Induced Dendritic Cells Activation Research Articles

Self-adjuvant Astragalus polysaccharide-based nanovaccines for enhanced tumor immunotherapy: a novel delivery system candidate for tumor vaccines.

The study of tumor nanovaccines (NVs) has gained interest because they specifically recognize and eliminate tumor cells. However, the poor recognition and internalization by dendritic cells (DCs) and insufficient immunogenicity restricted the vaccine efficacy. Herein, we extracted two molecular-weight Astragalus polysaccharides (APS, 12.19 kD; APSHMw, 135.67 kD) from Radix Astragali and made them self-assemble with OVA257-264 directly forming OVA/APS integrated nanocomplexes through the microfluidic method. The nanocomplexes were wrapped with a sheddable calcium phosphate layer to improve stability. APS in the formed nanocomplexes served as drug carriers and immune adjuvants for potent tumor immunotherapy. The optimal APS-NVs were approximately 160 nm with uniform size distribution and could remain stable in physiological saline solution. The FITC-OVA in APS-NVs could be effectively taken up by DCs, and APS-NVs could stimulate the maturation of DCs, improving the antigen cross-presentation efficiency in vitro. The possible mechanism was that APS can induce DC activation via multiple receptors such as dectin-1 and Toll-like receptors 2 and 4. Enhanced accumulation of APS-NVs both in draining and distal lymph nodes were observed following s.c. injection. Smaller APS-NVs could easily access the lymph nodes. Furthermore, APS-NVs could markedly promote antigen delivery efficiency to DCs and activate cytotoxic T cells. In addition, APS-NVs achieve a better antitumor effect in established B16-OVA melanoma tumors compared with the OVA+Alum treatment group. The antitumor mechanism correlated with the increase in cytotoxic T cells in the tumor region. Subsequently, the poor tumor inhibitory effect of APS-NVs on the nude mouse model of melanoma also confirmed the participation of antitumor adaptive immune response induced by NVs. Therefore, this study developed a promising APS-based tumor NV that is an efficient tumor immunotherapy without systemic side effects.

Artificial Immunogenic Cell Death Lipid Nanoparticle Functions as a Therapeutic Vaccine for Cancer

AbstractAnticancer drug‐mediated induction of immunogenic cell death (ICD) blocks metastasis or recurrence in cancer cells by promoting specific immune activity against cancer antigens. However, this strategy has failed to afford adequate treatment efficiency. Overcoming the failure of ICD‐mediated cancer therapy, lipid nanoparticles (LNPs) containing cancer cell surface proteins are synthesized using sonication and extrusion without microfluidics. In addition, these LNPs are decorated with high‐mobility group box 1 protein and calreticulin, indicators of ICD, and named artificial ICD LNPs (AiLNPs). Administration of AiLNPs effectively targets dendritic cells (DCs) and induces DC activation in mice. Moreover, treating CT‐26 tumor‐bearing mice with AiLNPs inhibits tumor growth by inducing CT‐26 antigen‐specific T‐cell immunity. Furthermore, AiLNPs containing Lewis lung carcinoma (LLC1) membrane proteins can prevent metastatic LLC1 tumor growth in the lung via LLC1 antigen‐specific T‐cell activation. Finally, AiLNPs synthesized with human breast cancer membrane proteins activate DC‐mediated antigen‐specific T‐cell immunity, effectively killing tumor cells. Therefore, AiLNPs are expected to be developed as a patient‐specific cancer treatment to prevent cancer recurrence and metastasis.

Exploring chitosan-shelled nanobubbles to improve HER2 + immunotherapy via dendritic cell targeting

Immunotherapy is a valuable approach to cancer treatment as it is able to activate the immune system. However, the curative methods currently in clinical practice, including immune checkpoint inhibitors, present some limitations. Dendritic cell vaccination has been investigated as an immunotherapeutic strategy, and nanotechnology-based delivery systems have emerged as powerful tools for improving immunotherapy and vaccine development. A number of nanodelivery systems have therefore been proposed to promote cancer immunotherapy. This work aims to design a novel immunotherapy nanoplatform for the treatment of HER2 + breast cancer, and specially tailored chitosan-shelled nanobubbles (NBs) have been developed for the delivery of a DNA vaccine. The NBs have been functionalized with anti-CD1a antibodies to target dendritic cells (DCs). The NB formulations possess dimensions of approximately 300 nm and positive surface charge, and also show good physical stability up to 6 months under storage at 4 °C. In vitro characterization has confirmed that these NBs are capable of loading DNA with good encapsulation efficiency (82%). The antiCD1a-functionalized NBs are designed to target DCs, and demonstrated the ability to induce DC activation in both human and mouse cell models, and also elicited a specific immune response that was capable of slowing tumor growth in mice in vivo. These findings are the proof of concept that loading a tumor vaccine into DC-targeted chitosan nanobubbles may become an attractive nanotechnology approach for the future immunotherapeutic treatment of cancer.Graphical abstract

Dendritic cells activated by cimetidine induce Th1/Th17 polarization in vitro and in vivo

Dendritic cells (DCs) are powerful antigen presentation cells and the initiator of adaptive immune response. Cimetidine, a widely used drug for gastric ulcers treatment, has significant immunomodulatory ability. However, the effects of cimetidine on DC-mediated T cell activation need to be further explored. In this study, we constructed the in vitro and in vivo model of cimetidine exposure, and our data showed that cimetidine stimulated the maturity of immature DCs, and further enhanced its T cell priming capacity. In vivo, the number of rat splenic CD103+ DC were not altered after cimetidine exposure, but the expression of surface markers CD54, CD11c, and MHC-II of which were up-regulated. Importantly, cimetidine interfered with DC-mediated T cell polarization, which was reflected in the up-regulation of Th1 and Th17 cells and the down-regulation of Th2 and Treg cells in vitro and in vivo. These results indicate that cimetidine can induce DC activation and promote DC mediated pro-inflammatory T cell response while weaken immunosuppressive T cell response.

Crude polysaccharides from Cistanche deserticola Y.C. Ma as an immunoregulator and an adjuvant for foot-and-mouth disease vaccine

Cistanche deserticola Y. C. Ma (CD) are ideal herbal remedies for health problems. In this study, modulator activities of CPCD on activating dendritic cells (DCs) and the adjuvant potential for foot-and-mouth disease vaccine (FMDV) was evaluated. CPCD without cytotoxicity induced DC activation via upregulating the phenotypic markers and antigen uptake. TLR-2 or TLR-4 antibodies suppressed levels of CPCD-mediated CD40 and CD86 as well as IL-6 and IL-1β in DCs. CPCD induced the phosphorylation of MAPKs related molecules and NF-κB. When co-injected intramuscularly with FMDV in mice, CPCD increased FMDV-specific antibody, lymphocyte proliferation, effector T-cells. Specifically, CPCD elicited Th1 response, as indicated by the higher expressions of CD4+ IFN-γ, CD8+ IFN-γ and CTL response. CPCD enhanced the expressions of MHC-II, CD80, CD86 and CD40 in DCs and lowered Tregs frequency. Therefore, CPCD could effectively stimulate stronger humoral and cellular responses by modulating DC activation through TLR-2/TLR-4 related MAPKs and NF-κB pathways.

Leptin promotes glycolytic metabolism to induce dendritic cells activation via STAT3-HK2 pathway

Leptin is over-secreted in many autoimmune diseases, which can promote dendritic cells (DCs) maturation and up-regulate the expression of inflammatory cytokines, but the underlying mechanisms are not fully elucidated. Considering the major role of leptin in maintaining energy balance and the significant role of glycolysis in DCs activation, our study aims to investigate whether leptin promotes the activation of DCs via glycolysis and its underlying mechanisms. We demonstrated that leptin promoted the activation of DCs, including up-regulating the expression of co-stimulatory molecules and inflammatory cytokines, enhancing the proliferation and T helper 17 (Th17) cell ratio in peripheral blood mononuclear cells (PBMC) co-cultured with leptin-stimulated DCs. Leptin also enhanced DCs glycolysis with increased glucose consumption, lactate production, and the expression of hexokinase 2 (HK2). In addition, the activation of DCs stimulated by leptin could be inhibited by the glycolysis inhibitor 2-deoxy-d-glucose (2-DG). To explore the signaling pathways involved in leptin-induced HK2 expression, we observed that the inhibitors of STAT3 (NSC74859) could repress the enhancement of HK2 triggered by leptin stimulation. Therefore, our results indicated that leptin promoted glycolytic metabolism to induce DCs activation via STAT3-HK2 pathway.

Aqueous extracts from cultivated Cistanche deserticola Y.C. Ma as polysaccharide adjuvant promote immune responses via facilitating dendritic cell activation

Ethnopharmacological relevanceHerbal polysaccharides have exhibited great immune-enhancing potential. Adjuvants are a key tool for developing efficacious vaccines. In our previous study, a water-soluble polysaccharide extracted from wild Cistanche deserticola Y.C. Ma showed potent immunostimulatory activity. Aim of studyIn this study, the immune profiles and efficacy of aqueous extracts of cultivated Cistanche deserticola Y.C. Ma (AECCD) on ICR mice against ovalbumin (OVA) were investigated. In vitro experiments, the possible DC activation mechanism by AECCD was evaluated. Materials and methodsAECCD were extracted using hot water after which the crude polysaccharides were precipitated by ethanol. Mice were firstly immunized subcutaneously with OVA (10 μg per mouse) alone or OVA (10 μg per mouse) respectively containing different dose of AECCD (200, 400 and 800 μg per mouse) on Days 1 and 14 and the magnitude and kinetics of antibodies and cell-mediated responses were then assessed. ResultsAECCD elicited vigorous and long-term IgG responses with mixed Th1/Th2 responses and up-regulated levels of Th-associated cytokines (CD4+IL-4, CD4+IFN-γ and CD8+IFN-γ). Moreover, AECCD induced the strong cellular immune response characterized by increased splenocyte proliferation as well as the activated T cell response. Notably, AECCD significantly enhanced the maturation of dendritic cells (DCs) and inhibited Tregs. In vitro experiments, Preliminary tests indicated that AECCD induced DC activation by promoting phenotypic maturation, cytokine section and allostimulatory activity. Toll-like receptor 4 (TLR4) was an essential receptor for DCs to directly bind AECCD. The inhibitors of NF-κB decreased the expression levels of CD40, CD80, CD86 and MHC-II and the production of IFN-γ, TNF-α and IL-6 through DCs. ConclusionsFinally, these findings suggested that AECCD could elicit potent and durable antigen specific immune responses through DC activation, which was involved in the regulation of maturation markers and cytokine expression via TLR4-related NF-κB pathway. The study indicates that AECCD is a potential immunomodulator.

The role of intracellular Pattern Recognition Receptors (PRRs) in recognition and immunomodulation of Dendritic cells (DC) in response to Daptomycin (Dap) resistant MRSA

Abstract Methicillin resistant Staphylococcus aureus (MRSA) infection is one of the leading causes of hospital-acquired infections and is more difficult to treat than other bacterial infections. In recent years, resistance to last line anti-staphylococcal antibiotic daptomycin has been observed during treatment of MRSA bacteremia. Protection against MRSA infection is challenging and requires both innate and adaptive immune effector mechanisms. Being at the interface between innate and adaptive immune responses, DCs are thus central to the immune protection against S. aureus. We have previously shown that distinct and paired clinical isolates of MRSA have differential capacity to induce DC activation, with MRSA strains resistant to the last_line antibiotic daptomycin inducing a compromised response to MRSA. However, this study explores the molecular mechanism by which MRSA strains impede DC activation. We explored the role of intracellular PRRs in the recognition and immunomodulation of these clinical isolates. We found that the cytosolic sensors cGAS and STING are implicated in the sensing of clinical isolates of MRSA, but their use is more prominent in response to DapS strains. We also determined that the TLR3 ligand poly(I:C) induced a highly synergistic increase in the activation of DC in response to both Daptomycin sensitive (DapS) and Daptomycin resistant (DapR) MRSA. Our work, thus provides important insights for the mechanism of differential recognition of clinical isolates of MRSA and immunomodulation through engagement of PRR pathways which are critical to achieve effective and comprehensive immune defense against this pathogen.

Liposomal Dendritic Cell Vaccine in Breast Cancer Immunotherapy.

Cancer vaccine is well recognized as a promising approach for immunotherapy of cancers. Since dendritic cells (DCs) are capable of processing and presenting antigens to initiate the immune response cascade, the development of DC vaccines is considered as a good choice for the treatment of cancer. Herein, a folic acid (FA)-modified liposome was constructed and loaded with chlorin e6 (Ce6) as a DC vaccine (FA-Lipo-Ce6). It was suggested that the loaded Ce6 within FA-Lipo-Ce6 can be activated under laser irradiation. The photodynamic therapy (PDT) of Ce6 was expected to create on-demand reactive oxygen species (ROS) in situ, which causes cell death and trigger the exposure of tumor-associated antigen (TAA). In addition, the produced ROS can mimic the inflammatory responses for the employment of DC for better antigen presentation and immune response. Most importantly, the employment of DC can recognize the exposed TAA to stimulate DC for effective vaccination in situ. Our results demonstrated the powerful capacity of FA-Lipo-Ce6 to induce DC activation, leading to effective suppression of the growth of breast cancers.

Skin sensitization to fragrance hydroperoxides: interplay between dendritic cells, keratinocytes and free radicals.

Skin sensitization to hydroperoxides (R-OOHs) of the commonly used fragrance terpenes limonene, linalool and citronellol is frequently reported. R-OOHs are believed to initiate the process leading to sensitization and allergic contact dermatitis through mechanisms involving radical intermediates. Thus, radical intermediates, keratinocytes and dendritic cells (DCs) may act in concert to initiate the process. To evaluate individual DC activation profiles by R-OOHs in the context of keratinocytes with regard to frequency, specificity and magnitude of upregulation. We used 2D and 3D cocultures with keratinocytes/reconstructed human epidermis (RHE) and DCs to evaluate cell surface levels of the costimulatory molecules CD86, CD80 and the adhesion molecule CD54 on cocultured DCs. Analysis of radical formation from limonene hydroperoxides in RHE was performed using electron paramagnetic resonance combined with the spin trapping technique. R-OOHs induce donor-dependent DC activation. Major differences were found between the limonene-OOHs. Limonene-1-OOH was stronger with respect to both frequency and magnitude of response. Using a 3D coculture model, no DC activation was detected after topical application of 0·2% limonene-OOHs (20µgcm-2 ), while 1·2% limonene-1-OOH or 2% limonene-2-OOH induced DC activation. Furthermore, we demonstrated differences in the carbon and oxygen radicals formed from the limonene-OOHs using RHE, mimicking what may happen in vivo. We report clear individual differences in DC maturation induced by the most important hydroperoxides. Response rates and magnitude of response both indicate that very small structural alterations in the hydroperoxides are translated into specific DC responses. In addition, we provide more insight into the amounts of hydroperoxides that can activate DCs and induce sensitization.

Growth Hormone Aggregates Activation of Human Dendritic Cells Is Controlled by Rac1 and PI3 Kinase Signaling Pathways.

The presence of protein aggregates in biological products is suggested to promote immunogenicity, leading to the production of anti-drug antibodies with neutralizing capacities. This suggests a CD4+ T-cell dependent adaptive immune response, thus a pivotal role for antigen-presenting cells, such as dendritic cells (DCs). We previously showed that human growth hormone aggregates induced DC maturation, with notably an increase in CXCL10 production. DC phenotypic modifications were sufficient to promote allogeneic CD4+ T-cell proliferation with Th1 polarization. In this work, we identified the main intracellular signaling pathways involved in DC activation by human growth hormone aggregates, showing that aggregates induced p38 mitogen-activated protein kinase, extracellular signal-regulated kinase, and c-Jun N-terminal kinase phosphorylation, as well as nuclear factor κB subunit p65 nuclear translocation. Next, investigating the implication of Rho GTPases and phosphoinositide 3-kinase (PI3K) in activated DC showed that Rac1 and Cdc42 regulated the phosphorylation of MAP kinases, whereas PI3K was only implicated in c-Jun N-terminal kinase phosphorylation. Furthermore, we showed that Rac1 and PI3K pathways, but not Cdc42, regulated the production of CXCL10 via the MAP kinases and nuclear factor κB. Taken together, our results bring new insight on how protein aggregates could induce DC activation, leading to a better understanding of aggregates role in therapeutic proteins immunogenicity.

The role of dendritic cells regulated by HMGB1/TLR4 signalling pathway in myocardial ischaemia reperfusion injury.

Inflammatory response plays an important role in ischaemia reperfusion injury (IRI) through a variety of inflammatory cells. Apart from neutrophils, macrophages and lymphocytes, the role of dendritic cells (DCs) in IRI has been noticed. The study was aimed at investigating whether the high‐mobility group protein box‐1/toll like receptor 4 (HMGB1/TLR4) signalling pathway regulate the migration, adhesion and aggregation of DCs to the myocardium, induce DCs activation and maturation, stimulate the expression of surface costimulatory molecules and participate in myocardial IRI. In vivo, migration, adhesion, and aggregation of DCs was enhanced; the expression of peripheral blood DCs CD80 and CD86, myocardial adhesion molecules were increased; and the infarct size was increased during myocardial ischaemia reperfusion injury myocardial ischemic/reperfusion injury (MI/RI). These responses induced by MI/RI were significantly inhibited by HMGB1 specific neutralizing antibody treatment. Cellular experiments confirmed that HMGB1 promoted the release of inflammatory cytokines through TLR4/MyD88/NF‐κB, upregulated CD80 and CD86 expression, mediated the damage of cardiomyocytes and accelerated the apoptosis. Our results indicate that DCs activation and maturation, stimulate the expression of surface costimulatory molecules by promoting the release of inflammatory factors through NF‐κB pathway and participate in myocardial IRI.

Dendritic cell-targeted recombinantLactobacilli induce DC activation and elicit specific immune responses against G57 genotype of avian H9N2 influenza virus infection

H9N2 avian influenza viruses are of significance in poultry and public health for the past two decades. Vaccination plays an important role in preventing the infection in domestic poultry. Current H9N2 vaccines have not yet offered ideal protection and eliminated shedding of G57 genotype viruses responsible for H9N2 outbreaks during 2010–2013. Targeted vaccination is a promising strategy to improve vaccine effectiveness. Such a vaccine strategy can be achieved if it is targeted to dendritic cells (DCs) that directly elicit mucosal and adaptive immune responses against microbe challenge. For this purpose, we develop a DC-targeted mucosal vaccine for the oral delivery of the HA protein fused to a DCpep by using Lactobacillus plantarum as an antigen delivery system against G57 virus infection. It showed that Lactobacillus plantarum expressing HA-DCpep confers efficient protection against G57 H9N2 infection, due to have the potential to activate DCs by the TLR-induced NF-κB pathway, to promote DC migration by the CCR7-CCL19/CCL21 axis, thereby enhancing the presentation of immunogen to T and B lymphocytes, resulting in skewing T cells polarization towards Th1, Th2 and Treg cells and evoking more efficient mucosal and adaptive immunity responses. The presented oral mucosal vaccine strategy illustrates the feasibility and efficacy of antigen targeting to DCs through genetic fusion of vaccines to DC-targeting peptides and aids in the design and selection of indications that could be used with this oral vaccine platform against influenza.

Inflammatory responses of primary human dendritic cells towards polydimethylsiloxane and polytetrafluoroethylene.

Although frequently used as implants materials, both polydimethylsiloxane (PDMS) and polytetrafluoroethylene (PTFE) are often associated with adverse effects including foreign body responses. Dendritic cells (DC) are crucial for the initiation of immune reactions and could also play a role in foreign body associated inflammations. Therefore, the interaction of DC with PDMS and PTFE was investigated regarding their capacity to induce undesired cell activation. Medical grade PDMS and PTFE films were embedded into polystyrene PS inserts via injection molding to prevent the DC from migrating below the substrate and thereby, interacting not only with the test sample but also with the culture vessel material. The viability, the expression of co-stimulatory molecules, and the cytokine/chemokine profiles were determined after 24 hours incubation of the DC with PDMS or PTFE. Blank PS inserts and tissue culture polystyrene (TCP) served as reference materials. The viability of DC was not substantially influenced after incubation with PDMS and PTFE. However, both polymers induced DC activation indicated by the upregulation of co-stimulatory molecules. The release profiles of 14 soluble inflammatory mediators showed substantial differences between PDMS, PTFE, PS, and TCP. This study showed the potential of PTFE and PDMS to activate primary human dendritic cells, which could be an explanation for the often observed inflammatory events associated with the implantation of these polymers.

Inhibition of H9N2 Virus Invasion into Dendritic Cells by the S-Layer Protein from L. acidophilus ATCC 4356.

Probiotics are essential for the prevention of virus invasion and the maintenance of the immune balance. However, the mechanism of competition between probiotics and virus are unknown. The objectives of this study were to isolate the surface layer (S-layer) protein from L. acidophilus ATCC 4356 as a new antiviral material, to evaluate the stimulatory effects of the S-layer protein on mouse dendritic cells (DCs) and to verify its ability to inhibit the invasion of H9N2 avian influenza virus (AIV) in DCs. We found that the S-layer protein induced DCs activation and up-regulated the IL-10 secretion. The invasion and replication of the H9N2 virus in mouse DCs was successfully demonstrated. However, the invasion of H9N2 virus into DCs could be inhibited by treatment with the S-layer protein prior to infection, which was verified by the reduced hemagglutinin (HA) and neuraminidase (NA) mRNA expression, and nucleoprotein (NP) protein expression in the DCs. Furthermore, treatment with the S-layer protein increases the Mx1, Isg15, and Ddx58 mRNA expressions, and remits the inflammatory process to inhibit H9N2 AIV infection. In conclusion, the S-layer protein stimulates the activation of mouse DCs, inhibits H9N2 virus invasion of DCs, and stimulates the IFN-I signaling pathway. Thus, the S-layer protein from Lactobacillus is a promising biological antiviral material for AIV prevention.

Attenuated Streptococcus pneumoniae vaccine candidate SPY1 promotes dendritic cell activation and drives a Th1/Th17 response

Streptococcus pneumoniae is one of the causative agent of pneumonia, meningitis, otitis media and sepsis. Vaccination is an effective strategy to combat S. pneumoniae invasion. We previously reported that SPY1, a novel attenuated vaccine candidate against S. pneumoniae, induces a protective immune response against pneumococcal infection in mice. However, underlying mechanisms have yet to be fully illustrated. To explore the mechanism of innate and adaptive immunities induced by SPY1. In this study, bone marrow-derived dendritic cells (DCs) of mice were infected with SPY1 and its parental wild-type strain D39, SPY1-infected DCs were co-cultured with homologous CD4+T cells or adoptive transfer to C57BL/6 mice. Results showed that SPY1 promoted DCs maturation with increased levels of surface molecules such as CD40, CD86, and MHC II, and upregulated the expression of proinflammatory cytokines, including TNF-α, IL-6, IL-12p40, IL-12p70 and IL-23. By contrast, D39 did not efficiently induce DCs activation and maturation. SPY1 could also activate MAPK and NF-κB signaling pathways in DC, but D39 unlikely affected this pathways. SPY1 treated DCs also induced Th1 and Th17 responses in vitro and in vivo. Our results supported the potential of SPY1 as a novel attenuated pneumococcus vaccine, because SPY1-activated DCs exhibit fully matured phenotype, initiated an adaptive immune response, and orchestrated Th1 and Th17 responses.

Abstract 1648: Ionizing radiation switches the function of tumor-derived exosomes from messengers of tolerance to inducers of antitumor immunity

Abstract Tumor-derived exosomes (TEX) are constantly shed by cancer cells and have been shown to carry tumor-derived antigens to dendritic cells (DCs). However, presentation of tumor antigens derived from TEX in the absence of activation signals for DCs leads to immune tolerance. Our lab has demonstrated that ionizing radiotherapy (RT) can convert the irradiated tumor into an in situ vaccine, leading to anti-tumor immune responses. We hypothesized that TEX derived from irradiated cancer cells play a role in RT-induced anti-tumor immunity by delivering tumor antigens together with pro-inflammatory signals that activate DCs and prime tumor-specific effector T cells. To begin to address this hypothesis we first studied how RT modifies TEX molecular composition and their ability to induce DC activation in vitro. Mouse carcinoma cells TSA cultured in exosome-free media were treated in vitro with sham RT (control TEX), or 3 fractions of 8Gy (RT-TEX). TEX were isolated from supernatants 48 hr later using differential ultracentrifugation and purified with a sucrose gradient. Electron microscopy was used to confirm TEX size and morphology. Mass spectrometry (LFQ-MS) followed by MS/MS analyses was used to characterize protein composition. miRNA were analyzed by nanoString nCounter Mouse miRNA expression assay kit using a panel of 578 mouse miRNAs. Normalized results were analyzed with MultiExperiment Viewer. Mouse bone marrow-derived dendritic cells (BMDC) were cultured with TEX for 48 hours. Cells were analyzed by flow cytometry for expression of activation markers. Significant changes in microRNA and protein compositions were seen in RT-TEX compared to control TEX. Specifically, RT-TEX showed increase in proteins involved in the Antigen Processing and Presentation pathway. Additionally, 17 unique proteins were present only in RT-TEX and included proteins involved in T cell development, MHC class I peptide processing, and pro-inflammatory lipid signaling. Analysis of DCs cultured with RT-TEX, but not control TEX, revealed an increase in cell surface expression of CD80 (1428 MFI vs. 963 MFI) and CD86 (3487 MFI vs. 2578 MFI). Interestingly, culturing BMDC with RT-TEX also resulted in an increase in DCs expressing CD103 and CD8a (2% vs 1.2% of CD11c+ cells), suggesting that RT-TEX may influence differentiation of BMDC towards this subset, which is critical for cross-presentation of tumor antigens to T cells. Obtained data support the hypothesis that RT-TEX may switch tolerogenic DCs into activated DCs by providing activation signals together with tumor-derived antigens. Further in vivo experiments are ongoing to determine the ability of RT-TEX to stimulate anti-tumor immune responses. Citation Format: Julie M. Diamond, Jessica R. Chapman, Beatrix Ueberheide, Silvia C. Formenti, Sandra Demaria. Ionizing radiation switches the function of tumor-derived exosomes from messengers of tolerance to inducers of antitumor immunity. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1648.

Hyperoxic Exposure of Immature Mice Increases the Inflammatory Response to Subsequent Rhinovirus Infection: Association with Danger Signals.

Infants with a history of prematurity and bronchopulmonary dysplasia have a high risk of asthma and viral-induced exacerbations later in life. We hypothesized that hyperoxic exposure, a predisposing factor to bronchopulmonary dysplasia, modulates the innate immune response, producing an exaggerated proinflammatory reaction to viral infection. Two- to 3-d-old C57BL/6J mice were exposed to air or 75% oxygen for 14 d. Mice were infected intranasally with rhinovirus (RV) immediately after O2 exposure. Lung mRNA and protein expression, histology, dendritic cells (DCs), and airway responsiveness were assessed 1-12 d postinfection. Tracheal aspirates from premature human infants were collected for mRNA detection. Hyperoxia increased lung IL-12 expression, which persisted up to 12 d postexposure. Hyperoxia-exposed RV-infected mice showed further increases in IL-12 and increased expression of IFN-γ, TNF-α, CCL2, CCL3, and CCL4, as well as increased airway inflammation and responsiveness. In RV-infected, air-exposed mice, the response was not significant. Induced IL-12 expression in hyperoxia-exposed, RV-infected mice was associated with increased IL-12-producing CD103(+) lung DCs. Hyperoxia also increased expression of Clec9a, a CD103(+) DC-specific damaged cell-recognition molecule. Hyperoxia increased levels of ATP metabolites and expression of adenosine receptor A1, further evidence of cell damage and related signaling. In human preterm infants, tracheal aspirate Clec9a expression positively correlated with the level of prematurity. Hyperoxic exposure increases the activation of CD103(+), Clec9a(+) DCs, leading to increased inflammation and airway hyperresponsiveness upon RV infection. In premature infants, danger signal-induced DC activation may promote proinflammatory airway responses, thereby increasing respiratory morbidity.

Ascophyllan functions as an adjuvant to promote anti-cancer effect by dendritic cell activation.

Our previous study demonstrated that ascophyllan, a sulfated polysaccharide purified from brown alga, has immune-activating effects. In this study, we evaluated ascophyllan as an adjuvant for its therapeutic and preventive effect on tumor in a mouse melanoma model. Ascophyllan induced migration of DCs to spleen and tumor-draining lymph node (drLN) in a mouse B16 melanoma model. Moreover, ascophyllan induced activation of dendritic cells (DCs), and promoted IFN-γ- and TNF-α-producing Th1 immune responses in tumor-bearing mice. In addition, treatment with a combination of ascophyllan and ovalbumin (OVA) in the tumor-bearing mice promoted proliferation of OVA-specific CD4 and CD8 T cells and migration of those cells into the tumor, consequently inhibiting the tumor growth. Immunization with the combination of ascophyllan and OVA caused enhanced OVA-specific antibody production and memory T cell responses compared to OVA immunization alone, and almost completely prevented B16-OVA tumor growth upon subsequent tumor challenge. Finally, the combination of ascophyllan and OVA prevented B16-OVA tumor invasion and metastasis into the liver. Thus, these results demonstrate that ascophyllan can function as an adjuvant to induce DC activation, antigen specific CTL activation, Th1 immune response and antibody production, and hence may be useful as a therapeutic and preventive tumor vaccine.

β-(1,3)-glucan isolated from Agrobacterium species induces maturation of bone marrow-derived dendritic cells and drives Th1 immune responses

The β-(1,3)-glucan originally isolated from Agrobacterium species was investigated for prompting maturation of dendritic cells (DCs) and driving Th1 immune responses. Bone-marrow derived DCs separated from mice were analyzed for augmentation of cell surface molecule (CD80, CD86, and major histo-compatability complex (MHC) class I/II) expression and pro-inflammatory cytokine production (tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6). β-Glucan functionally induced DCs activation via augmentation of CD80, CD86, and MHC class I/II expression and cytokine production (TNF-α, IL-1β, and IL-6). β-Glucan induced secretion of IL-12p70, but not IL-10. Both mitogen-activated protein kinases and nuclear factor (NF)-κB signaling mediated production of pro-inflammatory cytokine induced by β-glucan. β-Glucan-treated DCs exhibited accelerated proliferation of murine splenocytes with increased levels of interferon (IFN)-γ. β-Glucan regulates innate and adaptive immunity via DCs activation and Th1 polarization of immune responses.