Mast Cells Differentiated in Synovial Fluid and Resident in Osteophytes Exalt the Inflammatory Pathology of Osteoarthritis.

INTRODUCTION: Osteophytes are a prominent feature of osteoarthritis (OA) joints and one of the clinical hallmarks of the disease progression. Research on osteophytes is fragmentary and modes of its contribution to OA pathology are obscure. AIM: To elucidate the role of osteophytes in OA pathology from a perspective of molecular and cellular events. METHODS: RNA-seq of fully grown osteophytes, collected from tibial plateau of six OA patients revealed patterns corresponding to active extracellular matrix re-modulation and prominent participation of mast cells. Presence of mast cells was further confirmed by immunohistochemistry, performed on the sections of the osteophytes using anti-tryptase alpha/beta-1 and anti-FC epsilon RI antibodies and the related key up-regulated genes were validated by qRT-PCR. To test the role of OA synovial fluid (SF) in mast cell maturation as proposed by the authors, hematopoietic stem cells (HSCs) and ThP1 cells were cultured in a media supplemented with 10% SF samples, obtained from various grades of OA patients and were monitored using specific cell surface markers by flow cytometry. Proteomics analysis of SF samples was performed to detect additional markers specific to mast cells and inflammation that drive the cell differentiation and maturation. RESULTS: Transcriptomics of osteophytes revealed a significant upregulation of mast cells specific genes such as chymase 1 (CMA1; 5-fold) carboxypeptidase A3 (CPA3; 4-fold), MS4A2/FCERI (FCERI; 4.2-fold) and interleukin 1 receptor-like 1 (IL1RL1; 2.5-fold) indicating their prominent involvement. (In IHC, anti-tryptase alpha/beta-1 and anti- FC epsilon RI-stained active mast cells were seen populated in cartilage, subchondral bone, and trabecular bone.) Based on these outcomes and previous learnings, the authors claim a possibility of mast cells invasion into osteophytes is mediated by SF and present in vitro cell differentiation assay results, wherein ThP1 and HSCs showed differentiation into HLA-DR+/CD206+ and FCERI+ phenotype, respectively, after exposing them to medium containing 10% SF for 9 days. Proteomics analysis of these SF samples showed an accumulation of mast cell-specific inflammatory proteins. CONCLUSIONS: RNA-seq analysis followed by IHC study on osteophyte samples showed a population of mast cells resident in them and may further accentuate inflammatory pathology of OA. Besides subchondral bone, the authors propose an alternative passage of mast cells invasion in osteophytes, wherein OA SF was found to be necessary and sufficient for maturation of mast cell precursor into effector cells.

Decoding the intricacies of the mast cell compartment.

Historically, understanding of the human mast cell (MC) compartment has lagged behind the appreciation of other cell lineages. MCs exist in vascularised tissues but do not under normal circumstances circulate in blood, and there has been no pharmacological agent identified that totally and selectively inhibits human MC function. There are no substantiated accounts of an apparently healthy individual who is severely lacking in MCs. Thus, some of the approaches employed to understand the function of a specific immune cell are not available to the MC biologist. The disease categories that have provided the greatest insight into MC biology have been monoclonal and IgE-mediated MC disorders. This has led to the categorisation of MC diseases as intrinsic or extrinsic to the MC compartment and to the recognition of the role of mediators in MC activation disorders. Mastocytosis as a clonal disorder not only impacts the MC compartment through changes intrinsic to the MC, but also by the effects of episodes of significant release of MC mediators. The availability of newer therapeutic approaches developed to treat monoclonal MC disorders offer insights into how to more selectively approach management of MC centric diseases.

Ultramicronized Palmitoylethanolamide in the Management of Sepsis-Induced Coagulopathy and Disseminated Intravascular Coagulation.

Disseminated intravascular coagulation (DIC) is a severe condition characterized by the systemic formation of microthrombi complicated with bleeding tendency and organ dysfunction. In the last years, it represents one of the most frequent consequences of coronavirus disease 2019 (COVID-19). The pathogenesis of DIC is complex, with cross-talk between the coagulant and inflammatory pathways. The objective of this study is to investigate the anti-inflammatory action of ultramicronized palmitoylethanolamide (um-PEA) in a lipopolysaccharide (LPS)-induced DIC model in rats. Experimental DIC was induced by continual infusion of LPS (30 mg/kg) for 4 h through the tail vein. Um-PEA (30 mg/kg) was given orally 30 min before and 1 h after the start of intravenous infusion of LPS. Results showed that um-PEA reduced alteration of coagulation markers, as well as proinflammatory cytokine release in plasma and lung samples, induced by LPS infusion. Furthermore, um-PEA also has the effect of preventing the formation of fibrin deposition and lung damage. Moreover, um-PEA was able to reduce the number of mast cells (MCs) and the release of its serine proteases, which are also necessary for SARS-CoV-2 infection. These results suggest that um-PEA could be considered as a potential therapeutic approach in the management of DIC and in clinical implications associated to coagulopathy and lung dysfunction, such as COVID-19.

Identification of key genes and pathways associated with resting mast cells in meningioma.

BACKGROUND: To identify candidate key genes and pathways related to resting mast cells in meningioma and the underlying molecular mechanisms of meningioma. METHODS: Gene expression profiles of the used microarray datasets were obtained from the Gene Expression Omnibus (GEO) database. GO and KEGG pathway enrichments of DEGs were analyzed using the ClusterProfiler package in R. The protein-protein interaction network (PPI), and TF-miRNA- mRNA co-expression networks were constructed. Further, the difference in immune infiltration was investigated using the CIBERSORT algorithm. RESULTS: A total of 1499 DEGs were identified between tumor and normal controls. The analysis of the immune cell infiltration landscape showed that the probability of distribution of memory B cells, regulatory T cells (Tregs), and resting mast cells in tumor samples were significantly higher than those in the controls. Moreover, through WGCNA analysis, the module related to resting mast cells contained 158 DEGs, and KEGG pathway analysis revealed that the DEGs were dominant in the TNF signaling pathway, cytokine-cytokine receptor interaction, and IL-17 signaling pathway. Survival analysis of hub genes related to resting mast cells showed that the risk model was constructed based on 9 key genes. The TF-miRNA- mRNA co-regulation network, including MYC-miR-145-5p, TNFAIP3-miR-29c-3p, and TNFAIP3-hsa-miR-335-3p, were obtained. Further, 36 nodes and 197 interactions in the PPI network were identified. CONCLUSION: The results of this study revealed candidate key genes, miRNAs, and pathways related to resting mast cells involved in meningioma development, providing potential therapeutic targets for meningioma treatment.

Sex differences in anatomic plasticity of gut neuronal-mast cell interactions.

The gut wall houses mast cells that are anatomically situated near enteric neuronal fibers. Roles of specific neuropeptides in modulating function of immune components like mast cells in response to challenge with bacterial components are relatively unknown. Investigating such interactions requires models that include diverse cellular elements in native anatomic arrangements. Using an organotypic slice model that maintains gut wall cellular diversity ex vivo, the present study compared responses between tissues derived from male and female mice to examine neural-immune signaling in the gut wall after selected treatments. Ileum slices were treated with pharmacological reagents that block neuronal function (e.g., tetrodotoxin) or vasoactive intestinal peptide (VIP) receptors prior to challenge with lipopolysaccharide (LPS) to assess their influence on anatomic plasticity of VIP fibers and activation of mast cells. Sex differences were observed in the number of mucosal mast cells (c-kit/ACK2 immunoreactive) at baseline, regardless of treatment, with female ileum tissue having 46% more ACK2-IR mast cells than males. After challenge with LPS, male mast cell counts rose to female levels. Furthermore, sex differences were observed in the percentage of ACK2-IR cells within 1 µm of a VIP+ neuronal fiber, and mast cell size, a metric previously tied to activation, with females having larger cells at baseline. Male mast cell sizes reached female levels after LPS challenge. This study suggests sex differences in neural-immune plasticity and in mast cell activation both basally and in response to challenge with LPS. These sex differences could potentially impact functional neuroimmune response to pathogens.

The Evolutionary History of the Chymase Locus -a Locus Encoding Several of the Major Hematopoietic Serine Proteases.

Several hematopoietic cells of the immune system store large amounts of proteases in cytoplasmic granules. The absolute majority of these proteases belong to the large family of chymotrypsin-related serine proteases. The chymase locus is one of four loci encoding these granule-associated serine proteases in mammals. The chymase locus encodes only four genes in primates, (1) the gene for a mast-cell-specific chymotryptic enzyme, the chymase; (2) a T-cell-expressed asp-ase, granzyme B; (3) a neutrophil-expressed chymotryptic enzyme, cathepsin G; and (4) a T-cell-expressed chymotryptic enzyme named granzyme H. Interestingly, this locus has experienced a number of quite dramatic expansions during mammalian evolution. This is illustrated by the very large number of functional protease genes found in the chymase locus of mice (15 genes) and rats (18 genes). A separate expansion has also occurred in ruminants, where we find a new class of protease genes, the duodenases, which are expressed in the intestinal region. In contrast, the opossum has only two functional genes in this locus, the mast cell (MC) chymase and granzyme B. This low number of genes may be the result of an inversion, which may have hindered unequal crossing over, a mechanism which may have been a major factor in the expansion within the rodent lineage. The chymase locus can be traced back to early tetrapods as genes that cluster with the mammalian genes in phylogenetic trees can be found in frogs, alligators and turtles, but appear to have been lost in birds. We here present the collected data concerning the evolution of this rapidly evolving locus, and how these changes in gene numbers and specificities may have affected the immune functions in the various tetrapod species.

Anti-Allergic and Anti-Inflammatory Effects of Neferine on RBL-2H3 Cells.

Mast cells play a very important role in skin allergy and inflammation, including atopic dermatitis and psoriasis. In the past, it was found that neferine has anti-inflammatory and anti-aging effects on the skin, but its effect on mast cells has not yet been studied in detail. In this study, we used mast cells (RBL-2H3 cells) and mouse models to study the anti-allergic and inflammatory effects of neferine. First, we found that neferine inhibits the degranulation of mast cells and the expression of cytokines. In addition, we observed that when mast cells were stimulated by A23187/phorbol 12-myristate-13-acetate (PMA), the elevation of intracellular calcium was inhibited by neferine. The phosphorylation of the MAPK/NF-κB pathway is also reduced by pretreatment of neferine. The results of in vivo studies show that neferine can improve the appearance of dermatitis and mast cell infiltration caused by dinitrochlorobenzene (DNCB). Moreover, the expressions of barrier proteins in the skin are also restored. Finally, it was found that neferine can reduce the scratching behavior caused by compound 48/80. Taken together, our results indicate that neferine is a very good anti-allergic and anti-inflammatory natural product. Its effect on mast cells contributes to its pharmacological mechanism.

Basophils and Mast Cells in COVID-19 Pathogenesis.

Basophils and mast cells are among the principal inducers of Th2 responses and have a crucial role in allergic and anti-parasitic protective immunity. Basophils can function as antigen-presenting cells that bind antigens on their surface and boost humoral immune responses, inducing Th2 cell differentiation. Their depletion results in lower humoral memory activation and greater infection susceptibility. Basophils seem to have an active role upon immune response to SARS-CoV-2. In fact, a coordinate adaptive immune response to SARS-CoV-2 is magnified by basophils. It has been observed that basophil amount is lower during acute disease with respect to the recovery phase and that the grade of this depletion is an important determinant of the antibody response to the virus. Moreover, mast cells, present in a great quantity in the nasal epithelial and lung cells, participate in the first immune response to SARS-CoV-2. Their activation results in a hyperinflammatory syndrome through the release of inflammatory molecules, participating to the "cytokine storm" and, in a longer period, inducing pulmonary fibrosis. The literature data suggest that basophil counts may be a useful prognostic tool for COVID-19, since their reduction is associated with a worse prognosis. Mast cells, on the other hand, represent a possible therapeutic target for reducing the airway inflammation characteristic of the hyperacute phase of the disease.

[Elucidation of Mast Cell Activation Mechanism Mediated by Purinergic Signaling].

Mast cells (MCs) are immune cells that are distributed in all tissues throughout the body, and their cytoplasm is rich in granules containing histamine and tryptase. When MCs recognize antigens through IgE bound to FcεRI, they release these mediators by degranulation. Because degranulation induces various type I allergic reactions, such as anaphylactic shock and hay fever, elucidation of the control mechanism of degranulation is important to the development of a therapeutic strategy for allergic diseases. It is known that the antigen-induced degranulation response is fine-tuned by various humoral factors via the activation of G protein-coupled receptors. We found that extracellular ATP enhanced antigen-dependent and -independent MC degranulation via activation of ionotropic P2X4 receptors. P2X4 receptor activation itself had no effect on MC degranulation, but significantly enhanced antigen-triggered degranulation. Stimulation of the P2X4 receptor potentiated the FcεRI-mediated tyrosine kinase signaling cascade. In addition to antigen-induced responses, P2X4 receptor signaling also affected antigen-independent MC responses. Thus, co-stimulation of ATP and Gi-coupled receptor agonists, such as prostaglandin E2 (PGE2) and adenosine, resulted in synergistic degranulation. The significance of P2X4 receptor signaling in allergic and inflammatory responses in vivo was confirmed by impaired responses of antigen-induced passive anaphylaxis and PGE2-induced increases in vascular permeability in P2rx4 knockout mice compared to that of wild-type mice. These results suggest that the P2X4 receptor is a potential therapeutic target for both antigen-dependent and -independent allergic reactions.

Role of mast cells in oral inflammation.

Mast cells play important roles in the maintenance of many physiological functions as well as in the pathophysiology of diseases. Mast cells are involved in the inflammatory mechanisms of many systemic diseases. In this pandemic period, their role in physiological and pathological host inflammatory reactions in tissue disruption following SARS-CoV-2 infection has been stressed. A review of the literature was carried out by entering the key words "Mast Cells" AND "Oral Diseases" AND "Role of Mast Cells in Periodontitis". The results show us that mast cells are definitely involved in many oral diseases including periodontitis. Further in vivo and in vitro studies are needed to further investigate the specific role of the cells in physiological and pathological inflammation.

Mast cell tryptase enhances wound healing by promoting migration in human bronchial epithelial cells.

Epithelial damage and increase of intraepithelial mast cells (MC) are characteristics of asthma. The role of MC mediator tryptase and the protease-activated receptor-2 (PAR2) on epithelial wound healing is not fully investigated. Stimulation of bronchial epithelial cells (BECs) with tryptase promoted gap closure, migration and cellular speed compared to controls. Stimulated BECs had higher expression of migration marker CD151 compared to controls. Proliferation marker KI67 was upregulated in tryptase-stimulated BECs compared to controls. Treatment with PAR2 antagonist I-191 reduced gap closure, migration and cell speed compared to BECs stimulated with tryptase. We found that tryptase enhances epithelial wound healing by increased migration and proliferation, which is in part regulated via PAR2. Our data suggest that tryptase might be beneficial in tissue repair under baseline conditions. However, in a pathological context such as asthma with increased numbers of activated MCs, it might lead to epithelial remodeling and loss of function.

Use of omalizumab for management of idiopathic anaphylaxis: A systematic review and retrospective case series.

BACKGROUND: Patients with idiopathic anaphylaxis (IA) may fail to respond to a combination of high-dose H1 and H2 antihistamines and mast cell stabilizers. Treatment options for these patients are currently limited. OBJECTIVE: To describe the clinical experience of omalizumab use for the treatment of patients with IA with no evidence of underlying clonal mast cell disorders. METHODS: We performed a retrospective review at 2 separate institutions of medical records of patients with a diagnosis of IA without evidence of mast cell clonality who had received treatment with omalizumab. We searched PubMed for studies describing omalizumab use in similar patients. Information on symptoms and omalizumab therapy was compiled, and response pattern of anaphylaxis was determined. RESULTS: A total of 35 patients with IA and no evidence of mast cell clonality who received omalizumab were identified. The median age was 36 years at the start of omalizumab (range, 11-54 years; n = 29). The frequency of anaphylaxis episodes before omalizumab treatment varied from 2 total episodes to several episodes per month. The most often used initial omalizumab dose was 300 mg every 4 weeks (n = 16). Most patients ultimately achieved clinical response after starting omalizumab: complete response (63%, n = 22), partial response (28.5%, n = 10), with 3 nonresponders. CONCLUSION: Omalizumab may be an effective treatment option for patients with IA who do not have evidence of mast cell clonality and fail to respond to antihistamines and mast cell stabilizers.

PKCε controls the fusion of secretory vesicles in mast cells in a phosphatidic acid-dependent mode.

PKCε is highly expressed in mast cells and plays a fundamental role in the antigen-triggered activation of the allergic reaction. Although its regulation by diacylglycerols has been described, its regulation by acidic phospholipids and how this regulation leads to the control of downstream vesicle secretion is barely known. Here, we used structural and evolutionary studies to find the molecular mechanism that explains the selectivity of the C1B domain of PKCε by Phosphatidic Acid (PA). This resided in a collection of Arg residues that form a specific rim on the outer surface of the C1B domain, around the diacylglycerol binding cleft. In RBL-2H3 cells, this basic rim allowed the kinase to respond specifically to phosphatidic acid signals that induced its translocation to the plasma membrane and subsequent activation. Further experiments in cells that overexpress PKCε and a mutant of the PA binding site, showed that PA-dependent PKCε activation increased vesicle degranulation in RBL-2H3 cells, and this correlated with increased SNAP23 phosphorylation. Over-expression of PKCε in these cells also induced an increase in the number of docked vesicles containing SNAP23, when stimulated with PA. This accumulation could be attributed to the stabilizing effect of phosphorylation on the formation of the SNARE complex, which ultimately led to increased release of content in the presence of Ca2+ during the fusion process. Therefore, these findings reinforce the importance of PA signaling in the activation of PKCε, which could be an important target to inhibit the exacerbated responses of these cells in the allergic reaction.

Bone marrow derived mast cells injected into the osteoarthritic knee joints of mice induced by sodium monoiodoacetate enhanced spontaneous pain through activation of PAR2 and action of extracellular ATP.

Conditions that resemble osteoarthritis (OA) were produced by injection of sodium monoiodoacetate (MIA) into the knee joints of mice. Bone marrow derived mast cells (BMMCs) injected into the OA knee joints enhanced spontaneous pain. Since no spontaneous pain was observed when BMMCs were injected into the knee joints of control mice that had not been treated with MIA, BMMCs should be activated within the OA knee joints and release some pain-inducible factors. Protease activated receptor-2 (PAR2) antagonist (FSLLRY-NH2) almost abolished the pain-enhancing effects of BMMCs injected into the OA knee joints, suggesting that tryptase, a mast cell protease that is capable of activating PAR2, should be released from the injected BMMCs and enhance pain through activation of PAR2. When PAR2 agonist (SLIGKV-NH2) instead of BMMCs was injected into the OA knee joints, it was also enhanced pain. Apyrase, an ATP degrading enzyme, injected into the OA knee joints before BMMCs suppressed the pain enhanced by BMMCs. We showed that purinoceptors (P2X4 and P2X7) were expressed in BMMCs and that extracellular ATP stimulated the release of tryptase from BMMCs. These observations suggest that ATP may stimulate degranulation of BMMCs and thereby enhanced pain. BMMCs injected into the OA knee joints stimulated expression of IL-1ß, IL-6, TNF-α, CCL2, and MMP9 genes in the infrapatellar fat pads, and PAR2 antagonist suppressed the stimulatory effects of BMMCs. Our study suggests that intermittent pain frequently observed in OA knee joints may be due, at least partly, to mast cells through activation of PAR2 and action of ATP, and that intraarticular injection of BMMCs into the OA knee joints may provide a useful experimental system for investigating molecular mechanisms by which pain is induced in OA knee joints.

Mast Cells: A New Frontier for Cancer Immunotherapy.

Mast cells are unique tissue-resident immune cells of the myeloid lineage that have long been implicated in the pathogenesis of allergic and autoimmune disorders. More recently, mast cells have been recognized as key orchestrators of anti-tumor immunity, modulators of the cancer stroma, and have also been implicated in cancer cell intrinsic properties. As such, mast cells are an underrecognized but very promising target for cancer immunotherapy. In this review, we discuss the role of mast cells in shaping cancer and its microenvironment, the interaction between mast cells and cancer therapies, and strategies to target mast cells to improve cancer outcomes. Specifically, we address (1) decreasing cell numbers through c-KIT inhibition, (2) modulating mast cell activation and phenotype (through mast cell stabilizers, FcεR1 signaling pathway activators/inhibitors, antibodies targeting inhibitory receptors and ligands, toll like receptor agonists), and (3) altering secreted mast cell mediators and their downstream effects. Finally, we discuss the importance of translational research using patient samples to advance the field of mast cell targeting to optimally improve patient outcomes. As we aim to expand the successes of existing cancer immunotherapies, focused clinical and translational studies targeting mast cells in different cancer contexts are now warranted.

IL-10 in Mast Cell-Mediated Immune Responses: Anti-Inflammatory and Proinflammatory Roles.

Mast cells (MCs) play critical roles in Th2 immune responses, including the defense against parasitic infections and the initiation of type I allergic reactions. In addition, MCs are involved in several immune-related responses, including those in bacterial infections, autoimmune diseases, inflammatory bowel diseases, cancers, allograft rejections, and lifestyle diseases. Whereas antigen-specific IgE is a well-known activator of MCs, which express FcεRI on the cell surface, other receptors for cytokines, growth factors, pathogen-associated molecular patterns, and damage-associated molecular patterns also function as triggers of MC stimulation, resulting in the release of chemical mediators, eicosanoids, and various cytokines. In this review, we focus on the role of interleukin (IL)-10, an anti-inflammatory cytokine, in MC-mediated immune responses, in which MCs play roles not only as initiators of the immune response but also as suppressors of excessive inflammation. IL-10 exhibits diverse effects on the proliferation, differentiation, survival, and activation of MCs in vivo and in vitro. Furthermore, IL-10 derived from MCs exerts beneficial and detrimental effects on the maintenance of tissue homeostasis and in several immune-related diseases including contact hypersensitivity, auto-immune diseases, and infections. This review introduces the effects of IL-10 on various events in MCs, and the roles of MCs in IL-10-related immune responses and as a source of IL-10.

Does Sugammadex Suppress Allergic Inflammation Due to Rocuronium in Animal Model of Rat?

INTRODUCTION AND OBJECTIVES: There are a few reports in the literature about the successful use of sugammadex in the treatment of hypersensitivity reactions caused by rocuronium; however, the pathophysiological mechanism is still unknown. This study aims to investigate the changes caused by rocuronium in the lung and the effect of sugammadex on these changes with biochemical, light microscopic and immunohistochemical parameters on a rat model. MATERIALS AND METHODS: For the study, 28-male Sprague-Dawley rats were randomly divided, seven of each, into four groups. Group C (control) received only 0. 9 % NaCl without any drug. Group R received rocuronium alone 1mg/kg. Group S received sugammadex alone 96 mg/kg. Group RS received rocuronium 1mg/kg and sugammadex 96 mg/kg. After 24 h later, the animals were sacrificed and their tissues were removed. Biochemical (IgE/CRP), light microscopic and immunohistochemical findings were recorded. RESULTS: Immunoglobulin E and CRP levels, peribronchial, alveolar septal lymphocytic infiltration, thickening of the alveolar membranes and bleeding sites in Group R were significantly higher than all the other groups. In Group RS, while these parameters were significantly lower than that of Group R and Group S, it was significantly higher than that of Group C. Total mast cells and tryptase-positive mast cells counts were significantly higher in Group R than in all other groups. In Group RS, these parameters were statistically lower than that of Group R and Group S, but higher than that of Group C. CONCLUSIONS: This study shows that allergic inflammatory changes due to rocuronium in the lungs of rats are reduced with sugammadex. These results support cases of anaphylaxis due to rocuronium which improved with sugammadex.

Selective expression of a C-type lectin receptor, Clec12b, on skin mast cells.

Mast cells (MCs) are present in various organs including the skin, peritoneal cavity, lung, and intestine and involved in the development of allergic diseases and host defense against infection. However, the regulatory mechanism of mast cell activation remains incompletely understood. We found in a database that Clec12b encoding a C-type lectin receptor Clec12b is preferentially expressed in skin MCs in mice. However, neither MCs in other tissues such as trachea, tongue, esophagus, or peritoneal cavity nor most lymphocytes and myeloid cells express Clec12b. To analyze the protein expression of Clec12b, we newly generated a monoclonal antibody (named TX109), which recognizes both mouse and human Clec12b. Consistent with the gene expression profile, flow cytometry analysis demonstrated that Clec12b is expressed only on MCs in the skin, but not on any other immune cell types in various tissues, in mice. Similarly, Clec12b is also expressed on skin MCs, but not on circulating lymphocytes and myeloid cells, in humans. Our results suggest that Clec12b plays an important role in the regulation of MCs activation in the skin.

Thapsigargin-Stimulated LAD2 Human Mast Cell Line Is a Potent Cellular Adjuvant for the Maturation of Monocyte-Derived Dendritic Cells for Adoptive Cellular Immunotherapy.

The preparation of dendritic cells (DCs) for adoptive cellular immunotherapy (ACI) requires the maturation of ex vivo-produced immature(i) DCs. This maturation ensures that the antigen presentation triggers an immune response towards the antigen-expressing cells. Although there is a large number of maturation agents capable of inducing strong DC maturation, there is still only a very limited number of these agents approved for use in the production of DCs for ACI. In seeking novel DC maturation agents, we used differentially activated human mast cell (MC) line LAD2 as a cellular adjuvant to elicit or modulate the maturation of ex vivo-produced monocyte-derived iDCs. We found that co-culture of iDCs with differentially activated LAD2 MCs in serum-containing media significantly modulated polyinosinic:polycytidylic acid (poly I:C)-elicited DC maturation as determined through the surface expression of the maturation markers CD80, CD83, CD86, and human leukocyte antigen(HLA)-DR. Once iDCs were generated in serum-free conditions, they became refractory to the maturation with poly I:C, and the LAD2 MC modulatory potential was minimized. However, the maturation-refractory phenotype of the serum-free generated iDCs was largely overcome by co-culture with thapsigargin-stimulated LAD2 MCs. Our data suggest that differentially stimulated mast cells could be novel and highly potent cellular adjuvants for the maturation of DCs for ACI.

Cytokines Stimulated by IL-33 in Human Skin Mast Cells: Involvement of NF-κB and p38 at Distinct Levels and Potent Co-Operation with FcεRI and MRGPRX2.

The IL-1 family cytokine IL-33 activates and re-shapes mast cells (MCs), but whether and by what mechanisms it elicits cytokines in MCs from human skin remains poorly understood. The current study found that IL-33 activates CCL1, CCL2, IL-5, IL-8, IL-13, and TNF-α, while IL-1ß, IL-6, IL-31, and VEGFA remain unaffected in cutaneous MCs, highlighting that each MC subset responds to IL-33 with a unique cytokine profile. Mechanistically, IL-33 induced the rapid (1-2 min) and durable (2 h) phosphorylation of p38, whereas the phosphorylation of JNK was weaker and more transient. Moreover, the NF-κB pathway was potently activated, as revealed by IκB degradation, increased nuclear abundance of p50/p65, and vigorous phosphorylation of p65. The activation of NF-κB occurred independently of p38 or JNK. The induced transcription of the cytokines selected for further study (CCL1, CCL2, IL-8, TNF-α) was abolished by interference with NF-κB, while p38/JNK had only some cytokine-selective effects. Surprisingly, at the level of the secreted protein products, p38 was nearly as effective as NF-κB for all entities, suggesting post-transcriptional involvement. IL-33 did not only instruct skin MCs to produce selected cytokines, but it also efficiently co-operated with the allergic and pseudo-allergic/neurogenic activation networks in the production of IL-8, TNF-α, CCL1, and CCL2. Synergism was more pronounced at the protein than at the mRNA level and appeared stronger for MRGPRX2 ligands than for FcεRI. Our results underscore the pro-inflammatory nature of an acute IL-33 stimulus and imply that especially in combination with allergens or MRGPRX2 agonists, IL-33 will efficiently amplify skin inflammation and thereby aggravate inflammatory dermatoses.