Antigen delivery to dendritic cell populations: Internalization, antigen presentation and functional consequences for regulating immune responses

Dendritic cells (DCs) prime naive CD4+ (helper) and CD8+ (cytolytic) lymphocytes to produce armed, determinant-specific immune effector populations. DCs exist in immature and then mature states of differentiation during their life cycle. Immature DCs are thought to have a high capacity for capturing exogenous antigens for intracellular processing, and mature DCs, by contrast, mainly present epitopes derived from these internalized ligands. Understanding how specific antigen capturing processes are regulated within immature and mature DCs in vitro and in vivo will give insight into both DC physiology and optimal pathways to deliver antigenic constructs to DCs.; We have investigated murine DC subpopulations in phenotypically distinct states and microenvironments for their ability to capture soluble and particulate forms of antigen. We discovered that activated bone marrow-derived DC (BMDCs) were able to internalize 1 micron diameter particulate antigens in an unexpectedly efficient and inhibitable manner. Phagocytosis of particles was equivalent for DCs activated following exposure to LPS, CD40-ligation, and reculture. Uptake of these relatively large antigens into intracellular compartments leads to the remodeling of Lamp2+ lysosomal clusters that are prominent in all mature BMDC populations. Despite these similarities, presentation of antigens attached to particulates was dependent on the context in which maturation was induced.; We also sought to understand uptake and processing of particles by both CD8− and CD8+ DCs generated in vivo. Using Flt3L gene delivery to expand DCs in vivo, we have extensively characterized particulate antigen uptake and presentation within immature and mature DCs situated in the spleen. We find that particulate antigens are actively internalized by CD8− DCs, and are only weakly internalized by immature CD8+ DCs. Similarly, presentation of particulate antigens pulsed onto DCs in vitro occurred only with immature and mature CD8− DCs. To correlate our results with unpertubed DC populations residing in situ, we intravenously injected latex microspheres conjugated to OVA protein in vivo. We report that CD8− DCs in vivo primarily phagocytose beads, but that MHC-dependent presentation was observed in both CD8− and CD8+ DCs, providing evidence that capacities of CD8+ DCs to regulate immune responses in vivo may differ from what is observed in vitro.