Murine Bone Marrow-derived Dendritic Cells Research Articles

Phosphoproteomics of cAMP signaling of Bordetella adenylate cyclase toxin in mouse dendritic cells

The adenylate cyclase toxin (CyaA) of the whooping cough agent Bordetella pertussis subverts immune functions of host myeloid cells expressing the αMβ2 integrin (CD11b/CD18, CR3 or Mac-1). CyaA delivers into cytosol of cells an extremely catalytically active adenylyl cyclase enzyme, which disrupts the innate and adaptive immune functions of phagocytes through unregulated production of the key signaling molecule cAMP. We have used phosphoproteomics to analyze cAMP signaling of CyaA in murine bone marrow-derived dendritic cells. CyaA action resulted in alterations of phosphorylation state of a number of proteins that regulate actin cytoskeleton homeostasis, including Mena, Talin-1 and VASP. CyaA action repressed mTOR signaling through activation of mTORC1 inhibitors TSC2 and PRAS40 and altered phosphorylation of multiple chromatin remodelers, including the class II histone deacetylase HDAC5. CyaA toxin action further elicited inhibitory phosphorylation of SIK family kinases involved in modulation of immune response and provoked dephosphorylation of the transcriptional coactivator CRTC3, indicating that CyaA-promoted nuclear translocation of CRTC3 may account for CyaA-induced IL-10 production. These findings document the complexity of subversive physiological manipulation of myeloid phagocytes by the CyaA toxin, serving in immune evasion of the pertussis agent.

GM-CSF Monocyte-Derived Cells and Langerhans Cells As Part of the Dendritic Cell Family.

Dendritic cells (DCs) and macrophages (Mph) share many characteristics as components of the innate immune system. The criteria to classify the multitude of subsets within the mononuclear phagocyte system are currently phenotype, ontogeny, transcription patterns, epigenetic adaptations, and function. More recently, ontogenetic, transcriptional, and proteomic research approaches uncovered major developmental differences between Flt3L-dependent conventional DCs as compared with Mphs and monocyte-derived DCs (MoDCs), the latter mainly generated in vitro from murine bone marrow-derived DCs (BM-DCs) or human CD14+ peripheral blood monocytes. Conversely, in vitro GM-CSF-dependent monocyte-derived Mphs largely resemble MoDCs whereas tissue-resident Mphs show a common embryonic origin from yolk sac and fetal liver with Langerhans cells (LCs). The novel ontogenetic findings opened discussions on the terminology of DCs versus Mphs. Here, we bring forward arguments to facilitate definitions of BM-DCs, MoDCs, and LCs. We propose a group model of terminology for all DC subsets that attempts to encompass both ontogeny and function.

Constitutive interaction between 4-1BB and 4-1BBL on murine LPS-activated bone marrow dendritic cells masks detection of 4-1BBL by TKS-1 but not 19H3 antibody

4-1BB is a TNFR family member associated with NF-κB mediated survival signaling. 4-1BB is widely expressed on activated cells of the immune system, including activated T cells, NK cells and dendritic cells. Its ligand, 4-1BBL, is transiently expressed on activated antigen presenting cells and at low levels on activated T cells. Although 4-1BBL-deficient mice clearly demonstrate a role for 4-1BBL in CD8 T cell responses to viruses such as influenza, 4-1BBL can be difficult to detect following infection of mice. Here we provide evidence for a constitutive interaction between endogenous 4-1BB and 4-1BBL on LPS activated bone marrow-derived murine dendritic cells that can mask its detection, with implications for measurement of 4-1BBL expression. The masking of 4-1BBL by its receptor results in loss of reactivity to the anti-4-1BBL antibody TKS-1, whereas the 19H3 antibody binds to 4-1BBL in the presence or absence of 4-1BB. Moreover, 4-1BB/4-1BBL interaction can occur in trans between 4-1BB+/+ and 4-1BB−/− dendritic cells in culture. These data suggest that 19H3 is the preferable antibody to use to detect 4-1BBL in the presence of its receptor.

Dengue virus type 2 infection inhibits the expression of toll-like receptors 4 and 7 in murine bone marrow-derived dendritic cells

Objective To analyze the changes in the expression of Toll-like receptors 4 (TLR4) and 7 on the surface of murine bone marrow-derived dendritic cells following dengue virus type 2 (DEN2) infection. Methods DEN2 NGC strain was infected into BALB/c suckling mice through intracranial injection and injected into C6/36 cells to induce the in vivo and in vitro proliferation of DEN2, respectively. RT-PCR was performed to identify DEN2 RNA. Reed-Muench method was used to determine the 50% tissue culture infective dose (TCID50) of DEN2. Dendritic cells (DCs) were prepared by stimulating bone marrow cells isolated form C57BL/6 mice with IL-4 and GM-CSF and then identified by flow cytometry. The prepared murine bone marrow-derived DCs were infected with DEN2 and observed with direct immunofluorescence assay. Dynamic changes in the expression of CD11c, CD86 and I-A/I-E molecules on DCs after DEN2 infection were detected by flow cytometry. Levels of DEN2 RNA and the expression of TLR4 and TLR7 at mRNA level were dynamically detected by real-time quantitative PCR. Results The TCID50 of DEN2 to C6/36 cells was 10-5.8. Murine bone marrow-derived DCs were acquired with a purity of 70% and could be infected with DEN2 in vitro. The percentages of CD86 and I-A/I-E molecules on the surface of DCs infected with 1×105 TCID50 of DEN2 were statistically different from those of the negative control group. Neither of the two groups showed a significant difference in the percentages of membrane molecules over time. However, the percentages of membrane molecules on DCs increased with increasing viral load. Compared with the negative control group, the levels of DEN2 RNA in infection groups were increased with increasing virus load, while the expression of TLR4 and TLR7 on DEN2 infected-DCs at mRNA level was decreased with increasing viral load. Conclusion DEN2 infection promotes the maturation of DCs. Expression of TLR4 and TLR7 on DEN2 infected-DCs at mRNA level decreases with increasing viral load, which suggests that TLR4 and TLR7 are closely related to viral infection and play a certain role in the pathogenesis of DEN infection. Key words: Dengue virus type 2; DC; TLR4; TLR7

Murine bone marrow-derived DCs activated by porcine rotavirus stimulate the Th1 subtype response invitro.

Rotavirus (RV) infection causes acute, watery dehydrating diarrhea and even death in infants and other young animals, resulting in a severe economic burden; however, little is known about the innate immune mechanisms associated with RV infection. Dendritic cells (DCs), which are professional antigen-presenting cells (APCs), serve as a bridge connecting the innate and adaptive immune system. In this study, the interaction between murine bone marrow-derived DCs (BMDCs) and porcine rotavirus (PRV) was investigated invitro. Upon stimulation, the expression levels of MHC-II, CD40, CD80, CD86 and CD83 in BMDCs increased in a time-dependent manner, indicating activation and maturation by PRV. In addition, up-regulated Toll-like receptor 2 (TLR2), TLR3 and NF-κB increased the production of interleukin-12 and interferon-γ. The PRV-stimulated BMDCs also showed increased stimulatory capacity in mixed lymphocyte reactions and promoted the Th1 subtype response.

Synergistic combination of murine bone marrow-derived dendritic cells loaded ex vivo with whole tumor lysate and systemic chemotherapy mediates antitumor immune responses in vivo

In order to get mature dendritic cells (DC) that is a crucial prerequisite for success in tumor immunotherapy protocols. Herein, we assumed that administration of murine bone marrow (BM)-derived DC (BM-DC), loaded ex vivo with whole Ehrlich ascites carcinoma (EAC) lysate, in the context of systemic chemotherapy cyclophosphamide (CTX) to induce antitumor immune responses, may be a good strategy to improve the presentation of tumor-specific antigens to the immune system. In the first series of experiments, BM cells generated either from BM of naïve mice or from BM of EAC-bearing mice were cultured in the presence of GM-CSF and IL-4 for 6days. At day 7, cells were loaded for 48h with one of the following maturation agents: EAC lysate (1mg/ml), poly-inosinic: polycytidylic acid [poly(I:C)] (25μg/ml) or mRNA encoding human telomerase reverse transcriptase (hTERT-mRNA) (2μg/ml). In the second series of experiments, EAC-bearing mice were intraperitoneally (i.p.) injected with CTX followed by i.p. vaccination with DC, loaded ex vivo with EAC lysate. DC yield and the phenotypic expression of maturity-related surface markers of DC (i.e. CD11b and CD11c) in both series of experiments were investigated. As a result, a significant decrease in the number of DC generated from poly(I:C)-supplemented BM culture from EAC-bearing mice has been detected. Loading of BM cells with poly(I:C), EAC lysate or hTERT-mRNA could induce the expression of CD11b and CD11c. Additionally, vaccination of EAC-bearing mice with DC loaded ex vivo with EAC lysate following CTX treatment, resulted in increases in the percentage of multiple populations of CD11b+CD11c+ in BM, spleen and peripheral blood (PB). To conclude, further researches to clarify the mechanism involved in DC maturation are crucial not only to comprehend DC biology but also to optimize DC immunotherapy protocols.

Dietary, nondigestible oligosaccharides and Bifidobacterium breve M-16V suppress allergic inflammation in intestine via targeting dendritic cell maturation.

Dietary intervention with short-chain galacto-oligosaccharides (scGOS), long-chain fructo-oligosaccharides (lcFOS) and Bifidobacterium breve M-16V (Bb) (GF/Bb) suppresses food allergic symptoms in mice, potentially via intestinal epithelial cell (IEC)-derived galectin-9. Furthermore, in vitro studies showed galacto- and fructo-oligosaccharides (GF) to enhance the immunomodulatory capacity of a TLR9 ligand representing bacterial CpG DNA when exposed to IEC. In this study, we investigated whether GF/Bb modulates dendritic cells (DCs) and subsequent Th2 and regulatory T cell (Treg) frequency in the small intestinal lamina propria (SI-LP). BALB/c mice were fed GF/Bb during oral OVA sensitization. DC and T cell phenotype were determined in SI-LP mononuclear cells using flow cytometry. Murine bone marrow-derived DCs (BMDCs) were exposed to recombinant galectin-9 or human monocyte-derived DCs (moDCs) and were cultured in IEC-conditioned medium from GF and TLR9 ligand-exposed HT-29 cells. GF/Bb reduced allergic symptoms and enhanced serum galectin-9 levels, while suppressing activation, restoring phagocytic capacity, and normalizing CD103 expression of SI-LP DCs of OVA-allergic mice. In vitro, galectin-9 suppressed LPS-induced activation markers and cytokine secretion by BMDCs, and IEC-conditioned medium suppressed moDC activation in a galectin-9-dependent manner. Besides suppression of SI-LP DC activation, dietary GF/Bb also lowered the frequency of activated Th2 cells, while enhancing Treg in the SI-LP of OVA-allergic mice compared to the control diet. Dietary intervention with GF/Bb enhances galectin-9 and suppresses allergic symptoms of OVA-allergic mice in association with reduced intestinal DC and Th2 activation and increased Treg frequency in these mice.

Combination of Peptide-based Vaccine and Chinese Herbal Medicine as a Strategy to Enhance Immune responses in Virus-associated Cancer

Abstract Virus infections may account for the development of several cancers, such as HPV16/18 are known to cause around 70% of cervical cancer cases. Until now, several immunotherapeutic approaches for virus-induced cancer are under development. Peptide-based vaccines have several advantages over conventional whole-protein vaccines in terms of purity, lot-to-lot consistency, production costs, and the high antigenic specificity. However, the use of peptide antigens in vaccine development has been hampered by problems, such as weak immunogenicity coupled with a paucity of potent adjuvants. Specific T cell immune response is crucial for anti-tumor immunity. Manipulating specific T cell immunity by Chinese herbal medicine (CHM) is a promising field to explore. Therefore, we aimed to investigate whether CHM can modulate specific T cell responses to apply for the development of cancer vaccine. We have identified a CHM extract that could increase expression of maturation cytokines and activation markers of murine bone marrow-derived dendritic cells. Furthermore, in a murine TC-1 tumor-bearing model, we found that the CHM extract could act as an adjuvant to induce cellular immune responses and anti-tumor effect in peptide vaccine strategy. We suggested that immune-stimulator CHM combined with cancer vaccine endows them with increased immunologic activity, which may be used to bypass the requirement for the conditional adjuvant. Further delineation of the mechanism may provide new clues for vaccination strategy.

Molecular chaperone is required for gut tolerogenic dendritic cell development and function

Abstract The intestinal mucosa is continuously inundated with foreign antigens. Therefore, immune cells need to be tightly regulated to maintain tolerance against harmless dietary antigens and microbiota while responding to pathogens. Intestinal dendritic cells (DCs) play an integral role in regulating immunity and tolerance because of their unique position at the interface of innate and adaptive immunity. Multiple populations of intestinal DCs have been reported based on the expression of CD103, CD11b, and CX3CR1. However, the mechanism by which different DC subsets regulate intestinal homeostasis is still unclear. Heat shock protein gp96 is an essential chaperone for most TLRs, integrins, and other proteins important for DC functions. However, the role of DC-intrinsic gp96 in regulating gut tolerance has not been studied. By a genetic strategy, we discovered that selective deletion of gp96 from CD11c+ cells in mice results in alteration of DC and T cell subsets in the gut as well as the loss of antigen-specific regulatory T cell (Treg) induction. Strikingly DC-specific gp96 knockout mice develop spontaneous colitis by 24 weeks of age. Furthermore, differentiation of murine bone marrow-derived DCs revealed that gp96 depletion selectively decreased cell maturation in response to FLT3L compared to CSF2. Also, gp96 depletion reduced DC migration and cross-presentation. Taken together, our results demonstrated that gp96 depletion from CD11c+ cells selectively inhibits tolerogenic DCs in the gut leading to decreased Treg cells, which indicates that DC-intrinsic gp96 plays essential roles in maintaining gut tolerance.

The immunosuppressive characteristics of FB1 by inhibition of maturation and function of BMDCs

Fumonisin B1 (FB1) is one kind of mycotoxins that has the neurotoxicity, carcinogenicity, hepatotoxicity and immunotoxicity produced by the fungus Fusarium verticillioides, which commonly infects corn and other crops and is harmful to animal and human health upon consumption of FB1-contaminated feed or food. However, the mechanism of immunotoxicity, especially the immunosuppression induced by FB1 is still unclear. The most pivotal cells in the induction of immune responses are dendritic cells (DCs). In this study, we used murine bone marrow-derived dendritic cells (BMDCs) as a model system to elucidate the effect of FB1 on the function of BMDCs through biological methods. We found that FB1 reversed the morphological changes and enhanced the endocytosis of FITC-dextran in LPS-treated BMDCs. At the same time, FB1 decreased the LPS-induced expressions of MHC II, C[1]D80 and CD86 molecules in BMDCs (p<0.05), as well as the T-cell stimulatory capacity of BMDCs (p<0.01). Moreover, the secretions of IL-6, IL-10 and IL-12, but not TNF-α induced by LPS exposure were suppressed by FB1 in a dose dependent (p<0.01). It was considered that the immunosuppressive effects of FB1 were mainly caused by changing the morphology and interfering with the process of antigen uptake, processing and presentation. The results highlighted that FB1 had the capacity to modulate the immune responses of BMDCs.

Monoolein, isolated from Ishige sinicola, inhibits lipopolysaccharide-induced inflammatory response by attenuating mitogen-activated protein kinase and NF-κB pathways.

Ishige sinicola (I. sinicola) is an edible brown alga native to South Korea. In the present study, we screened the anti-inflammatory activity of monoolein isolated from I. sinicola. Monoolein pretreatment in lipopolysaccharide (LPS)-stimulated primary murine bone marrow-derived dendritic cells (BMDCs) showed strong dose-dependent inhibition of interleukin (IL)-12 p40, IL-6, and TNF-α cytokine production with IC50 values of 1.69±0.02, 6.87±0.37, and 5.19±0.56 μM, respectively. Pretreatment of monoolein attenuated the activation of MAPK and NF-κB pathways in the LPS-stimulated BMDCs by inhibiting the phosphorylation of p38, ERK1/2, JNK1/2, and IκBα. Furthermore, monoolein inhibited the production of NO and iNOS in RAW264.7 cells. Overall, our findings indicate that monoolein has a significant anti-inflammatory activity, and further studies regarding the potential of monoolein for medicinal use is warranted.

Paeonol ameliorates imiquimod-induced psoriasis-like skin lesions in BALB/c mice by inhibiting the maturation and activation of dendritic cells.

Paeonol, an active component derived from the traditional Chinese medicine Cortex Moutan, possesses anti-inflammatory, analgesic, antioxidant and anti-allergic properties. Psoriasis is a chronic, recurrent, inflammatory dermatosis accompanied by excessive activation of Toll-like receptors (TLRs) in dendritic cells (DCs), which are primarily responsible for initiating an immune response. We investigated the effect of paeonol on inflammation in an imiquimod (IMQ)-induced psoriasis-like mouse model and murine bone marrow-derived dendritic cells (BMDCs) stimulated by R848. Mice were intragastrically administered 100 mg/kg (high), 50 mg/kg (medium) and 25 mg/kg (low) paeonol, respectively. We evaluated inflammation of psoriasis-like lesions based on histological changes, protein levels of myeloid differentiation factor 88 (MyD88) and TLR8 in skin lesions by western blotting, and levels of CD11c+ DCs in skin by immunoassay and in spleens by flow cytometry. Inflammatory cytokines [interleukin (IL)-23, IL-12 and IL-1β] in skin lesions and BMDCs were also assessed by RT-PCR and ELISA. Application of paeonol decreased IMQ-induced keratinocyte proliferation, and infiltration of CD3+ cells, while the treatment ameliorated CD11c+ cells in the spleen and skin, and reduced MyD88 and TLR8 proteins in skin lesions. Paeonol inhibited IMQ-induced mRNA expression of IL-23, but not IL-12 and IL-1β in BMDCs, along with significantly lower levels of DCs expressing MHCII, CD80 and CD86 in vitro. These results indicate that paeonol suppresses the maturation and activation of DCs by decreasing MyD88 and TLR8 proteins in the TLR7/8 signaling pathway which finally alleviates psoriasis-like skin lesions. The TLR7/8 signaling pathway in DCs provides an important insight into the mechanism of psoriasis, and paeonol may be a potent therapeutic drug for psoriasis.

Effect of Matrix Metallopeptidase 13 on the Function of Mouse Bone Marrow-derived Dendritic Cells.

Background:Dendritic cells are professional antigen-presenting cells found in an immature state in epithelia and interstitial space, where they capture antigens such as pathogens or damaged tissue. Matrix metallopeptidase 13 (MMP-13), a member of the collagenase subfamily, is involved in many different cellular processes and is expressed in murine bone marrow-derived dendritic cells (DCs). The function of MMP-13 in DCs is not well understood. Here, we investigated the effect of MMP-13 on DC maturation, apoptosis, and phagocytosis.Methods:Bone marrow-derived dendritic cells were obtained from C57BL/6 mice. One short-interfering RNA specific for MMP-13 was used to transfect DCs. MMP-13-silenced DCs and control DCs were prepared, and apoptosis was measured using real-time polymerase chain reaction and Western blotting. MMP-13-silenced DCs and control DCs were analyzed for surface expression of CD80 and CD86 and phagocytosis capability using flow cytometry.Results:Compared to the control DCs, MMP-13-silenced DCs increased expression of anti-apoptosis-related genes, BAG1 (control group vs. MMP-13-silenced group: 4.08 ± 0.60 vs. 6.11 ± 0.87, P = 0.008), BCL-2 (control group vs. MMP-13-silenced group: 7.54 ± 0.76 vs. 9.54 ± 1.29, P = 0.036), and TP73 (control group vs. MMP-13-silenced group: 4.33 ± 0.29 vs. 5.60 ± 0.32, P = 0.001) and decreased apoptosis-related genes, CASP1 (control group vs. MMP-13-silenced group: 3.79 ± 0.67 vs. 2.54 ± 0.39, P = 0.019), LTBR (control group vs. MMP-13-silenced group: 9.23 ± 1.25 vs. 6.24 ± 1.15, P = 0.012), and CASP4 (control group vs. MMP-13-silenced group: 2.07 ± 0.56 vs. 0.35 ± 0.35, P = 0.002). Protein levels confirmed the same expression pattern. MMP-13-silenced groups decreased expression of CD86 on DCs; however, there was no statistical difference in CD80 surface expression. Furthermore, MMP-13-silenced groups exhibited weaker phagocytosis capability.Conclusion:These results indicate that MMP-13 inhibition dampens DC maturation, apoptosis, and phagocytosis.

The protection role of Atg16l1 in CD11c+dendritic cells in murine colitis

The autophagy-related 16-like 1 gene (Atg16l1) is associated with inflammatory bowel disease (IBD) and has been shown to play an essential role in paneth cell function and intestinal homeostasis. However, the functional consequences of Atg16l1 deficiency in myeloid cells, particularly in dendritic cells (DCs), are not fully characterized. The aim of this study is to investigate the functional consequence of Atg16l1 in CD11c+DCs in murine colitis. We generated mice deficient in Atg16l1 in CD11c+DCs. Dextran Sulfate Sodium (DSS) and S. typhimurium infection induced colitis was used to assess the role of DCs specific Atg16l1 deficiency in vivo in murine colitis. Bone marrow derived dendritic cells (BMDC) were isolated and autophagy function was assessed with microtubule-associated protein 1 light chain 3β (Map1lc3b or LC3) by western blot. Uptake of Salmonella enteric serovar typhimurium (S. typhimurium) was assessed by flow cytometry and transmission electron microscopy (TEM). The production of reactive oxygen species (ROS) and intracellular S. typhimurium killing in BMDCs were assessed. We showed worsened colonic inflammation in Atg16l1 deficiency mice in DSS induced murine colitis with increased proinflammatory cytokines of IL-1β and TNF-α. Mechanistic studies performed in primary murine BMDCs showed that Atg16l1 deficiency increased ROS production, reduced microbial killing and impaired antigen processing for altered intracellular trafficking. Together, these results indicate impaired CD11c+DCs function with Atg16l1 deficiency contributes to the severity of murine colitis.

Immunogenicity is preferentially induced in sparse dendritic cell cultures

We have previously shown that human monocyte-derived dendritic cells (DCs) acquired different characteristics in dense or sparse cell cultures. Sparsity promoted the development of IL-12 producing migratory DCs, whereas dense cultures increased IL-10 production. Here we analysed whether the density-dependent endogenous breaks could modulate DC-based vaccines. Using murine bone marrow-derived DC models we show that sparse cultures were essential to achieve several key functions required for immunogenic DC vaccines, including mobility to draining lymph nodes, recruitment and massive proliferation of antigen-specific CD4+ T cells, in addition to their TH1 polarization. Transcription analyses confirmed higher commitment in sparse cultures towards T cell activation, whereas DCs obtained from dense cultures up-regulated immunosuppressive pathway components and genes suggesting higher differentiation plasticity towards osteoclasts. Interestingly, we detected a striking up-regulation of fatty acid and cholesterol biosynthesis pathways in sparse cultures, suggesting an important link between DC immunogenicity and lipid homeostasis regulation.

Virosome-bound antigen enhances DC-dependent specific CD4+ T cell stimulation, inducing a Th1 and Treg profile in vitro

There is considerable interest to develop antigen-carriers for immune-modulatory clinical applications, but insufficient information is available on their effects on antigen-presenting cells. We employed virosomes coupled to ovalbumin (OVA) to study their interaction with murine bone marrow-derived dendritic cells (BMDCs) and modulation of downstream T cell responses. BMDCs were treated in vitro with virosomes or liposomes prior to determining BMDC phenotype, viability, and intracellular trafficking. Antigen-specific CD4+ T cell activation was measured by co-culture of BMDCs with DO11.10 CD4+ T cells. Compared to liposomes, virosomes were rapidly taken up. Neither nanocarrier type affected BMDC viability, nor did a moderate degree of activation differ for markers such as CD40, CD80, CD86. Virosome uptake occurred via clathrin-mediated endocytosis and phagocytosis, with co-localization in late endosomes. Only BMDCs treated with OVA-coupled virosomes induced enhanced OVA-specific CD4+ T cell proliferation. Antigen-coupled virosomes are endowed with an intrinsic ability to modulate DC-dependent adaptive immune responses.

Effect of Different Titanium Surfaces on Maturation of Murine Bone Marrow-Derived Dendritic Cells

Dendritic cells (DCs) play a pivotal role in the host response to implanted biomaterials. Osseointegration of titanium (Ti) implant is an immunological and inflammatory-driven process. However, the role of DCs in this complex process is largely unknown. This study aimed to investigate the effect of different Ti surfaces on DC maturation, and evaluate its subsequent potential on osteogenic differentiation of preosteoblasts. Murine bone marrow-derived DCs were seeded on Ti disks with different surface treatments, including pretreatment (PT), sandblasted/acid-etched (SLA) and modified SLA (modSLA) surface. Compared with DCs cultured on PT and SLA surfaces, the cells seeded on modSLA surface demonstrated a more round morphology with lower expression of CD86 and MHC-II, the DC maturation markers. Those cells also secreted high levels of anti-inflammatory cytokine IL-10 and TGF-β. Notably, addition of conditioned medium (CM) from modSLA-induced DCs significantly increased the mRNA expression of Runx2 and ALP as well as ALP activity by murine preosteoblast MC3T3-E1 cells. Our data demonstrated that Ti disks with different surfaces lead to differential DCs responses. PT and SLA surfaces induce DCs mature, while DCs seeded on modSLA-Ti surface maintain an immature phenotype and exhibit a potential of promoting osteogenic differentiation of MC3T3-E1 cells.

Species-specific immunity to Helicobacter suis.

Helicobacter (H.) suis is mainly associated with pigs, but is also the most prevalent gastric non-H.pylori Helicobacter species found in humans. Both H.pylori and H.suis may cause persistent infection of the stomach. Several immune evasion mechanisms have been proposed for H.pylori, which focus to a great extent on its major virulence factors, which are absent in H.suis. The aim of this study was to gain more knowledge on immune evasion by H.suis. Cytokine expression kinetics were monitored in the stomach of BALB/c mice experimentally infected with H.suis. The cytokine expression profile in the stomach of naturally H.suis-infected pigs was also determined. Subsequently, the effect of H.suis on murine and porcine dendritic cell (DC) maturation and their ability to elicit T-cell effector responses was analyzed. Despite a Th17/Th2 response in the murine stomach, the inflammatory cell influx was unable to clear H.suis infection. H.suis-stimulated murine bone marrow-derived dendritic cells induced IL-17 secretion by CD4+ cells in vitro. Natural H.suis infection in pigs evoked increased expression levels of IL-17 mRNA in the antrum and IL-10 mRNA in the fundus. In contrast to mice, H.suis-stimulated porcine monocyte-derived dendritic cells were unable to express MHCII molecules on their cell surface. These semimature DCs induced proliferation of T-cells, which showed an increased expression of TGF-β and FoxP3 mRNA levels. Helicobacter suis might evade host immune responses by skewing toward a Treg-biased response.

Selenizing Hericium erinaceus polysaccharides induces dendritic cells maturation through MAPK and NF-κB signaling pathways

In this study, polysaccharides extracted from Hericium erinaceus were modified to obtain its nine selenium derivatives, sHEP1-sHEP9. Their structures were identified, yields and selenium contents were determined, the phenotypic and functional maturation of murine bone marrow-derived dendritic cells (DCs) and relevant mechanisms were compared taking unmodified HEP as control. The results revealed that the selenylation were successful. sHEP1, sHEP2 and sHEP8 treatment of DCs increased their surface expression of MHC-II and CD86 and indicated that sHEP1, sHEP2 and sHEP8 induced DC maturation. Furthermore, sHEP2 and sHEP8 also significantly decreased DCs endocytosis and significantly enhanced cytokine (IL-12 and IFN-γ) production. In line with TLR4 activation, sHEP2 increased the phosphorylation of ERK, p38, and JNK, and the nuclear translocation of p-c-Jun, p-CREB, and c-Fos. sHEP2 also activated NF-κB signaling, as evidenced by degradation of IκBα/β and nuclear translocation of p65 and p50. Together, these results suggest that sHEP is a strong immunostimulant.

Proteomic Analysis of Excretory-Secretory Products of Mesocestoides corti Metacestodes Reveals Potential Suppressors of Dendritic Cell Functions.

Accumulating evidences have assigned a central role to parasite-derived proteins in immunomodulation. Here, we report on the proteomic identification and characterization of immunomodulatory excretory-secretory (ES) products from the metacestode larva (tetrathyridium) of the tapeworm Mesocestoides corti (syn. M. vogae). We demonstrate that ES products but not larval homogenates inhibit the stimuli-driven release of the pro-inflammatory, Th1-inducing cytokine IL-12p70 by murine bone marrow-derived dendritic cells (BMDCs). Within the ES fraction, we biochemically narrowed down the immunosuppressive activity to glycoproteins since active components were lipid-free, but sensitive to heat- and carbohydrate-treatment. Finally, using bioassay-guided chromatographic analyses assisted by comparative proteomics of active and inactive fractions of the ES products, we defined a comprehensive list of candidate proteins released by M. corti tetrathyridia as potential suppressors of DC functions. Our study provides a comprehensive library of somatic and ES products and highlight some candidate parasite factors that might drive the subversion of DC functions to facilitate the persistence of M. corti tetrathyridia in their hosts.