Activation Of Myeloid Dendritic Cells Research Articles

Vitamin D3 mitigates autoimmune inflammation caused by activation of myeloid dendritic cells in SLE.

Systemic lupus erythematosus (SLE) is an autoimmune disease in which defective T cells, immune complex deposition and other immune system alterations contribute to pathological changes of multiple organ systems. The vitamin D metabolite c is a critical immunomodulator playing pivotal roles in the immune system. Epidemiological evidence indicates that vitamin D deficiency is correlated with the severity of SLE. Our aim is to investigate the effects of 1,25(OH)2D3 (VitD3) on the activation of myeloid dendritic cells (mDCs) by autologous DNA-containing immune complex (DNA-ICs), and the effects of VitD3 on immune system balance during SLE. We purified DNA-ICs from the serum of SLE patients and isolated mDCs from normal subjects. In vitro studies showed that DNA-ICs were internalized and consumed by mDCs. VitD3 blocked the effects of DNA-ICs on RelB, IL-10 and TNF-α in mDCs. Further analysis indicated that DNA-ICs stimulated histone acetylation in the RelB promoter region, which was inhibited by VitD3. Knockdown of the histone deacetylase 3 gene (HDAC3) blocked these VitD3-mediated effects. Co-culture of mDCs and CD4+ T cells showed that VitD3 inhibited multiple processes mediated by DNA-ICs, including proliferation, downregulation of IL-10, TGF-β and upregulation of TNF-α. Moreover, VitD3 could also reverse the effects of DNA-IC-induced imbalance of CD4+ CD127- Foxp3+ T cells and CD4+ IL17+ T cells. Taken together, our results indicated that autologous DNA-ICs stimulate the activation of mDCs in the pathogenesis of SLE, and VitD3 inhibits this stimulatory effects of DNA-ICs by negative transcriptional regulation of RelB gene and maintaining the Treg/Th17 immune cell balance. These results suggest that vitamin D may have therapeutic value for the treatment of SLE.

Integrated bioinformatics analysis of IFITM1 as a prognostic biomarker and investigation of its immunological role in prostate adenocarcinoma.

Prostate adenocarcinoma (PRAD) is a highly aggressive malignancy with high mortality and poor prognosis, and its potential mechanism remains unclear. Our study aimed to identify novel markers for the prognosis of PRAD using bioinformatics technology. The GSE32571 dataset was downloaded from the GEO database, and analyzed via the limma R package to identify differentially expressed genes (DEGs) and differentially expressed immune score-related genes (DEISRGs). The immune-related genes (IRGs) were further obtained by overlapping DEISRGs and DEGs, and the core gene was identified via survival analysis. Furthermore, the expression level, prognostic value, and potential functions of the core gene were evaluated via multiple bioinformatics databases. A total of 301 IRGs were identified from the GSE32571 dataset, and IFITM1 was a down-regulated gene in several types of cancer, including PRAD. Besides, low expression of IFITM1 was associated with a poor prognosis in PRAD. GSEA indicated that the vital pathways of IFITM1-associated genes were mainly enriched in primary immunodeficiency, Th17 cell differentiation, Th1, and Th2 cell differentiation, natural killer cell-mediated cytotoxicity, myeloid dendritic cell activation, regulation of leukocyte activation, etc. Furthermore, IFITM1 was closely correlated with 22 types of tumor-infiltrating immune cells. IFITM1 was a prognostic biomarker for PRAD patients, and it can be acted as a potential immune therapy target in PRAD.

Selected Articles from This Issue

Davis and colleagues report the first systematic assessment of the safety and pharmacokinetics of dual immune checkpoint inhibition with ipilimumab and nivolumab in pediatric, adolescent, and young adult patients with relapsed or refractory sarcoma. They establish a recommended phase II dose of nivolumab (3 mg/kg) plus ipilimumab (1 mg/kg) and found the combination to be generally well tolerated. In expanded phase II cohorts, the authors observed two sustained partial responses. They further demonstrate that increased ipilimumab dosing (3 mg/kg) in combination with nivolumab (1 mg/kg) carries higher toxicity without clinical benefit in this population.Advanced stage head and neck squamous cell carcinoma (HNSCC) patients are at a high risk of recurrent disease. Due to dismal 5-year survival rates, such patients are in dire need of effective adjuvant therapy. Everolimus, an mTOR inhibitor, has documented activity in HNSCC and is well tolerated with minimal long-term toxicity. In this placebo-controlled phase II trial, Nathan and colleagues are the first to show promising results using everolimus as adjuvant therapy after complete response to definitive treatment in a subset of HPV-negative patients with advanced stage disease. In particular, HPV-negative TP53 mutated tumors appear to yield the best benefit. Thus, subsequent trials using everolimus in this patient population are warranted.TLR9 agonists demonstrated efficacy in activation of the innate and adaptive immune system in preclinical models. Babiker and colleagues investigated the activity of tilsotolimod, an investigational synthetic TLR9 agonist, in patients with refractory solid tumors after progression on standard of care therapies. This early phase clinical trial established safety, maximum tolerated dose, and demonstrated early signs of efficacy of tilsotolimod. In addition, translational data revealed activation of the innate and adaptive immune system through the type I interferon (IFN) pathway, IFNγ and IFNα gene upregulation, increase in inflammatory chemokines, activation of myeloid dendritic cells and antigen presentation, and an increase in genes for checkpoint and costimulatory proteins. The translational results suggest further investigation of tilsotolimod in combination with checkpoint inhibitors in solid tumors.Biomarkers are urgently needed to select chemotherapy regimens in pancreatic ductal adenocarcinoma (PDAC). In this study, Perera and colleagues investigated whether mRNA expression levels of hENT1 predict responsiveness to gemcitabinenab-paclitaxel (GnP) in patients with advanced PDAC. The results of this prospective observational clinical trial using tumor-enriched RNA sequencing revealed higher response rates and improved survival in patients receiving GnP, when tumors were considered hENT1high compared to those that were hENT1low. No difference in response or survival were seen in patients receiving modified FOLFIRINOX. An interaction analysis confirmed hENT1 as a predictive biomarker for GnP response. This suggests that hENT1 expression can be used to identify patients who will respond to GnP.

Deletion of Ppard in CD11c+ cells attenuates atherosclerosis in ApoE knockout mice.

Ppardδ, one of the lipid-activated nuclear receptor expressed in many cell typesto activate gene transcription, also regulates cellular functions other than lipid metabolism. The mechanism regulating the function of antigen-presenting cells during the development of atherosclerosis is not fully understood. Here we aimed to study the involvement of PPARδ in CD11c+ cells in atherosclerosis. We used the Cre-loxP approach to make conditional deletion of Ppard in CD11c+ cells in mice on Apoe-/- background, which were fed with high cholesterol diet to develop atherosclerosis. Ppard deficiency in CD11c+ cells attenuated atherosclerotic plaque formation and infiltration of myeloid-derived dendritic cells (DCs) and T lymphocytes. Reduced lesion was accompanied by reduced activation of dendritic cells, and also a reduction of activation and differentiation of T cells to Th1 cells. In addition, DC migration to lymph node was also attenuated with Ppard deletion. In bone marrow-derived DCs, Ppard deficiency reduced palmitic acid-induced upregulation of co-stimulatory molecules and pro-inflammatory cytokine IL12 and TNFα. Our results indicated PPARδ activation by fatty acid resulted in the activation of myeloid DCs and subsequent polarization of T lymphocytes, which contributed to atherosclerosis in Apoe-/- mice. These findings also reveal the potential regulatory role of PPARδ in antigen presentation to orchestrate the immune responses during atherosclerosis.

Abnormal Activation of Myeloid Dendritic Cells in JORRP Patients and Associated with Disease Activity

BackgroundRepresenting the first line of host defense against virus infections and an essential link between innate and adaptive immune response, the role of dendritic cells (DCs) in peripheral blood of juvenile-onset recurrent respiratory papillomatosis (JORRP) patients and association with disease activity were still not established. Materials and MethodsIn our present study, 28 JORRP patients and 28 age and sex matched healthy controls were enrolled. The percentage, phenotype and cytokine secretion of DC and was measured by flow cytometry. Plasma cytokine were detected by the enzyme-linked immunosorbent assay (ELISA). ResultsWe found that the percentage of myeloid DC (mDC) was significantly lower in JORRP patients compared to healthy controls and was negatively correlated with interval times, but not surgical times or disease onset. Moreover, the activation marker, CD40 and CD86 was significantly up-regulated on the surfaces of mDC in JORRP patients compared with healthy controls. Neither the percentage nor activation of plasmacytoid DC (pDC) showed statistical difference between JORRP patients and healthy controls. HLA-DR expression on both mDC and pDC was down-regulated in JORRP group and negatively correlated with surgical times. Antigen presenting ability of DC was greatly impaired in JORRP patients of higher number of operations and shorter interval time. Plasma IL-10 as well as IL-10 secreted by mDC was higher in JORRP patients compared with healthy control. Finally, we detected an up-regulated TLR2 and TLR4 expression on mDCs and TLR4 expression was positively correlated with HLA-DR expression on mDC of JORRP patients. ConclusionOur results demonstrate an abnormal TLR2 and TLR4 expression in mDCs may contribute to suppressive immune response to HPV6 or HPV11 infection and associated with disease activity in JORRP patients.

1468P - Effector CD4+ T-cell induction by thoracic radiotherapy for patients with NSCLC

BackgroundA recent clinical study indicated that anti-programmed death-ligand 1 antibody durvalumab consolidation therapy for patients with non-small cell lung cancer (NSCLC) who received chemoradiotherapy (CRT) resulted in robust improvement in progression-free survival (PFS) and prevented distant metastasis. It has been demonstrated that cluster of differentiation (CD)8+ T cell priming is facilitated by immunogenic cancer cell death by radiotherapy. On the other hand, accumulating evidence indicates that CD4+ T cells are required for effective antitumor T cell responses because CD4+ T cells help regulate priming, migration, invasive potential, and antitumor killing activity of cytotoxic lymphocytes. It remains uncertain how thoracic CRT evokes antitumor T cell immune responses in patients with NSCLC. MethodsThe present study comprised 56 consecutive patients with locally advanced NSCLC who received curative CRT or thoracic radiotherapy in the Saitama Medical University International Medical Center. Twenty-two patients received durvalumab consolidation therapy after CRT. Peripheral blood samples were collected before and sequentially after radiotherapy. Peripheral blood mononuclear cells were analyzed using LSR FortessaTM and CyTOFTM. ResultsAfter radiotherapy, significant increases in the percentages of CD62Llow CD4+ T cells and human leukocyte antigen-DR+ myeloid dendritic cells were observed. Mass cytometry analysis revealed that the increased CD62Llow CD4+ T cells consisted of T-bet+ CXCR3+ CD27- type 1 T helper (Th1) cells. Programmed death-1, lymphocyte-activation gene-3, and inducible T-cell costimulator expression in CD62Llow CD4+ T cells also significantly increased. Interestingly, CD62Llow CD4+ T cells initiated decreasing 4 weeks after radiotherapy in half of the patients. ConclusionsRadiotherapy induced myeloid dendritic cell activation and subsequent increase in T-bet+ CXCR3+ CD27- CD62Llow CD4+ Th1 cells. The correlation between CD4+ T-cell responses and clinical responses of durvalumab is under examination. Legal entity responsible for the studyThe authors. FundingHas not received any funding. DisclosureO. Yamaguchi: Research grant / Funding (institution): Nihon Medi-Physics Co., Ltd.; Speaker Bureau / Expert testimony: Ono Pharmaceutical; Speaker Bureau / Expert testimony: Bristol-Myers Squibb; Speaker Bureau / Expert testimony: AstraZeneca. A. Mouri: Speaker Bureau / Expert testimony: Ono Pharmaceutical; Speaker Bureau / Expert testimony: AstraZeneca; Speaker Bureau / Expert testimony: Eli Lilly; Speaker Bureau / Expert testimony: Boehringer Ingelheim. K. Kobayashi: Speaker Bureau / Expert testimony: AstraZeneca; Speaker Bureau / Expert testimony: Boehringer Ingelheim; Speaker Bureau / Expert testimony: Taiho Pharmaceutical Company; Speaker Bureau / Expert testimony: Ono Pharmaceutical Company. K. Kaira: Research grant / Funding (institution): AstraZeneca; Speaker Bureau / Expert testimony: Boehringer Ingelheim; Speaker Bureau / Expert testimony: Ono Pharmaceutical Company; Speaker Bureau / Expert testimony: Eli Lilly. H. Kagamu: Honoraria (self): AstraZeneca; Honoraria (self), Research grant / Funding (institution): Eli Lilly; Honoraria (self), Research grant / Funding (institution): Boehringer Ingelheim; Honoraria (self), Research grant / Funding (institution): Ono Pharmaceutical; Honoraria (self), Research grant / Funding (institution): Taiho Pharmaceutical; Honoraria (self), Research grant / Funding (institution): Chugai Pharmaceutical. All other authors have declared no conflicts of interest.

The Role of PKM2 in the Activation of Myeloid Dendritic Cells from Patients with Severe Aplastic Anemia

Severe aplastic anemia (SAA) is a hematologic disease characterized by severe bone marrow failure. The abnormal activation of myeloid dendritic cells (mDCs) in SAA might accelerate the polarization of Th0 cells to Th1 cells ,thus causing the over-function of T lymphocytes and apoptosis of hematopoietic cells. Hence, mDCs played an important role in the primary stage of the immune response.Here, we analyze the protein components of mDC from SAA as well as normal control to explore the possible reason that activate mDC. The protein components of mDCs was detected by proteomics technology. Furtherly we applied both realtime PCR and western blot from a perspective of mRNA and protein to validate our discovery. Results revealed that expression of pyruvate kinase M2(PKM2) were enhanced in mDCs from SAA patients at an early stage of the onset. Concurrently, the level of PKM2 in mDCs of SAA patients was conspicuously higher than that in normal control not only at a protein level but also at the gene level.To clarify the role of PKM2 in the activation of mDCs, we investigate the effect of targeted PKM2 siRNA -knockout in mDCs at the cellular level. We measured the levels of costimulatory molecules CD80 and CD86 on mDCs by flow cytometry and identified significantly higher expression of CD86 in the control group relative to the transfected group.To quantify antigen uptake and processing capacities of mDCs, cells were treated with carboxylate-modified yellow-green (YG) microspheres and then analyzed by flow cytometry. The percentage of phagocytosis (PP) was determined by the following formula: PP = M1 + M2 + M3 + M4, where M1, M2, M3, and M4 correspond to the number of cells with one, two, three and four microspheres. Phagocytosis index(PI)=(intracellular cpms/ intracellular + extracellular cpms)x100%.After PKM2 siRNA transfection, we observed lower phagocytic ability of mDCs. Additionally, the level of PP of mDCs from control group was higher than that of PKM2 siRNA -knockout group. Through scanning electron microscopy (SEM), mDCs before siRNA transfection from SAA patients presented longer and more branched protrusions than those seen in PKM2 siRNA transfected mDCs.Next, the effects of PKM2 on cytotoxic T cells(CTL) activation by mDCs from SAA patients were also determined. mDCs before and after PKM2 siRNA transfection were co cultured with CTL cells for 72 hours. After cell co culture, the mRNA expressions of perforin were identified significantly higher in the control group relative to the transfected group. Culture supernatant were collected for quantification of IFN-γ and IL-4 by commercial ELISA kits. The levels of IFN-γ and IL-4 in contral group were much higher than that of transfected group .Additionally, flow cytometry experiments assessing annexin V/propidium iodide staining showed that high PKM2 expression in mDCs was significantly associated with reduced apoptosis rate of CTLs. Therefore, the enhanced function of mDCs from SAA could activate the downstream CTL function and then enhance the killing effect to target cells.Our findings demonstrated that elevated levels of PKM2 in mDCs play an important role in the activation of mDCs and thereby affecting the immune status of SAA by enhancing the function of mDCs and downstream CTLs. DisclosuresNo relevant conflicts of interest to declare.

B cell-derived transforming growth factor-β1 expression limits the induction phase of autoimmune neuroinflammation.

Studies in experimental autoimmune encephalomyelitis (EAE), a murine model of multiple sclerosis (MS), have shown that regulatory B cells modulate the course of the disease via the production of suppressive cytokines. While data indicate a role for transforming growth factor (TGF)-β1 expression in regulatory B cell functions, this mechanism has not yet been tested in autoimmune neuroinflammation. Transgenic mice deficient for TGF-β1 expression in B cells (B–TGF-β1−/−) were tested in EAE induced by recombinant mouse myelin oligodendrocyte glycoprotein (rmMOG). In this model, B–TGF-β1−/− mice showed an earlier onset of neurologic impairment compared to their littermate controls. Exacerbated EAE susceptibility in B–TGF-β1−/− mice was associated with augmented CNS T helper (Th)1/17 responses. Moreover, selective B cell TGF-β1–deficiency increased the frequencies and activation of myeloid dendritic cells, potent professional antigen-presenting cells (APCs), suggesting that B cell-derived TGF-β1 can constrain Th1/17 responses through inhibition of APC activity. Collectively our data suggest that B cells can down-regulate the function of APCs, and in turn encephalitogenic Th1/17 responses, via TGF-β1, findings that may be relevant to B cell-targeted therapies.

Activation of myeloid dendritic cells, effector cells and regulatory T cells in lichen planus.

BackgroundLichen planus (LP) is a chronic mucocutaneous inflammatory disease. Evaluating the balance between regulatory T cells and effector T cells could be useful for monitoring the proinflammatory profile of LP. Therefore, this study aimed to assess populations of dendritic cells (DCs) and regulatory and effector T cells in peripheral blood samples collected from patients with LP to evaluate the polyfunctionality of T cells upon toll-like receptor (TLR) activation.MethodsPeripheral blood mononuclear cells collected from 18 patients with LP and 22 healthy control subjects were stimulated with agonists of TLR4, TLR7, TLR7/TLR8 or TLR9. Frequencies of circulating IFN-α+ plasmacytoid DCs (pDCs); TNF-α+ myeloid DCs (mDCs); regulatory T cells (Tregs); and IL-17-, IL-10-, IL-22-, TNF-, and IFN-γ-secreting T cells were assessed via flow cytometry.ResultsThe frequencies of regulatory CD4+ and CD8+CD25+Foxp3+CD127low/− T cells and TNF-α+ mDCs were induced following activation with TLR4, TLR7 and TLR8 agonists in the LP group. Moreover, increased baseline frequencies of CD4+IL-10+ T cells and CD8+IL-22+ or IFN-γ+T cells were found. In the LP group, TLR4 activation induced an increased frequency of CD4+IFN-γ+ T cells, while TLR7/8 and staphylococcal enterotoxin B (SEB) activation induced an increased frequency of CD8+ IL-22+ T cells. An increased frequency of polyfunctional CD4+ T cells that simultaneously secreted 3 of the evaluated cytokines (not including IL-10) was verified upon TLR7/8/9 activation, while polyfunctional CD8+ T cells were already detectable at baseline.ConclusionsTLR-mediated activation of the innate immune response induced the production of proinflammatory mDCs, Tregs and polyfunctional T cells in patients with LP. Therefore, TLR activation has an adjuvant role in inducing both innate and adaptive immune responses.Electronic supplementary materialThe online version of this article (doi:10.1186/s12967-016-0938-1) contains supplementary material, which is available to authorized users.

Comparison of the TLR7 and TLR9 innate immune signaling pathways in african green monkeys and rhesus macaques

Abstract Innate immunity is an evolutionary mechanism which is involved in the initial detection of pathogens and stimulates the first line of host defense. The Toll-like receptor (TLR) family has been the most extensively studied innate immune pathway and it has been shown that TLRs can be activated in response to virtually any microbe which invades the host. These innate immune responses play a critical role in the development of pathogen-specific adaptive immune responses and disease pathology and are targeted by the use of adjuvants in modern vaccines. Importantly, divergent TLR7 and TLR9 signaling pathways between rhesus macaques and sooty mangabeys have been shown to distinguish between pathogenic and nonpathogenic AIDS virus infections in those species. Therefore, it is important to understand the TLR signaling pathways of non-human primate (NHP) species that are currently used for infectious disease and vaccine research. In the present study, we administered defined TLR7 and TLR9 agonists to two distinct NHP species, african green monkeys and rhesus macaques. We evaluated B cell, T cell, NK cell, monocyte, plasmacytoid dendritic cell and myeloid dendritic cell activation at 0, 24, 72 and 168 hours after dosing. Gene expression profiling and full pathway analysis was also evaluated to illuminate fundamental aspects of innate immune responses that underlie the mechanism of action of TLR-agonist adjuvants in these two NHP species.

8. Examining the Effects of Peptide-Gold Nanoparticle Hybrids on the Activation of Myeloid Dendritic Cells

Dendritic cells (DCs) are professional antigen-presenting cells that play an important role in innate and adaptive immunity. They have become a major target for immune modulation based therapeutic strategies. Gold nanoparticles (GNPs) are biocompatible materials. The surface of GNPs can be easily modified with peptides via thiol-gold linkage to alter their surface chemistry and to modulate their cellular uptake. The purpose of this present study was to investigate the ability of peptide-GNP hybrids to modulate the DC immune response in vitro. To assess the activation effects of peptide-GNP hybrids, DCs were differentiated from bone marrow (BM) cells of black B57BL/6 mice by culture with granulocyte macrophage colony stimulating factor (GM-CSF). At day 9 of culture, BM-derived DCs (BMDCs) were exposed to GNP hybrids with a diameter of approximately 10 nm. The purity of the resulting DCs was determined by the CD11c positive cell population measured by Flow Cytometry. We found that the peptide-GNP hybrids were internalized by immature BMDC via bright field microscope imaging. Furthermore, the addition of GNPs to immature BMDCs was found to influence both phenotype and function of immature BMDCs. Stimulation by peptide-GNP hybrids on the immature BMDCs led to an enhanced expression of co-stimulatory molecules on the cell surface, including CD40, CD80 and CD86, and a secretion of cytokines in the culture medium. This observation suggests that peptide-GNP stimulation can lead to the activation of BMDCs, and implies a potential application for strengthening vaccine delivery devices or cancer immune therapy.

Persistence of asthma following allergen avoidance is associated with proTh2 myeloid dendritic cell activation

BackgroundThe natural history of asthma includes in some patients periods of disease remission, but the underlying mechanisms are unknown.ObjectivesWe explored whether type 1 myeloid dendritic cell (mDC) dysfunction could be...

Intact dendritic cell pathogen-recognition receptor functions associate with chronic hepatitis C treatment-induced viral clearance.

Although studies have addressed the exhaustion of the host's immune response to HCV and its role in treatment, there is little information about the possible contribution of innate immunity to treatment-induced clearance. We hypothesized that because intact myeloid dendritic cell (MDC) pathogen sensing functions are associated with improved HCV-specific CD8+ T cell functionality in some chronically infected patients, it might enhance HCV clearance rate under standard interferon therapy. To investigate this hypothesis, TLR-induced MDC activation and HCV-specific CD8+ T cell response quality were monitored longitudinally at the single-cell level using polychromatic flow cytometry in chronically infected patients undergoing interferon therapy. We correlated the immunological, biochemical and virological data with response to treatment. We demonstrate that the clinical efficacy of interferon-induced viral clearance is influenced by the extent to which HCV inhibits MDC functions before treatment, rather than solely on a breakdown of the extrinsic T cell immunosuppressive environment. Thus, viral inhibition of MDC functions before treatment emerges as a co-determining factor in the clinical efficacy of interferon therapy during chronic HCV infection.

Thymic stromal lymphopoietin receptor blockade reduces allergic inflammation in a cynomolgus monkey model of asthma

Thymic stromal lymphopoietin (TSLP) pathway blockade is a potential strategy for asthma treatment because the main activities of TSLP are activation of myeloid dendritic cells (mDCs) and modulation of cytokine production by mast cells. TSLP-activated mDCs prime the differentiation of naive T cells into inflammatory TH2 cells. We sought to investigate mechanisms underlying the development of allergic lung inflammation in cynomolgus monkeys using gene expression profiling and to assess the effect of thymic stromal lymphopoietin receptor (TSLPR) blockade in this model. An mAb against human TSLPR was generated and confirmed to be cross-reactive to cynomolgus monkey. Animals were dosed weekly with either vehicle or anti-TSLPR mAb for 6 weeks, and their responses to allergen challenge at baseline, week 2, and week 6 were assessed. After 6 weeks of treatment, anti-TSLPR mAb-treated animals showed reduced bronchoalveolar lavage (BAL) fluid eosinophil counts, reduced airway resistance in response to allergen challenge, and reduced IL-13 cytokine levels in BAL fluid compared with values seen in vehicle-treated animals. Expression profiling of BAL fluid cells collected before and after challenge showed a group of genes upregulated by allergen challenge that strongly overlapped with 11 genes upregulated in dendritic cells (DCs) when invitro stimulated by TSLP (TSLP-DC gene signature). The number of genes differentially expressed in response to challenge was reduced in antibody-treated animals after 6 weeks relative to vehicle-treated animals. Expression of the TSLP-DC gene signature was also significantly reduced in antibody-treated animals. These results demonstrate promising efficacy for TSLPR blockade in an allergic lung inflammation model in which TSLP activation of mDCs might play a key role.

OP0222 TSLP expression is increased in RA joints and causes increased activation of intra-articular myeloid dendritic cells with enhanced stimulation of TH1 and TH17 activity

BackgroundThymic stromal lymphopoietin (TSLP) is well known for its potent activation of myeloid dendritic cells (mDCs) resulting in Th2-mediated immune responses. TSLP signals cells via the IL-7 receptor-alpha chain (IL-7Rα),...

Monocyte and Myeloid Dendritic Cell Activation Occurs Throughout HIV Type 2 Infection, an Attenuated Form of HIV Disease

Monocytes and myeloid dendritic cells (mDCs) are important orchestrators of innate and human immunodeficiency virus (HIV)-specific immune responses and of the generalized inflammation that characterizes AIDS progression. To our knowledge, we are the first to investigate monocyte and mDC imbalances in HIV type 2 (HIV-2)-positive patients, who typically feature reduced viremia and slow disease progression despite the recognized ability of HIV-2 to establish viral reservoirs and overcome host restriction factors in myeloid cells. We found a heightened state of monocyte and mDC activation throughout HIV-2 infection (characterized by CD14(bright)CD16(+) expansion, as well as increased levels of soluble CD14, HLA-DR, and CD86), together with progressive mDC depletion. Importantly, HIV-2-positive patients also featured overexpression of the inhibitory molecule PD-L1 on monocytes and mDCs, which may act by limiting the production of proinflammatory molecules. These data, from patients with a naturally occurring form of attenuated HIV disease, challenge current paradigms regarding the role of monocytes in HIV/AIDS and open new perspectives regarding potential strategies to modulate inflammatory states.

Hepatic enrichment and activation of myeloid dendritic cells during chronic hepatitis C virus infection

Chronic hepatitis C virus (HCV) infection is a serious disease that can result in numerous long-term complications leading to liver failure or death. Approximately 80% of people fail to clear their infection, largely as the result of weak, narrowly targeting or waning antiviral T-cell responses. Although professional antigen presenting cells (APCs) like dendritic cells (DCs) might serve as targets for modulation of T-cell immunity, the particular role of DCs in immunity to HCV is not known. Moreover the identity, phenotype, and functional characteristics of such populations in the liver, the site of HCV replication, have proven difficult to elucidate. Using a multicolor flow-based approach, we identified six distinct populations of professional APCs among liver interstitial leukocytes isolated from uninfected and HCV-infected patients. Although a generalized enrichment of DCs in the liver compared to blood was observed for all patients, HCV infection was characterized by a significant increase in the frequency of intrahepatic myeloid DCs (both CD1c+ and CD141+). Phenotypic analyses of liver plasmacytoid (pDC) and myeloid DCs (mDC) further revealed the HCV-induced expression of maturation molecules CD80, CD83, CD40, and programmed death ligand-1. Importantly, pDC and mDCs from HCV-infected liver were capable of secreting effector cytokines, interferon-alpha and interleukin-12, respectively, in response to Toll-like receptor stimulation in vitro. Chronic HCV infection facilitates the "customized" recruitment of liver DC subsets with established functional roles in antigen presentation. These DCs are characterized by a mature, activated phenotype and are functionally responsive to antigenic stimulation in vitro. Such findings highlight an important paradox surrounding liver DC recruitment during HCV infection, where despite their activation these cells do not provide adequate protection from the virus.

Distinct TLR adjuvants differentially stimulate systemic and local innate immune responses in nonhuman primates

AbstractTLR ligands (TLR-Ls) represent novel vaccine adjuvants, but their immunologic effects in humans remain poorly defined in vivo. In the present study, we analyzed the innate responses stimulated by different TLR-Ls in rhesus macaques. MPL (TLR4-L), R-848 (TLR7/8-L), or cytosine-phosphate-guanine oligodeoxynucleotide (TLR9-L) induced a rapid and robust expansion of blood neutrophils, with a concomitant reduction in PBMCs. Furthermore, all TLR-Ls induced rapid (3-8 hours) expansion of CD14+ monocytes, but only TLR7/8-L and TLR9-L mobilized the CD14+CD16+ and CD14dimCD16++ monocytes, and only TLR7/8-L and TLR9-L induced activation of myeloid dendritic cells (mDCs) and plasmacytoid DCs (pDCs), production of IP-10 and type-I IFN, and expression of type-I IFN–related and chemokine genes in the blood. In the draining lymph nodes (LNs), consistent with the effects in blood, all TLR-Ls induced expansion of CD14+ monocytes, but only TLR7/8-L and TLR9-L expanded the activated CD14+CD16+ cells. TLR4-L and TLR9-L differentially induced the expansion of mDCs and pDCs (1-3 days), but did not activate DCs. In contrast, TLR7/8-L did not induce DC expansion, but did activate mDCs. Finally, both TLR9-L and TLR7/8-L induced the expression of genes related to chemokines and type-I IFNs in LNs. Thus different TLR-Ls mediate distinct signatures of early innate responses both locally and systemically.

Protease-activated receptor 2 activation of myeloid dendritic cells regulates allergic airway inflammation

BackgroundA common characteristic of allergens is that they contain proteases that can activate protease-activated receptor (PAR-2); however the mechanism by which PAR-2 regulates allergic airway inflammation is unclear.MethodsMice (wild type and PAR-2-deficient) were sensitized using German cockroach (GC) feces (frass), the isolated protease from GC frass, or through adoptive transfer of GC frass-treated bone marrow-derived dendritic cells (BMDC) and measurements of airway inflammation (cellular infiltration, cytokine expression, and mucin production), serum IgE levels and airway hyperresponsiveness (AHR) were assessed. BMDC were cultured, treated with GC frass and assessed for cytokine production. PAR-2 expression on pulmonary mDCs was determined by flow cytometry.ResultsExposure to GC frass induced AHR and airway inflammation in wild type mice; however PAR-2-deficient mice had significantly attenuated responses. To directly investigate the role of the protease, we isolated the protease from GC frass and administered the endotoxin-free protease into the airways of mice in the presence of OVA. GC frass proteases were sufficient to promote the development of AHR, serum IgE, and Th2 cytokine production. PAR-2 expression on mDC was upregulated following GC frass exposure, but the presence of a functional PAR-2 did not alter antigen uptake. To determine if PAR-2 activation led to differential cytokine production, we cultured BMDC in the presence of GM-CSF and treated these cells ex vivo with GC frass. PAR-2-deficient BMDC released significantly less IL-6, IL-23 and TNFα compared to BMDC from wild type mice, suggesting PAR-2 activation was important in Th2/Th17 skewing cytokine production. To determine the role for PAR-2 on mDCs on the initiation of allergic airway inflammation, BMDCs from wild type and PAR-2-deficient mice were treated in the presence or absence of GC frass and then adoptively transferred into the airway of wild type mice. Importantly, GC frass-stimulated wild type BMDCs were sufficient to induce AHR and allergic airway inflammation, while GC frass-stimulated PAR-2-deficient BMDC had attenuated responses.ConclusionsTogether these data suggest an important role for allergen activation of PAR-2 on mDCs in mediating Th2/Th17 cytokine production and allergic airway responses.

Decreased Dengue Replication and an Increased Anti-viral Humoral Response with the use of Combined Toll-Like Receptor 3 and 7/8 Agonists in Macaques

BackgroundPathogenic versus protective outcomes to Dengue virus (DENV) infection are associated with innate immune function. This study aimed to determine the role of increased TLR3- and TLR7/8-mediated innate signaling after Dengue infection of rhesus macaques in vivo to evaluate its impact on disease and anti-DENV immune responses.Methodology/Principal FindingsTLR3 and TLR7/8 agonists (emulsified in Montanide) were administered subcutaneously to rhesus macaques at 48 hours and 7 days after DENV infection. The Frequency and activation of myeloid dendritic cells, plasmacytoid dendritic cells, and B cells were measured by flow cytometry while the serum levels of 14 different cytokines and chemokines were quantified. Adaptive immune responses were measured by DENV-specific antibody subtype measurements. Results showed that the combined TLR agonists reduced viral replication and induced the development of a proinflammatory reaction, otherwise absent in Dengue infection alone, without any clear signs of exacerbated disease. Specifically, the TLR-induced response was characterized by activation changes in mDC subsets concurrent with higher serum levels of CXCL-10 and IL-1Ra. TLR stimulation also induced higher titers of anti-DENV antibodies and acted to increase the IgG2/IgG1 ratio of anti-DENV to favor the subtype associated with DENV control. We also observed an effect of DENV-mediated suppression of mDC activation consistent with prior in vitro studies.Conclusions/SignificanceThese data show that concurrent TLR3/7/8 activation of the innate immune response after DENV infection in vivo acts to increase antiviral mechanisms via increased inflammatory and humoral responses in rhesus macaques, resulting in decreased viremia and melioration of the infection. These findings underscore an in vivo protective rather than a pathogenic role for combined TLR3/7/8-mediated activation in Dengue infection of rhesus macaques. Our study provides definitive proof-of-concept into the mechanism by which DENV evades immune recognition and activation in vivo.