Differentiation and Assessment of Monocyte-Derived Dendritic Cells in the Presence of Endothelial Cells Conditioned Media and Maturation Factors

Dendritic cells (DCs) migrate from the periphery to the T cell zone of secondary lymphoid organs. Thus, activated DCs are specialized cells that obtain information from the periphery and transfer it to lymphoid organs. During their migration from the blood into peripheral tissues and then into the lymph nodes, DCs interact with endothelial cells and their soluble mediators. Therefore, we studied the effects of the endothelial condition medium (ECM) on phenotypic and functional characteristics of monocyte- driven dendritic cells. DCs were generated by culture of monocyte cells for 5 days in RPMI medium (RPMI 1640) supplemented with 10% FCS (fetal calf serum), IL-4 (interlukin-4), and GM-CSF (granulocyte, macrophage-colony stimulating factor). They were then incubated for 48 hours in MCM (monocyte conditioned medium), TNF-α (tumor necrotic factor alpha), and polyinosinic-polycytidylic acid (polyIC; control). ECM was added on day 5 and the beginning of the culture (day-0) to the control. The maturation of harvested DCs on day 7 was evaluated via flow cytometry, beta-counter and ELISA kits. DCs generated from peripheral blood monocytes specifically interact with ECM via soluble mediators. This induced the phenotypic maturation of DC via increased expression of CD83, CD80, HLA-DR, down-regulation of CD14, decreased phagocytic activity, and producing stimulatory cytokines in comparison with maturation factors alone. Moreover, ECM-matured DCs potently induced T cell activation reflected by increased T cell proliferation. Our results demonstrated that human endothelial cells with which DCs can encounter during their trafficking from tissue to lymph nodes can act as potent regulators of DC differentiation and function.