A Comparison of The Expansion and Functional Capabilities of Dendritic Cells Derived from Blood Monocytes vs Mobilized Blood CD34+ Cells, Matured With Either CD40L or TNF-α/IFN-α
B. Chen, P. Stiff, G. Sloan, J. Kash, R. Manjunath, B. Nickoloff
Departments of Medicine and Pathology, Loyola University Medical Center, Maywood, IL
The potent antigen presenting capacities of dendritic cells (DCs) provide novel strategies for tumor immunotherapy. While DCs can be generated ex vivo from blood monocytes (PBMC) or CD34+ stem cells, the optimal cell source for DCs, based on expansion and functional abilities is unknown. We compared the phenotype and the functional properties of these two sources of DCs, with the intent of establishing an optimal culture system allowing large-scale clinical generation of DCs. PBMC DCs were expanded and differentiated in a serum-free medium (X Vivo 10) for 7 days in GM-CSF (100 ng/ml) and IL-4 (25 ng/ml). CD34-derived DCs were expanded for 14 days in the presence ofGM--CSF (100 ng/ml), IL-4 (25 ng/mI) and - Flt3L (1000 ng/ml) in either X Vivo 10, or X Vivo 10 with 5% autologous plasma or 2% albumin. Expanded DCs from both cell sources were matured for 7 days with either CD40L (1000 ng/ml) or the combination of IFN-α (1000 u/ml) + TNF- α (1000 u/ml). Cells were analyzed by flowcytometry on days 0, 7, 14, and 21 of culture for expression of CD34, CD14, CD1a, CD80, CD83, CD86 and HLA-DR. Function was analyzed using mixed lymphocyte response (MLR) assays and the ability to of the DCs to phagocytose apoptotic melanoma and ovarian tumor cells pre- and post-maturation. Optimal PBMC DC generation occurred with 14 days of expansion/maturation that used CD40L: from 20 x 106 PBMCs a mean 2.3 x 106 mature, CD86+ DCs were generated.
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