Surface Of Dendritic Cells Research Articles

Activation of dendritic cells by crosslinked collagen hydrogels (artificial corneas) varies with their composition

Activated T cells are known to promote fibrosis, a major complication limiting the range of polymeric hydrogels as artificial corneal implants. As T cells are activated by dendritic cells (DC), minimally activating hydrogels would be optimal. In this study, we evaluated the ability of a series of engineered (manufactured/fabricated) and natural collagen matrices to either activate DC or conversely induce DC apoptosis in vitro. Bone marrow DC were cultured on a series of singly and doubly crosslinked hydrogels (made from recombinant human collagen III [RHCIII] or collagen mimetic peptide [CMP]) or on natural collagen-containing matrices, MatrigelTM and de-cellularised mouse corneal stroma. DC surface expression of major histocompatibility complex Class II and CD86 as well as apoptosis markers were examined. Natural matrices induced low levels of DC activation and maintained a "tolerogenic" phenotype. The same applied to singly crosslinked CMP-PEG gels. RHCIII gels singly crosslinked using either N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide with the coinitiator N-hydroxy succinimide (EDC-NHS) or N-cyclohexyl-N-(2-morpholinoethyl)carbodiimide metho-p-toulenesulfonate with NHS (CMC-NHS) induced varying levels of DC activation. In contrast, however, RHCIII hydrogels incorporating an additional polymeric network of 2-methacryloyloxyethyl phosphorylcholine did not activate DC but instead induced DC apoptosis, a phenomenon observed in natural matrices. This correlated with increased DC expression of leukocyte-associated immunoglobulin-like receptor-1. Despite low immunogenic potential, viable tolerogenic DC migrated into and through both natural and manufactured RHCIII gels. These data show that the immunogenic potential of RHCIII gels varies with the nature and composition of the gel. Preclinical evaluation of hydrogel immunogenic/fibrogenic potential is recommended.

Proteasomal degradation within endocytic organelles mediates antigen cross-presentation.

During MHC-I-restricted antigen processing, peptides generated by cytosolic proteasomes are translocated by the transporter associated with antigen processing (TAP) into the endoplasmic reticulum, where they bind to newly synthesized MHC-I molecules. Dendritic cells and other cell types can also generate MHC-I complexes with peptides derived from internalized proteins, a process called cross-presentation. Here, we show that active proteasomes within cross-presenting cell phagosomes can generate these peptides. Active proteasomes are detectable within endocytic compartments in mouse bone marrow-derived dendritic cells. In TAP-deficient mouse dendritic cells, cross-presentation is enhanced by the introduction of human β2 -microglobulin, which increases surface expression of MHC-I and suggests a role for recycling MHC-I molecules. In addition, surface MHC-I can be reduced by proteasome inhibition and stabilized by MHC-I-restricted peptides. This is consistent with constitutive proteasome-dependent but TAP-independent peptide loading in the endocytic pathway. Rab-GTPase mutants that restrain phagosome maturation increase proteasome recruitment and enhance TAP-independent cross-presentation. Thus, phagosomal/endosomal binding of peptides locally generated by proteasomes allows cross-presentation to generate MHC-I-peptide complexes identical to those produced by conventional antigen processing.

Targeting Conventional Dendritic Cells to Fine-Tune Antibody Responses.

Dendritic cells (DCs) facilitate cross talk between the innate and adaptive immune system. They sense and phagocytose invading pathogens, and are not only capable of activating naïve T cells, but can also determine the polarization of T cell responses into different effector subtypes. Polarized T cells in turn have a crucial role in antibody class switching and affinity maturation, and consequently the quality of the resulting humoral immunity. Targeting vaccines to DCs thus provides a great deal of opportunities for influencing the humoral immune responses, by fine-tuning the T cell response as well as regulating antigen availability for B cells. In this review we aim to outline how different DC targeted vaccination strategies can be utilized to induce a desired humoral immune response. A range of factors, including route of vaccine administration, use of adjuvants, choice of DC subset and surface receptor to target have been reported to influence the resulting immune response and will be reviewed herein. Finally, we will discuss opportunities for designing improved vaccines and challenges with translating this knowledge into clinical or veterinary medicine.

Abstract 1559: TLR7/8 immune-stimulating antibody conjugates elicit robust myeloid activation leading to enhanced effector function and anti-tumor immunity in pre-clinical models

Abstract Deficient anti-tumor immunity often results from an immunosuppressive tumor microenvironment (TME) that renders antigen presenting cells (APCs) unable to effectively stimulate T cells. Recent studies indicate that local delivery of immunostimulatory adjuvants can activate tumor resident APCs, driving uptake, processing and presentation of tumor neoantigens to T cells that mediate anti-tumor immunity. To overcome challenges associated with intratumoral delivery of such adjuvants, we developed a novel class of TLR immune-stimulating antibody conjugates (TAC) that comprise a TLR7/8 agonist conjugated to tumor-targeting monoclonal antibodies. In vitro co-cultures with human cancer cell lines and leukocytes revealed that TACs potently activate primary APCs, leading to increased co-stimulatory molecule expression (e.g. CD40, CD86) and secretion of pro-inflammatory cytokines (e.g. TNFα). The TACs also enhanced antibody-mediated effector functions such as ADCC and ADCP. Surprisingly, these constructs also induced dendritic cell (DC) differentiation from monocytes, as measured by changes in cellular morphology and DC surface markers (e.g. CD14 downregulation). CyTOF-based analysis of intracellular signaling in human leukocytes revealed a unique signaling signature of the conjugate compared to a mixture of its components, suggesting a novel biological mechanism by which the conjugate stimulates APCs. Finally, we demonstrated in vivo efficacy in syngeneic tumor models in which TAC treatment led to tumor clearance and development of immunologic memory. These results provide a strong rationale for this technology as a platform for cancer immunotherapy. Citation Format: Shelley E. Ackerman, Joseph C. Gonzalez, Josh D. Gregorio, Jason C. Paik, Felix J. Hartmann, Justin A. Kenkel, Arthur Lee, Angela Luo, Cecelia I. Pearson, Murray L. Nguyen, Benjamin Ackerman, Lauren Y. Sheu, Richard P. Laura, Steven J. Chapin, Brian S. Safina, Sean C. Bendall, David Dornan, Edgar G. Engleman, Michael N. Alonso. TLR7/8 immune-stimulating antibody conjugates elicit robust myeloid activation leading to enhanced effector function and anti-tumor immunity in pre-clinical models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1559.

The Cross-Talk between miR-511-3p and C-Type Lectin Receptors on Dendritic Cells Affects Dendritic Cell Function.

MicroRNAs are small, noncoding RNAs that function as posttranscriptional modulators of gene expression by binding target mRNAs and inhibiting translation. They are therefore crucial regulators of several biological as well as immunological events. Recently, miR-511-3p has been implicated in the development and differentiation of APCs, such as dendritic cells (DCs), and regulating several human diseases. Interestingly, miR-511-3p is embedded within the human MRC1 gene that encodes the mannose receptor. In this study, we sought to examine the impact of miR-511-3p up- or downregulation on human DC surface phenotype, cytokine profile, immunogenicity (using IDO activity as a surrogate), and downstream T cell polarization. Using gene silencing and a selection of microRNA mimics, we could successfully suppress or induce the expression of miR-511-3p in DCs. Consequently, we show for the first time, to our knowledge, that inhibition and/or overexpression of miR-511-3p has opposing effects on the expression levels of two key C-type lectin receptors, namely the mannose receptor and DC-specific ICAM 3 nonintegrin at protein and mRNA levels, thereby affecting C-type lectin receptor-induced modulation of IDO activity in DCs. Furthermore, we show that downregulation of miR-511-3p drives an anti-inflammatory DC response characterized by IL-10 production. Interestingly, the miR-511-3plow DCs also promoted IL-4 secretion and suppressed IL-17 in cocultures with autologous T cells. Together, our data highlight the potential role of miR-511 in regulating DC function and downstream events leading to Th polarization and immune modulation.

Costimulatory Molecules and Immune Checkpoints Are Differentially Expressed on Different Subsets of Dendritic Cells.

Dendritic cells (DCs) play a crucial role in initiating and shaping immune responses. The effects of DCs on adaptive immune responses depend partly on functional specialization of distinct DC subsets, and partly on the activation state of DCs, which is largely dictated by environmental signals. Fully activated immunostimulatory DCs express high levels of costimulatory molecules, produce pro-inflammatory cytokines, and stimulate T cell proliferation, whereas tolerogenic DCs express low levels of costimulatory molecules, produce immunomodulatory cytokines and impair T cell proliferation. Relevant to the increasing use of immune checkpoint blockade in cancer treatment, signals generated from inhibitory checkpoint molecules on DC surface may also contribute to the inhibitory properties of tolerogenic DCs. Yet, our knowledge on the expression of inhibitory molecules on human DC subsets is fragmentary. Therefore, in this study, we investigated the expression of three immune checkpoints on peripheral blood DC subsets, in basal conditions and upon exposure to pro-inflammatory and anti-inflammatory stimuli, by using a flow cytometric panel that allows a direct comparison of the activatory/inhibitory phenotype of DC-lineage and inflammatory DC subsets. We demonstrated that functionally distinct DC subsets are characterized by differential expression of activatory and inhibitory molecules, and that cDC1s in particular are endowed with a unique immune checkpoint repertoire characterized by high TIM-3 expression, scarce PD-L1 expression and lack of ILT2. Notably, this unique cDC1 repertoire was subverted in a group of patients with myelodysplastic syndromes included in the study. Applied to the characterization of DCs in the tumor microenvironment, this panel has the potential to provide valuable information to be used for investigating the role of DC subsets in cancer, guiding DC-targeting treatments, and possibly identifying predictive biomarkers for clinical response to cancer immunotherapy.

A Sampling of Highlights from the Literature

Capping PD-L1 with cis CD80 blocks inhibition (by Trounce via Wikimedia Commons)PD-L1 and CD80 only interact in cis. Dendritic cells (DCs) can express excessive amounts of CD80, which is necessary for costimulation of antitumor T cells. However, another role that CD80 plays is to tie up any PD-L1 expressed on the DC surface. In doing so, PD-1 on T cells is prevented from engaging its ligand PD-L1 on DCs, allowing for CD28 costimulation and thus, the consequent activation of specific T cells can commence.Sugiura D, …, Okazaki T. Science 2019 May 10;364:558–66.Toxic stems keep controls at bay (E. Aardvark via Wikimedia Commons)Miao and colleagues developed an inducible epidermal squamous cell carcinoma model and find that cancer stem cells responding to TGFβ can evade immune responses through upregulation of CD80, which then binds to CTLA-4 on T cells, dampening the response. Miranda and colleagues find other mechanisms to explain how cancer stem cells can keep tumors "cold", which include increased expression of immune checkpoints, decreased type I IFN signaling, and reduced expression of endogenous retroviruses.Miao Y, …, Fuchs E. Cell 2019 April 25. DOI:10.1016/j.cell.2019.03.025.Miranda A, …, Nelson BH. Proc Natl Acad Sci U S A 2019 Apr 30;116:9020–9.Releasing cDC2s (by E. Alcinoe via OpenPhoto)Migratory conventional dendritic cells (cDC2s) in tumor draining lymph nodes were identified by single-cell analysis from the heterogeneous myeloid population as critical for the proliferation of conventional CD4+ T cells. cDC2s can only fully prime CD4+ T cells if regulatory T cells are depleted or if anti–CTLA-4 treatment is given. Human tumors also have extensive heterogeneity among myeloid cells, and patients whose tumors had more cDC2s and fewer Tregs had the best prognoses, suggesting usefulness as a biomarker.Binnewies M, …, Krummel MF. Cell 2019 Apr 18;177:556–71.Allies with iron-dependent death (by E. Serrao via Flickr)An iron-dependent form of cell death, ferroptosis, results from the accumulation of lipid peroxidation in tumor cells. CD8+ T cells secrete IFNγ, which decreases the amount of two subunits of a cystine transporter necessary to fend off this accumulation in tumor cells. Mice depleted of cystine or cysteine have enhanced responses to immune checkpoint blockade (ICB), and patients whose tumors have intact cystine transport have poorer prognoses. Patient response to ICB correlated with reduced expression of the transporters and more IFNγ-producing CD8+ T cells.Wang W, …, Zou W. Nature 2019 May 1;569:270–4.Comparing landscapes (from LJ Miller)Biomarkers of TAMs associated with survival, recruitment, and metastasis are identified by differential gene expression. Breast and endometrial tumors secrete undetermined factors that stimulate a regulatory loop whereby tumor cells secrete CSF1 and TNFα inducing TAMs to express SIGLEC1 and CCL8, attracting not only more monocytes but also stimulating tumor cells to make more CSF1 and TNFα, as well as inducing cancer cell motility.Cassetta L, …, Pollard JW. Cancer Cell 2019 Apr 15;35:588–602.Fusions stimulate responses (by N. Hotsuma via Flickr)A patient with an HPV− metastatic squamous cell carcinoma (SCC) responded completely to anti–PD-1. T cells recognized and were activated by a neoepitope formed by a fusion of two genes in the cancer cells. T-cell clones specific for this fusion-derived epitope increased with anti–PD-1 therapy, then decreased as the tumor regressed and stabilized at low numbers. Other patients' T cells were found to recognize other immunostimulatory fusion neoepitopes. Analysis of data sets from patients who received PD-1 blockade revealed that the frequency of fusions predicted to be immunogenic declined in responders, suggestive of immunoediting.Yang W, …, Morris LGT. Nat Med 2019 Apr 22;25:767–75.

Characteristics of Impaired Dendritic Cell Function in Patients With Hepatitis B Virus Infection.

Dendritic cells (DCs) are antigen-presenting cells with a central role in host immune response. This study analyzed gene expression and DC function in hepatitis B virus (HBV) patients, functions impaired because of HBV, and identified the genes related to these functions. Peripheral blood mononuclear cells from 64 HBV patients and 19 healthy controls were analyzed. Peripheral blood DCs were stained with antibodies against human leukocyte antigen-DR/Lin-1/CD123/CD11c and separated into plasmacytoid DCs (pDCs) and myeloid DCs by fluorescence-activated cell sorting. Using an interferon-gamma enzyme-linked immunospot assay, we analyzed antigen-specific response in HBV-infected patients. Regarding DC function, we analyzed antigen-presenting capacity, cell migration capacity, phagocytic capacity, and cytokine production capacity. DC gene expression was analyzed by microarray to identify genes related to DC function. No difference was found in the number of DCs in peripheral blood between healthy participants and HBV patients. In cell-surface marker analysis, CD80, CD83, CD86, CD40, and C-C motif chemokine receptor 7 expression levels in pDCs were related to the HBV-specific T-cell response. DCs from HBV patients exhibited decreases in antigen-presenting capacity, migration capacity, and cytokine production capacity. In gene expression analysis, immune-related genes with greatly reduced expression levels in chronic hepatitis B patients were identified. Of these genes, interleukin (IL)-6 signal transducer (IL6ST) expression level positively correlated with DC surface marker expression level. Adjustment of IL6ST expression level in DCs and treatment with oncostatin M resulted in recovery of DC function. Conclusion: IL6ST expression was identified as one cause of decline in DC function in HBV patients. Adjustment of IL6 family cytokine signaling may be useful for recovering reduced DC function in HBV infection.

Mechanisms of PTPσ-Mediated Presynaptic Differentiation.

Formation of synapses between neurons depends in part on binding between axonal and dendritic cell surface synaptic organizing proteins, which recruit components of the developing presynaptic and postsynaptic specializations. One of these presynaptic organizing molecules is protein tyrosine phosphatase σ (PTPσ). Although the protein domains involved in adhesion between PTPσ and its postsynaptic binding partners are known, the mechanisms by which it signals into the presynaptic neuron to recruit synaptic vesicles and other necessary components for regulated transmitter release are not well understood. One attractive candidate to mediate this function is liprin-α, a scaffolding protein with well-established roles at the synapse. We systematically mutated residues of the PTPσ intracellular region (ICR) and used the yeast dihydrofolate reductase (DHFR) protein complementation assay to screen for disrupted interactions between these mutant forms of PTPσ and its various binding partners. Using a molecular replacement strategy, we show that disrupting the interaction between PTPσ and liprin-α, but not between PTPσ and itself or another binding partner, caskin, abolishes presynaptic differentiation. Furthermore, phosphatase activity of PTPσ and binding to extracellular heparan sulfate (HS) proteoglycans are dispensable for presynaptic induction. Previous reports have suggested that binding between PTPσ and liprin-α is mediated by the PTPσ membrane-distal phosphatase-like domain. However, we provide evidence here that both of the PTPσ phosphatase-like domains mediate binding to liprin-α and are required for PTPσ-mediated presynaptic differentiation. These findings further our understanding of the mechanistic basis by which PTPσ acts as a presynaptic organizer.

Poly Lactic-co-Glycolic Acid Nanoparticles Containing Human Gastric Tumor Lysates as Antigen Delivery Vehicles for Dendritic Cell-Based Antitumor Immunotherapy

ABSTRACTStimulating antitumor T cells using dendritic cells (DCs) is a novel and promising method in cancer therapy. Poly lactic-co-glycolic acid is one of the best-known polymers used for encapsulating antigen to protect them against proteolytic enzymes. In this study, poly lactic-co-glycolic acid nanoparticles (NPs) were used as DC antigen delivery vehicles in a preclinical model of immunotherapy of gastric cancer. The DCs were generated from peripheral blood monocytes by conventional in vitro differentiation and loaded with either soluble tumor lysate or lysate encapsulated in NPs using a double emulsion/solvent evaporation technique. Morphology of NPs was determined by scanning electron microscopy. Tumor lysate, either in the soluble form or encapsulated in NPs, was loaded into DC and stimulatory capacity was compared using patient-derived autologous CD3+ T cells as responders. The amount of relevant cytokines produced by Ag-loaded DC and in DC/T cell cocultures was evaluated as a measure of initial DC stimulation and T-cell responses, respectively. Significance increases in expression of DC surface molecules (i.e., CD80, CD83, CD86, and Human Leukocyte Antigen–DR (HLA-DR)) and cytokine production by both DC and DC/T cell cocultures (i.e., interleukin (IL)-12:IL-10 and interferon [IFN]-γ:IL-4 ratios) was observed following loading with lysate NP versus controls. The results suggest that NP-encapsulated antigen can shift antitumor T-cell responses toward a Th1 bias, which potentially increases DC vaccine potency in clinical settings.

Sjogren's Syndrome and TAM Receptors: A Possible Contribution to Disease Onset.

Sjogren's syndrome (SS) is a chronic, progressive autoimmune disease featuring both organ-specific and systemic manifestations, the most frequent being dry mouth and dry eyes resulting from lymphocytic infiltration into the salivary and lacrimal glands. Like the related autoimmune disease systemic lupus erythematosus (SLE), SS patients and mouse models display accumulation of apoptotic cells and a Type I interferon (IFN) signature. Receptor tyrosine kinases (RTKs) of the Tyro3, Axl, and Mer (TAM) family are present on the surface of macrophages and dendritic cells and participate in phagocytosis of apoptotic cells (efferocytosis) and inhibition of Type I IFN signaling. This review examines the relationship between TAM receptor dysfunction and SS and explores the potential contributions of TAM defects on macrophages to SS development.

The effect of genetic complementation on the fitness and diversity of viruses spreading as collective infectious units

Viruses can spread collectively using different types of structures such as extracellular vesicles, virion aggregates, polyploid capsids, occlusion bodies, and even cells that accumulate virions at their surface, such as bacteria and dendritic cells. Despite the mounting evidence for collective spread, its implications for viral fitness and diversity remain poorly understood. It has been postulated that, by increasing the cellular multiplicity of infection, collective spread could enable mutually beneficial interactions among different viral genetic variants. One such interaction is genetic complementation, whereby deleterious mutations carried by different genomes are compensated. Here, we used simulations to evaluate whether complementation is likely to increase the fitness of viruses spreading collectively. We show that complementation among co-spreading viruses initially buffers the deleterious effects of mutations, but has no positive effect on mean population fitness over the long term, and even promotes error catastrophe at high mutation rates. Additionally, we found that collective spread increases the risk of invasion by social cheaters such as defective interfering particles. We also show that mutation accumulation depends on the type of collective infectious units considered. Co-spreading viral genomes produced in the same cell (e.g. extracellular vesicles, polyploid capsids, occlusion bodies) should exhibit higher genetic relatedness than groups formed extracellularly by viruses released from different cells (aggregates, binding to bacterial or dendritic cell surfaces), and we found that increased relatedness limits the adverse effects of complementation as well cheater invasion risk. Finally, we found that the costs of complementation can be offset by recombination. Based on our results, we suggest that alternative factors promoting collective spread should be considered.

Glycans in the roles of parasitological diagnosis and host-parasite interplay.

The investigation of the glycan repertoire of several organisms has revealed a wide variation in terms of structures and abundance of glycan moieties. Among the parasites, it is possible to observe different sets of glycoconjugates across taxa and developmental stages within a species. The presence of distinct glycoconjugates throughout the life cycle of a parasite could relate to the ability of that organism to adapt and survive in different hosts and environments. Carbohydrates on the surface, and in excretory-secretory products of parasites, play essential roles in host-parasite interactions. Carbohydrate portions of complex molecules of parasites stimulate and modulate host immune responses, mainly through interactions with specific receptors on the surface of dendritic cells, leading to the generation of a pattern of response that may benefit parasite survival. Available data reviewed here also show the frequent aspect of parasite immunomodulation of mammalian responses through specific glycan interactions, which ultimately makes these molecules promising in the fields of diagnostics and vaccinology.

Construction and expression of a single-chain variable fragment antibody against chicken DEC 205 for targeting the bacterial expressed hemagglutinin-neuraminidase of Newcastle disease virus

Targeting antigens to endocytic receptors on the surface of dendritic cells is a new strategy for increasing the adaptive immune response. The objective of the current study was the construction and bacterial expression of a recombinant antibody single-chain fragment variable (ScFv) directed against chicken DEC 205, an endocytic receptor, for use in the genetic fusion of antigens. In particular, we use as antigen the hemagglutinin-neuraminidase (HN) of Newcastle disease virus. Our results show that inoculation of chickens with HN genetically fused to the ScFv anti-DEC 205 induced an evidently higher immune response against HN, in contrast to inoculation with unconjugated HN. In addition, neutralizing antibodies against Newcastle disease virus were detected only in the serum from chickens immunized with HN fused to ScFv anti-DEC 205. Inoculated fused antigens to ScFv against endocytic receptor DEC 205 resulted in a greater antibody-specific anti-HN production compared with antigens applied alone. The results of this study show that the strategy described here has the potential to be used in the development of more effective vaccines against infectious diseases in chickens.

Dendritic cells present cancer-derived p:MHCI molecules to prime anti-tumor CD8 T cells

Abstract Migratory CD103+ dendritic cells (DCs) initiate anti-tumor CD8+ T cell responses by presenting tumor antigens on class I major histocompatibility complex (MHCI) molecules. Given that CD103+ DCs are adept at cross-presenting antigens, the current paradigm is that they present tumor antigens via classical TAP-dependent cross-presentation. However, recent studies have shown that DCs can present cellular antigens by other means, including by acquiring and presenting intact peptide:MHC (p:MHC) complexes from other cells, a mechanism known as cross-dressing. To assess the role of antigen cross-dressing in priming spontaneous anti-tumor CD8+ T cell responses in vivo, the murine MHCI molecule H2-Kb was knocked out in B16 melanoma and C1498 leukemia cell lines. Engraftment of these syngeneic tumors expressing Kb-restricted model antigens into C57BL/6 mice revealed a defect in antigen-specific CD8+ T cell priming when the cancer cells lacked Kb. While the absence of Kb expression by the tumor did not alter the level of costimulatory molecules on DCs or the cytokine milieu of the draining lymph node, it was correlated with a reduction in p:MHCI molecules for a model tumor antigen, SIINFEKL, presented by CD103+ DCs. Further, tumor-derived GFP-tagged Kb molecules were readily observed on the surface of DCs isolated from the tumor, and the presentation of acquired tumor-derived p:MHCI complexes by migratory DCs from tumor-draining lymph nodes of Kb/Db−/− and Tap1−/− mice was sufficient for ex vivo CD8+ T cell priming. Together, these data address a critical knowledge gap in tumor antigen presentation, and highlight a previously under-appreciated pathway as a crucial and non-redundant mechanism of CD8+ T cell activation by DCs.

Profile of Dendritic Cells Surface Biomarkers in Nasal and Oral Mucosa of Human Subjects Reactive to Grass Pollen and House Dust Mites

Abstract Dendritic cells (DCs) play key roles in human allergic pathophysiology at different anatomic sites, such as the respiratory tract. To identify relationships between DCs subpopulations in the upper respiratory tract and atopic status, a public-available dataset (http://doi.org/10.5281/zenodo.50228) on nasal and oral DCs cell-surface biomarkers expression following allergen challenge (through skin prick test) was re-analyzed, with regression and principal component analysis (PCA), to identify profiles of DC cell-surface biomarkers based on the allergen reactivity. The dataset included immunohistochemical measures on CD207 (langerin), CD1a, BDCA-1 to -4 nasal and oral epithelium and lamina propia, as well as demographic information (age, gender), history of respiratory disease, and allergen skin provocation (grass pollen or house dust mite). Subjects with reactivity to grass pollen had different DCs cell-surface biomarker profiles than subjects reactive to house dust mite in both nasal and oral mucosal epitheium and lamina propia. Except for BDCA-3 and 4, reactivity to grass pollen yielded higher expression of DC cell-surface biomarkers in nasal epithelium. In nasal lamina propia, subject reactive to grass pollen had higher expression of BDCA-1 and -2. In the oral epithelium and lamina propia, subjects reactive to house dust mite had higher expression to only CD1a. Graphical evaluation with PCA of both nasal and oral mucosa of DC cell-surface biomarkers yielded principal components explaining three clusters in oral mucosa; two in nasal mucosa. These findings suggest that different allergen activate differently specific innate immune cells, including different DCs subpopulations at different anatomical sites.

Silica Particles Mediate Phenotypic and Functional Alteration of Dendritic Cells and Induce Th2 Cell Polarization.

During silicosis, immune cells, including macrophages, T cells, B cells, and NK cells, participate in fibrosis development through alteration of the immune status. Dendritic cells (DCs) are professional antigen-presenting cells (APCs) with a key role in initiating immune responses and sustaining immune tolerance to maintain homeostasis. The relative contribution of DCs to silicosis progression is not well-documented. In the current study, we investigated the phenotypic and functional alterations of peripheral blood mononuclear cell (PBMC)-derived DCs of Sprague–Dawley (SD) rat during immune responses to silica exposure. We established models for direct and indirect exposure of DCs to silica by either treating DCs with silica or coculturing them with alveolar macrophages (AMs) treated with silica, respectively. The functional activity of DCs was analyzed by measuring their expression of costimulatory molecules, fluorescent microparticle uptake, cytokine production, and ability to mediate T cell polarization in vitro. In vivo, we demonstrated that silica could induce DC migration in response to silica exposure. Our results show that cytokine production by DCs was increased in response to direct silica direct exposure, while indirect silica exposure led to reduced cytokine levels. Moreover, the phagocytic capacity of DCs increased in cocultures after silica exposure. Gene and protein expression analyses showed that silica exposure altered the expression levels of Toll-like receptor pathway proteins and inflammatory factors. DC surface expression of the costimulatory molecules, CD80, CD86, and major histocompatibility complex, was inhibited by exposure to silica, which mediated a Th2-polarizing response in vitro. In rats, silica exposure induced migration of DCs from the peripheral blood into the alveoli. These results demonstrate that direct and indirect exposure to silica particles alter the phenotype and function of DCs, thereby regulating immune responses. Such changes may contribute to the development of silicosis by altering DC phenotype, function, and migration and by influencing the balance between Th1 and Th2 cells.

Amelioration of CIA by Asarinin Is Associated to a Downregulation of TLR9/NF-κB and Regulation of Th1/Th2/Treg Expression.

To study the role of asarinin on collagen-induced arthritis (CIA) and its treatment mechanism on dendritic cells (DCs) and T cells. Before the onset of arthritis, asarinin were given orally to CIA mouse. Macroscopic scoring and micrometer caliper measurement were used to assess arthritis. The occurrence of cartilage destruction and bone erosion were assessed by histology of knee. Sandwich enzyme-linked immunosorbent assay (ELISA) and PCR were used to assess the level of cytokines in hindpaw and arthritic joint. The CD11c MicroBeads were employed to isolate CD11c+ cells from the spleen. Quantitative PCR was used to determine DCs surface molecules of spleen. Macroscopic score and the frequency of arthritis were inhibited by asarinin. Swelling of hindpaws, inflammatory cell infiltration in the synovium, cartilage destruction, and bone erosion were delayed with asarinin. Asarinin treatment suppressed the expression of T helper type 1 (Th1) cytokines and increased the levels of Th2 cytokines (interleukin (IL)-10), transforming growth factor (TGF)-β and Foxp3 in the synovium and hindpaw, however T-bet mRNA levels in synovium decreased. Lower expression of toll-like receptor 9 (TLR9) and nuclear factor-kappaB (NF-κB) were found in DCs after asarinin treatment. There was no difference in the expression of intercellular cell adhesion molecule-1(ICAM-1), OX40-L, and 4-1BBL in spleen DCs between the asarinin group and model control group. Asarinin can treat CIA. TLR9/NF-κB pathway may be involved in the asarinin treatment of CIA by skewing the balance of Th1/Th2/regulatory T (Treg) to a Th2 type.

Role of Programmed Death‐Ligand 1 in Cancer

Programmed death‐1 (PD‐1) is an immunoregulatory receptor on cytotoxic T cells that suppresses the immune system upon interaction with its two ligands, programmed death ligand‐1 (PD‐L1) and programmed death ligand‐2 (PD‐L2). Programmed death‐ligand 1 (PD‐L1), previously known as “CD279”, is made up of 268 amino acids with a crystal structure. It is part of the immunoglobulin superfamily with a transmembrane domain and a tail which contains two phosphorylation sites. PD‐L1 is located on the cell surface of monocytes, B cells, T cells, and dendritic cells, and works in conjunction with PD‐1 to modulate an immune response. In binding with its ligand PD‐1 on an activated T cell, PD‐L1 suppresses the immune response of the T cell, rendering it inactive. PD‐L1's other ligand, the PD‐L2, also plays a role in deregulating the T cell immune response. Many cancer cells express PD‐L1, attenuating immunity and allowing cancer cell proliferation. This distorted relationship provides a focal point for cancer research. Attempts to inhibit the effects of PD‐L1 are concentrated on blocking its interaction to PD‐1, resurrecting the immune response, and thereby allowing CD8+ cytotoxic cells to attack tumor cells. Potential treatments include the development of an anti‐PD‐L1 antibody therapy to induce tumor remissions for patients with different types and stages of cancer. Supporting promising PD‐L1 research, the FDA has approved the PD‐L1 biomarker to accelerate investigation into a potential cure for cancer. In 2018, James P. Allison and Tasuku Hojo received the Nobel Prize in Medicine for research on PD‐L1 that lead to the discovery of checkpoint inhibitors that release immune responses, thereby reactivating T‐cells. The Walton High School MSOE Center for Biomolecular Modeling SMART Team has designed a 3‐D model of PD‐1 in interaction with its ligand PDL‐1 using JMOL to investigate the relationship between the structure and function of the proteins in relation to its current role in cancer development and its future in potential treatment.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The Coincidence Between Increasing Age, Immunosuppression, and the Incidence of Patients With Glioblastoma.

Background: Glioblastoma (GBM) is the most aggressive primary brain tumor in adults and is associated with a median overall survival (mOS) of 16–21 months. Our previous work found a negative association between advanced aging and the survival benefit after treatment with immunotherapy in an experimental brain tumor model. Given the recent phase III clinical success of immunotherapy in patients with many types of cancer, but not for patients with GBM, we hypothesize that aging enhances immunosuppression in the brain and contributes to the lack of efficacy for immunotherapy to improve mOS in patients with malignant glioma. Herein, we compare epidemiological data for the incidence and mortality of patients with central nervous system (CNS) cancers, in addition to immune-related gene expression in the normal human brain, as well as peripheral blood immunological changes across the adult lifespan.Methods: Data were extracted from the National Cancer Institute’s surveillance, epidemiology, and end results (SEER)-, the Broad Institute’s Genotype Tissue Expression project (GTEx)-, and the University of California San Francisco’s 10k Immunomes-databases and analyzed for associations with aging.Results: The proportion of elderly individuals, defined as ≥65 years of age, has predominantly increased for more than 100 years in the United States. Over time, the rise in elderly United States citizens has correlated with an increased incidence and mortality rate associated with primary brain and other CNS cancer. With advanced aging, human mRNA expression for factors associated with immunoregulation including immunosuppressive indoleamine 2,3 dioxygenase 1 (IDO) and programmed death-ligand 1 (PD-L1), as well as the dendritic cell surface marker, CD11c, increase in the brain of normal human subjects, coincident with increased circulating immunosuppressive Tregs and decreased cytolytic CD8+ T cells in the peripheral blood. Strikingly, these changes are maximally pronounced in the 60–69 year old group; consistent with the median age of a diagnosis for GBM.Conclusion: These data demonstrate a significant association between normal human aging and increased immunosuppression in the circulation and CNS; particularly late in life. Our data raise several hypotheses including that, aging: (i) progressively suppresses normal immunosurveillance and thereby contributes to GBM cell initiation and/or outgrowth; (ii) decreases immunotherapeutic efficacy against malignant glioma.