboxylate Cement; D013601:T-Lymphocytes / Q000276:immunology; D013997:Time Factors
D016207:Cytokines; D006680:HLA Antigens; D008567:Membranes, Artificial; D011076:Polycarboxylate Cement; C005506:polycarbonate