Dendritic Cells In Tolerance Research Articles

CCL22 Induces the Polarization of Immature Dendritic Cells into Tolerogenic Dendritic Cells in Radiation-Induced Lung Injury through the CCR4-Dectin2-PLC-γ2-NFATC2-Nr4a2-PD-L1 Signaling Pathway.

Recruitment of immune cells to the injury site plays a pivotal role in the pathology of radiation-associated diseases. In this study, we investigated the impact of the chemokine CCL22 released from alveolar type II epithelial (AT2) cells after irradiation on the recruitment and functional changes of dendritic cells (DCs) in the development of radiation-induced lung injury (RILI). By examining changes in CCL22 protein levels in lung tissue of C57BL/6N mice with RILI, we discovered that ionizing radiation increased CCL22 expression in irradiated alveolar AT2 cells, as did MLE-12 cells after irradiation. A transwell migration assay revealed that CCL22 promoted the migration of CCR4-positive DCs to the injury site, which explained the migration of pulmonary CCR4-positive DCs in RILI mice invivo. Coculture experiments demonstrated that, consistent with the response of regulatory T cells in the lung tissue of RILI mice, exogenous CCL22-induced DCs promoted regulatory T cell proliferation. Mechanistically, we demonstrated that Dectin2 and Nr4a2 are key targets in the CCL22 signaling pathway, which was confirmed in pulmonary DCs of RILI mice. As a result, CCL22 upregulated the expression of PD-L1, IL-6, and IL-10 in DCs. Consequently, we identified a mechanism in which CCL22 induced DC tolerance through the CCR4-Dectin2-PLC-γ2-NFATC2-Nr4a2-PD-L1 pathway. Collectively, these findings demonstrated that ionizing radiation stimulates the expression of CCL22 in AT2 cells to recruit DCs to the injury site and further polarizes them into a tolerant subgroup of CCL22 DCs to regulate lung immunity, ultimately providing potential therapeutic targets for DC-mediated RILI.

Immunosuppressive microvesicles-mimetic derived from tolerant dendritic cells to target T-lymphocytes for inflammation diseases therapy

The utilization of extracellular vesicles (EV) in immunotherapy, aiming at suppressing peripheral immune cells responsible for inflammation, has demonstrated significant efficacy in treating various inflammatory diseases. However, the clinical application of EV has faced challenges due to their inadequate targeting ability. In addition, most of the circulating EV would be cleared by the liver, resulting in a short biological half-life after systemic administration. Inspired by the natural microvesicles (MV, as a subset of large size EV) are originated and shed from the plasma membrane, we developed the immunosuppressive MV-mimetic (MVM) from endotoxin tolerant dendritic cells (DC) by a straightforward and effective extrusion approach, in which DC surface proteins were inherited for providing the homing ability to the spleen, while αCD3 antibodies were conjugated to the MVM membranes for specific targeting of T cells. The engineered MVM carried a large number of bioactive cargos from the parental cells, which exhibited a remarkable ability to promote the induction of regulatory T cells (Treg) and polarization of anti-inflammatory M2 macrophages. Mechanistically, the elevated Treg level by MVM was mediated due to the upregulation of miR-155-3p. Furthermore, it was observed that systemic and local immunosuppression was induced by MVM in models of sepsis and rheumatoid arthritis through the improvement of Treg and M2 macrophages. These findings reveal a promising cell-free strategy for managing inflammatory responses to infections or tissue injury, thereby maintaining immune homeostasis.

Autoantigenic Peptide and Immunomodulator Codelivery System for Rheumatoid Arthritis Treatment by Reestablishing Immune Tolerance.

Rheumatoid arthritis (RA) is a systemic autoimmune disease characterized by abnormal activation of CD4+ T cells and an imbalance of T helper 17 (Th17) and regulatory T (Treg) cells. Tolerogenic therapy via administration of self-antigens is a promising strategy for RA treatment, but delivery of autoantigens alone may exacerbate disease conditions. Current studies indicated that codelivery of autoantigens with immunomodulators can lead to a more tolerogenic immune response. Here, we constructed an autoantigen type II collagen peptide (CII250-270)- and immunomodulator leflunomide (LEF)-coloaded phosphatidylserine liposome vaccine (CII250-270-LEF-PSL) for RA treatment via induction of tolerant dendritic cells (tolDC) for further activation of Treg cells. The in vivo results showed that CII250-270-LEF-PSL can effectively induce tolDC, regulate the balance of Th1/Th2 and Th17/Treg, and reduce the secretion of pro-inflammatory cytokines (IFN-γ, IL-1β, and IL-17A) and IgG antibodies to inhibit synovial inflammation and bone erosion. Furthermore, our study also suggested that LEF regulated Th1 cell differentiation by inhibiting the activation of the JAK1/STAT1 signaling pathway, further alleviating RA. Overall, this work proved that the combination of autoantigenic peptides and immunomodulators was a promising modality for RA treatment by reestablishing antigen-specific immune tolerance, which also inspired additional insights into the development of combination therapies for the tolerability of RA.

Immunomodulatory and Anti-Inflammatory Effects of Vitamin A and Tryptophan on Monocyte-Derived Dendritic Cells Stimulated with Gliadin in Celiac Disease Patients.

Celiac Disease (CeD) is an autoimmune disorder with various symptoms upon gluten exposure. Dendritic cells (DCs) play a crucial role in gliadin-induced inflammation. Vitamin A (retinol; Ret) and its metabolite, retinoic acid (RA), along with tryptophan (Trp) and its metabolite, kynurenic acid (KYNA), are known to influence the immune function of DCs and enhance their tolerogenicity. This research aims to assess the impact of gliadin on DC maturation and the potential of vitamin A and tryptophan to induce immune tolerance in DCs. The monocyte cells obtained from peripheral blood mononuclear cells (PBMCs) of celiac disease patients were differentiated into DCs in the absence or presence of Ret, RA, Trp, KYNA, and then stimulated with peptic and tryptic (PT) digested of gliadin. We used flow cytometry to analyze CD11c, CD14, HLA-DR, CD83, CD86, and CD103 expression. ELISA was carried out to measure TGF-β, IL-10, IL-12, and TNF-α levels. qRT-PCR was used to assess the mRNA expression of retinaldehyde dehydrogenase 2 (RALDH2) and integrin αE (CD103). The mRNA and protein levels of Indoleamine 2, 3-dioxygenase (IDO) was analyzed by qRT-PCR and Western blot assays, respectively. Our findings demonstrate that PT-gliadin enhances the expression of maturation markers, i.e. CD83, CD86 and HLA-DR and promote the secretion of TNF-α and IL-12 in DCs. Interestingly, vitamin A, tryptophan, and their metabolites increase the expression of CD103, while limiting the expression of HLA-DR, CD83, and CD86. They also enhance RALDH2 and IDO expression and promote the secretion of TGF-β and IL-10, while limiting IL-12 and TNF-α secretion. These findings suggest that vitamin A and tryptophan have beneficial effects on PT-gliadin-stimulated DCs, highlighting their potential as therapeutic agents for celiac disease. However, further research is needed to fully understand their underlying mechanisms of action in these cells.

TDO2-overexpressed Dendritic Cells Possess Tolerogenicity and Ameliorate Collagen-induced Arthritis by Modulating the Th17/Regulatory T Cell Balance.

Tolerogenic dendritic cells are promising for restoring immune homeostasis and may be an alternative therapy for autoimmune diseases such as rheumatoid arthritis. The kynurenine pathway is a vital mechanism that induces tolerance in dendritic cells (DCs). Tryptophan 2,3-dioxygenase (TDO2) is an important rate-limiting enzyme in the kynurenine pathway and participates in immune regulation. However, the role of TDO2 in shaping the tolerogenic phenotypes of DCs remains unclear. In this study, we investigated the effects and mechanisms of TDO2-overexpressed DCs in regulating the T cell balance both invivo and invitro. TDO2-overexpressed DC2.4 and TDO2-/- mouse bone marrow-derived DCs (BMDCs) were generated to verify the role of TDO2 in DC maturation and functionality. TDO2 overexpression in BMDCs via PGE2 treatment exhibited an immature phenotype and tolerogenic state, whereas TDO2-/- BMDCs exhibited a mature phenotype and a proinflammatory state. Furthermore, transplant of TDO2-overexpressed BMDCs alleviated collagen-induced arthritis severity in mice, which was correlated with a reduction in Th17 populations and an increase in regulatory T cells. Collectively, these results indicate that TDO2 plays an important role in the tolerogenic phenotype and may be a promising target for the generation tolerogenic DCs for rheumatoid arthritis treatment.

Matrix metalloproteinase-8 regulates dendritic cell tolerance in late polymicrobial sepsis via the nuclear factor kappa-B p65/β-catenin pathway.

Tolerogenic dendritic cells (DCs) are associated with poor prognosis of sepsis. Matrix metalloproteinases (MMPs) have been shown to have immunomodulatory effects. However, whether MMPs are involved in the functional reprogramming of DCs is unknown. The study aims to investigate the role of MMPs in sepsis-induced DCs tolerance and the potential mechanisms. A murine model of late sepsis was induced by cecal ligation and puncture (CLP). The expression levels of members of the MMP family were detected in sepsis-induced tolerogenic DCs by using microarray assessment. The potential roles and mechanisms underlying MMP8 in the differentiation, maturation and functional reprogramming of DCs during late sepsis were assessed both in vitro and in vivo. DCs from late septic mice expressed higher levels of MMP8, MMP9, MMP14, MMP19, MMP25 and MMP27, and MMP8 levels were the highest. MMP8 deficiency significantly alleviated sepsis-induced immune tolerance of DCs both in vivo and in vitro. Adoptive transfer of MMP8 knockdown post-septic bone marrow-derived DCs protected mice against sepsis-associated lethality and organ dysfunction, inhibited regulatory T-cell expansion and enhanced Th1 response. Furthermore, the effect of MMP8 on DC tolerance was found to be associated with the nuclear factor kappa-B p65/β-catenin pathway. Increased MMP8 levels in septic DCs might serve as a negative feedback loop, thereby suppressing the proinflammatory response and inducing DC tolerance.

Metformin induces tolerogenicity of dendritic cells by promoting metabolic reprogramming

Dendritic cells (DCs) can mediate immune responses or immune tolerance depending on their immunophenotype and functional status. Remodeling of DCs’ immune functions can develop proper therapeutic regimens for different immune-mediated diseases. In the immunopathology of autoimmune diseases (ADs), activated DCs notably promote effector T-cell polarization and exacerbate the disease. Recent evidence indicates that metformin can attenuate the clinical symptoms of ADs due to its anti-inflammatory properties. Whether and how the therapeutic effects of metformin on ADs are associated with DCs remain unknown. In this study, metformin was added to a culture system of LPS-induced DC maturation. The results revealed that metformin shifted DC into a tolerant phenotype, resulting in reduced surface expression of MHC-II, costimulatory molecules and CCR7, decreased levels of proinflammatory cytokines (TNF-α and IFN-γ), increased level of IL-10, upregulated immunomodulatory molecules (ICOSL and PD-L) and an enhanced capacity to promote regulatory T-cell (Treg) differentiation. Further results demonstrated that the anti-inflammatory effects of metformin in vivo were closely related to remodeling the immunophenotype of DCs. Mechanistically, metformin could mediate the metabolic reprogramming of DCs through FoxO3a signaling pathways, including disturbing the balance of fatty acid synthesis (FAS) and fatty acid oxidation (FAO), increasing glycolysis but inhibiting the tricarboxylic acid cycle (TAC) and pentose phosphate pathway (PPP), which resulted in the accumulation of fatty acids (FAs) and lactic acid, as well as low anabolism in DCs. Our findings indicated that metformin could induce tolerance in DCs by reprogramming their metabolic patterns and play anti-inflammatory roles in vitro and in vivo.

Slimming and Reinvigorating Tumor-Associated Dendritic Cells with Hierarchical Lipid Rewiring Nanoparticles.

Dendritic cells (DCs) are crucial mediators of innate and adaptive antitumor immunity, whereas exogenously and endogenously driven lipid accumulation causes immune tolerance of tumor-associated DCs (TADCs) and thereby diminishes tumor responsiveness to various therapies. Herein, a type of multilevel lipid rewiring nanoparticles (NPs) for TADC revitalization is designed. These self-assembled NPs specifically bind to the lipid transport receptor Msr1 on the TADC surface and orchestrate the restriction of extracellular lipid uptake, cytoplasmic de novo lipid biosynthesis and nuclear lipogenic gene transcription. It is found that the slimming of TADCs via the three-in-one lipid metabolic reprogramming substantially promotes their maturation and rehabilitate their functions in inflammatory cytokine production, cytotoxic T cell recruitment, and tumor inhibition. Significantly, tumor resistance to immune checkpoint blockade therapy is further overcome. The study presents a non-canonical strategy to remodel tumor-infiltrating immune cells and paves a new path for improving the efficacy of cancer immunotherapy.

Age-mediated gut microbiota dysbiosis promotes the loss of dendritic cells tolerance.

The old age-related loss of immune tolerance inflicts a person with a wide range of autoimmune and inflammatory diseases. Dendritic cells (DCs) are the sentinels of the immune system that maintain immune tolerance through cytokines and regulatory T-cells generation. Aging disturbs the microbial composition of the gut, causing immune system dysregulation. However, the vis-à-vis role of gut dysbiosis on DCs tolerance remains highly elusive. Consequently, we studied the influence of aging on gut dysbiosis and its impact on the loss of DC tolerance. We show that DCs generated from either the aged (DCOld ) or gut-dysbiotic young (DCDysbiotic ) but not young (DCYoung ) mice exhibited loss of tolerance, as evidenced by their failure to optimally induce the generation of Tregs and control the overactivation of CD4+ T cells. The mechanism deciphered for the loss of DCOld and DCDysbiotic tolerance was chiefly through the overactivation of NF-κB, impaired frequency of Tregs, upregulation in the level of pro-inflammatory molecules (IL-6, IL-1β, TNF-α, IL-12, IFN-γ), and decline in the anti-inflammatory moieties (IL-10, TGF-β, IL-4, IDO, arginase, NO, IRF-4, IRF-8, PDL1, BTLA4, ALDH2). Importantly, a significant decline in the frequency of the Lactobacillus genus was noticed in the gut. Replenishing the gut of old mice with the Lactobacillus plantarum reinvigorated the tolerogenic function of DCs through the rewiring of inflammatory and metabolic pathways. Thus, for the first time, we demonstrate the impact of age-related gut dysbiosis on the loss of DC tolerance. This finding may open avenues for therapeutic intervention for treating age-associated disorders with the Lactobacillus plantarum.

Tumor Suppressor Adenomatous Polyposis Coli Sustains Dendritic Cell Tolerance through IL-10 in a β-Catenin-Dependent Manner.

Dendritic cells (DC) play important roles in balancing immunity and tolerance, in which β-catenin signaling plays an important role, yet the underlying mechanisms remain elusive. In this study, we investigated the functions of the tumor suppressor adenomatous polyposis coli (APC), also a key component of the β-catenin upstream destruction complex in DC. APC depletion in DC does not alter DC and T cell homeostasis under resting conditions. However, APC deficiency in DC leads to attenuated antitumor immunity in mice, which exhibit fewer CD8+ T cells and more Foxp3+ regulatory T cells in tumor and draining lymph nodes. Loss of APC in DC does not affect the expression levels of costimulatory molecules. However, APC-deficient DC produce more IL-10 and exhibit a higher ability of inducing regulatory T cells but a lower ability of priming CD8+ T cells, both of which can be reversed by IL-10 inhibition. Lastly, β-catenin depletion in APC-deficient DC rescues their antitumor immunity and reverses elevated IL-10 production. Taken together, our results identify that APC drives DC tolerance via the β-catenin/IL-10 axis.

NCoR1 controls immune tolerance in conventional dendritic cells by fine-tuning glycolysis and fatty acid oxidation

Dendritic cells (DCs) undergo rapid metabolic reprogramming to generate signal-specific immune responses. The fine control of cellular metabolism underlying DC immune tolerance remains elusive. We have recently reported that NCoR1 ablation generates immune-tolerant DCs through enhanced IL-10, IL-27 and SOCS3 expression. In this study, we did comprehensive metabolic profiling of these tolerogenic DCs and identified that they meet their energy requirements through enhanced glycolysis and oxidative phosphorylation (OXPHOS), supported by fatty acid oxidation-driven oxygen consumption. In addition, the reduced pyruvate and glutamine oxidation with a broken TCA cycle maintains the tolerogenic state of the cells. Mechanistically, the AKT-mTOR–HIF–1α-axis mediated glycolysis and CPT1a-driven β-oxidation were enhanced in these tolerogenic DCs. To confirm these observations, we used synthetic metabolic inhibitors and found that the combined inhibition of HIF-1α and CPT1a using KC7F2 and etomoxir, respectively, compromised the overall transcriptional signature of immunological tolerance including the regulatory cytokines IL-10 and IL-27. Functionally, treatment of tolerogenic DCs with dual KC7F2 and etomoxir treatment perturbed the polarization of co-cultured naïve CD4+ T helper (Th) cells towards Th1 than Tregs, ex vivo and in vivo. Physiologically, the Mycobacterium tuberculosis (Mtb) infection model depicted significantly reduced bacterial burden in BMcDC1 ex vivo and in CD103+ lung DCs in Mtb infected NCoR1DC−/−mice. The spleen of these infected animals also showed increased Th1-mediated responses in the inhibitor-treated group. These findings suggested strong involvement of NCoR1 in immune tolerance. Our validation in primary human monocyte-derived DCs (moDCs) showed diminished NCOR1 expression in dexamethasone-derived tolerogenic moDCs along with suppression of CD4+T cell proliferation and Th1 polarization. Furthermore, the combined KC7F2 and etomoxir treatment rescued the decreased T cell proliferative capacity and the Th1 phenotype. Overall, for the first time, we demonstrated here that NCoR1 mediated control of glycolysis and fatty acid oxidation fine-tunes immune tolerance versus inflammation balance in murine and human DCs.

Methylation changes induced by a glycodendropeptide immunotherapy and associated to tolerance in mice.

Allergen-specific immunotherapy (AIT) is applied as treatment to rise tolerance in patients with food allergies. Although AIT is thoroughly used, the underlying epigenetic events related to tolerant induction are still unknown. Thus, we aim to investigate epigenetic changes that could be related to tolerance in dendritic cells (DCs) from anaphylactic mice to lipid transfer proteins, Pru p 3, in the context of a sublingual immunotherapy (SLIT) with a glycodendropeptide (D1ManPrup3) that has demonstrated tolerant or desensitization responses depending on the treatment dose. Changes in DNA methylation in CpG context were determined comparing Sensitized (Antigen-only) animals and two groups receiving SLIT with the D1ManPrup3 nanostructure (D1ManPrup3-SLIT): Tolerant (2nM D1ManPrup3) and Desensitized (5nM D1ManPrup3), against anaphylactic animals. DNA from lymph nodes-DCs were isolated and then, Whole Genome Bisulphite Sequencing was performed to analyze methylation. Most differentially methylated regions were found on the area of influence of gene promoters (DMPRs). Compared to the Anaphylactic group, the highest value was found in Desensitized mice (n = 7,713 DMPRs), followed by Tolerant (n = 4,091 DMPRs) and Sensitized (n = 3,931 DMPRs) mice. Moreover, many of these epigenetic changes were found in genes involved in immune and tolerance responses (Il1b, Il12b, Il1a, Ifng, and Tnf) as shown by functional enrichment (DCs regulation, B cell-mediated immunity, and effector mechanisms). In conclusion, different doses of D1ManPrup3-SLIT induce different DNA methylation changes, which are reflected in the induction of distinct responses, tolerance, or desensitization.

Interleukin-27 Ameliorates Atherosclerosis in ApoE-/- Mice through Regulatory T Cell Augmentation and Dendritic Cell Tolerance.

Atherosclerosis, which is characterized by chronic inflammation in the arterial wall, is driven by immune cells and cytokines. Recent evidence indicated that interleukin (IL)-27 showed pleiotropic properties in immune diseases. However, precise mechanisms of IL-27, especially in atherosclerosis remains unknown. In our research, we examined the influence of the administration of IL-27 and an anti-IL-27p28 antibody (anti-IL-27p28-Ab) on both the initiation and the progression of atherosclerosis. In the groups (both the initiation and the progression) receiving recombinant IL-27 administration, the formation of atherosclerotic plaques was suspended, and the percentage of regulatory T cells (LAP+ or Foxp3+) in the spleen and peripheral blood was increased. Meanwhile, the number of T helper 1 (Th1) and T helper 17 (Th17) cells was decreased. In the peripheral blood plasma, TGF-β and IL-10 expression were increased, while the levels of IFN-γ and IL-17 were reduced. As for lesions, the mRNA expression of Foxp3, TGF-β, and IL-10 was increased, while that of IFN-γ and IL-17 was reduced. In the anti-IL-27p28 antibody groups, we obtained opposite results. We also observed that DCs treated with IL-27 display a tolerogenic phenotype and that IL-27-treated tolerogenic DCs (tDCs) are likely to play a protective role during atherosclerosis. Our study indicates that IL-27 or adoptive transfer of IL-27 loaded tDCs may be a new therapeutic approach in atherosclerosis.

Echinococcus granulosus Protoscoleces-Derived Exosome-like Vesicles and Egr-miR-277a-3p Promote Dendritic Cell Maturation and Differentiation.

Cystic echinococcosis, a major parasitic disease caused by Echinococcus granulosus, seriously threatens human health. The excretory–secretory (ES) products of E. granulosus can induce immune tolerance in dendritic cells (DCs) to downregulate the host’s immune response; however, the effect of exosomes in the ES products on the DCs has remained unclear. This study showed that E. granulosus protoscoleces-derived exosome-like vesicles (PSC-ELVs) could be internalized by bone marrow-derived dendritic cells (BMDCs), allowing for the delivery of the parasite microRNAs to the BMDCs. Moreover, PSC-ELVs induced BMDCs to produce the proinflammatory cytokinesinterleukin (IL)-6, IL-12, IL-β, tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ). PSC-ELVs also upregulated the BMDCs surface marker major histocompatibility complex class II (MHC II), as well as costimulatory molecules CD40, CD80, and CD86. PSC-ELV-derived egr-miR-277a-3p upregulated the IL-6, IL-12, and TNF-α mRNA levels in BMDCs. Moreover, egr-miR-277a-3p directly targeted Nfkb1 (encoding nuclear factor kappa B 1) to significantly suppress the mRNA and protein levels of NF-κB1 in BMDCs, while the expression of NF-κB p65 significantly increased, suggesting that egr-miR-277a-3p induces the production of proinflammatory cytokines by the modification of the NF-kB p65/p50 ratio in BMDCs. These results demonstrated that PSC-ELVs and egr-miR-277a-3p might enhance DCs maturation and differentiation in a cross-species manner, which in turn may modulate the host immune responses and offer a new approach to echinococcosis prevention and treatment.

Differential Response of Dendritic Cells and Macrophages to Signals from Airway Epithelial Cells in Humans: Changes with Age

Abstract Advancing age significantly enhances the risk of respiratory diseases such as COPD, fibrosis etc. Immune response to respiratory viral infections is also impaired often leading to hospitalization. The underlying mechanisms are not well understood. Airway epithelial cells (AECs) and dendritic cells (DCs) and alveolar macrophages are the primary cells involved in respiratory immunity and tolerance. Since DCs and macrophages exist in close proximity to AECs, the function of both these cells is influenced by AECs. Using a 2D model to study the interaction of human primary bronchial epithelial cells (representative of AECs) with DCs we have recently demonstrated that AECs enhance the immune surveillance capacity of myeloid DCs (mDCs) from the circulation at homeostasis. Culture of mDCs with AECs resulted in the upregulation of pathogen recognition receptors (PRRs) and downstream signaling molecules. In contrast to circulatory mDCs, culture of macrophages with AECs led to a significant inhibition of LPS induced pro-inflammatory responses. More importantly, AECs induced the secretion of TGF-β and IL-10 from macrophages even in the absence of LPS stimulation. Thus, it seems AECs induce tolerance in macrophages but enhance the immunogenicity of mDCs suggesting a division of labor between DCs and macrophages in the respiratory mucosa. Investigation into the possible mechanisms for this differential response indicates that metabolites and growth factors from AECs may be responsible for inducing tolerance in macrophages and priming DCs. We also find that the crosstalk between AECs and macrophages and DCs is altered with age which may contribute to the increased susceptibility of elderly to respiratory diseases and infections. This study was supported by grant from the NIH AG045216 (to AA), and from the National Center for Research Resources and the National Center for Advancing Translational Sciences # UL1 TR000153.

Overexpression of toll-like receptors and co-stimulatory molecules on immature dendritic cells of Crohn's disease

IntroductionCrohn's disease (CD) is a chronic, relapsing inflammatory disorder of the gastrointestinal tract resulting from a loss of homeostasis between the intestinal immune system and the gut microbiome. Also, evidence shows dendritic cells (DC) are the main antigen-presenting cells (APC) in the intestinal mucosa; they can regulate innate and adaptive immunity that is pivotal for maintaining immunological tolerance in the gut. The maturation of immature DC (iDCs) regulate by their surface molecules such as Toll-Like Receptors (TLRs) and co-stimulatory molecules (CD80 and CD86). The level of TLRs on iDCs plays a critical role in developing adaptive responses to antigens. Crohn's disease has a dysfunctional immune system and low tolerance of dendritic cells. This study aimed to investigate the expression of TLR2, TLR4, TLR5, and TLR9 and co-stimulatory molecules (CD80 and CD86). MethodologyHuman peripheral blood mononuclear cells (PBMCs) were collected from 15 CD patients and 12 healthy controls. PBMCs were isolated by density gradient centrifugation, and then iDCs were derived for 6 days. Flow cytometry was performed on iDCs to characterize and distinguish the phenotype of iDC populations. iDCs were harvested for RNA extraction, and the mRNA expression level of TLRs was examined by quantitative RT-PCR using SYBR Green chemistry. ResultThe results showed that TLR2, TLR4, TLR5, and TLR9 mRNA expression significantly increased in the iDCs isolated from patients with CD compared with healthy control P ≤ 0.0001. Although, TLR2 had the lowest expression among other TLRs on iDCs in CD patients. Also, the expression level of CD80 and CD86 were significantly higher in iDCs isolated from CD patients compared to healthy controls P < 0.001 OR = 0.75 CI (−8.76 to −3.17). ConclusionTLR2, TLR4, TLR5, and TLR9 expression on iDCs may be critical in the biological pathogenesis of CD. TLR alterations in the iDCs may contribute to determining dendritic cell maturation pathway lead to increases inflammatory response.

Enhanced uptake and anti-maturation effect of celastrol-loaded mannosylated liposomes on dendritic cells for psoriasis treatment

Psoriasis is an autoimmune skin disease in which dendritic cells (DCs) trigger the progression of psoriasis by complex interactions with keratinocytes and other immune cells. In the present study, we aimed to load celastrol, an anti-inflammatory ingredient isolated from Chinese herbs, on mannosylated liposomes to enhance DC uptake as well as to induce DC tolerance in an imiquimod-induced psoriasis-like mouse model. Mannose was grafted onto liposomes to target mannose receptors on DCs. The results demonstrated that compared with unmodified liposomes, DCs preferred to take up more fluorescence-labeled mannosylated liposomes. After loading celastrol into mannose-modified liposomes, they effectively inhibited the expression of maturation markers, including CD80, CD86 and MHC-II, on DCs both in vitro and in vivo. Additionally, after intradermal injection with a microneedle, celastrol-loaded mannose-modified liposomes (CEL-MAN-LPs) achieved a superior therapeutic effect compared with free drug and celastrol-loaded unmodified liposomes in the psoriasis mouse model in terms of the psoriasis area and severity index, histology evaluation, spleen weight, and expression of inflammatory cytokines. In conclusion, our results clearly revealed that CEL-MAN-LPs was an effective formulation for psoriasis treatment and suggested that this treatment has the potential to be applied to other inflammatory diseases triggered by activated DCs.

Rapamycin treated tol-dendritic cells derived from BM-MSCs reversed graft rejection in a rat liver transplantation model by inducing CD8+CD45RC−Treg

ObjectiveTo investigate the influence of tolerance dendritic cells (tolDCs), generated from Bone marrow mesenchymal stem cells (BM-MSCs) treated with rapamycin (Rapa) on liver allograft survival in a rat acute liver transplantation model. MethodsDifferent GM-CSF induction project was used to obtain immature DCs (imDCs), mature DCs (matDCs) or tolDCs from BM-MSCs. First, MLR was performed to analyze the activity of tolDCs on polyclonaly stimulated total T cells. Then, co-cultured imDCs, matDCs and tolDCs with CD8+T cells isolated by magnetic activated cell sorting to analyze the influence on its regulatory characteristic. Last, the established rat acute liver transplantation model were adoptive transfused with imDCs, matDCs or tolDCs isolated by anti-CD11c immunomagnetic beads. The phenotype of DC cells and level of CD8+Treg in the culture system and in vivo, the expression of CD8 and CD45RC in the tissues were analyzed by flow cytometry and immunohistochemistry, respectively. ResultsThe loGM-CSF plus IL-4 decreased the costimulatory molecules of CD80/86 and MHC class II of DCs comparison with hiGM-CSF from BM-MSCs no matter whether stimulation by LPS (P<0.05). Rapa treated not only reduced the expression of CD80/86 and MHC class II but also down-regulated the expression of CD11c after LPS stimulation which was more obviously in tolDCs by loGM-CSF project (P<0.05). Moreover, tolDCs displayed a rather higher level of IL-10 and low level of IL-12p70 than others (P<0.01), which shown a rather lower stimulative effect on the proliferation of T cells comparison with matDCs and imDCs. Co-cultured with CD8+Treg showed an improvement on induction of CD8+TCR+CD45RC−T cells (CD8+Treg) in ex vivo. The rats transfused with tolDCs has a delayed survival benefits with high level of CD8+Tregs (P<0.01) and high expression of CD45RC in liver tissue (P<0.01) and spleen when comparison with other groups. The infused tolDCs improved a mean survival time (MST) of 32 days comparison with a MTS of 9.5 days and 15.75 days displayed by rat that per-infused with matDCs and imDCs, respectively. ConclusionRapa modified tolDCs derived from BM-MSCs reversed graft rejection by improve tolerance characteristics of CD8+CD45RC−Treg in acute liver rat transplantation.

Pharmacological Wnt ligand inhibition overcomes key tumor-mediated resistance pathways to anti-PD-1 immunotherapy.

SUMMARYWhile immune checkpoint blockade is associated with prolonged responses in multiple cancers, most patients still do not benefit from this therapeutic strategy. The Wnt-β-catenin pathway is associated with diminished T cell infiltration; however, activating mutations are rare, implicating a role for autocrine/paracrine Wnt ligand-driven signaling in immune evasion. In this study, we show that proximal mediators of the Wnt signaling pathway are associated with anti-PD-1 resistance, and pharmacologic inhibition of Wnt ligand signaling supports anti-PD-1 efficacy by reversing dendritic cell tolerization and the recruitment of granulocytic myeloid-derived suppressor cells in autochthonous tumor models. We further demonstrate that the inhibition of Wnt signaling promotes the development of a tumor microenvironment that is more conducive to favorable responses to checkpoint blockade in cancer patients. These findings support a rationale for Wnt ligand-focused treatment approaches in future immunotherapy clinical trials and suggest a strategy for selecting those tumors more responsive to Wnt inhibition.

Tolerogenic Immunotherapy: Targeting DC Surface Receptors to Induce Antigen-Specific Tolerance.

Dendritic cells (DCs) are well-established as major players in the regulation of immune responses. They either induce inflammatory or tolerogenic responses, depending on the DC-subtype and stimuli they receive from the local environment. This dual capacity of DCs has raised therapeutic interest for their use to modify immune-activation via the generation of tolerogenic DCs (tolDCs). Several compounds such as vitamin D3, retinoic acid, dexamethasone, or IL-10 and TGF-β have shown potency in the induction of tolDCs. However, an increasing interest exists in defining tolerance inducing receptors on DCs for new targeting strategies aimed to develop tolerance inducing immunotherapies, on which we focus particular in this review. Ligation of specific cell surface molecules on DCs can result in antigen presentation to T cells in the presence of inhibitory costimulatory molecules and tolerogenic cytokines, giving rise to regulatory T cells. The combination of factors such as antigen structure and conformation, delivery method, and receptor specificity is of paramount importance. During the last decades, research provided many tools that can specifically target various receptors on DCs to induce a tolerogenic phenotype. Based on advances in the knowledge of pathogen recognition receptor expression profiles in human DC subsets, the most promising cell surface receptors that are currently being explored as possible targets for the induction of tolerance in DCs will be discussed. We also review the different strategies that are being tested to target DC receptors such as antigen-carbohydrate conjugates, antibody-antigen fusion proteins and antigen-adjuvant conjugates.