BACKGROUND: In vitro generated dendritic cells (DCs) are widely used as adjuvants in cancer immunotherapy. The major sources for DC generation are monocytes and CD34+ cells. CD34+-derived DCs are less frequently used in clinical applications because it requires complex generation methods. Here a simple method for the large-scale generation of mature functional DCs from umbilical cord blood-derived CD34+ cells is described. STUDY DESIGN AND METHODS: CD34+ cells were first expanded with a combination of early acting growth factors in a medium containing autologous plasma. In the second step the DC precursors were further either enriched by plastic adherence or sorted on a cell sorter and differentiated as DCs. DCs generated by both methods were compared for their morphology, phenotype, and different functional variables. RESULTS: This culture system provided a large-scale expansion of CD34+ cells giving a mean fold increase of 615. The majority of the expanded cells were interstitial DC precursors, that is, CD14+-positive cells. In vitro generated immature DCs could be matured into functional DCs by appropriate maturation stimuli. DCs generated by the plastic adherence method had a better cytokine profile and strong mixed leukocyte reaction compared to those generated by cell sorting. CONCLUSION: A two-step culture system provides a large-scale expansion of CD34+ cells with a preferential lineage commitment toward CD14+ cells. Enrichment of these precursors with a simple plastic adherence technique results in generation of large numbers of mature, functional DCs. This method of in vitro DC generation will have applications in cancer immunotherapy.
BACKGROUND: In vitro generated dendritic cells (DCs) are widely used as adjuvants in cancer immunotherapy. The major sources for DC generation are monocytes and CD34+ cells. CD34+-derived DCs are less frequently used in clinical applications because it requires complex generation methods. Here a simple method for the large-scale generation of mature functional DCs from umbilical cord blood-derived CD34+ cells is described. STUDY DESIGN AND METHODS: CD34+ cells were first expanded with a combination of early acting growth factors in a medium containing autologous plasma. In the second step the DC precursors were further either enriched by plastic adherence or sorted on a cell sorter and differentiated as DCs. DCs generated by both methods were compared for their morphology, phenotype, and different functional variables. RESULTS: This culture system provided a large-scale expansion of CD34+ cells giving a mean fold increase of 615. The majority of the expanded cells were interstitial DC precursors, that is, CD14+-positive cells. In vitro generated immature DCs could be matured into functional DCs by appropriate maturation stimuli. DCs generated by the plastic adherence method had a better cytokine profile and strong mixed leukocyte reaction compared to those generated by cell sorting. CONCLUSION: A two-step culture system provides a large-scale expansion of CD34+ cells with a preferential lineage commitment toward CD14+ cells. Enrichment of these precursors with a simple plastic adherence technique results in generation of large numbers of mature, functional DCs. This method of in vitro DC generation will have applications in cancer immunotherapy.
Authors: Colin de Haar; Maud Plantinga; Nina Jg Blokland; Niek P van Til; Thijs Wh Flinsenberg; Viggo F Van Tendeloo; Evelien L Smits; Louis Boon; Lotte Spel; Marianne Boes; Jaap Jan Boelens; Stefan Nierkens Journal: Oncoimmunology Date: 2015-05-27 Impact factor: 8.110
Authors: Nicole Bedke; Emily J Swindle; Camelia Molnar; Patrick G Holt; Deborah H Strickland; Graham C Roberts; Ruth Morris; Stephen T Holgate; Donna E Davies; Cornelia Blume Journal: J Immunol Methods Date: 2019-11-09 Impact factor: 2.303