Extracellular vesicles from mature dendritic cells (DC) differentiate monocytes into immature DC

During inflammation, murine and human monocytes can develop into dendritic cells (DC), but this process is not entirely understood. Here, we demonstrate that extracellular vesicles (EV) secreted by mature human DC (maDC) differentiate peripheral monocytes into immature DC, expressing a unique marker pattern, including 6-sulfo LacNAc (slan), Zbtb46, CD64, and CD14. While EV from both maDC and immature DC differentiated monocytes similar to GM-CSF/IL-4 stimulation, only maDC-EV produced precursors, which upon maturation stimulus developed into T-cell–activating and IL-12p70–secreting maDC. Mechanistically, maDC-EV induced cell signaling through GM-CSF, which was abundant in EV as were IL-4 and other cytokines and chemokines. When injected into the mouse skin, murine maDC-EV attracted immune cells including monocytes that developed activation markers typical for inflammatory cells. Skin-injected EV also reached lymph nodes, causing a similar immune cell infiltration. We conclude that DC-derived EV likely serve to perpetuate an immune reaction and may contribute to chronic inflammation.

Prostaglandin E2 enhances T-cell proliferation by inducing the costimulatory molecules OX40L, CD70, and 4-1BBL on dendritic cells

Dendritic cell (DC)–based immunotherapy of malignant diseases relies on 2 critical parameters: antigen transport from the periphery to draining lymph nodes and efficient priming of primary and stimulation of secondary immune responses. Prostaglandin E2 (PGE2) signaling has been shown to be pivotal for DC migration toward lymph node–derived chemokines in vitro and in vivo. Here, we demonstrate that PGE2 induced the expression of the costimulatory molecules OX40L, CD70, and 4-1BBL on human DCs. Short triggering by PGE2 early during DC maturation was sufficient to induce the costimulatory molecules. The expression of the costimulatory molecules was independent of the maturation stimulus but strictly dependent on PGE2 on both monocyte-derived (Mo) DCs and peripheral blood myeloid (PB) DCs. PGE2-matured MoDCs showed enhanced costimulatory capacities resulting in augmented antigen-specific CD4+ and CD8+ T-cell proliferation in primary and recall T-cell responses. Blocking OX40/OX40L signaling impaired the enhanced T-cell proliferation induced by PGE2-matured MoDCs. Moreover, MoDCs matured in the presence of PGE2 induced the expression of OX40, OX40L, and CD70 on T cells facilitating T-cell/T-cell interaction that warrant long-lasting costimulation. This newly identified parameter will help to further optimize DC-based immunotherapy.

Antigen targeting to dendritic cells elicits long-lived T cell help for antibody responses

Resistance to several prevalent infectious diseases requires both cellular and humoral immune responses. T cell immunity is initiated by mature dendritic cells (DCs) in lymphoid organs, whereas humoral responses to most antigens require further collaboration between primed, antigen-specific helper T cells and naive or memory B cells. To determine whether antigens delivered to DCs in lymphoid organs induce T cell help for antibody responses, we targeted a carrier protein, ovalbumin (OVA), to DCs in the presence of a maturation stimulus and assayed for antibodies to a hapten, (4-hydroxy-3-nitrophenyl) acetyl (NP), after boosting with OVA-NP. A single DC-targeted immunization elicited long-lived T cell helper responses to the carrier protein, leading to large numbers of antibody-secreting cells and high titers of high-affinity antihapten immunoglobulin Gs. Small doses of DC-targeted OVA induced higher titers and a broader spectrum of anti-NP antibody isotypes than large doses of OVA in alum adjuvant. Similar results were obtained when the circumsporozoite protein of Plasmodium yoelii was delivered to DCs. We conclude that antigen targeting to DCs combined with a maturation stimulus produces broad-based and long-lived T cell help for humoral immune responses.

Differential Effects of Dengue Virus on Infected and Bystander Dendritic Cells

ABSTRACT Dendritic cells (DCs) play a central role as major targets of dengue virus (DV) infections and initiators of antiviral immune responses. Previous observations showed that DCs are activated by infection, presumably acquiring the capacity to promote cell-mediated immunity. However, separate evaluations of the maturation profiles of infected and uninfected bystander cells show that infection impairs the ability of DCs to upregulate cell surface expression of costimulatory, maturation, and major histocompatibility complex molecules, resulting in reduced T-cell stimulatory capacity. Infected DCs failed to respond to tumor necrosis factor alpha as an additional maturation stimulus and were apoptotic. Interleukin 10 (IL-10) was detected in supernatants from cultures of DV-infected DCs and cocultures of DCs and T cells. Taken together, these results constitute an immune evasion strategy used by DV that directly impairs antigen-presenting cell function by maturation blockade and induction of apoptosis.

Killed but Metabolically Active Recombinant Listeria monocytogenes as an Antigen Delivery and Activation Platform for Human Dendritic Cell-Based Cancer Immunotherapy.

Dendritic cells (DCs) are bone marrow-derived professional antigen presenting cells which play a pivotal role in initiating immune responses. To achieve optimal potency for priming of naive T cells, DCs must not only present high levels of MHC peptide complexes, but must also undergo maturation. An ideal DC-targeting vaccine formulation, therefore, should simultaneously deliver multiple antigens and activate DCs to increase expression of surface MHC-peptide complexes, costimulatory molecules, and production of immuno-stimulatory cytokines. Despite the development of a variety of ex vivo methods, no standard efficient and cost effective methods combining antigen loading and activation/maturation of DCs for clinical use currently exist. Here we report the development of a novel DC antigen-loading platform based on killed, but metabolically active (KBMA) recombinant Listeria monocytogenes (Lm). While recombinant Lm has a relatively long history as a cancer vaccine platform, its utility as a DC activation and loading platform has not been fully explored. KBMA Lm were 8-logs more sensitive to photochemical inactivation by the combined treatment of S-59 psoralen (amotosalen HCl) and long-wave ultraviolet light (S-59/UVA), by virtue of abrogation of nucleotide excision repair (NER) capacity, through engineered deletion of the bacterial UvrAB genes. Due to random distribution of infrequent psoralen adducts, KBMA Lm cannot propagate, but are metabolically active and can express their genetic repertoire, and program presentation of expressed heterologous antigens via the MHC class I and class II pathways. Infection of human monocyte-derived or blood myeloid DCs with KBMA Listeria induced their maturation and production of pro-inflammatory and pro- Th1 cytokines. The DCs achieved a fully mature phenotype, comparable to that induced by the standard maturation stimulus MCM-mimic. To determine whether KBMA Lm could program presentation of encoded antigens by the MHC class I pathway, we constructed a recombinant Lm vaccine expressing CD8+ T cell epitopes of influenza matrix protein (MP58–66) and MelanA/Mart-1 protein (Mart126–35). DCs infected with the recombinant KBMA Lm vaccine presented the recombinant class I epitopes and induced IFN-γ production by MP58–66- or Mart126–35-specific human CD8+ T cell clones. These studies demonstrate that KBMA Listeria is a novel and powerful ex vivo DC antigen loading platform combining a potent DC-maturation stimulus with the cytosolic delivery of recombinant antigen for presentation.

The CD16+ (FcγRIII+) Subset of Human Monocytes Preferentially Becomes Migratory Dendritic Cells in a Model Tissue Setting

Much remains to be learned about the physiologic events that promote monocytes to become lymph-homing dendritic cells (DCs). In a model of transendothelial trafficking, some monocytes become DCs in response to endogenous signals. These DCs migrate across endothelium in the ablumenal-to-lumenal direction (reverse transmigration), reminiscent of the migration into lymphatic vessels. Here we show that the subpopulation of monocytes that expresses CD16 (Fcγ receptor III) is predisposed to become migratory DCs. The vast majority of cells derived from CD16+ monocytes reverse transmigrated, and their presence was associated with migratory cells expressing high levels of CD86 and human histocompatibility leukocyte antigen (HLA)-DR, and robust capacity to induce allogeneic T cell proliferation. A minority of CD16− monocytes reverse transmigrated, and these cells stimulated T cell proliferation less efficiently. CD16 was not functionally required for reverse transmigration, but promoted cell survival when yeast particles (zymosan) were present as a maturation stimulus in the subendothelial matrix. The cell surface phenotype and migratory characteristics of CD16+ monocytes were inducible in CD16− monocytes by preincubation with TGFβ1. We propose that CD16+ monocytes may contribute significantly to precursors for DCs that transiently survey tissues and migrate to lymph nodes via afferent lymphatic vessels.