Cytokine Production In Dendritic Cells Research Articles

Oral vitamin D3 supplementation reduces monocyte-derived dendritic cell maturation and cytokine production in Crohn’s disease patients

Low serum vitamin D levels may provoke or aggravate Crohn's disease (CrD). Vitamin D3 is a well-known immune modulator that affects immune functions in vitro and may prevent CrD flares. Dendritic cells (DC) are key mediators of vitamin D3 effects. In this study, we describe changes in monocyte-derived DC (mo-DC) maturation marker expression and cytokine production following 26 weeks of oral vitamin D3 supplementation in CrD patients. Ten CrD patients who had increased serum 25-hydroxy vitamin D levels after oral vitamin D3 and calcium treatment and ten seasonally matched placebo-treated patients were selected for this study. Mo-DC were generated before and after the 26 weeks and induced to mature upon lipopolysaccharide (LPS) stimulation. Maturation marker expression and cytokine production were analysed. Mo-DC function was analysed in a mixed leucocyte reaction (MLR). Compared with baseline values, LPS-matured mo-DC exhibited reduced expression of CD80 and reduced production of the cytokines IL-10, IL-1β, and IL-6 following 26 weeks of oral vitamin D3 supplementation. Mo-DC performance in an allogeneic MLR was unchanged after vitamin D3 supplementation. Treatment with the placebo did not affect maturation markers, cytokine production, or the MLR. Vitamin D3 treatment in CrD patients led to hypo-responsive LPS-stimulated mo-DC. This finding indicates that vitamin D3 levels have an impact on the monocytic precursors of mo-DC in vivo and may explain the positive effects of vitamin D3 supplementation on CrD patients. Alternatively, CrD patients with high serum vitamin D3 levels may represent a subgroup with low disease activity.

Genetic dissection of dendritic cell homeostasis and function: lessons from cell type-specific gene ablation

Dendritic cells (DCs) are a heterogeneous cell population of great importance in the immune system. The emergence of new genetic technology utilizing the CD11c promoter and Cre recombinase has facilitated the dissection of functional significance and molecular regulation of DCs in immune responses and homeostasis in vivo. For the first time, this strategy allows observation of the effects of DC-specific gene deletion on immune system function in an intact organism. In this review, we present the latest findings from studies using the Cre recombinase system for cell type-specific deletion of key molecules that mediate DC homeostasis and function. Our focus is on the molecular pathways that orchestrate DC life span, migration, antigen presentation, pattern recognition, and cytokine production and signaling.

TLR4 is essential for dendritic cell activation and anti-tumor T-cell response enhancement by DAMPs released from chemically stressed cancer cells

The combination of immunotherapy and chemotherapy is regarded as a promising approach for the treatment of certain types of cancer. However, the underlying mechanisms need to be fully investigated to guide the design of more efficient protocols for cancer chemoimmunotherapy. It is well known that danger-associated molecular patterns (DAMPs) can activate immune cells, including dendritic cells (DCs), via Toll-like receptors (TLRs); however, the role of DAMPs released from chemical drug-treated tumor cells in the activation of the immune response needs to be further elucidated. Here, we found that colorectal cancer (CRC) cells treated with oxaliplatin (OXA) and/or 5-fluorouracil (5-Fu) released high levels of high-mobility group box 1 (HMGB1) and heat shock protein 70 (HSP70). After OXA/5-Fu therapy, the sera of CRC patients also exhibited increased levels of HMGB1 and HSP70, both of which are well-known DAMPs. The supernatants of dying CRC cells treated with OXA/5-Fu promoted mouse and human DC maturation, with upregulation of HLA-DR, CD80 and CD86 expression and enhancement of IL-1β, TNF-α, MIP-1α, MIP-1β, RANTES and IP-10 production. Vaccines composed of DCs pulsed with the supernatants of chemically stressed CRC cells induced a more significant IFN-γ-producing Th1 response both in vitro and in vivo. However, the supernatants of chemically stressed CRC cells failed to induce phenotypic maturation and cytokine production in TLR4-deficient DCs, indicating an essential role of TLR4 in DAMP-induced DC maturation and activation. Furthermore, pulsing with the supernatants of chemically stressed CRC cells did not efficiently induce an IFN-γ-producing Th1 response in TLR4-deficient DCs. Collectively, these results demonstrate that DAMPs released from chemically stressed cancer cells can activate DCs via TLR4 and enhance the induction of an anti-tumor T-cell immune response, delineating a clinically relevant immuno-adjuvant pathway triggered by DAMPs.

The potential role of IL-37 in atherosclerosis.

Atherosclerosis is an inflammatory disease characterized by extensive lipid deposition and atherosclerotic plaque formation in the intima. Interleukin (IL)-37 is anti-inflammatory cytokine in the IL-1 ligand family. Given that IL-37 plays an important function in the development and progression of inflammatory and autoimmune diseases, it may be associated with the development of atherosclerosis. IL-37, which is normally expressed at low levels in peripheral blood mononuclear cells (PBMC), mainly monocytes, and dendritic cells (DC), is rapidly up-regulated in the inflammatory context, and therefore IL-37 conversely inhibits the production of inflammatory cytokines in PBMC and DC. In addition, IL-37 effectively suppresses the activation of macrophage and DC. It is not controversial that the activation of macrophage and DC and the over-expression of inflammatory cytokines are critical component elements in inflammatory process of atherosclerosis. Therefore, IL-37 may play a protective role in atherosclerosis through inhibition of inflammatory cytokines production and suppression of macrophage and DC activation.

Synergistic Effect between Amoxicillin and TLR Ligands on Dendritic Cells from Amoxicillin-Delayed Allergic Patients

Amoxicillin, a low-molecular-weight compound, is able to interact with dendritic cells inducing semi-maturation in vitro. Specific antigens and TLR ligands can synergistically interact with dendritic cells (DC), leading to complete maturation and more efficient T-cell stimulation. The aim of the study was to evaluate the synergistic effect of amoxicillin and the TLR2, 4 and 7/8 agonists (PAM, LPS and R848, respectively) in TLR expression, DC maturation and specific T-cell response in patients with delayed-type hypersensitivity (DTH) reactions to amoxicillin. Monocyte-derived DC from 15 patients with DTH to amoxicillin and 15 controls were cultured with amoxicillin in the presence or absence of TLR2, 4 and 7/8 agonists (PAM, LPS and R848, respectively). We studied TLR1-9 gene expression by RT-qPCR, and DC maturation, lymphocyte proliferation and cytokine production by flow cytometry. DC from both patients and controls expressed all TLRs except TLR9. The amoxicillin plus TLR2/4 or TLR7/8 ligands showed significant differences, mainly in patients: AX+PAM+LPS induced a decrease in TLR2 and AX+R848 in TLR2, 4, 7 and 8 mRNA levels. AX+PAM+LPS significantly increased the percentage of maturation in patients (75%) vs. controls (40%) (p=0.036) and T-cell proliferation (80.7% vs. 27.3% of cases; p=0.001). Moreover, the combinations AX+PAM+LPS and AX+R848 produced a significant increase in IL-12p70 during both DC maturation and T-cell proliferation. These results indicate that in amoxicillin-induced maculopapular exanthema, the presence of different TLR agonists could be critical for the induction of the innate and adaptive immune responses and this should be taken into account when evaluating allergic reactions to these drugs.

Ovatodiolide inhibits the maturation of allergen-induced bone marrow-derived dendritic cells and induction of Th2 cell differentiation

Ovatodiolide was a unique macrocyclic diterpenoid isolated from the traditional Chinese medicinal herb Anisomeles indica. The present study attempted to examine the ovatodiolide effects on dendritic cell (DC) maturation and immuno-stimulatory activities. The effects of ovatodiolide on DC surface molecule expression, cytokine production, and capacity to induce T-cell differentiation were examined in ovalbumin (OVA)/thymic stromal lymphopoietin (TSLP)-stimulated DCs. Ovatodiolide attenuated the expression of DC surface molecules CD80, CD86, histocompatibility complex (MHC) class II, and Th2 subset of CD4(+) T cells co-stimulatory molecule-OX40 ligand production. Additionally, ovatodiolide suppressed the CD4(+) T cells proliferation, and production of inflammatory cytokines interleukin (IL)-4, IL-5, and tumor necrosis factor (TNF)-α. This study may be useful to develop ovatodiolide as a therapeutic adjuvant.

24: STING, not type I IFN signaling, is required for the mucosal adjuvant activity of Cyclic-di-GMP in vivo

Mucosal vaccination prevents the onset of disease, blocks early colonization, and reduces the risk of horizontal transmission. Currently, there is no vaccine formulation containing a mucosal adjuvant approved for human use. Cyclic-di-GMP (CDG) is a small molecule found only in bacteria, regulating bacterial growth and virulence. CDG exhibits strong immunostimulatory properties in mammalian cells. Karaolis et al. first discovered that CDG activates human immature Dendritic cells (DC), and enhancing DC’s ability to stimulate T cells. Importantly, intranasal co-immunization of CDG and antigens protects host from pathogen infections, including H5N1 influenza, and Streptococcus pneumonia infections in mice. Thus, CDG is a promising mucosal adjuvant candidate. The mechanism by which CDG acts as a mucosal adjuvant is unknown. STING (stimulator of interferon genes) has been identified as a ligand for CDG in mammalian cells activating TBK1-IRF3-type I IFN signaling. Here, using a STING-/- mouse (Tmem173), we investigated the in vivo role of STING in the mucosal adjuvant activity of CDG. We found that STING-/- mice fail to generate antigen-specific antibody response after intranasal immunization of an antigen and CDG. Furthermore, the productions of Th1/Th2/Th17 cytokines, proinflammatory cytokines and type I IFN are all missing in CDG immunized STING-/- mice. Surprisingly, we found that type I IFN signaling is not required for the mucosal adjuvant activity of CDG because IFNAR1-/- mice have the same antigen-specific antibody response as the wild-type mice. We further found that CDG activates STING-dependent but TBK1-type I IFN signaling-independent proinflammatory cytokine production in macrophages and DCs. This is distinct from STING-mediated DNA vaccine adjuvant activity, which requires type I IFN signaling. Thus, our results establish an essential role of STING in mediating the mucosal adjuvant activity of CDG in vivo and reveal novel TBK1-type I IFN stimulation-independent function of STING.

Galectin-3 Modulates Th17 Responses by Regulating Dendritic Cell Cytokines

Galectin-3 is a β-galactoside-binding animal lectin with diverse functions, including regulation of T helper (Th) 1 and Th2 responses. Current data indicate that galectin-3 expressed in dendritic cells (DCs) may be contributory. Th17 cells have emerged as critical inducers of tissue inflammation in autoimmune disease and important mediators of host defense against fungal pathogens, although little is known about galectin-3 involvement in Th17 development. We investigated the role of galectin-3 in the induction of Th17 immunity in galectin-3-deficient (gal3(-/-)) and gal3(+/+) mouse bone marrow-derived DCs. We demonstrate that intracellular galectin-3 negatively regulates Th17 polarization in response to the dectin-1 agonist curdlan (a β-glucan present on the cell wall of fungal species) and lipopolysaccharide, agents that prime DCs for Th17 differentiation. On activation of dectin-1, gal3(-/-) DCs secreted higher levels of the Th17-axis cytokine IL-23 compared with gal3(+/+) DCs and contained higher levels of activated c-Rel, an NF-κB subunit that promotes IL-23 expression. Levels of active Raf-1, a kinase that participates in downstream inhibition of c-Rel binding to the IL23A promoter, were impaired in gal3(-/-) DCs. Modulation of Th17 by galectin-3 in DCs also occurred in vivo because adoptive transfer of gal3(-/-) DCs exposed to Candida albicans conferred higher Th17 responses and protection against fungal infection. We conclude that galectin-3 suppresses Th17 responses by regulating DC cytokine production.

The Second Messenger Phosphatidylinositol-5-Phosphate Facilitates Antiviral Innate Immune Signaling

Innate immune receptors, notably Toll-like receptors (TLRs) and RIG-I-like receptors (RLRs), sense viral infection and activate transcription factors, including interferon regulatory factor-3 (IRF3), to induce type I interferon (IFN). We demonstrate that the lipid phosphatidylinositol-5-phosphate (PtdIns5P) is increased upon viral infection and facilitates type I IFN production by binding to IRF3 and its upstream kinase TBK1 and promoting TBK1-mediated IRF3 phosphorylation and activation. Additionally, we determine that PtdIns5P is produced through the kinase PIKfyve, which phosphorylates PtdIns to generate PtdIns5P. Accordingly, PIKfyve knockdown or pharamoclogical inhibition decreases PtdIns5P levels and type I IFN production after TLR or RLR stimulation, and results in increased viral replication. A synthetic PtdIns5P, C8-PtdIns5P, promotes IRF3 phosphorylation and cytokine production in dendritic cells and acts as an adjuvant to boost immune responses in immunized mice. Thus, PtdIns5P produced during viral infection is a second messenger that targets the TBK1-IRF3 axis to elicit antiviral immunity.

Epstein-Barr Virus Encoded dUTPase Containing Exosomes Modulate Innate and Adaptive Immune Responses in Human Dendritic Cells and Peripheral Blood Mononuclear Cells

We have recently demonstrated that Epstein-Barr virus (EBV)-encoded deoxyuridine triphosphate nucleotidohydrolase (dUTPase) modulates innate immunity in human primary monocyte-derived macrophages through toll-like receptor (TLR) 2 leading to NF-κB activation and the production of pro-inflammatory cytokines. Our previous depletion studies indicated that dendritic cells (DCs) may also be a target of the EBV-encoded dUTPase. However, the role of EBV-encoded dUTPase in DC activation/function and its potential contribution to the inflammatory cellular milieu characteristic of EBV-associated diseases remains poorly understood. In the present study, we demonstrate that EBV-encoded dUTPase significantly altered the expression of genes involved in oncogenesis, inflammation and viral defense mechanisms in human primary DCs by microarray analysis. Proteome array studies revealed that EBV-encoded dUTPase modulates DC immune responses by inducing the secretion of pro-inflammatory TH1/TH17 cytokines. More importantly, we demonstrate that EBV-encoded dUTPase is secreted in exosomes from chemically induced Raji cells at sufficient levels to induce NF-κB activation and cytokine secretion in primary DCs and peripheral blood mononuclear cells (PBMCs). Interestingly, the production of pro-inflammatory cytokines in DCs and PBMCs was TLR2-dependent. Together these findings suggest that the EBV-encoded dUTPase may act as an intercellular signaling molecule capable of modulating the cellular microenvironment and thus, it may be important in the pathophysiology of EBV related diseases.

LAB/NTAL facilitates fungal/PAMP-induced IL-12 and IFN-γ production by repressing β-catenin activation in dendritic cells.

Fungal pathogens elicit cytokine responses downstream of immunoreceptor tyrosine-based activation motif (ITAM)-coupled or hemiITAM-containing receptors and TLRs. The Linker for Activation of B cells/Non-T cell Activating Linker (LAB/NTAL) encoded by Lat2, is a known regulator of ITAM-coupled receptors and TLR-associated cytokine responses. Here we demonstrate that LAB is involved in anti-fungal immunity. We show that Lat2 −/− mice are more susceptible to C. albicans infection than wild type (WT) mice. Dendritic cells (DCs) express LAB and we show that it is basally phosphorylated by the growth factor M-CSF or following engagement of Dectin-2, but not Dectin-1. Our data revealed a unique mechanism whereby LAB controls basal and fungal/pathogen-associated molecular patterns (PAMP)-induced nuclear β-catenin levels. This in turn is important for controlling fungal/PAMP-induced cytokine production in DCs. C. albicans- and LPS-induced IL-12 and IL-23 production was blunted in Lat2−/− DCs. Accordingly, Lat2−/− DCs directed reduced Th1 polarization in vitro and Lat2 −/− mice displayed reduced Natural Killer (NK) and T cell-mediated IFN-γ production in vivo/ex vivo. Thus our data define a novel link between LAB and β-catenin nuclear accumulation in DCs that facilitates IFN-γ responses during anti-fungal immunity. In addition, these findings are likely to be relevant to other infectious diseases that require IL-12 family cytokines and an IFN-γ response for pathogen clearance.

Lysosomal transporter SLC15A4 regulates Toll like receptor 7/9-mediated antibody production (P4195)

Abstract Solute Carrier family (SLC) 15A4 is known to be a proton-coupled transporter expressed by the immune and nervous systems, specifically carrying free histidine and certain di- or tri-peptides. Previous studies reported that SLC15A4 gene is associated with the autoimmune disorders such as inflammatory bowel diseases and systemic lupus erythematosus. However, the precise mechanism of how this transporter contributes to pathogenesis of these diseases remains unclear. We have investigated the functional involvement of SLC15A4 in the innate immune system, as this transporter is highly expressed in innate immune cells, and is known to reside on the lysosomal membranes. We previously reported that SLC15A4 plays crucial roles in TLR9- and NOD1-mediated cytokine production in dendritic cells (DCs). Here we show that SLC15A4 is also important for TLR7-triggered immune responses not only in DCs but also in B cells. We found that SLC15A4 is also expressed in B cells, and activation of naïve SLC15A4-deficient B cells was attenuated when triggered by TLR7 or TLR9 ligand, showing impaired IFN-beta or IL-6 production. Moreover, SLC15A4-deficient B cells failed to produce IgG2c antibody in vitro and in vivo. Although further investigation is necessary to understand the precise mechanism, our findings suggest that TLR7 or 9 activity, such as antibody production could be regulated in lysosomes by SLC15A4.

Early events in immunologic tolerance to apoptotic cells are dependent on marginal zone macrophage CCL22 production. (P4081)

Abstract Apoptotic cells (ACs) promote immunologic tolerance but the early innate mechanism(s) involved in the process are not known. Here we report that administration of ACs i/v induced rapid splenic expression of the regulatory T cell chemokine CCL22 by CD169+ marginal zone macrophages (MZMs). Similarly, in-vitro culture with ACs lead to expression of CCL22 in purified MZMs, but not CD11c+ dendritic cells (DCs) and conditioned media from MZM cocultures induced migration of Tregs in a CCL22 dependent manner. Administration of a soluble antagonist for the CCL22 receptor (i.e. CCR4) skewed the early innate immune response to ACs in-vivo shifting the balance from regulatory (IL-10, TGF-β) to inflammatory (TNF-α, IL-6, IL-12) cytokine production in splenic DCs and macrophages. Similarly, CCR4 blockade inhibited splenic accumulation of Tregs after apoptotic cell injection i/v and enhanced effector T cell responses to AC-associated antigens. Short-term CCR4 inhibition at the time of i/v apoptotic cell challenge (i.e. blockade only at the time of apoptotic cell administration) reversed AC-mediated tolerance to skin allografts in both primary recipients and in Treg adoptive transfers to secondary recipients instead promoting more rapid rejection. Suggesting a primary role for CCL22 in AC tolerance. Thus, we show for the first time that selective induction of CCL22 by MZMs is an essential early innate step in the generation of infectious immune tolerance to apoptotic self.

Tu1159 Unravelling the Mechanism of Action of Infliximab in Crohn's Disease and Healthy Controls Following In Vitro Culture With Blood-Enriched Dendritic Cells

Introduction Dendritic cells (DC) play a key role in discriminating between commensal microorganisms and harmful pathogens. DC phenotype and cytokine production determine the type of immune response elicited by T-cells following antigen presentation. DC also direct the T-cells to target tissues to perform their function via imprinting tissue-specific homing markers. In CD, dysregulation of the immune response to gut microbiota & aberrant immune cell trafficking play a role in disease pathogenesis. Infliximab (IFX) is an effective treatment for CD but its mechanism of action is unclear. In this study we investigated the in-vitro effect of IFX on phenotype & ongoing cytokine production of human blood-enriched DC from patients with active CD and healthy controls (HC) Methods Low density cells (LDC), enriched for DC, were obtained following Ficoll & Nycoprep separation of blood from patients with active CD (CDAI > 220) and HC. LDC were cultured (0.5x106cells/ml) with IFX (1.10&100μg/ml&basal) for 24hr. Activation marker (CD40, CD80, HLADR), TLR receptor (TLR2.4) and homing marker (CCR4.5.7.8.9.10, β7) expression was quantified by flow cytometry. Natural ongoing intracellular cytokine production (TNFα, TGFβ&IL-6.10.12.15) was assessed via intracellular staining and flow cytometry. Cytokine secretion was measured on cell-free culture supernatants via Multiplex. Unpaired t-test and one-way ANOVA statistical analyses were applied Results TNFα and IL-6 were increased in culture supernatants from CD although their intracellular ongoing cytokine production was decreased. LDC from CD had decreased β7 (gut-homing integrin) expression. Following IFX culture, LDC decreased β7 expression & CCR9 intensity ratio (dose-dependent). There was a trend towards reduction in TLR2 and 4 expression (not statistically significant). IL-12 production by LDC from HC was increased following IFX culture. There was a marked reduction in TNFα and IL-12 in cell supernatant following culture with IFX Conclusion Increased TNFα and IL-6 in culture supernatants from CD patients coupled with a decrease in ongoing production by DC suggests a negative cytokine feedback system. Reduced β7 expression on LDC from CD patients may suggest that DC have already been recruited to the site of mucosal inflammation. Further experiments are needed to confirm this. Reduced expression of CCR9 and elevated production of IL-12 in LDC cultured with IFX, shows reduced affinity for gut-homing and increased immunogenicity and may suggest a possible mechanism for IFX-induced paradoxical inflammation. The most dramatic effect of IFX was a reduction in TNFα in the supernatant, which is likely to represent neutralisation of the cytokine Disclosure of Interest None Declared.

Nano-Decoration of the Hemagglutinating Virus of Japan Envelope (HVJ-E) Using a Layer-by-Layer Assembly Technique

In this study, we created a nanoscale layer of hyaluronic acid (HA) on the inactivated Hemagglutinating Virus of Japan envelope (HVJ-E) via a layer-by-layer (LbL) assembly technique for CD-44 targeted delivery. HVJ-E was selected as the template virus because it has shown a tumor-suppressing ability by eliciting inflammatory cytokine production in dendritic cells. Although it has been required to increase the tumor-targeting ability and reduce nonspecific binding because HVJ-E fuses with virtually all cells and induces hemagglutination in the bloodstream, complete modifications of single-envelope-type viruses with HA have been difficult. Therefore, we studied the surface ζ potential of HVJ-E at different pH values and carefully examined the deposition conditions for the first layer using three cationic polymers: poly-L-lysine (PLL), chitosan (CH), and glycol chitosan (GC). GC-coated HVJ-E particles showed the highest disperse ability under physiological pH and salt conditions without aggregation. An HA layer was then prepared via alternating deposition of HA and GC. The successive decoration of multilayers on HVJ-E has been confirmed by dynamic light scattering (DLS), ζ potentials, and transmission electron microscopy (TEM). An enzymatic degradation assay revealed that only the outermost HA layer was selectively degraded by hyaluronidase. However, entire layers were destabilized at lower pH. Therefore, the HA/GC-coated HVJ-E describe here can be thought of as a potential bomb for cancer immunotherapy because of the ability of targeting CD44 as well as the explosion of nanodecorated HA/GC layers at endosomal pH while preventing nonspecific binding at physiological pH and salt conditions such as in the bloodstream or normal tissues.

Mammal‐Derived Respiratory Lipocalin Allergens do not Exhibit Dendritic Cell‐Activating Capacity

Most mammal-derived respiratory allergens belong to the lipocalin family of proteins. Determinants of their allergenic capacity are still unknown. Innate immune cells, in particular dendritic cells, have been shown to be involved in the allergenicity of some proteins. As recognition by dendritic cells is one of the few plausible mechanisms for the allergenicity of proteins, we wanted to investigate their role in the allergenicity of lipocalin allergens. Therefore, we first incubated human monocyte-derived dendritic cells with immunologically functional recombinant allergens mouse Mus m 1, dog Can f 1 and 2, cow Bos d 2, horse Equ c 1 and natural Bos d 2. Then, the surface marker expression and cytokine production of dendritic cells and their capacity to promote T cell proliferation and Th2 immune deviation in naïve CD4(+) T cells were examined in vitro. We found that near to endotoxin-free lipocalin allergens had no effect on the activation, allostimulatory capacity or cytokine production of dendritic cells. The dendritic cells could not induce immune deviation in naïve CD4(+) T cells. In contrast, lipopolysaccharide activated the dendritic cells efficiently. However, lipocalin allergens were not able to modify the lipopolysaccharide-induced responses. We conclude that an important group of mammal-derived respiratory allergens, lipocalins, appear not to be able to activate dendritic cells, a major component involved in the allergenicity of some proteins. It is conceivable that this incapacity of lipocalin allergens to arouse innate immunity may be associated with their poor capacity to induce a strong T cell response, verified in several studies.

Heat-inactivated Lactobacillus rhamnosus and Lactobacillus delbrueckii induce efficient maturation and differential cytokine production in human monocyte derived dendritic cells

Probiotics' strain specific immunoregulatory properties in development of protective immune outcomes are believed to be induced by targeting and polarising dendritic cells (DCs) as the key determinants of the immune response. Therefore, we studied the effects of a heat-inactivated form of Lactobacillus rhamnosus GG (LGG) and Lactobacillus delbrueckii (L.del) on maturation pattern and extracellular cytokine production profile of monocyte derived dendritic cells. Expression of specific maturation markers and induction of extracellular cytokines were detected by flow cytometry. Up-regulation of CD86, CD80 and CD83 and down-regulation of DCSIGN was observed. In addition, LGG induced secretion of high levels IL-10, INF-γ and IL-1β whereas L.del seemed to be more potent in induction of TNF-α in a dose dependent manner. These results indicated that non-viable forms of both tested strains are able to induce divergent immune regulatory effects via the induction of phenotypic changes and cytokine production of DCs.

Alginate-coated chitosan nanogel capacity to modulate the effect of TLR ligands on blood dendritic cells

Biodegradable nanoparticles have been employed for vaccine delivery, frequently admixed with adjuvants. Surprisingly, there is little information on their modulation of immune responses, speculated to be negligible. We analyzed the immunomodulatory capacity of alginate-coated chitosan nanogels (Ng), on porcine and human blood dendritic cells (DCs), when applied with defined adjuvants targeting different DC subpopulations. DC maturation, cytokine production and cell migration were assessed. Ng differentially influenced the immunomodulatory characteristics of individual Toll-like receptor (TLR) ligands: Pam3Cys-SK4-induced IL-1β was enhanced; CpG-oligodeoxynucleotides (CpG-ODN)-induced IFN-α, IL-6 and TNFα were impaired; CpG-ODN-induced CD86 and CCR7, and cell migration, were diminished—plasmacytoid DCs (pDCs) were particularly sensitive. Therein, the Ng influence on DC endocytosis of the TLR ligands was apparently a major contributory element. This demonstrates the importance of predefining the interplay between delivery vehicles and admixed immunostimulatory moieties, for ensuring appropriate immune activation and efficacious combinations. From the Clinical EditorBiodegradable nanoparticles have been utilized in vaccine delivery; however, there is little information available on their immunomodulatory properties, which are thought to be negligible. This study clearly demonstrates that nanogels do influence the developing immune response, which needs to be taken into consideration when utilizing these otherwise very efficacious vaccine delivery approaches.

Inflammatory cytokine and microRNA responses of primary human dendritic cells cultured with Helicobacter pylori strains

Primary human dendritic cells (DC) were used to explore the inflammatory effectors, including cytokines and microRNAs, regulated by Helicobacter pylori. In a 48 h ex-vivo co-culture system, both H. pylori B38 and B45 strains activated human DCs and promoted a strong inflammatory response characterized by the early production of pro-inflammatory TNFα and IL-6 cytokines, followed by IL-10, IL-1β, and IL-23 secretion. IL-23 was the only cytokine dependent on the cag pathogenicity island status of the bacterial strains. DC activation and cytokine production were accompanied by an early miR-146a upregulation followed by a strong miR-155 induction, which mainly controlled TNFα production. These results pave the way for further investigations into the nature of H. pylori antigens and the subsequently activated signaling pathways involved in the inflammatory response to H. pylori infection, the deregulation of which may likely contribute to gastric lymphomagenesis.

MiR-451 Regulates Dendritic Cell Cytokine Responses to Influenza Infection

MicroRNAs (miRNAs) are important posttranscriptional regulators in immune cells, but how viral infection regulates miRNA expression to shape dendritic cell (DC) responses has not been well characterized. We identified 20 miRNAs that were differentially expressed in primary murine DCs in response to the dsRNA agonist polyinosinic-polycytidylic acid, a subset of which were modestly regulated by influenza infection. miR-451 was unique because it was induced more strongly in primary splenic and lung DCs by live viral infection than by purified agonists of pattern recognition receptors. We determined that miR-451 regulates a subset of proinflammatory cytokine responses. Three types of primary DCs treated with antisense RNA antagomirs directed against miR-451 secreted elevated levels of IL-6, TNF, CCL5/RANTES, and CCL3/MIP1α, and these results were confirmed using miR-451(null) cells. miR-451 negatively regulates YWHAZ/14-3-3ζ protein levels in various cell types, and we measured a similar inhibition of YWHAZ levels in DCs. It is known that YWHAZ can control the activity of two negative regulators of cytokine production: FOXO3, which is an inhibitory transcription factor, and ZFP36/Tristetraprolin, which binds to AU-rich elements within 3'-untranslated regions to destabilize cytokine mRNAs. Inhibition of miR-451 expression correlated with increased YWHAZ protein expression and decreased ZFP36 expression, providing a possible mechanism for the elevated secretion of IL-6, TNF, CCL5/RANTES, and CCL3/MIP1α. miR-451 levels are themselves increased by IL-6 and type I IFN, potentially forming a regulatory loop. These data suggest that viral infection specifically induces a miRNA that directs a negative regulatory cascade to tune DC cytokine production.