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Arthritis is a disorder that is characterized by joint inflammation and other symptoms. Rheumatoid arthritis (RA), an autoimmune disease, is one of the most common arthritis in worldwide. Inflammation of the synovium is the main factor that triggers bone erosion in the joints in RA, but the pathogenesis of RA is not clearly understood. Kefir grain-fermented products have been demonstrated to enhance immune function and exhibit immune-modulating bioactivities. This study aims to explore the role of kefir peptides (KPs) on the regulation of dendritic cell, which are found in RA synovial fluid, and the protection effects of KPs on mice with collagen-induced arthritis (CIA). Immature mouse bone marrow-derived dendritic cells (BMDCs) were treated with KPs (2.2 and 4.4 mg/ml) and then exposed to lipopolysaccharide (LPS) to study the immune regulation function of KPs in dendritic cells. Mice with CIA (n = 5 per group) were orally administrated KPs (3.75 and 7.5 mg/day/kg) for 21 days and therapeutic effect of KPs on mice with arthritis were assessed. In this study, we found that KPs could inhibit surface molecule expression, reduce inflammatory cytokine release, and repress NF-κB and MAPK signaling in LPS-stimulated mouse BMDCs. In addition, a high dose of KPs (7.5 mg/kg) significantly alleviated arthritis symptoms, decreased inflammatory cytokine expression, suppressed splenic DC maturation and decrease the percentage of Th1 and Th17 in the spleens on mice with CIA. Our findings demonstrated that KPs ameliorate CIA in mice through the mechanism of suppressing DC maturation and inflammatory cytokine releases.

Extra virgin olive oil (EVOO) represents one of the most important health-promoting foods whose antioxidant and anti-inflammatory activities are mainly associated to its polyphenols content. To date, studies exploring the effect of EVOO polyphenols on dendritic cells (DCs), acting as a crosstalk between the innate and the adaptive immune response, are scanty. Therefore, we studied the ability of three EVOO extracts (cv. Coratina, Cima di Mola/Coratina, and Casaliva), characterized by different polyphenols amount, to regulate DCs maturation in resting conditions or after an inflammatory stimulus. Cima di Mola/Coratina and Casaliva extracts were demonstrated to be the most effective in modulating DCs toward an anti-inflammatory profile by reduction of TNF and IL-6 secretion and CD86 expression, along with a down-modulation of Il-1β and iNOS expression. From factorial analysis results, 9 polyphenols were tentatively established to play a synergistic role in modulating DCs inflammatory ability, thus reducing the risk of chronic inflammation.

Background: Lipopolysaccharide (LPS) produced by Gram-negative bacteria effectively stimulates the maturation of bone marrow (BM)-derived dendritic cells (BMDCs). Previous studies have shown that LPS-activated BMDCs (DClps) might induce tolerance in autoimmune diseases and cancer in vivo, whereas it remains unclear whether DClps can modulate the immune microenvironment in allergic asthma. Objectives: We sought to elucidate the potential effects of DClps on OVA-sensitized/challenged airway inflammation in a mouse model of asthma, which may help facilitate the application of specific tolerogenic dendritic cells (tolDCs) in allergic asthma patients in the future. Methods: We generated and obtained DClps from wild-type mice to evaluate their functional characteristics by ELISA and FACS. We also induced OVA-sensitized/challenged asthmatic mice and intraperitoneally treated these mice with DClps to assess the effects of these injected cells by histopathologic analysis and performing inflammatory cell counts in bronchoalveolar lavage fluid (BALF). Changes in memory CD4+ T cells, regulatory T cells (Tregs) and phosphorylated protein in lung digests were analyzed. Results: DClps exhibited lower levels of CD80 and MHC class II molecules (MHCII) and increased levels of anti-inflammatory cytokines such as IL-10 and TGF-β than immature DCs (DCia). Additionally, DClps treatment dramatically ameliorated airway inflammation and diminished the infiltration of pulmonary inflammatory cells. In addition, we prolonged the modeling time of asthmatic mice and demonstrated that DClps treatment decreased the proliferation activity of pulmonary memory CD4+ T cells, which further rendered the downregulation of type 2 T helper (Th2) cytokines. However, the number of pulmonary Tregs did not discernibly change. DClps treatment also markedly reduced the phosphorylation level of STAT6 protein. Conclusion: Our findings demonstrate that LPS stimulation may lead to a tolerogenic phenotype in BMDCs, which can induce tolerance and regulate the microenvironment of asthmatic mice, possibly through the upregulation of anti-inflammatory cytokines, inhibition of pulmonary memory CD4+ T cells and downregulation of STAT6 phosphorylation.

Silver nanoparticles (AgNP) are one of the most studied nanoparticles due to their anti-bacterial, -fungal, -viral, -parasitic, and -inflammatory properties. This raises the need to evaluate the toxicity and biological effects of AgNP in the immune system in order to develop new safer biomedical products. In this study, an AgNP formulation currently approved for veterinary applications was applied to mouse bone marrow-derived dendritic cells (BMDC), considered important antigen-presenting cells of the immune system, to evaluate cytotoxicity, genotoxicity, and any significant influence on expression of cellular markers associated with BMDC phenotype and maturation status. The results showed that after 12 h of AgNP exposure, a significant decrease in BMDC viability occurred at the highest concentration tested (1.0 µg AgNP/ml) and at lower doses, the cells maintained membrane integrity and metabolic activity. DNA damage was not significant with any AgNP level aside from the 1.0 µg AgNP/ml level. Regarding phenotype, no differences in expression of CD40 (co-stimulatory molecule highly present in mature BMDC) or in CD273 (a marker for inhibitory T-cell response) were observed. The current results showed that the toxicity of this AgNP formulation was dose-related. The findings also suggest BMDC could maintain structural conservation of co-stimulatory/co-inhibitory surface molecules after 12 h of exposure to this AgNP. This work represents the first step in identifying the toxic effects of this AgNP formulation on dendritic cells.

Leucine-rich repeat kinase 2 (LRRK2) is the causal molecule of familial Parkinson&rsquo, s disease. Although the characteristics of LRRK2 have gradually been revealed, its true physiological functions remain unknown. LRRK2 is highly expressed in immune cells such as B2 cells and macrophages, suggesting that it plays important roles in the immune system. In the present study, we investigate the roles of LRRK2 in the immune functions of dendritic cells (DCs). Bone marrow-derived DCs from both C57BL/6 wild-type (WT) and LRRK2 knockout (KO) mice were induced by culture with granulocyte/macrophage-colony stimulating factor (GM/CSF) in vitro. We observed the differentiation of DCs, the phosphorylation of the transcriptional factors NF-&kappa, B, Erk1/2, and p-38 after lipopolysaccharide (LPS) stimulation and antigen-presenting ability by flow cytometry. We also analyzed the production of inflammatory cytokines by ELISA. During the observation period, there was no difference in DC differentiation between WT and LRRK2-KO mice. After LPS stimulation, phosphorylation of NF-&kappa, B was significantly increased in DCs from the KO mice. Large amounts of inflammatory cytokines were produced by DCs from KO mice after both stimulation with LPS and infection with Leishmania. CD4+ T-cells isolated from antigen-immunized mice proliferated to a significantly greater degree upon coculture with antigen-stimulated DCs from KO mice than upon coculture with DCs from WT mice. These results suggest that LRRK2 may play important roles in signal transduction and antigen presentation by DCs.

Background: Peste des petits ruminants virus (PPRV) is the causative agent of PPR, which can cause an acute, highly contagious and fatal disease of sheep and goats, resulting in significant economic losses for commercial animal husbandry due to its high mortality and morbidity. As professional antigen-presenting cells, dendritic cells (DCs) play a unique role in innate immunity. This study aimed to gain a deeper understanding of the transcriptional response of bone marrow-derived dendritic cells (BMDCs) stimulated with PPRV. Results: Transcriptional profiling was performed using RNA sequencing. Herein, we reported that compared to untreatedBMDCs, 4492 differentially expressed genes (DEGs) were identified following PPRV stimulation, out of these DEGs 2311 were upregulated and 2181 were downregulated, respectively. A total of three gene ontology (GO) term clusters of biological process, cell component and molecular function were significantly enriched in 963 GO terms in the PPRV-stimulated BMDCs. These GO clusters were related to inflammatory response, cell division and vacuole, anchoring junction, positive regulation of cellular component and nucleoside binding. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways of DEGs were enriched in a chemokine signaling pathway, protein processing in endoplasmic reticulum, cell cycle and mTOR signaling pathway. Additionally, identified DEGs of BMDCs were further validated by qRT-PCR and the results were in accordance with the change of the genes. This study suggested the effects of PPRV stimulation on the maturation and function of BMDCs. Conclusion: We found that the dramatic BMDCs transcriptome changes triggered were predominantly related to an inflammatory response and chemokine signaling pathway.

We found that resveratrol enhances interferon (IFN)-γ-induced tryptophanyl-tRNA-synthetase (TTS) expression in bone marrow-derived dendritic cells (BMDCs). Resveratrol-induced TTS expression is associated with glycogen synthase kinase-3β (GSK-3β) activity. In addition, we found that resveratrol regulates naïve CD8+ T-cell polarization by modulating GSK-3β activity in IFN-γ-stimulated BMDCs, and that resveratol induces upregulation of TTS in CD8+ T-cells in the in vivo tumor environment. Taken together, resveratrol upregulates IFN-γ-induced TTS expression in a GSK-3β-dependent manner, and this TTS modulation is crucial for DC-mediated T-cell modulation. [BMB Reports 2015; 48(5): 283-288]

Stem cell factor (SCF), the ligand of c-kit, is a key cytokine for hematopoiesis. Hematopoietic precursors express c-kit, whereas differentiated cells of hematopoietic lineage are negative for this receptor, with the exception of NK cells, mast cells, and a few others. While it has long been recognized that dendritic cells (DCs) can express c-kit, several questions remain concerning the SCF/c-kit axis in DCs. This is particularly relevant for DCs found in those organs wherein SCF is highly expressed, including the bone marrow (BM). We characterized c-kit expression by conventional DCs (cDCs) from BM and demonstrated a higher proportion of c-kit+ cells among type 1 cDC subsets (cDC1s) than type 2 cDC subsets (cDC2s) in both humans and mice, whereas similar levels of c-kit expression were observed in cDC1s and cDC2s from mouse spleen. To further study c-kit regulation, DCs were generated with granulocyte-macrophage colony-stimulating factor (GM-CSF) from mouse BM, a widely used protocol. CD11c+ cells were purified from pooled non-adherent and slightly adherent cells collected after 7 days of culture, thus obtaining highly purified BM-derived DCs (BMdDCs). BMdDCs contained a small fraction of c-kit+ cells, and by replating them for 2 days with GM-CSF, we obtained a homogeneous population of c-kit+ CD40hi MHCIIhi cells. Not only did BMdDCs express c-kit but they also produced SCF, and both were striking upregulated if GM-CSF was omitted after replating. Furthermore, a small but significant reduction in BMdDC survival was observed upon SCF silencing. Incubation of BMdDCs with SCF did not modulate antigen presentation ability of these cells, nor it did regulate their membrane expression of the chemokine receptor CXCR4. We conclude that the SCF/c-kit-mediated prosurvival circuit may have been overlooked because of the prominent use of GM-CSF in DC cultures in vitro, including those human DC cultures destined for the clinics. We speculate that DCs more prominently rely on SCF in vivo in some microenvironments, with potential implications for graft-versus-host disease and antitumor immunity.

BACKGROUND: The E2 multimeric scaffold represents a powerful delivery system able to elicit robust humoral and cellular immune responses upon systemic administrations. Here recombinant E2 scaffold displaying the third variable loop of HIV-1 Envelope gp120 glycoprotein was administered via mucosa, and the mucosal and systemic immune responses were analysed. To gain further insights into the molecular mechanisms that orchestrate the immune response upon E2 vaccination, we analysed the transcriptome profile of dendritic cells (DCs) exposed to the E2 scaffold with the aim to define a specific gene expression signature for E2-primed immune responses. RESULTS: The in vivo immunogenicity and the potential of E2 scaffold as a mucosal vaccine candidate were investigated in BALB/c mice vaccinated via the intranasal route. Fecal and systemic antigen-specific IgA antibodies, cytokine-producing CD4(+) and CD8(+) cells were induced assessing the immunogenicity of E2 particles via intranasal administration. The cytokine analysis identified a mixed T-helper cell response, while the systemic antibody response showed a prevalence of IgG1 isotype indicative of a polarized Th2-type immune response. RNA-Sequencing analysis revealed that E2 scaffold up-regulates in DCs transcriptional regulators of the Th2-polarizing cell response, defining a type 2 DC transcriptomic signature. CONCLUSIONS: The current study provides experimental evidence to the possible application of E2 scaffold as antigen delivery system for mucosal immunization and taking advantages of genome-wide approach dissects the type of response induced by E2 particles.

Dendritic cells (DC) have been investigated as a cell-based therapy for Type 1 Diabetes (T1D). BM-DC expanded ex vivo with GM-CSF and IL-4 is typically cultured with fetal bovine serum (FBS). The effect of FBS on NOD BM-DC has not been extensively studied. In the present study we compare BM-DC generated in serum-free culture media (X-VIVO20; FBS−) with BM-DC generated in media containing 10% FBS (RPMI1640/10%FBS; FBS+). We show that FBS− BM-DC display a phenotype and cytokine-producing profile distinct from FBS+ BMDC. Additionally, compared to FBS+ BM-DC, we show evidence of an altered Th cell response induced by FBS− BM-DC. Finally, we demonstrate that only FBS− BM-DC prevent the onset of T1D and induce increased levels of CD4+Foxp3+ regulatory T cells as well as a long-lasting β cell-specific T cell response. This study indicates that serum-free media generates a more tolerogenic BM-DC capable of preventing T1D in the NOD mice.

Pseudomonas (P.) aeruginosa frequently colonizes the respiratory tract of patients with chronic respiratory tract infections such as diffuse panbronchiolitis (DPB). The number of dendritic cells (DC5) that play a central role in immune functions as antigen-presenting cells is reportedly increased in the bronchiolar tissues of patients with DPB. However, the functions of DCs in chronic P. aeruginosa respiratory tract infection have not been defined. Here, we assessed the functions of DCs and the effect of macrolide antibiotics that are therapeutic agents for DPB, in a murine model of DPB caused by P. aeruginosa. Mice were intubated with either P. aeruginosa- or saline-precoated tubes for 80 days. Thereafter, the expression of T-cell co-stimulatory molecules (CD40, CD80, and CD86) and cytokine secretion (interleukin (IL)-i 0, IL-6, IL-12p40, and tumor necrosis factor (TNF)-a) on bone marrow-derived DCs stimulated by lipopolysaccharide were examined by flow cytometry and enzyme-linked immunosorbent assays. The expression of co-stimulatory molecules was significantly decreased in mice infected with P. aeruginosa compared to the saline-treated control mice, but production of these cytokines did not significantly differ between the two groups. Pretreatment with clarithromycin ex vivo decreased CD4O expression on DCs obtained from P. aeruginosa-infected mice and also decreased the production of IL-6, IL-i 2p40 and TNF-a by DCs. These findings suggest that chronic P. aeruginosa infection alters DC functions and that macrolides function as anti-inflammatory agents by modulating the functions of DCs in chronic P. aeruginosa infection., The Tohoku Journal of Experimental Medicine, 220(1), pp.59-65; 2010

Anticancer immunotherapy is a promising treatment modality since it bears the potential of being highly specific, but effective clinical applications are still under development. We have recently described an exosome-based strategy for co-delivery of α-galactosylceramide and a tumor-associated antigen that synergistically potentiates tumor-specific adaptive immune responses while preventing the anergy of invariant natural killer T (iNKT) cells. We propose that the next generation of exosome-based immunotherapies should involve iNKT-cell ligands to induce a broad, amplified and sustainable antitumor immune response.

Background: HemoHIM, which is an herbal preparation of three edible herbs (Angelicam gigas Nakai, Cnidium offinale Makino, and Peaonia japonica Miyabe), is known to have various biological and immunological activities, but the modulatory effects of this preparation on dendritic cells (DCs)-mediated immune responses have not been examined previously. DCs are a unique group of white blood cells that initiate primary immune responses by capturing, processing, and presenting antigens to T cells. Results: In the present study, we investigated the effect of HemoHIM on the functional and phenotypic maturation of murine bone marrow-derived dendritic cells (BMDCs) both in vitro and in vivo. The expression of co-stimulatory molecules (CD40, CD80, CD86, MHC I, and MHC II) and the production of cytokines (IL-1 beta, IL-6, IL-12p70, and TNF-alpha) were increased by HemoHIM in BMDCs. Furthermore, the antigen-uptake ability of BMDCs was decreased by HemoHIM, and the antigen-presenting ability of HemoHIM-treated mature BMDCs increased TLR4-dependent CD4(+) and CD8(+) T cell responses. Conclusions: Our findings demonstrated that HemoHIM induces TLR4-mediated BMDCs functional and phenotypic maturation through in vivo and in vitro. And our study showed the antigen-presenting ability that HemoHIM-treated mature BMDCs increase CD4(+) and CD8(+) T cell responses by in vitro. These results suggest that HemoHIM has the potential to mediate DC immune responses.