Ag-presenting Dendritic Cells Research Articles

Dendritic cell ICAM-1 strengthens synapses with CD8 T cells but is not required for their early differentiation

Lymphocyte priming in lymph nodes (LNs) was postulated to depend on the formation of stable Tcell receptor (TCR)-specific immune synapses (ISs) with antigen (Ag)-presenting dendritic cells (DCs). The high-affinity LFA-1 ligand ICAM-1 was implicated in different ISs studied invitro. We dissect the invivo roles of endogenous DC ICAM-1 in Ag-stimulated Tcell proliferation and differentiation and find that under type 1 polarizing conditions in vaccinated or vaccinia virus-infected skin-draining LNs, Ag-presenting DCs engage in ICAM-1-dependent stable conjugates with a subset of Ag-specific CD8 blasts. Nevertheless, in the absence of these conjugates, CD8 lymphocyte proliferation and differentiation into functional cytotoxic Tcells (CTLs) and skin homing effector lymphocytes takes place normally. Our results suggest that although CD8 Tcell blasts engage in tight ICAM-1-dependent DC-T ISs, firm ISs are dispensable for TCR-triggered proliferation and differentiation into productive effector lymphocytes.

Nanoparticle dose and antigen loading attenuate antigen-specific T-cell responses.

Immune-mediated hypersensitivities such as autoimmunity, allergy, and allogeneic graft rejection are treated with therapeutics that suppress the immune system, and the lack of specificity is associated with significant side effects. The delivery of disease-relevant antigens (Ags) by carrier systems such as poly(lactide-co-glycolide) nanoparticles (PLG-Ag) and carbodiimide (ECDI)-fixed splenocytes (SP-Ag) has demonstrated Ag-specific tolerance induction in model systems of these diseases. Despite therapeutic outcomes by both platforms, tolerance is conferred with different efficacy. This investigation evaluated Ag loading and total particle dose of PLG-Ag on Ag presentation in a coculture system of dendritic cells (DCs) and Ag-restricted T cells, with SP-Ag employed as a control. CD25 expression was observed in nearly all T cells even at low concentrations of PLG-Ag, indicating efficient presentation of Ag by dendritic cells. However, the secretion of IL-2, Th1, and Th2 cytokines (IFNγ and IL-4, respectively) varied depending on PLG-Ag concentration and Ag loading. Concentration escalation of soluble Ag resulted in an increase in IL-2 and IFNγ and a decrease in IL-4. Treatment with PLG-Ag followed a similar trend but with lower levels of IL-2 and IFNγ secreted. Transcriptional Activity CEll ARrays (TRACER) were employed to measure the real-time transcription factor (TF) activity in Ag-presenting DCs. The kinetics and magnitude of TF activity was dependent on the Ag delivery method, concentration, and Ag loading. Ag positively regulated IRF1 activity and, as carriers, NPs and ECDI-treated SP negatively regulated this signaling. The effect of Ag loading and dose on tolerance induction were corroborated in vivo using the delayed-type hypersensitivity (DTH) and experimental autoimmune encephalomyelitis (EAE) mouse models where a threshold of 8 μg/mg Ag loading and 0.5 mg PLG-Ag dose were required for tolerance. Together, the effect of Ag loading and dosing on in vitro and in vivo immune regulation provide useful insights for translating Ag-carrier systems for the clinical treatment of immune disorders.

Compartmentalization of dendritic cell and T-cell interactions in the lymph node: Anatomy of T-cell fate decisions.

Upon receiving cognate and co-stimulatory priming signals from antigen (Ag)-presenting dendritic cells (DCs) in secondary lymphoid tissues, naïve CD4+ T cells differentiate into distinct effector and memory populations. These alternate cell fate decisions, which ultimately control the T-cell functional attributes, are dictated by programming signals provided by Ag-bearing DCs and by other cells that are present in the microenvironment in which T-cell priming occurs. We know that DCs can be subdivided into multiple populations and that the various DC subsets exhibit differential capacities to initiate development of the different CD4+ T-helper populations. What is less well understood is why different subanatomic regions of secondary lymphoid tissues are colonized by distinct populations of Ag-presenting DCs and how the location of these DCs influences the type of T-cell response that will be generated. Here we review how chemokine receptors and their ligands, which position allergen and nematode-activated DCs within different microdomains of secondary lymphoid tissues, contribute to the establishment of IL-4 committed follicular helper T and type 2 helper cell responses.

The second and third amino acids of Pam2 lipopeptides are key for the proliferation of cytotoxic T cells.

The TLR2 agonist, dipalmitoyl lipopeptide (Pam2LP), has been used as an immune adjuvant without much success. Pam2LP is recognised by TLR2/6 receptors in humans and in mice. This study examined the proliferative activity of cytotoxic T lymphocytes (CTL) using mouse Ag-presenting dendritic cells (DCs) and OT-I assay system, where a library of synthetic Pam2LP was utilised from the Staphylococcus aureus database. Ag-specific CTL expansion and IFN-γ levels largely depended on the Pam2LP peptide sequence. The first Aa is cysteine (Cys), which has an active SH residue to bridge fatty acids, and the second and third Aa are hydrophilic or non-polar. The sequence structurally adapted to the residual constitution of the reported TLR2/6 pocket. The inactive sequence contained proline or leucine/isoleucine after the first Cys. Notably, no direct activation of OT-I cells was detected without DCs by stimulation with the active Pam2LP having the Cys-Ser sequence. MyD88, but not TICAM-1 or IFN pathways, in DCs participates in DC maturation characterised by upregulation of CD40, CD80 and CD86. Hence, the active Pam2LPs appear suitable for dimeric TLR2/6 on DCs, resulting in induction of DC maturation.

Peripheral Deletion of CD8 T Cells Requires p38 MAPK in Cross-Presenting Dendritic Cells.

Peripheral tolerance mechanisms exist to prevent autoimmune destruction by self-reactive T cells that escape thymic deletion. Dominant tolerance imposed by CD4+Foxp3+ T regulatory cells can actively control autoaggressive T cell responses. Tolerance mechanisms that act endogenous to the T cell also exist. These mechanisms include T cell inactivation (anergy) and deletion. A major difference between anergic T cells and T cells undergoing peripheral deletion is the capacity of the latter to still signal through MAPKs upon TCR stimulation, suggesting these signals may be required for T deletion. In this study, we used several different models of CD8 T cell deletion to investigate the contribution of MAPK activation. Using chemical inhibitors, we established that inhibition of p38, but not ERK or JNK, rescue T cells from undergoing peripheral deletion both in vitro and in vivo. Using T cell-specific murine lines genetically altered in expression of p38α, and mice in which p38α was deleted only in CD11c-expressing cells, we surprisingly found that CD8 T cell-intrinsic p38α activation was not responsible for increased survival, but rather that inhibition of p38α in the Ag-presenting dendritic cells prevented CD8 T cell deletion.

The neurokinin 1 receptor and its agonists recruited at the DC-T cell synapse are necessary to promote the survival of activated T cells in vivo

Abstract Initiation of cellular immunity relies on T cell activation by Ag-presenting dendritic cells (DCs). By signaling via the neurokinin-1 receptor (NK1R), the neuropeptides hemokinin-1 (HK-1) and substance P (SP) potentiate cellular immunity by mechanisms not fully elucidated. Although SP is mainly secreted by sensory nerves, it is also synthesized and released by T cells. Here, we investigated the role of NK1R and autocrine SP and HK-1 on T cell activation. Using Imagestream flow cytometry (Amnis) to visualize couplets of OVA loaded WT or SP/HK1double KO DCs and responder OT-II CD4 T cells, we observed that the NK1R, SP and HK-1 colocalize with phalloidin at the site of DC-T cell contact. Following CD3/CD28 activation in vitro, we demonstrate that T cells express the full-length NK1R, and that NK1R signaling induces Ca2 flux, activation of calcineurin, NFAT1/2 and NFκB, which results in IL-2 secretion and survival of CD4 and CD8 T cells. These effects were abrogated in NK1RKO orSP/HK1double KO T cells, or by soluble NK1R antagonists. Addition of exogenous SP or HK-1 to CD3/CD28 activated WT T cells did not increase IL-2 secretion indicating that autocrine SP and HK1 released at the DC-T cell synapse suffice to promote T cell survival. In a model of skin DTH in NK1RKO or HK-1/SPdouble KOT cell chimeras, 73 ± 5% of activated (CD44high) CD4 and CD8 T cells die in the draining lymph nodes after priming, and 79%± 7% of the remaining T cells recruited to the skin die in situ following elicitation. In summary, NK1R signaling by autocrine SP and HK1 at the DC-T cell synapse is necessary for the survival of activated T cells and the development of potent cellular immunity.

Age-Specific Adjuvant Synergy: Dual TLR7/8 and Mincle Activation of Human Newborn Dendritic Cells Enables Th1 Polarization.

Due to functionally distinct cell-mediated immunity, newborns and infants are highly susceptible to infection with intracellular pathogens. Indeed, neonatal Ag-presenting dendritic cells (DCs) demonstrate impaired Th1 responses to many candidate adjuvants, including most TLR agonists (TLRAs). Combination adjuvantation systems may provide enhanced immune activation but have typically been developed without regard to the age of the target population. We posited that distinct combinations of TLRAs and C-type lectin receptor agonists may enhance Th1 responses of newborn DCs. TLRA/C-type lectin receptor agonist combinations were screened for enhancement of TNF production by human newborn and adult monocyte-derived DCs cultured in 10% autologous plasma or in newborn cord, infant, adult, and elderly whole blood. Monocyte-derived DC activation was characterized by targeted gene expression analysis, caspase-1 and NF-κB studies, cytokine multiplex and naive autologous CD4+ T cell activation. Dual activation of newborn DCs via the C-type lectin receptor, macrophage-inducible C-type lectin (trehalose-6,6-dibehenate), and TLR7/8 (R848) greatly enhanced caspase-1 and NF-κB activation, Th1 polarizing cytokine production and autologous Th1 polarization. Combined activation via TLR4 (glycopyranosyl lipid adjuvant aqueous formulation) and Dectin-1 (β-glucan peptide) acted synergistically in newborns and adults, but to a lesser extent. The degree of synergy varied dramatically with age, and was the greatest in newborns and infants with less synergy in adults and elders. Overall, combination adjuvant systems demonstrate markedly different immune activation with age, with combined DC activation via Macrophage-inducible C-type lectin and TLR7/8 representing a novel approach to enhance the efficacy of early-life vaccines.

Modulation of Protective Efficacy of Fracisella tularensis Immunogens by Differential Cultivation in Mueler Hinton Broth Versus Brain Heart Infusion Medium

Abstract Francisella tularensis (Ft) can survive in a variety of habitats with a remarkable ability to adapt to changing environmental conditions. For example, Ft expresses a distinct set of antigens (Ags) in the macrophage, which are not expressed in Ft grown in vitro with Mueler Hinton Broth (MHB) medium. In contrast, Ft grown in Brain Heart Infusion (BHI) medium exhibits an antigenic profile more closely resembling that of in vivo growth conditions. In addition, studies also suggest an alteration in immunological properties of Ft when grown in BHI versus MHB medium. Thus, we investigated the protective efficacy of BHI and MHB-grown Ft, when inactivated and used as an immunogen to protect against Ft LVS challenge. Inactivated Ft (iFt) generated in MHB (iFt-MHB) was found to have superior protective efficacy as compared to that of the iFt generated in BHI (iFt-BHI). The enhanced protection generated by iFt- MHB-immunized mice was also associated with lower bacterial burden and sepsis-like conditions in the lungs and spleens of iFt-MHB versus iFt-BHI-immunized mice. Further evaluation of early immunological events revealed, in the case of iFt-MHB immunogen, superior Ag binding to Ag-presenting dendritic cells (DCs), increased MHC class II expression by DCs, and enhanced Ag presentation by DCs, as apposed to the use of iFt-BHI. Also, iFt-MHB induced increased levels of Ft-specific IgG as well as IFNg and IL17 production versus that of iFt-BHI. These studies thus demonstrate the critical importance of considering bacterial growth medium in Ft vaccine development.

Adjuvant for vaccine immunotherapy of cancer – focusing on Toll‐like receptor 2 and 3 agonists for safely enhancing antitumor immunity

Immune-enhancing adjuvants usually targets antigen (Ag)-presenting cells to tune up cellular and humoral immunity. CD141(+) dendritic cells (DC) represent the professional Ag-presenting cells in humans. In response to microbial pattern molecules, these DCs upgrade the maturation stage sufficient to improve cross-presentation of exogenous Ag, and upregulation of MHC and costimulators, allowing CD4/CD8 T cells to proliferate and liberating cytokines/chemokines that support lymphocyte attraction and survival. These DCs also facilitate natural killer-mediated cell damage. Toll-like receptors (TLRs) and their signaling pathways in DCs play a pivotal role in DC maturation. Therefore, providing adjuvants in addition to Ag is indispensable for successful vaccine immunotherapy for cancer, which has been approved in comparison with antimicrobial vaccines. Mouse CD8α(+) DCs express TLR7 and TLR9 in addition to the TLR2 family (TLR1, 2, and 6) and TLR3, whereas human CD141(+) DCs exclusively express the TLR2 family and TLR3. Although human and mouse plasmacytoid DCs commonly express TLR7/9 to respond to their agonists, the results on mouse adjuvant studies using TLR7/9 agonists cannot be simply extrapolated to human adjuvant immunotherapy. In contrast, TLR2 and TLR3 are similarly expressed in both human and mouse Ag-presenting DCs. Bacillus Calmette-Guerin peptidoglycan and polyinosinic-polycytidylic acid are representative agonists for TLR2 and TLR3, respectively, although they additionally stimulate cytoplasmic sensors: their functional specificities may not be limited to the relevant TLRs. These adjuvants have been posted up to a certain achievement in immunotherapy in some cancers. We herein summarize the history and perspectives of TLR2 and TLR3 agonists in vaccine-adjuvant immunotherapy for cancer.

Mutual Interaction of Basophils and T Cells in Chronic Inflammatory Diseases.

Basophils are, together with mast cells, typical innate effector cells of allergen-induced IgE-dependent allergic diseases. Both cell types express the high-affinity receptor for IgE (FcεR1), release histamine, inflammatory mediators, and cytokines following FcεR1 cross-linking. Basophils are rare granulocytes in blood, lymphoid, and non-lymphoid tissues, and the difficulties to detect and isolate these cells has hampered the study of their biology and the understanding of their possible role in pathology. Furthermore, the existence of other FcεR1-expressing cells, including professional Ag-presenting dendritic cells, generated some controversy regarding the ability of basophils to express MHC Class II molecules, present Ag and drive naïve T cell differentiation into Th2 cells. The focus of this review is to present the recent advances on the interactions between basophils and peripheral blood and tissue memory Th1, Th2, and Th17 cells, as well as their potential role in IgE-independent non-allergic chronic inflammatory disorders, including human inflammatory bowel diseases. Basophils interactions with the innate players of IgE-dependent allergic inflammation, particularly innate lymphoid cells, will also be considered. The previously unrecognized function for basophils in skewing adaptive immune responses opens novel perspectives for the understanding of their contribution to the pathogenesis of inflammatory diseases.

Histamine Promotes the Development of Monocyte-Derived Dendritic Cells and Reduces Tumor Growth by Targeting the Myeloid NADPH Oxidase

The efficiency of immune-mediated clearance of cancer cells is hampered by immunosuppressive mediators in the malignant microenvironment, including NADPH oxidase-derived reactive oxygen species. We aimed at defining the effects of histamine, an inhibitor of the myeloid NADPH oxidase/NOX2, on the development of Ag-presenting dendritic cells (DCs) from myeloid precursors and the impact of these mechanisms for tumor growth. Histamine was found to promote the maturation of human DCs from monocytes by increasing the expression of HLA-DR and costimulatory molecules, which resulted in improved induction of Th cells with Th0 polarity. Experiments using wild-type and NOX2-deficient myelomonoblastic cells showed that histamine facilitated myeloid cell maturation only in cells capable of generating reactive oxygen species. Treatment of mice with histamine reduced the growth of murine EL-4 lymphomas in parallel with an increment of tumor-infiltrating DCs in NOX2-sufficient mice but not in NOX2-deficient (gp91(phox) (-/-)) mice. We propose that strategies to target the myeloid NADPH oxidase may facilitate the development of endogenous DCs in cancer.

Enhancement of adaptive immunity by the human vaccine adjuvant AS01 depends on activated dendritic cells.

Adjuvant System AS01 is a liposome-based vaccine adjuvant containing 3-O-desacyl-4'-monophosphoryl lipid A and the saponin QS-21. AS01 has been selected for the clinical development of several candidate vaccines including the RTS,S malaria vaccine and the subunit glycoprotein E varicella zoster vaccine (both currently in phase III). Given the known immunostimulatory properties of MPL and QS-21, the objective of this study was to describe the early immune response parameters after immunization with an AS01-adjuvanted vaccine and to identify relationships with the vaccine-specific adaptive immune response. Cytokine production and innate immune cell recruitment occurred rapidly and transiently at the muscle injection site and draining lymph node postinjection, consistent with the rapid drainage of the vaccine components to the draining lymph node. The induction of Ag-specific Ab and T cell responses was dependent on the Ag being injected at the same time or within 24 h after AS01, suggesting that the early events occurring postinjection were required for these elevated adaptive responses. In the draining lymph node, after 24 h, the numbers of activated and Ag-loaded monocytes and MHCII(high) dendritic cells were higher after the injection of the AS01-adjuvanted vaccine than after Ag alone. However, only MHCII(high) dendritic cells appeared efficient at and necessary for direct Ag presentation to T cells. These data suggest that the ability of AS01 to improve adaptive immune responses, as has been demonstrated in clinical trials, is linked to a transient stimulation of the innate immune system leading to the generation of high number of efficient Ag-presenting dendritic cells.

Enhanced function of cytotoxic T lymphocytes induced by dendritic cells modified with truncated PSMA and 4-1BBL

Interactions between costimulatory molecules and their receptors are vital for Ag-presenting dendritic cells (DCs) to initiate T cells activation, expansion and their antitumor immune responses. Augmentation of costimulatory signal due to the interaction of DCs and T cells may amplify, sustain and drive diversity of cytotoxic T lymphocytes (CTLs) and consequently enhance the antitumor response. 4-1BBL/4-1BB is such a pair of costimulatory ligand and receptor, playing an important role in the co-stimulation of CTLs. Previously, we demonstrated that DCs transduced with recombinant adenovirus encoding truncated PSMA (tPSMA) and m4-1BBL could induce prostate cancer regression in mouse models. In the present study, we further explored the adjuvant role of 4-1BBL in modulating CTLs activation induced by tPSMA gene-pulsed DCs. The apoptosis and cytotoxicity against tPSMA expressing RM-1 cells of CTLs were determined. Results showed that tPSMA gene-pulsed DCs effectively induced T lymphocyte activation and cytotoxicity, which was enhanced by upregulated expression of 4-1BBL, displaying better cell viability, lower CTLs apoptosis, higher expression anti-apoptotic protein of Bcl-xL and phosphorylation of P38, enhanced NF-κB activation, as well as more IFN-γ production. These results demonstrated that 4-1BBL may play a significant role in the co-stimulation pathway for Ag-presenting DCs-mediated CTLs activity, which might be a beneficial adjuvant factor for DCs-based cancer immunotherapy.

SB1578, a Novel Inhibitor of JAK2, FLT3, and c-Fms for the Treatment of Rheumatoid Arthritis

SB1578 is a novel, orally bioavailable JAK2 inhibitor with specificity for JAK2 within the JAK family and also potent activity against FLT3 and c-Fms. These three tyrosine kinases play a pivotal role in activation of pathways that underlie the pathogenesis of rheumatoid arthritis. SB1578 blocks the activation of these kinases and their downstream signaling in pertinent cells, leading to inhibition of pathological cellular responses. The biochemical and cellular activities of SB1578 translate into its high efficacy in two rodent models of arthritis. SB1578 not only prevents the onset of arthritis but is also potent in treating established disease in collagen-induced arthritis mice with beneficial effects on histopathological parameters of bone resorption and cartilage damage. SB1578 abrogates the inflammatory response and prevents the infiltration of macrophages and neutrophils into affected joints. It also leads to inhibition of Ag-presenting dendritic cells and inhibits the autoimmune component of the disease. In summary, SB1578 has a unique kinase spectrum, and its pharmacological profile provides a strong rationale for the ongoing clinical development in autoimmune diseases.

Suppressor of Cytokine Signaling 1 DNA Administration Inhibits Inflammatory and Pathogenic Responses in Autoimmune Myocarditis

Myocarditis and subsequent dilated cardiomyopathy are major causes of heart failure in young adults. Myocarditis in humans is highly heterogeneous in etiology. Recent studies have indicated that a subgroup of myocarditis patients may benefit from immune-targeted therapies, because autoimmunity plays an important role in myocarditis as well as contributing to the progression to cardiomyopathy and heart failure. Suppressor of cytokine signaling (SOCS) 1 plays a key role in the negative regulation of both TLR- and cytokine receptor-mediated signaling, which is involved in innate immunity and subsequent adaptive immunity. In this study, we investigated the therapeutic effect of SOCS1 DNA administration on experimental autoimmune myocarditis (EAM) in mice. EAM was induced by s.c. immunization with cardiac-specific peptides derived from α myosin H chain in BALB/c mice. In contrast to control myocarditis mice, SOCS1 DNA-injected mice were protected from development of EAM and heart failure. SOCS1 DNA administration was effective for reducing the activation of autoreactive CD4(+) T cells by inhibition of the function of Ag-presenting dendritic cells. Our findings suggest that SOCS1 DNA administration has considerable therapeutic potential in individuals with autoimmune myocarditis and dilated cardiomyopathy.

The Adaptor Protein Bam32 in Human Dendritic Cells Participates in the Regulation of MHC Class I-Induced CD8+ T Cell Activation

The B lymphocyte adaptor molecule of 32 kDa (Bam32) is strongly induced during the maturation of dendritic cells (DC). Most known functions of Bam32 are related to the signaling of the B cell receptor for Ag. Because DC do not express receptors specific for Ags, we aim at characterizing the role of Bam32 in human monocyte-derived DC in this study. Our results show that binding of allogeneic T cells to mature DC causes accumulation of Bam32 on the contact sites and that this translocation is mimicked by Ab-mediated engagement of MHC class I. Silencing of Bam32 in mature monocyte-derived DC results in an enhanced proliferation of CD8(+) T cells in an Ag-specific T cell proliferation assay. Further studies identify galectin-1 as an intracellular binding partner of Bam32. Regulating immune responses via regulatory T cell (Treg) modulation is one of the many immunological activities attributed to galectin-1. Therefore, we assayed mixed leukocyte reactions for Treg expansion and found fewer Treg in reactions stimulated with DC silenced for Bam32 compared to reactions stimulated with DC treated with a nontarget control. Based on our findings, we propose a role for Bam32 in the signaling of MHC class I molecules in professional Ag-presenting DC for the regulation of CD8(+) T cell activation. It is distinct from that of MHC class I recognized by CD8(+) T cells leading to target [corrected] cell death. Thus, our data pinpoint a novel level of T cell regulation that may be of biological relevance.

Proapoptotic and Antiapoptotic Actions of Stat1 versus Stat3 Underlie Neuroprotective and Immunoregulatory Functions of IL-11

Current therapies for multiple sclerosis target inflammation but do not directly address oligodendrocyte protection or myelin repair. The gp130 family cytokines ciliary neurotrophic factor, leukemia inhibitory factor, and IL-11 have been identified as oligodendrocyte growth factors, and IL-11 is also strongly immunoregulatory, but their underlying mechanisms of action are incompletely characterized. In this study, we demonstrate that these effects of IL-11 are mediated via differential regulation of apoptosis in oligodendrocytes versus Ag-presenting dendritic cells (DCs), and are dependent on lineage-specific activity of the transcription factors Stat1 versus Stat3. Focal demyelinating lesions induced in cerebral cortices of IL-11Rα(-/-) mice using stereotactic microinjection of lysolecithin were larger than in controls, and remyelination was delayed. In IL-11Rα(-/-) mice, lesions displayed extensive oligodendrocyte loss and axonal transection, and increased infiltration by inflammatory cells including CD11c(+) DCs, CD3(+) lymphocytes, and CD11b(+) phagocytes. In oligodendrocyte progenitor cell (OPC) cultures, IL-11 restricted caspase 9 activation and apoptosis, and it increased myelination in OPC-neuron cocultures. Importantly, siRNA inhibition of Stat1 enhanced the antiapoptotic effects of IL-11 on OPCs, but IL-11 induced apoptosis in the presence of Stat3 silencing. In contrast, IL-11 augmented caspase activation and apoptosis in cultures of CD11c(+) DCs, but not in CD11b(+) or CD3(+) cells. Inhibition of Stat3 exacerbated the proapoptotic effects of IL-11 on DCs, whereas they were ablated in Stat1(-/-) cultures. Collectively, these findings reveal novel mechanisms underlying the actions of a neuroprotective and immunoregulatory member of the gp130 cytokine family, suggesting avenues to enhance oligodendrocyte viability and restrict CNS inflammation in multiple sclerosis.

Basophils as TH2‐inducing APCs: the dog can sing but is it a diva?

Basophils as T_H2‐inducing APCs: the dog can sing but is it a diva?

Cutting Edge: CD28 Engagement Releases Antigen-Activated Invariant NKT Cells from the Inhibitory Effects of PD-1

Costimulatory and coinhibitory signals are important for the maintenance of immune homeostasis both in the steady state and during immune responses. In this study, we explore the relative contributions of these signals to the rapid production of large amounts of cytokines by activated invariant NKT cells (iNKT cells). We find that upon antigenic stimulation, iNKT cells rapidly up-regulate programmed death (PD)-1 and induce high levels of PD ligand 1 and costimulatory molecules on the surface of cognate Ag-presenting dendritic cells and that iNKT cells require a CD28 signal to secrete cytokines in the presence of a PD-1/PD ligand 1 interaction. CD28-deficient iNKT cells synthesized but failed to secrete cytokines during activation, and blockade of the PD-1 pathway restored the ability of CD28-deficient iNKT cells to secrete cytokines. The opposing functions of CD28 and PD-1 thus tightly regulate the unique effector function iNKT cells.

IFN-α Skews Monocytes into CD56+-Expressing Dendritic Cells with Potent Functional Activities In Vitro and In Vivo

The antitumor effect of IFN-alpha is mediated by the activation of CTLs, NK cells, and the generation of highly potent Ag-presenting dendritic cells (IFN-DCs). In this study, we show that IFN-DCs generated in vitro from monocytes express CD56 on their surface, a marker which has been thought to be specific for NK cells. FACS analyses of CD56(+) and CD56(-) IFN-DCs showed a nearly identical pattern for most of the classical DC markers. Importantly, however, only CD56(+) IFN-DCs exhibited cytolytic activity up to 24% that could almost completely be blocked (-81%) after coincubation with anti-TRAIL. Intracytoplasmatic cytokine staining revealed that the majority of IFN-DCs independently of their CD56 expression were IFN-gamma positive as well. In contrast, CD56(+) IFN-DCs showed stronger capacity in stimulating allogenic T cells compared with CD56(-) IFN-DC. Based on these results, five patients with metastasized medullary thyroid carcinoma were treated for the first time with monocyte-derived tumor Ag-pulsed IFN-DCs. After a long term follow-up (in mean 37 mo) all patients are alive. Immunohistochemical analyses of delayed-type hypersensitivity skin reaction showed a strong infiltration with CD8(+) cells. In two patients no substantial change in tumor morphology was detected. Importantly, by analyzing PBMCs, these patients also showed an increase of Ag-specific IFN-gamma-secreting T cells. In summary, we here describe for the first time that cytotoxic activity of IFN-DCs is mainly mediated by an IFN-DC subset showing partial phenotypic and functional characteristics of NK cells. These cells represent another mechanism of the antitumor effect induced by IFN-alpha.