Function Of Human Dendritic Cells Research Articles

Mapping human dendritic cell and macrophage populations in health and disease

Dendritic cells (DC) are a class of bone marrow derived cells arising from lympho-myeloid haematopoiesis that form an essential interface between the innate sensing of pathogens and the activation of adaptive immunity. This task requires a wide range of mechanisms and responses, which are divided between three major DC subsets: plasmacytoid DC (pDC), myeloid/conventional DC1 (cDC1) and myeloid/conventional DC2 (cDC2). Each DC subset develops under the control of a specific repertoire of transcription factors involving differential levels of IRF8 and IRF4 in collaboration with PU.1, ID2, E2-2, ZEB2, KLF4, IKAROS and BATF3. DC haematopoiesis is conserved between mammalian species and distinct from monocyte development. Although monocytes can differentiate into DC during inflammation, most tissues are dominated by resident cells of the DC lineage during the steady state. Recent studies based on an increasing level of resolution of phenotype and gene expression have identified pre-DC in human blood and heterogeneity among cDC2. These advances facilitate the integration of mouse and human immunology, support efforts to unravel human DC function in vivo and continue to present new translational opportunities to medicine.

Substrate stiffness influences phenotype and function of human antigen-presenting dendritic cells

Dendritic cells (DCs) are specialized immune cells that scan peripheral tissues for foreign material or aberrant cells and, upon recognition of such danger signals, travel to lymph nodes to activate T cells and evoke an immune response. For this, DCs travel large distances through the body, encountering a variety of microenvironments with different mechanical properties such as tissue stiffness. While immune-related pathological conditions such as fibrosis or cancer are associated with tissue stiffening, the role of tissue stiffness in regulating key functions of DCs has not been studied yet. Here, we investigated the effect of substrate stiffness on the phenotype and function of DCs by conditioning DCs on polyacrylamide substrates of 2, 12 and 50 kPa. Interestingly, we found that C-type lectin expression on immature DCs (iDCs) is regulated by substrate stiffness, resulting in differential antigen internalization. Furthermore, we show that substrate stiffness affects β2 integrin expression and podosome formation by iDCs. Finally, we demonstrate that substrate stiffness influences CD83 and CCR7 expression on mature DCs, the latter leading to altered chemokine-directed migration. Together, our results indicate that DC phenotype and function are affected by substrate stiffness, suggesting that tissue stiffness is an important determinant for modulating immune responses.

IL10 Release upon PD-1 Blockade Sustains Immunosuppression in Ovarian Cancer.

Ligation of programmed cell death-1 (PD-1) in the tumor microenvironment is known to inhibit effective adaptive antitumor immunity. Blockade of PD-1 in humans has resulted in impressive, durable regression responses in select tumor types. However, durable responses have been elusive in ovarian cancer patients. PD-1 was recently shown to be expressed on and thereby impair the functions of tumor-infiltrating murine and human myeloid dendritic cells (TIDC) in ovarian cancer. In the present work, we characterize the regulation of PD-1 expression and the effects of PD-1 blockade on TIDC. Treatment of TIDC and bone marrow-derived dendritic cells (DC) with IL10 led to increased PD-1 expression. Both groups of DCs also responded to PD-1 blockade by increasing production of IL10. Similarly, treatment of ovarian tumor-bearing mice with PD-1 blocking antibody resulted in an increase in IL10 levels in both serum and ascites. While PD-1 blockade or IL10 neutralization as monotherapies were inefficient, combination of these two led to improved survival and delayed tumor growth; this was accompanied by augmented antitumor T- and B-cell responses and decreased infiltration of immunosuppressive MDSC. Taken together, our findings implicate compensatory release of IL10 as one of the adaptive resistance mechanisms that undermine the efficacy of anti-PD-1 (or anti-PD-L1) monotherapies and prompt further studies aimed at identifying such resistance mechanisms. Cancer Res; 77(23); 6667-78. ©2017 AACR.

Manipulating human dendritic cell phenotype and function with targeted porous silicon nanoparticles

Dendritic cells (DC) are the most potent antigen-presenting cells and are fundamental for the establishment of transplant tolerance. The Dendritic Cell-Specific Intracellular adhesion molecule-3-Grabbing Non-integrin (DC-SIGN; CD209) receptor provides a target for dendritic cell therapy. Biodegradable and high-surface area porous silicon (pSi) nanoparticles displaying anti-DC-SIGN antibodies and loaded with the immunosuppressant rapamycin (Sirolimus) serve as a fit-for-purpose platform to target and modify DC. Here, we describe the fabrication of rapamycin-loaded DC-SIGN displaying pSi nanoparticles, the uptake efficiency into DC and the extent of nanoparticle-induced modulation of phenotype and function. DC-SIGN antibody displaying pSi nanoparticles favourably targeted and were phagocytosed by monocyte-derived and myeloid DC in whole human blood in a time- and dose-dependent manner. DC preconditioning with rapamycin-loaded nanoparticles, resulted in a maturation resistant phenotype and significantly suppressed allogeneic T-cell proliferation.

Monomethyl fumarate treatment impairs maturation of human myeloid dendritic cells and their ability to activate T cells

Background: Dimethyl fumarate (DMF) and its active metabolite monomethyl fumarate (MMF) effectively lead to reduction in disease relapses and active magnetic resonance imaging (MRI) lesions. DMF and MMF are known to be effective in modulating T- and B-cell responses; however, their effect on the phenotype and function of human myeloid dendritic cells (mDCs) is not fully understood. Objective: To investigate the role of MMF on human mDCs maturation and function. Methods: mDCs from healthy controls were isolated and cultured in vitro with MMF. The effect of MMF on mDC gene expression was determined by polymerase chain reaction (PCR) array after in vitro MMF treatment. The ability of mDCs to activate T cells was assessed by in vitro co-culture system. mDCs from DMF-treated multiple sclerosis (MS) patients were analyzed by flow cytometry and PCR. Results: MMF treatment induced a less mature phenotype of mDCs with reduced expression of major histocompatibility complex class II (MHC-II), co-stimulatory molecules CD86, CD40, CD83, and expression of nuclear factor κB (NF-κB) subunits RELA and RELB. mDCs from DMF-treated MS patients also showed the same immature phenotype. T cells co-cultured with MMF-treated mDCs showed reduced proliferation with decreased production of interferon gamma (IFN-γ), interleukin-17 (IL-17), and granulocyte-macrophage colony-stimulating factor (GM-CSF) compared to untreated cells. Conclusion: We report that MMF can modulate immune response by affecting human mDC function.

Butyrate and propionate inhibit antigen-specific CD8+ T cell activation by suppressing IL-12 production by antigen-presenting cells

Short chain fatty acids (SCFAs), such as acetate, butyrate and propionate, are products of microbial macronutrients fermentation that distribute systemically and are believed to modulate host immune responses. Recent data have indicated that certain SCFAs, such as butyrate and propionate, directly modulate human dendritic cell (DC) function. Given the role of DCs in initiating and shaping the adaptive immune response, we now explore how SCFAs affect the activation of antigen-specific CD8+ T cells stimulated with autologous, MART1 peptide-pulsed DC. We show that butyrate reduces the frequency of peptide-specific CD8+ T cells and, together with propionate, inhibit the activity of those cells. On the contrary, acetate does not affect them. Importantly, butyrate and propionate inhibit the production of IL-12 and IL-23 in the DCs and exogenous IL-12 fully restores the activation of the MART-1-specific CD8+ T cells, whereas IL-23 has no effect. In conclusion, these results point to a pivotal role of butyrate and propionate in modulating CD8+ T cell activation via the inhibition of IL-12 secretion from DCs. These findings reveal a novel mechanism whereby bacterial fermentation products may modulate CD8+ T cell function with possible implications in anti-cancer immunotherapy.

Development, Diversity, and Function of Dendritic Cells in Mouse and Human.

The study of murine dendritic cell (DC) development has been integral to the identification of specialized DC subsets that have unique requirements for their form and function. Advances in the field have also provided a framework for the identification of human DC counterparts, which appear to have conserved mechanisms of development and function. Multiple transcription factors are expressed in unique combinations that direct the development of classical DCs (cDCs), which include two major subsets known as cDC1s and cDC2s, and plasmacytoid DCs (pDCs). pDCs are potent producers of type I interferons and thus these cells are implicated in immune responses that depend on this cytokine. Mouse models deficient in the cDC1 lineage have revealed their importance in directing immune responses to intracellular bacteria, viruses, and cancer through the cross-presentation of cell-associated antigen. Models of transcription factor deficiency have been used to identify subsets of cDC2 that are required for T helper (Th)2 and Th17 responses to certain pathogens; however, no single factor is known to be absolutely required for the development of the complete cDC2 lineage. In this review, we will discuss the current state of knowledge of mouse and human DC development and function and highlight areas in the field that remain unresolved.

Human dendritic cells activated with MV130 induce Th1, Th17 and IL-10 responses via RIPK2 and MyD88 signalling pathways.

Recurrent respiratory tract infections (RRTIs) are the first leading cause of community‐ and nosocomial‐acquired infections. Antibiotics remain the mainstay of treatment, enhancing the potential to develop antibiotic resistances. Therefore, the development of new alternative approaches to prevent and treat RRTIs is highly demanded. Daily sublingual administration of the whole heat‐inactivated polybacterial preparation (PBP) MV130 significantly reduced the rate of respiratory infections in RRTIs patients, however, the immunological mechanisms of action remain unknown. Herein, we study the capacity of MV130 to immunomodulate the function of human dendritic cells (DCs) as a potential mechanism that contribute to the clinical benefits. We demonstrate that DCs from RRTIs patients and healthy controls display similar ex vivo immunological responses to MV130. By combining systems biology and functional immunological approaches we show that MV130 promotes the generation of Th1/Th17 responses via receptor‐interacting serine/threonine‐protein kinase‐2 (RIPK2)‐ and myeloid‐differentiation primary‐response gene‐88 (MyD88)‐mediated signalling pathways under the control of IL‐10. In vivo BALB/c mice sublingually immunized with MV130 display potent systemic Th1/Th17 and IL‐10 responses against related and unrelated antigens. We elucidate immunological mechanisms underlying the potential way of action of MV130, which might help to design alternative treatments in other clinical conditions with high risk of recurrent infections.

Genome-Wide Analyses of MicroRNA Profiling in Interleukin-27 Treated Monocyte-Derived Human Dendritic Cells Using Deep Sequencing: A Pilot Study.

MicroRNAs (miRNAs) regulate gene expression and thereby influence cell fate and function. Recent studies suggest that an abundant class of miRNAs play important roles in immune cells, such as T cells, natural killer (NK) cells, B cells, and dendritic cells (DCs). Interleukin (IL)-27 is a member of the IL-12 family of cytokines with broad anti-viral effects. It is a potent inhibitor of HIV-1 infection in CD4+ T cells and macrophages, as well as monocyte-derived immature dendritic cells (iDCs). This pilot study compared miRNA profiles between iDCs and IL-27-treated iDCs (27DCs) using deep sequencing methods and identified 46 known miRNAs that were significantly differentially expressed in 27DCs: 36 were upregulated and 10 downregulated by IL-27. Many of the potential target genes of these miRNAs are involved in IL-27 associated pathways, such as JAK/STAT, MAPKs, and PI3K and several were also previously reported to be involved in the regulation of human DC function. This study found that these miRNAs also potentially target several viral genomes and therefore may have antiviral effects. Four of these differential miRNAs (miR-99a-5p, miR-222-3p, miR-138-5p, and miR-125b-5p) were validated using quantitative reverse transcription PCR (RT-qPCR). Twenty-two novel miRNAs were discovered from deep sequencing and confirmed using RT-qPCR. This study furthers the understanding of the role of IL-27 in immunity and lays a foundation for future characterization of the role of specific miRNAs in DCs.

Immunomodulatory Drugs (IMiDs), Lenalidomide and Pomalidomide, Suppress Th1-Inducing Capacity of Dendritic Cells but Enhance Th2-Mediated Allergic Response

Background: Analogs of thalidomide, lenalidomide (LEN) and pomalidomide (POM), are novel anti-multiple myeloma (MM) drugs (so-called Immunomodulatory Drugs; IMiDs). LEN has been reported to enhance function of effector immune cells such as T cells and NK cells and to induce selective reduction of regulatory T cells. Meanwhile, LEN and POM treatments lead to relatively high rate onset of itchy skin with rash like allergic reaction. However, cellular and molecular mechanisms underlying their immunomodulatory effects still remain largely unclear. Although there is evidence indicating the immunomodulatory actions of LEN on mouse conventional dendritic cells (DCs), there are few reports showing their effects on human DC subsets. DCs are pivotal in orchestrating both innate and acquire immunity as the center of the immune regulatory system, and series of analyses have clarified a functional plasticity of DCs to induce Th1 or Th2 response. Therefore, we focused on the effects of LEN and POM on the function of human myeloid DCs (mDCs), which are the major regulators to induce Th1 or Th2 responses and play a pathogenic role in allergy by their dysregulated Th2-inducing function (Ito T. J Exp Med 2005; 202: 1213).Methods: Using cell sorting, flow cytometry, real-time PCR, and ELISA methods, function and signaling were analyzed in blood human CD11+ mDCs from healthy adult volunteers. Sera were obtained from 24 MM patients with before and after LEN therapy. Written informed consent was obtained for all patients. This study was approved by the Institutional Review Board of Kansai Medical University.Results: We found that both agents at clinical in vivo plasma concentration of 0.1 µM to 1 µM did not affect the mDC survival and their CD86 and OX40-ligand expression in response to toll-like receptor (TLR)-ligands (LPS, poly IC, or R848) and Th2-inducing cytokine, thymic stromal lymphopoietin (TSLP) for 24 h culture. Either LEN or POM inhibited dose-dependently mDC-derived Th1-associated cytokines, IL-12 in response to R848 (1 µM LEN; 5.2 % of cont. and 0.3 µM POM; 12 % of cont.), IFN-λ in response to poly IC (1 µM LEN; 5.4 % of cont. and 0.3 µM POM; 10 % of cont.), and TNFα in response to R848 (1 µM LEN; 36 % of cont. and 0.3 µM POM; 17 % of cont.), but enhanced IL-10 production in response to R848 (1 µM LEN; 4.8-fold increase and 0.3 µM POM; 7.5-fold increase). When mDCs were stimulated with TSLP, both agents significantly enhanced the production of Th2-recruiting chemokine CCL17/TARC (1 µM LEN; 2.4 fold increase and 0.3 µM POM; 3.9 fold increase), which functions as chemoattractant for memory Th2 cells and contribute to aggravation of allergic inflammation. These capacities of POM were stronger than those of LEN. In addition, we revealed that, in response to LPS or R848, both LEN and POM downregulated not only IRF4 mRNA in human mDCs as in LEN-treated myeloma cells, but also STAT4 mRNA, which is important for DC-mediated Th1 response and IL-12 production. In mDCs treated with TSLP, both agents were found to upregulate STAT6 mRNA, which is responsible for TSLP-mediated CCL17 secretion (Arima K. Sci Signal2010;3:ra4). Clinically, serum CCL17/TARC levels are associated with the disease activity of atopic dermatitis and sensitively reflect short-term changes in skin conditions. We also found, in MM patients, serum CCL17 levels at the onset of LEN-associated rash during 4 weeks (n=9; 1271 ± 376 pg/ml) were significantly higher (p <0.001) compared to those without rash in LEN treatment (n=15; 476 ± 236 pg/ml) and those before treatment (296 ± 172 pg/ml).Conclusion: Our data suggest that the IMiDs, while suppress the Th1-inducing capacity of DCs, rather lead to enhance allergic response at DC phase through modulation of signaling pathways in this stream of action. Thus, IMiDs could have the potential to shift DC-mediated response from Th1 to Th2, and our findings provide a plausible explanation for pathogenesis of IMiD-associated rash in the treatment for MM. DisclosuresIto:Celgene corporation: Honoraria; GSK: Patents & Royalties.

Kluyveromyces marxianus and Saccharomyces boulardii Induce Distinct Levels of Dendritic Cell Cytokine Secretion and Significantly Different T Cell Responses In Vitro.

Interactions between members of the intestinal microbiota and the mucosal immune system can significantly impact human health, and in this context, fungi and food-related yeasts are known to influence intestinal inflammation through direct interactions with specialized immune cells in vivo. The aim of the present study was to characterize the immune modulating properties of the food-related yeast Kluyveromyces marxianus in terms of adaptive immune responses indicating inflammation versus tolerance and to explore the mechanisms behind the observed responses. Benchmarking against a Saccharomyces boulardii strain with probiotic effects documented in clinical trials, we evaluated the ability of K. marxianus to modulate human dendritic cell (DC) function in vitro. Further, we assessed yeast induced DC modulation of naive T cells toward effector responses dominated by secretion of IFNγ and IL-17 versus induction of a Treg response characterized by robust IL-10 secretion. In addition, we blocked relevant DC surface receptors and investigated the stimulating properties of β-glucan containing yeast cell wall extracts. K. marxianus and S. boulardii induced distinct levels of DC cytokine secretion, primarily driven by Dectin-1 recognition of β-glucan components in their cell walls. Upon co-incubation of yeast exposed DCs and naive T cells, S. boulardii induced a potent IFNγ response indicating TH1 mobilization. In contrast, K. marxianus induced a response dominated by Foxp3+ Treg cells, a characteristic that may benefit human health in conditions characterized by excessive inflammation and positions K. marxianus as a strong candidate for further development as a novel yeast probiotic.

The Safety Property of β-D-Mannuronic Acid (M2000) as a Novel Immunosuppressive Agent on Differentiation, Maturation and Function of Human Dendritic Cells.

The effect of M2000 on differentiation, maturation and function of dendritic cells was examined. To investigate how M2000 affects human dendritic cells (DC) in a defined inflammatory environment, human peripheral blood mononuclear cells (PBMC) were isolated from healthy blood and monocytes were purified using anti-CD14 microbeads. Monocytes were incubated with M2000 in two different doses (6 and 12 J.g/well) along with adding the granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 for inducing monocytes to immature DC and lipopolysaccharide for running DC maturation. The differentiation, maturation and function of dendritic cells were examined with flow cytometry and ELISA method. The results demonstrate that M2000 has no significant side on differentiation, maturation and function of dendritic cells in immature DC and mature DC process in vitro. Our findings show that β-D-mannuronic acid (m2000) as a safe agent had no adverse effect on differentiation, maturation and function of dendritic cells which might be recommended as a novel immunosuppressive agent with no or fewer side effects in increasing the risk of infectious diseases and cancers.

Propolis modulates miRNAs involved in TLR-4 pathway, NF-κB activation, cytokine production and in the bactericidal activity of human dendritic cells.

Dendritic cells (DCs) are antigen-presenting cells, essential for recognition and presentation of pathogens to T cells. Propolis, a resinous material produced by bees from various plants, exhibits numerous biological properties, highlighting its immunomodulatory action. Here, we assayed the effects of propolis on the maturation and function of human DCs. DCs were generated from human monocytes and incubated with propolis and LPS. NF-κB and cytokines production were determined by ELISA. microRNA's expression was analysed by RT-qPCR and cell markers detection by flow cytometry. Colony-forming units were obtained to assess the bactericidal activity of propolis-treated DCs. Propolis activated DCs in the presence of LPS, inducing NF-kB, TNF-α, IL-6 and IL-10 production. The inhibition of hsa-miR-148a and hsa-miR-148b abolished the inhibitory effects on HLA-DR and pro-inflammatory cytokines. The increased expression of hsa-miR-155 may be correlated to the increase in TLR-4 and CD86 expression, maintaining LPS-induced expression of HLA-DR and CD40. Such parameters may be involved in the increased bactericidal activity of DCs against Streptococcus mutans. Propolis modulated the maturation and function of DCs and may be useful in the initial steps of the immune response, providing a novel approach to the development of DC-based strategies and for the discovery of new immunomodulators.

Azithromycin modulates immune response of human monocyte-derived dendritic cells and CD4+ T cells

Azithromycin (AZM) is a macrolide antibiotic that exhibits anti-inflammatory activity aside from its antimicrobial effect, a feature that may ameliorate certain inflammatory disorders and prevent graft-versus-host disease in patients receiving stem cell transplantation. In the present study, we investigated the ability of AZM to influence the function of human monocyte-derived dendritic cells (DCs) and CD4+ T cells. We found that AZM down-regulated CD80, CD86, and HLA-DR expression in lipopolysaccharide (LPS)-stimulated DCs and suppressed interleukin (IL)-6, IL-10, IL-12, and tumor necrosis factor-alpha production in these cells. In addition, AZM increased endocytosis and/or expression of Toll-like receptor (TLR)2, TLR4, and TLR9 in DCs and suppressed anti-CD3/CD28–induced CD4+ T cell proliferation and interferon-gamma production, an effect that was synergistic with dexamethasone. Finally, AZM suppressed DC-induced allogeneic T cell proliferation and cytokine production. Our study demonstrates that AZM modulates DC and CD4+ T cell function and may be of therapeutic benefit in various inflammatory disorders.

Pleurotus citrinopileatus polysaccharide induces activation of human dendritic cells through multiple pathways

Many edible mushrooms have become attractive as “health foods” and as source materials for immunomodulators. To increase our insight in the immune-modulatory properties of a polysaccharide of the oyster mushroom Pleurotus citrinopileatus, PCPS, we analyzed its effects on the function of human dendritic cells (DCs). We showed that PCPS induces upregulation of the surface maturation markers CD80, CD86 and HLA-DR on DCs, indicating its potential to induce DC maturation. In addition, PCPS stimulates DCs to secrete the pro-inflammatory cytokines TNF, IL-1β, IL-6 and IL-12, as well as the anti-inflammatory cytokine IL-10, and induces enhanced mRNA levels of the chemokines CCL2, CCL3, CCL8, CXCL9, CXCL10, and LTA. The secretion of TNF and IL-12 by PCPS-activated DCs could significantly be decreased by an anti-Dectin-1 antibody, as well as by a Syk kinase and a Raf-1 inhibitor, indicating that PCPS induces Dectin-1 signaling at least partly through the Syk- and the Raf-1-dependent pathways in DCs. Structural analysis of PCPS suggests that this polysaccharide is a β-1,3-branched β-1,6-glucan, which is in line with its capacity to activate Dectin-1. We showed that PCPS can induce TLR2 and TLR4, but not TLR3, signaling using TLR-HEK293 reporter cell lines. In human DCs, the effect of PCPS was additively increased by TLR4 activation, and synergistically enhanced by stimulation of TLR2, suggesting that interaction of PCPS with these TLRs contributes to the observed DC modulation. In conclusion, PCPS has the capacity to activate human DCs via multiple pathways.

Synergistic production of TNFα and IFNα by human pDCs incubated with IFNλ3 and IL-3

In this study, we investigated whether IFNλ3 and IL-3 reciprocally influence their capacity to activate various functions of human plasmacytoid dendritic cells (pDCs). In fact, we preliminarily observed that IFNλ3 upregulates the expression of the IL-3Rα (CD123), while IL-3 augments the expression of IFNλR1 in pDCs. As a result, we found that combination of IFNλ3 and IL-3 induces a strong potentiation in the production of TNFα, IFNα, as well as in the expression of Interferon-Stimulated Gene (ISG) mRNAs by pDCs, as compared to either IFNλ3 or IL-3 alone. In such regard, we found that endogenous IFNα autocrinally promotes the expression of ISG mRNAs in IL-3-, but not in IFNλ3 plus IL-3-, treated pDCs. Moreover, we uncovered that the production of IFNα by IFNλ3 plus IL-3-treated pDCs is mostly dependent on endogenously produced TNFα. Altogether, our data demonstrate that IFNλ3 and IL-3 collaborate to promote, at maximal levels, discrete functional responses of human pDCs.

How genetically modified probiotic bacteria can modify human dendritic cell functions?

Abstract Selected lactic-acid bacterial strains such as Lactobacilli are classified as probiotics with beneficial effects on the human host. It has previously been shown that the group of Lactobacillus rhamnosus/casei/paracasei are able to prevent cytokine-induced apoptosis of intestinal epithelial cells by secreted factors identified as peptidoglycan-hydrolases. To exert their protective and immunomodulatory effects, these bacteria have to interact with the underlying epithelial and immune cells, such as dendritic cells (DCs). DCs recognize Lactobacilli and their components by pattern recognition receptors (PRR) that may induce stimulatory or inhibitory T-lymphocyte responses. Our results showed that the inflammatory response of human monocyte-derived DCs (moDCs) to bacterial stimuli and the ability to polarize T-lymphocytes to different directions was highly dependent on the intact structure of the bacterial cell wall, while the expression of the cell surface molecules of moDCs was independent on the modification of the peptidoglycan (PG) layer. It was also found that the secreted bacterial cell wall preparations were able to activate moDCs demonstrated by the increased levels of pro – and anti-inflammatory cytokines and elevated expression of cell surface markers independently on the genetic manipulation of the cell wall. Based on these results we conclude that the genetic modification of the bacterial cell wall is able to modulate the stimulatory potential of moDCs and has an impact on the outcome of T-lymphocyte responses at the level of the whole bacteria. In contrast to the effects of the specific PG extracts, the functions of moDCs are independent on the genetically modified bacteria or their cell wall and its components.

Dexmedetomidine Inhibits Maturation and Function of Human Cord Blood-Derived Dendritic Cells by Interfering with Synthesis and Secretion of IL-12 and IL-23.

AimsTo investigate the effects and underlying mechanism of dexmedetomidine on the cultured human dendritic cells (DCs).MethodsHuman DCs and cytotoxic T lymphocytes (CTLs) were obtained from human cord blood mononuclear cells by density gradient centrifugation. Cultured DCs were divided into three groups: dexmedetomidine group, dexmedetomidine plus yohimbine (dexmedetomidine inhibitor) group and control group. DCs in the three groups were treated with dexmedetomidine, dexmedetomidine plus yohimbine and culture medium, respectively. After washing, the DCs were co-incubated with cultured CTLs. The maturation degree of DCs was evaluated by detecting (1) the ratios of HLA-DR-, CD86-, and CD80-positive cells (flow cytometry), and (2) expression of IL-12 and IL-23 (PCR and Elisa). The function of DCs was evaluated by detecting the proliferation (MTS assay) and cytotoxicity activity (the Elisa of IFN-γ) of CTLs. In addition, in order to explore the mechanisms of dexmedetomidine modulating DCs, α2-adrenergic receptor and its downstream signals in DCs were also detected.ResultsThe ratios of HLA-DR-, CD86-, and CD80-positive cells to total cells were similar among the three groups (P>0.05). Compared to the control group, the protein levels of IL-12 and IL-23 in the culture medium and the mRNA levels of IL-12 p35, IL-12 p40 and IL-23 p19 in the DCs all decreased in dexmedetomidine group (P<0.05). In addition, the proliferation of CTLs and the secretion of IFN-γ also decreased in the dexmedetomidine group, compared with the control group (P<0.05). Moreover, these changes induced by dexmedetomidine in the dexmedetomidine group were reversed by α2-adrenergic receptor inhibitor yohimbine in the dexmedetomidine plus yohimbine group. It was also found the decrease of mRNA levels of IL-12 p35, IL-12 p40 and IL-23 p19 in the dexmedetomidine group could be reversed by ERK1/2 or AKT inhibitors.ConclusionDexmedetomidine could negatively modulate human immunity by inhibiting the maturation of DCs and then decreasing the proliferation and cytotoxicity activity of CTLs. The α2-adrenergic receptors and its downstream molecules ERK1/2 and AKT are closely involved in the modulation of dexmedetomidine on DCs.

Hypoxia attenuates anti‐Aspergillus fumigatus immune responses initiated by human dendritic cells

Aspergillus fumigatus is an opportunistic mould that causes invasive pulmonary aspergillosis (IPA), a life-threatening infection in immunocompromised patients. During the course of IPA, localised areas of tissue hypoxia occur. Bacterial infection models revealed that hypoxic microenvironments modulate the function of host immune cells. However, the influence of hypoxia on anti-fungal immunity has been largely unknown. We evaluated the impact of hypoxia on the human anti-A.fumigatus immune response. Human monocyte-derived dendritic cells (DCs) were stimulated invitro with germ tubes of A.fumigatus under normoxia or hypoxia (1% O2 ), followed by analysis of DC viability, maturation and cytokine release. While DC viability was unaffected, hypoxia attenuated cytokine release from DCs and maturation of DCs upon stimulation with A.fumigatus. These data suggest that hypoxia at the site of A.fumigatus infection inhibits full activation and function of human DCs. Thereby, this study identified hypoxia as a crucial immune-modulating factor in the human anti-fungal immune response that might influence the course and outcome of IPA in immunocompromised patients.

In vitro impact of bisphenols BPA, BPF, BPAF and 17β-estradiol (E2) on human monocyte-derived dendritic cell generation, maturation and function

Bisphenols (BPs) are widely spread pollutants that act as estrogen-like endocrine disruptors and are potentially affecting human health on a long run. We explored the effects of BPA, BPF and BPAF, on in vitro differentiation and maturation of MDDCs. Monocytes were treated with 17β-estradiol (E2) and each BP at the beginning of their differentiation into iMDDCs. We found that 10 and 50μM of BPA and BPF, 10 and 30μM of BPAF and 10 and 50nM of E2 did not affect cell viability. However, 50μM of BPA and BPF, as well as 10 and 30μM of BPAF, significantly decreased the endocytotic capacity of iMDDCs. Both, BPA (50μM) and BPAF (30μM) decreased the expression of CD1a and increased the amount of DC-SIGN molecules on iMDDCs. The E2 pre-treatment moderately decreased expression of CD80, CD86 and CD83 co-stimulatory molecules while increasing the numbers of HLA-DR on mMDDCs. Only BPAF significantly influenced the expression of CD80 and CD86 (both decreased), as well as CD83 and HLA-DR molecules (both increased) on mMDDCs. In addition, BPAF modulated DC maturation signaling pathways by lowering the phosphorylation of p65 NF-κB (nuclear factor-kappaB) and ERK (extracellular signal regulated kinase) 1/2 proteins. Consequently, the in vitro proliferation of allogeneic T cells, stimulated with differently pre-treated iMDDCs and mMDDCs, was significantly reduced only in case of BPAF.