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Bronchial and alveolar remodeling and impaired epithelial function are characteristics of chronic respiratory diseases. In these patients, an increased number of mast cells (MCs) positive for serine proteases, tryptase and chymase, infiltrate the epithelium and alveolar parenchyma. However, little is known regarding the implication of intraepithelial MCs on the local environment, such as epithelial cell function and properties. In this study, we investigated whether MC tryptase is involved in bronchial and alveolar remodeling and the mechanisms of regulation during inflammation. Using novel holographic live cell imaging, we found that MC tryptase enhanced human bronchial and alveolar epithelial cell growth and shortened the cell division intervals. The elevated cell growth induced by tryptase remained in a pro-inflammatory state. Tryptase also increased the expression of the anti-apoptotic protein BIRC3, as well as growth factor release in epithelial cells. Thus, our data imply that the intraepithelial and alveolar MC release of tryptase may play a critical role in disturbing bronchial epithelial and alveolar homeostasis by altering cell growth–death regulation. [ABSTRACT FROM AUTHOR]

Isatidis folium or Isatis tinctoria L. is a flowering plant of the Brassicaceae family, commonly known as woad, with an ancient and well-documented history as an indigo dye and medicinal plant. This study aimed to evaluate the anti-atopic dermatitis (AD) effects of Isatidis folium water extract (WIF) using a 2,4-dinitrochlorobenzene (DNCB)-induced AD-like mouse model and to investigate the underlying mechanism using tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ)-activated HaCaT cells. Oral administration of WIF reduced spleen weight, decreased serum IgE and TNF-α levels, reduced epidermal and dermal thickness, and inhibited eosinophil and mast cell recruitment to the dermis compared to DNCB-induced control groups. Furthermore, oral WIF administration suppressed extracellular signal-regulated kinase and p38 mitogen-activated protein kinase protein expression levels, p65 translocation from the cytoplasm to the nucleus, and mRNA expression of TNF-α, IFN-γ, interleukin (IL)-6, and IL-13 in skin lesion tissues. In HaCaT cells, WIF suppressed the production of regulated upon activation, normal T cell expressed and secreted (RANTES), thymus and activation-regulated chemokine (TARC), macrophage-derived chemokine (MDC), MCP-1, and MIP-3a, which are inflammatory cytokines and chemokines related to AD, and inhibited the mRNA expression of RANTES, TARC, and MDC in TNF-α/IFN-γ-stimulated HaCaT cells. Overall, the results revealed that WIF ameliorated AD-like skin inflammation by suppressing proinflammatory cytokine and chemokine production via nuclear factor-κB pathway inhibition, suggesting WIF as a potential candidate for AD treatment. [ABSTRACT FROM AUTHOR]

Background: Curcumin is a diketone compound extracted from the rhizomes of some plants in the Zingiberaceae and Araceae family. It possesses a variety of biological activities, including antioxidant, anti-inflammatory and anti-cancer properties. However, the cellular and molecular antipruritic mechanisms of curcumin remain to be explored. Objective: Our objective was to study the role of curcumin in pruritus and determine whether its antipruritic effect is related to MrgprB2 receptor. Methods: The effect of curcumin on pruritus in mice was examined by scratching behavior test. The antipruritic mechanism of curcumin was explored by using transgenic mice (MrgprB2−/− mice, MrgprB2CreTd/tomato mice), histological analysis, western blot and immunofluorescence. In addition, the relationship between curcumin and MrgprB2/X2 receptor was studied in vitro by using calcium imaging, plasmid transfection and molecular docking Results: In the current study, we found that curcumin had obvious antipruritic effect. Its antipruritic effect was related to the regulation of MrgprB2 receptor activation and mast cells tryptase release. In vitro, mouse peritoneal mast cells activated by compound 48/80 could be inhibited by curcumin. In addition, curcumin was also found to suppress the calcium flux in MrgprX2 or MrgprB2-overexpression HEK cells induced by compound 48/80, substance P, and PAMP 9-20, displaying the specific relation with the MrgprB2/X2 receptor. Moreover, molecular docking results showed that curcumin had affinity to MrgprX2 protein. Conclusions: Overall, these results indicated that curcumin has the potential to treat pruritus induced by mast cell MrgprB2 receptor. [ABSTRACT FROM AUTHOR]

Background: Serum histamine, immunoglobulin E, and tryptase are markers of allergic diseases. Despite the reported association between migraine and allergic diseases, differences in these marker levels between episodic and chronic migraines remain unelucidated. Methods: We investigated serum histamine, immunoglobulin E, and tryptase levels in 97 and 96 participants with episodic migraine and chronic migraine, respectively, and 56 controls according to the presence of allergic diseases. Results: Serum histamine levels in episodic migraine (median and interquartile ranges, 0.78 [0.65–1.25] ng/mL, p < 0.001) and chronic migraine (0.89 [0.67–1.28] ng/mL, p < 0.001) participants were significantly lower than those in healthy controls (1.19 [0.81–2.08] ng/mL) among the 160 participants without allergic diseases. Serum immunoglobulin E levels in episodic migraine and chronic migraine participants with allergic diseases negatively correlated with headache frequency (correlation coefficient = −0.263, p = 0.017). Serum histamine levels in participants with allergic diseases and serum immunoglobulin E levels in participants without allergic diseases were not significantly different among episodic migraine, chronic migraine, and control groups. Serum tryptase levels did not significantly differ among episodic migraine, chronic migraine, and control participants with and without allergic diseases. Conclusions: Altered serum histamine and immunoglobulin E levels in episodic migraine and chronic migraine and different profiles concerning allergic diseases suggest the involvement of allergic mechanisms in migraine pathogenesis. [ABSTRACT FROM AUTHOR]

Functional dyspepsia (FD) is a common functional gastrointestinal (GI) disorder, but its pathophysiology is poorly understood. Mast cells (MCs) may play a critical role in the development of FD. Therefore, the aim of this study was to investigate the effect of MCs on barrier function, tight junction (TJ) proteins and related signaling pathways. The expression of the TJ proteins claudin-8, ZO-1 and occludin in biopsy tissues from seven FD patients and five controls was assessed. Based on the in vivo results, we further investigated the effect of (1) MC degranulation in a coculture model of Caco-2/RBL-2H3 cells and tryptase in Caco-2 monolayers, (2) MC degranulation in the presence or absence of a PAR-2 antagonist and (3) MC degranulation in the presence or absence of an ERK1/2 signaling pathway inhibitor. The epithelial integrity of Caco-2 cell monolayers was assessed by measuring the transepithelial electrical resistance (TEER). The expression of TJ proteins was evaluated by western blotting, QT-PCR and immunostaining. Epithelial claudin-8, ZO-1 and occludin protein expression were significantly reduced in tissues from FD patients compared with controls. MC degranulation and tryptase decreased the TEER and reduced the expression of TJ proteins in Caco-2 cell monolayers. A PAR-2 antagonist and an ERK1/2 signaling pathway inhibitor significantly reduced the effect of MC degranulation on the TEER and TJ protein expression in Caco-2 cell monolayers. MCs disrupt duodenal barrier function by modulating the levels of TJ proteins, and the PAR-2 and ERK1/2 signaling pathways may mediate the pathogenesis of FD. [ABSTRACT FROM AUTHOR]

Although there is controversial evidence of innate immune cell activation mechanisms during liver inflammation, the accumulation of these innate immune cells promotes the recruitment of adaptive immune cells and progression of liver fibrosis and cirrhosis. [10][37] Kupffer cell-MC interactions were also recently demonstrated by the stabilization of MC membranes, or inhibition of degranulation, observed with increased expression of heme oxygenase-1 (HO-1) in Kupffer cells. [38][39] Both MCs and Kupffer cells interact with a variety of hepatic cells (such as cholangiocytes), mainly due to localization, and are involved in the necessary recruitment of immune cells (i.e., T cells) during injury/inflammation. Keywords: mast cells; cholestasis; innate immune cells EN mast cells cholestasis innate immune cells 226 233 8 07/18/23 20230501 NES 230501 Mast cells (MCs) are multifunctional immune cells that contribute to both pathogen clearance and inflammation. [Extracted from the article]

Background: Mastocytosis is a heterogeneous group of rare disorders characterized by the accumulation of clonal mast cells in organs such as the skin and bone marrow. The diagnosis of cutaneous mastocytosis (CM) is based on clinical findings, positive Darier's sign, and histopathology, if necessary. Methods: Medical records of 86 children with CM diagnosed during a 35‐year long period were reviewed. Most patients (93%) developed CM during the first year of life (median age 3 months). Clinical features at presentation and during the follow‐up period were analyzed. Baseline serum tryptase level was measured in 28 patients. Results: A total of 85% of patients had maculopapular cutaneous mastocytosis/urticaria pigmentosa (MPCM/UP), 9% had mastocytoma, and 6% had diffuse cutaneous mastocytosis (DCM). Boy to girl ratio was 1.1:1. Fifty‐four of 86 patients (63%) were followed from 2 to 37 years (median 13 years). Complete resolution was registered in 14% of mastocytoma cases, 14% of MCPM/UP, and in 25% of DCM patients. After the age of 18, skin lesion persisted in 14% mastocytoma, 7% MCPM/UP, and 25% children with DCM. Atopic dermatitis was diagnosed in 9.6% of patients with MPCM/UP. Three of 28 patients had elevated serum tryptase. Prognosis in all patients was good, and there were no signs of progression to systemic mastocytosis (SM). Conclusion: To the best of our knowledge, our results represent the longest single‐center follow‐up study of childhood‐onset CM. We found no complications of massive mast cell degranulation or progression to SM. [ABSTRACT FROM AUTHOR]

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Background: As an essential regulator of type I interferon (IFN) response, TMEM173 participates in immune regulation and cell death induction. In recent studies, activation of TMEM173 has been regarded as a promising strategy for cancer immunotherapy. However, transcriptomic features of TMEM173 in B-cell acute lymphoblastic leukemia (B-ALL) remain elusive. Methods: Quantitative real-time PCR (qRT-PCR) and western blotting (WB) were applied to determine the mRNA and protein levels of TMEM173 in peripheral blood mononuclear cells (PBMCs). TMEM173 mutation status was assessed by Sanger sequencing. Single-cell RNA sequencing (scRNA-seq) analysis was performed to explore the expression of TMEM173 in different types of bone marrow (BM) cells. Results: The mRNA and protein levels of TMEM173 were increased in PBMCs from B-ALL patients. Besides, frameshift mutation was presented in TMEM173 sequences of 2 B-ALL patients. ScRNA-seq analysis identified the specific transcriptome profiles of TMEM173 in the BM of high-risk B-ALL patients. Specifically, expression levels of TMEM173 in granulocytes, progenitor cells, mast cells, and plasmacytoid dendritic cells (pDCs) were higher than that in B cells, T cells, natural killer (NK) cells, and dendritic cells (DCs). Subset analysis further revealed that TMEM173 and pyroptosis effector gasdermin D (GSDMD) restrained in precursor-B (pre-B) cells with proliferative features, which expressed nuclear factor kappa-B (NF-κB), CD19, and Bruton's tyrosine kinase (BTK) during the progression of B-ALL. In addition, TMEM173 was associated with the functional activation of NK cells and DCs in B-ALL. Conclusions: Our findings provide insights into the transcriptomic features of TMEM173 in the BM of high-risk B-ALL patients. Targeted activation of TMEM173 in specific cells might provide new therapeutic strategies for B-ALL patients. [ABSTRACT FROM AUTHOR]

PRG4 is an extracellular matrix protein that maintains homeostasis through its boundary lubricating and anti-inflammatory properties. Altered expression and function of PRG4 have been associated with joint inflammatory diseases, including osteoarthritis. Here we show that mast cell tryptase β cleaves PRG4 in a dose- and time-dependent manner, which was confirmed by silver stain gel electrophoresis and mass spectrometry. Tryptase-treated PRG4 results in a reduction of lubrication. Compared to full-length, cleaved PRG4 further activates NF-κB expression in cells overexpressing TLR2, −4, and −5. In the destabilization of the medial meniscus model of osteoarthritis in rat, tryptase β and PRG4 colocalize at the site of injury in knee cartilage and is associated with disease severity. When human primary synovial fibroblasts from male osteoarthritis patients or male healthy subjects treated with tryptase β and/or PRG4 are subjected to a quantitative shotgun proteomics and proteome changes are characterized, it further supports the role of NF-κB activation. Here we show that tryptase β as a modulator of joint lubrication in osteoarthritis via the cleavage of PRG4. Altered expression and function of the extracellular matrix protein PRG4 have been associated with osteoarthritis. Here, the authors show that mast cell tryptase β cleaves PRG4, resulting in a reduction of lubrication and activation of inflammation in this context. [ABSTRACT FROM AUTHOR]

Inflammation is pathogenic to skin diseases, including atopic dermatitis (AD) and eczema. Treatment for AD remains mostly symptomatic with newer but costly options, tainted with adverse side effects. There is an unmet need for safe therapeutic and preventative strategies for AD. Resveratrol (R) is a natural compound known for its anti-inflammatory properties. However, animal and human R studies have yielded contrasting results. Mast cells (MCs) are innate immune skin-resident cells that initiate the development of inflammation and progression to overt disease. R's effects on MCs are also controversial. Using a human-like mouse model of AD development consisting of a single topical application of antigen ovalbumin (O) for 7 days, we previously established that the activation of MCs by a bioactive sphingolipid metabolite sphingosine-1-phosphate (S1P) initiated substantial skin remodeling compared to controls. Here, we show that daily R application normalized O-mediated epidermal thickening, ameliorated cell infiltration, and inhibited skin MC activation and chemokine expression. We unraveled R's multiple mechanisms of action, including decreased activation of the S1P-producing enzyme, sphingosine kinase 1 (SphK1), and of transcription factors Signal Transducer and Activator of Transcription 3 (Stat3) and NF-κBp65, involved in chemokine production. Thus, R may be poised for protection against MC-driven pathogenic skin inflammation. [ABSTRACT FROM AUTHOR]

Mast cells are responsible for IgE-dependent allergic responses, but they also produce various bioactive mediators and contribute to the pathogenesis of various cardiovascular diseases, including pulmonary hypertension (PH). The importance of lipid mediators in the pathogenesis of PH has become evident in recent years, as exemplified by prostaglandin I2, the most central therapeutic target in pulmonary arterial hypertension. New bioactive lipids other than eicosanoids have also been identified that are associated with the pathogenesis of PH. However, it remains largely unknown how mast cell-derived lipid mediators are involved in pulmonary vascular remodeling. Recently, it has been demonstrated that mast cells produce epoxidized n-3 fatty acid (n-3 epoxides) in a degranulation-independent manner, and that n-3 epoxides produced by mast cells regulate the abnormal activation of pulmonary fibroblasts and suppress the progression of pulmonary vascular remodeling. This review summarizes the role of mast cells and bioactive lipids in the pathogenesis of PH. In addition, we introduce the pathophysiological role and therapeutic potential of n-3 epoxides, a mast cell-derived novel lipid mediator, in the pulmonary vascular remodeling in PH. Further knowledge of mast cells and lipid mediators is expected to lead to the development of innovative therapies targeting pulmonary vascular remodeling. [ABSTRACT FROM AUTHOR]

The mechanisms of ovarian endometrioid cyst formation, or cystic ovarian endometriosis, still remain to be elucidated. To address this issue, we analyzed the involvement of mast cell (MC) tryptase and carboxypeptidase A3 (CPA3) in the development of endometriomas. It was found that the formation of endometrioid cysts was accompanied by an increased MC population in the ovarian medulla, as well as by an MC appearance in the cortical substance. The formation of MC subpopulations was associated with endometrioma wall structures. An active, targeted secretion of tryptase and CPA3 to the epithelium of endometrioid cysts, immunocompetent cells, and the cells of the cytogenic ovarian stroma was detected. The identification of specific proteases in the cell nuclei of the ovarian local tissue microenvironment suggests new mechanisms for the regulatory effects of MCs. The cytoplasmic outgrowths of MCs propagate in the structures of the stroma over a considerable distance; they offer new potentials for MC effects on the structures of the ovarian-specific tissue microenvironment under pathological conditions. Our findings indicate the potential roles of MC tryptase and CPA3 in the development of ovarian endometriomas and infer new perspectives on their uses as pharmacological targets in personalized medicine. [ABSTRACT FROM AUTHOR]

Tryptase, a protease implicated in asthma pathology, is secreted from mast cells upon activation during an inflammatory allergic response. MTPS9579A is a novel monoclonal antibody that inhibits tryptase activity by irreversibly dissociating the active tetramer into inactive monomers. This study assessed the relationship between MTPS9579A concentrations in healthy subjects and tryptase levels in serum and nasal mucosal lining fluid from healthy subjects and patients with moderate‐to‐severe asthma. These data were used to develop a mechanistic pharmacokinetic/pharmacodynamic (PK/PD) model that quantitatively inter‐relates MTPS9579A exposure and inhibition of active tryptase in the airway of patients with asthma. From initial estimates of airway tryptase levels and drug partitioning, the PK/PD model predicted almost complete neutralization of active tryptase in the airway of patients with asthma with MTPS9579A doses of 900 mg and greater, administered intravenously (i.v.) once every 4 weeks (q4w). Suppression of active tryptase during an asthma exacerbation event was also evaluated using the model by simulating the administration of MTPS9579A during a 100‐fold increase in tryptase secretion in the local tissue. The PK/PD model predicted that 1800 mg MTPS9579A i.v. q4w results in 95.7% suppression of active tryptase at the steady‐state trough concentration. Understanding how the exposure‐response relationship of MTPS9579A in healthy subjects translates to patients with asthma is critical for future clinical studies assessing tryptase inhibition in the airway of patients with moderate‐to‐severe asthma. [ABSTRACT FROM AUTHOR]

The association between Hymenoptera venom‐triggered anaphylaxis (HVA) and clonal mast cell‐related disorders (cMCD) has been known for decades. However, recent breakthroughs in peripheral blood screening for KIT p.D816V missense variant have revealed the true extent of this clinical association whilst adding to our understanding of the underlying aetiology. Thus, recent large studies highlighted the presence of KIT p.D816V among 18.2% and 23% of patients with severe Hymenoptera venom‐triggered anaphylaxis. A significant proportion of those patients have normal serum basal tryptase (BST) levels, with no cutaneous findings such as urticaria pigmentosa or other systemic findings such as organomegaly that would have suggested the presence of cMCD. These findings of an increased prevalence suggest that the impact of cMCD on anaphylaxis could be clinically underestimated and that the leading question for clinicians could be changed from 'how many patients with cMCD have anaphylaxis?' to 'how many patients with anaphylaxis have cMCD?'. The discovery of hereditary α‐tryptasemia (HαT)—a genetic trait caused by an increased copy number of the Tryptase Alpha/Beta 1 (TPSAB1) gene‐, first described in 2016, is now known to underlie the majority of cases of elevated BST outside of cMCD and chronic kidney disease. HαT is the first common heritable genetic modifier of anaphylaxis described, and it is associated with increased risk for severe HVA (relative risk = 2.0), idiopathic anaphylaxis, and an increased prevalence of anaphylaxis in patients with cMCD, possibly due to the unique activity profile of α/β ‐tryptase heterotetramers that may potentiate immediate hypersensitivity reaction severity. Our narrative review aims to highlight recent research to have increased our understanding of cMCD and HαT, through recent lessons learned from studying their association with HVA. Additionally, we examined the studies of mast cell‐related disorders in food and drug allergy in an effort to determine whether one should also consider cMCD and/or HαT in cases of severe anaphylaxis triggered by food or drugs. [ABSTRACT FROM AUTHOR]

Cationic and amphiphilic peptides can be used as homing devices to accumulate conjugated antibiotics to bacteria-enriched sites and promote efficient microbial killing. However, just as important as tackling bacterial infections, is themodulation of the immune response in this complex microenvironment. In the present report, we designed a peptide chimaera called Chim2, formed by a membrane-active module, an enzyme hydrolysis site and a formyl peptide receptor 2 (FPR2) agonist. Thismoleculewas designed to adsorb onto bacterialmembranes, promote their lysis, and upon hydrolysis by local enzymes, release the FPR2 agonist sequence for activation and recruitment of immune cells. We synthesized the isolated peptide modules of Chim2 and characterized their biological activities independently and as a single polypeptide chain. We conducted antimicrobial assays, alongwith other tests aiming at the analyses of the cellular and immunological responses. In addition, assays using vesicles as models of eukaryotic and prokaryotic membranes were conducted and solution structures of Chim2 were generated by 1HNMR.Chim2 is antimicrobial, adsorbs preferentially to negatively charged vesicles while adopting an a-helix structure and exposes its disorganized tail to the solvent, which facilitates hydrolysis by tryptase-like enzymes, allowing the release of the FPR2 agonist fragment. This fragmentwas shown to induce accumulationof the cellular activation marker, lipid bodies, in mouse macrophages and the release of immunomodulatory interleukins. In conclusion, these data demonstrate that peptides with antimicrobial and immunomodulatory activities can be considered for further development as drugs. [ABSTRACT FROM AUTHOR]

Mast cells are essential regulators of inflammation most recognized for their central role in allergic inflammatory disorders. Signaling via the high-affinity immunoglobulin E (IgE) receptor, FceRI, leads to rapid degranulation of preformed granules and the sustained release of newly synthesized proinflammatory mediators. Our group recently established rosemary extract as a potent regulator of mast cell functions, attenuating MAPK and NF-κB signaling. Carnosic acid (CA)--a major polyphenolic constituent of rosemary extract--has been shown to exhibit anti-inflammatory effects in other immune cell models, but its role as a potential modulator of mast cell activation is undefined. Therefore, we sought here to determine the modulatory effects of CA in a mast cell model of allergic inflammation. We sensitized bone marrow-derived mast cells with anti-trinitrophenyl IgE and activated with allergen (TNP-BSA) under stem cell factor potentiation, in addition to treatment with CA. Our results indicate that CA significantly inhibits allergen-induced early phase responses including Ca2+ mobilization, ROS production, and subsequent degranulation. We also show CA treatment reduced late phase responses, including the release of all cytokines and chemokines examined following IgE stimulation and corresponding gene expression excepting that of CCL2. Importantly, we determined that CA mediates its inhibitory effects through modulation of tyrosine kinase Syk and downstream effectors TAK1 (Ser412) and Akt (Ser473) as well as NFκB signaling, while phosphorylation of FceRI (γ chain) and MAPK proteins remained unaltered. These novel findings establish CA as a potent modulator of mast cell activation, warranting further investigation as a putative anti-allergy therapeutic. [ABSTRACT FROM AUTHOR]

Vaccination against COVID-19 is the main global preventive measure in the fight against SARS-CoV-2. As the COVID-19 vaccine will be administered to billions of individuals worldwide during the pandemic, there exists a reasonable fear of severe adverse reactions requiring constant vigilance and careful treatment, particularly in individuals with a history of severe allergy. The aim of our study was to develop and implement an algorithm for the safe vaccination against COVID-19 in patients with pathology associated with hypersensitivity reactions. From January to June 2022, after signing the informed consent, 126 adult patients with relevant diseases in remission were included in a prospective single-center study. Complaints and case histories were collected from all patients, including comorbidity and medications received, anthropometric data were studied, a standard clinical examination was performed, tryptase in venous blood was measured, the level of control was additionally assessed in patients with asthma using the asthma control test and spirography. According to the results of the study, it was proven that vaccination against COVID-19 in patients with pathology associated with hypersensitivity reactions is safe, on condition of prior consultation with an allergist and performing vaccination in a medical institution. The algorithm developed for preparing for vaccination in the mentioned above persons consists of a careful collection of case history; assessment and, in case of absence – achieving control of the underlying disease; analysis and correction of therapy with temporary replacement of medications that are potential cofactors of anaphylaxis; determination of blood tryptase level. For safe vaccination in patients with severe allergic anamnesis, administration of 20 mg of desloratadine before each dose of the vaccine with observation after vaccination within 30 minutes is suggested. [ABSTRACT FROM AUTHOR]

Mast cells are tissue-resident cells playing major roles in homeostasis and disease conditions. Lung mast cells are particularly important in airway inflammatory diseases such as asthma. Human mast cells are classically divided into the subsets MCT and MCTC, where MCT express the mast cell protease tryptase and MCTC in addition express chymase, carboxypeptidase A3 (CPA3) and cathepsin G. Apart from the disctintion of the MCT and MCTC subsets, little is known about the heterogeniety of human lung mast cells and a deep analysis of their heterogeniety has previously not been performed. We therefore performed single cell RNA sequencing on sorted human lung mast cells using SmartSeq2. The mast cells showed high expression of classical mast cell markers. The expression of several individual genes varied considerably among the cells, however, no subpopulations were detected by unbiased clustering. Variable genes included the protease-encoding transcripts CMA1 (chymase) and CTSG (cathepsin G). Human lung mast cells are predominantly of the MCT subset and consistent with this, the expression of CMA1 was only detectable in a small proportion of the cells, and correlated moderately to CTSG. However, in contrast to established data for the protein, CPA3 mRNA was high in all cells and the correlation of CPA3 to CMA1 was weak. [ABSTRACT FROM AUTHOR]

In addition to high-affinity IgE receptor (FceRI), a subtype of mouse mast cells (MCs) expresses a G protein-coupled receptor known as Mas-related G protein-coupled receptor (GPCR)-B2 (MRGPRB2; human ortholog MRGPRX2). GPCR kinase 2 (GRK2) is a Serine/Threonine kinase that phosphorylates GPCRs to promote their desensitization and internalization. We previously showed that silencing GRK2 expression in mouse bone marrow-derived MCs (BMMCs) blocks IgE-mediated degranulation. Compound 48/80 (C48/80), substance P (SP) and LL-37 cause degranulation in human and mouse MCs via MRGPRX2 and MRGPRB2, respectively. We also reported that C48/80 and SP cause desensitization and internalization of MRGPRX2, but LL-37 does not. Here, we generated mice with MC-specific deletion of Grk2 (Cpa3Cre+/Grk2fl/fl) to determine its role on IgE-mediated responses and to assess whether it differentially regulates degranulation in response to LL-37, C48/80 and SP. Absence of GRK2 substantially inhibited IgE-mediated tyrosine phosphorylation of STAT5, calcium mobilization, and degranulation in mouse primary lung-derived MCs (PLMCs). By contrast, peritoneal MCs (PMCs) from Cpa3Cre+/Grk2fl/fl mice demonstrated significant enhancement of degranulation in response to C48/80 and SP, but not LL-37. Deletion of Grk2 in MCs attenuated IgE-mediated passive cutaneous anaphylaxis (PCA) and itch but not passive systemic anaphylaxis (PSA). Surprisingly, PSA was significantly reduced in Mrgprb2-/- mice. These findings suggest that GRK2 contributes to PCA and itch but not PSA. By contrast, GRK2 desensitizes MRGPRX2/B2-mediated responses to C48/80 and SP but not LL-37. However, IgE-mediated PSA likely involves the activation of MRGPRB2 by LL-37 or a similar agonist, whose function is resistant to modulation by GRK2. [ABSTRACT FROM AUTHOR]

Background: We examined the distributions of 22 immune cell types and the responses to PD-1/PD-L1 inhibitors according to EGFR mutation profile, in three independent datasets of lung adenocarcinoma (LUAD). Methods: We used CIBERSORTx to analyze the distributions of immune cells, and tumor immune dysfunction and exclusion (TIDE) or tumor mutation burden (TMB) to analyze responses to anti-PD-1/PD-L1 therapy, in two public LUAD datasets. The results were verified with a validation set that included patients treated with PD-1/PD-L1 inhibitors. Results: Compared to EGFR mutants, EGFR wild-type carcinomas had higher numbers of CD8+ T cells, CD4 memory activated T cells and neutrophils, and lower numbers of resting dendritic cells and resting mast cells, in two of the datasets. In our subgroup analyses, CD8+ T cells and CD4 memory activated T cells were more numerous in EGFR rare variants than in wild-types, L858R mutants, and exon 19 deletion mutants. In our TIDE or TMB analyses, EGFR rare variants were predicted to respond better to PD-1/PD-L1 inhibitors than wild-types, L858R mutants, and exon 19 deletion mutants. In the validation set verified by immunohistochemical staining, levels of CD8+ T cells in the EGFR rare variant or wild-type groups were significantly higher than in the EGFR L858R and exon 19 deletion groups. In patients treated with PD-1/PD-L1 inhibitors, the survival rates of patients with EGFR wild-type and rare mutant carcinomas were higher than those with L858R and exon 19 deletion carcinomas. Conclusion: The EGFR rare mutation form of LUAD shows a higher immune activation state compared to wild-type, L858R, and exon 19 deletion variants, indicating it as a potential target for PD-1/PD-L1 inhibitor therapy. [ABSTRACT FROM AUTHOR]

Objective and methods: IL-33 is present in endothelial, epithelial, and fibroblast-like cells and released upon cell injury. IL-33 reportedly induces mast-cell degranulation and is involved in various diseases, including allergic diseases. So, IL-33-related diseases seem to overlap with histamine-related diseases. In addition to the release from mast cells, histamine is newly formed by the induction of histidine decarboxylase (HDC). Some inflammatory and/or hematopoietic cytokines (IL-1, IL-3, etc.) are known to induce HDC, and the histamine produced by HDC induction is released without storage. We examined the involvement of HDC and histamine in the effects of IL-33. Results: A single intraperitoneal injection of IL-33 into mice induced HDC directly and/or via other cytokines (including IL-5) within a few hours in various tissues, particularly strongly in hematopoietic organs. The major cells exhibiting HDC-induction were mast cells and c-kit+ cells in the bone marrow. HDC was also induced in non-mast cells in non-hematopoietic organs. HDC, histamine, and histamine H4 receptors (H4Rs) contributed to the suppression of IL-33-induced eosinophilia. Conclusion: IL-33 directly and indirectly (via IL-5) induces HDC in various cells, particularly potently in c-kit+ cells and mature mast cells, and the newly formed histamine contributes to the negative regulation of IL-33-induced eosinophilia via H4Rs. [ABSTRACT FROM AUTHOR]

Mast cells (MCs) are tissue cells that are derived from bone marrow stem cells that contribute to allergic reactions, inflammatory diseases, innate and adaptive immunity, autoimmunity, and mental disorders. MCs located near the meninges communicate with microglia through the production of mediators such as histamine and tryptase, but also through the secretion of IL-1, IL-6 and TNF, which can create pathological effects in the brain. Preformed chemical mediators of inflammation and tumor necrosis factor (TNF) are rapidly released from the granules of MCs, the only immune cells capable of storing the cytokine TNF, although it can also be produced later through mRNA. The role of MCs in nervous system diseases has been extensively studied and reported in the scientific literature; it is of great clinical interest. However, many of the published articles concern studies on animals (mainly rats or mice) and not on humans. MCs are known to interact with neuropeptides that mediate endothelial cell activation, resulting in central nervous system (CNS) inflammatory disorders. In the brain, MCs interact with neurons causing neuronal excitation with the production of neuropeptides and the release of inflammatory mediators such as cytokines and chemokines. This article explores the current understanding of MC activation by neuropeptide substance P (SP), corticotropin-releasing hormone (CRH), and neurotensin, and the role of pro-inflammatory cytokines, suggesting a therapeutic effect of the anti-inflammatory cytokines IL-37 and IL-38. [ABSTRACT FROM AUTHOR]

Hereditary alpha tryptasemia (HaT), an autosomal dominant condition first described in 2014, has previously been associated with multiple dermatologic, allergic, gastrointestinal, neuropsychiatric, autonomic, and connective tissue abnormalities. We describe a pediatric patient with predominantly mixed cutaneous inflammatory manifestations and atopic manifestations resistant to treatment who was found to have HaT. HaT should be considered in individuals with refractory inflammatory dermatologic disease and signs and/or symptoms concerning for mast cell activation. [ABSTRACT FROM AUTHOR]

Defining the immunological landscape of human tissue is an important area of research, but challenges include the impact of tissue disaggregation on cell phenotypes and the low abundance of immune cells in many tissues. Here, we describe methods to troubleshoot and standardize Cellular Indexing of Transcriptomes and Epitopes by sequencing (CITE-seq) for studies involving enzymatic digestion of human tissue. We tested epitope susceptibility of 92 antibodies commonly used to differentiate immune lineages and cell states on human peripheral blood mononuclear cells following treatment with an enzymatic digestion cocktail used to isolate islets. We observed CD4, CD8a, CD25, CD27, CD120b, CCR4, CCR6, and PD1 display significant sensitivity to enzymatic treatment, effects that often could not be overcome with alternate antibodies. Comparison of flow cytometry-based CITE-seq antibody titrations and sequencing data supports that for the majority of antibodies, flow cytometry accurately predicts optimal antibody concentrations for CITE-seq. Comparison by CITE-seq of immune cells in enzymatically digested islet tissue and donor-matched spleen not treated with enzymes revealed little digestion-induced epitope cleavage, suggesting increased sensitivity of CITE-seq and/or that the islet structure may protect resident immune cells from enzymes. Within islets, CITE-seq identified immune cells difficult to identify by transcriptional signatures alone, such as distinct tissue-resident T cell subsets, mast cells, and innate lymphoid cells (ILCs). Collectively this study identifies strategies for the rational design and testing of CITE-seq antibodies for single-cell studies of immune cells within islets and other tissues. [ABSTRACT FROM AUTHOR]

Systemic mastocytosis (SM) is a rare hematological neoplasm caused by the excessive proliferation of pathological mast cells that accumulate in the bone marrow (BM) and other extracutaneous organs leading to multi-organ damage and failure. Mast cell leukemia (MCL) is a rare form of systemic mastocytosis, accounting for < 1% of all cases of mastocytosis. MCL usually behaves aggressively with poor responses to current treatment options. Here, we report a diagnostic challenge with the leukemic subtype of MCL with a primary suspicion of pancreatic cancer. A cytomorphological, immunophenotypic, and histopathological examination of the bone marrow was performed. The diagnosis was based on the presence of ≥ 20% atypical and immature mast cells in the bone marrow and ≥ 10% mast cells among the peripheral white blood cells. The neoplastic cell population was identified as mast cell lineage by the expression of CD117 and tryptase. Only 3% of neoplastic cells displayed surface markers characteristic for clonal mast cells: CD25 and CD2. The D816V KIT mutation was not found. Neoplastic mast cells expressed CD30, a marker that is currently considered as a new minor criterion for SM. In the presented case, the primary suspicion of pancreatic cancer with osteosclerotic, lung, and pleural metastases was misleading, and a differential diagnosis based on hematological findings was performed. The patient's severe symptoms were likely the result of organ damage from mast cell infiltration. Despite the use of intensive acute myeloid leukemia (AML)-like polychemotherapy, the patient died during the course of post-induction myelosuppression due to bleeding complications. [ABSTRACT FROM AUTHOR]

Mast cells (MCs) as key players in the development of both physiological and pathological processes in the organism can form a specific tissue microenvironment. Having a rich secretion of biologically active substances, MCs can secrete tryptase and/or chymase and thereby participate in the regulation of processes such as inflammation, neoangiogenesis, allergic reactions, and oncogenesis. Reactive oxygen intermediates (ROI) play an essential role in regulation of MC degranulation, shown in vitro and in vivo models. Application of molecular hydrogen as a substance with antioxidant characteristics pathogenically appears to be an important mechanism decreasing MC secretory activity, and, as a consequence, a novel option to reduce an inflammatory background in the specific tissue microenvironment.

[Display omitted] Innate immune cells in the tumor microenvironment (TME) play an important role in breast cancer (BC) metastatic spread and influence patient survival. Macrophages differentiate along a proinflammatory M1 to protumorigenic M2 phenotype spectrum which affects distinct functions, like angiogenesis and cytokine production, and modulates BC aggressiveness and affects patient survival. Mast cells (MCs) are myeloid derived cells that serve as the first line of innate immune defense but their role in the TME of BC is not well understood. In this study, we have identified a subpopulation of innate immune cells that shows strong immunopositivity for the least studied adipokine CTRP8. Using a new and highly specific polyclonal antiserum on patient BC tissues, we identify a subset of tryptase + MCs and CD68 + macrophages co-expressing immunoreactive CTRP8. In M1 polarized THP-1 myeloid cells, this adipokine stimulated increased secretion of pro-inflammatory cytokines and elevated expression of the relaxin/ CTRP8 receptor RXFP1. Comparative analysis of secreted cytokine profiles in THP-1 M1 macrophages exposed to either CTRP8, relaxin-2 (RLN2), or the small molecule RXFP1 agonist ML-290 revealed ligand-specific cytokine signatures. Our study identified novel subsets of CTRP8 + myeloid derived innate immune cells and links this adipokine to pro-inflammatory events in the TME of BC. [ABSTRACT FROM AUTHOR]

Abstract Background Remimazolam was approved in Japan in January 2020. We report two cases of circulatory collapse due to suspected remimazolam anaphylaxis during anesthetic induction. Case presentation Case 1: A 74-year-old male was scheduled for debridement and skin grafting for a severe burn injury. We induced anesthesia with 4 mg of remimazolam and 20 mg of ketamine. The patient subsequently developed treatment-resistant severe hypotension. Case 2: A 59-year-old male was scheduled for laparoscopic-assisted sigmoid colectomy. We induced anesthesia with 9 mg of remimazolam. Within a few minutes, the patient developed treatment-resistant severe hypotension. As serum tryptase was elevated in both cases and only intravenous administration of adrenaline was effective, we considered the circulatory collapse might be due to anaphylaxis. Conclusion We experienced two cases of circulatory collapse due to suspected remimazolam anaphylaxis during anesthetic induction. The prevalence of remimazolam anaphylaxis is not yet known, and further research is needed.

Diffuse cutaneous mastocytosis (DCM) is a rare but potentially fatal condition when diagnosis and targeted treatments are delayed. This case illustrates the life-threatening complications in DCM and reviews the currently available treatments. To our knowledge, this is the first report of mastocytosis with somatic K509I mutation and concomitant tuberous sclerosis.

We aimed to investigate the relationship between mast cell (MC) infiltration into the bladder with urothelial barrier dysfunction and bladder hyperactivity in a chronic bladder ischemia (CBI) rat model. We compared CBI rats (CBI group; n = 10) with normal rats (control group; n = 10). We measured the expression of mast cell tryptase (MCT) and protease-activated receptor 2 (PAR2), which are correlated with C fiber activation via MCT, and Uroplakins (UP Ia, Ib, II and III), which are critical to urothelial barrier function, via Western blotting. The effects of FSLLRY-NH2, a PAR2 antagonist, administered intravenously, on the bladder function of CBI rats were evaluated with a cystometrogram. In the CBI group, the MC number in the bladder was significantly greater (p = 0.03), and the expression of MCT (p = 0.02) and PAR2 (p = 0.02) was significantly increased compared to that of the control group. The 10 μg/kg FSLLRY-NH2 injection significantly increased the micturition interval of CBI rats (p = 0.03). The percentage of UP-II-positive cells on the urothelium with immunohistochemical staining was significantly lower in the CBI group than in the control group (p < 0.01). Chronic ischemia induces urothelial barrier dysfunction via impairing UP II, consequently inducing MC infiltration into the bladder wall and increased PAR2 expression. PAR2 activation by MCT may contribute to bladder hyperactivity. [ABSTRACT FROM AUTHOR]

The study aimed to evaluate the effects and relationships between mast cells in the matrix, mast cell enzymes tryptase and chymase, epithelial proliferation, microvascular density, and bone destruction in cholesteatoma. Thirty-five biopsies diagnosed with cholesteatoma and seven healthy skin tissues taken from the retro-auricular region for control were evaluated. Immunohistochemical studies were performed with CD117, CD34, Ki-67, chymase, and tryptase antibodies, in a single session for all cases and the control group. The relationship between erosion size and antibody load was determined. The mean cholesteatoma epithelium Ki-67 was higher than the control group (p < 0.001). CD117-positive mast cells, chymase-positive mast cells, tryptase-positive mast cells, and microvessel density were significantly higher in the cholesteatoma matrix compared to the control group (p < 0.002, p < 0.001, p < 0.005). In the group with bone erosion scores of two and above, immunohistochemical markers tended to be higher. A positive correlation was found between CD117 and chymase, tryptase, and microvessel density; between tryptase, chymase, and microvessel density; and between chymase and microvessel density. CD117-positive mast cells and chymase-positive mast cells stimulate angiogenesis, increase the epithelium's proliferative capacity in the cholesteatoma matrix, and form cholesteatoma. The increased proliferation of cholesteatoma epithelium and increased vascular density in the matrix exacerbate bone erosion. [ABSTRACT FROM AUTHOR]

The article presents the study examines sources of differential selection in lung transplantation in the United States, which can undermine the objectivity of lung allocation scores (LAS) and impact who is transplanted. It discusses various parameters including transplant center's degree of risk tolerance and accountability, the success and fairness of the LAS, donor-organ availability and regional competition, patient health versus program health.

Leishmania parasites infect mammalian hosts through the bites of sand fly vectors. The response by mast cells (MC) to the parasite and vector-derived factors, delivered by sand fly bites, has not been characterized. We analyzed MC numbers and their mediators in BALB/c mice naturally infected in the ear with Leishmania major through the bite of the sand fly vector Phlebotomus duboscqi and compared them to non-infected sand fly bites. MC were found at the bite sites of infective and non-infected sand flies throughout 48 h, showing the release of granules with intense TNF-α, histamine, and tryptase staining. At 30 min and 48 h, the MC numbers were significantly higher (p < 0.001) in infected as compared to non-infected bites or controls. Neutrophil recruitment was intense during the first 6 h in the skin of infected and non-infected sand fly bites and decreased thereafter. An influx of neutrophils also occurred in lymph nodes, where a strong TNF-α stain was observed in mononuclear cells. Our data show that MC orchestrate an early inflammatory response after infected and non-infected sand fly bites, leading to neutrophilic recruitment, which potentially provides a safe passage for the parasite within the mammalian host. [ABSTRACT FROM AUTHOR]

Background: Allergic asthma is a chronic lung disease in which the lung inflammation and airway remodeling are orchestrated by both the inflammatory and the immune cells that creates a lung millieu that favors the perpetuation of clinical symptoms. The cell signaling in asthma involves the mast cells activation during initial contact with the allergen and, principally, the participation of eosinophils as well as Th2 cells which determine increased levels of IgE, exaggerated secretion of mucus and collagen, and bronchial hyperreactivity. Moreover, allergic asthma presents lower level of cytokines associated to the both Th1 and Treg cells response, and it implies in deficiency of anti-inflammatory response to counterregulate the exaggerated inflammation against allergen. Therefore, the equilibrium between cytokines as well as transcription factors associated to Th2, Th1, and Treg cells is compromised in allergic asthma. Imuno TF® is a food supplement with ability to interfere in immune system pathways. It has been previously demonstrated that Imuno TF® upregulated Th1 cell response whilst downregulated Th2 cell response in human lymphocytes. Objective: For this reason, we hypothesized that the Imuno TF effect could be restore the balance between Th1/Th2 CD4 T cells response in murine allergic asthma. Methods: Initially, animals were sensitized with OVA via i.p. and challenged with OVA i.n. on days 14, 15 and 16. Treatment with Imuno TF once a day was performed via orogastric from day 17 to day 20. Mice were euthanized on day 21. Results: The Imuno TF reduced eosinophilia, mucus production, and airway remodeling (collagen deposition) in asthma mice. Imuno TF influenced cellular signaling associated to allergic asthma once downregulated STAT6 expression as well as decreased IL-4, IL-5, and IL-13 in lung and serum. In addition, Imuno TF restored T-bet and Foxp3 expression as well as increased IL-12, IFN-γ, and IL-10. Conclusion: Ultimately, Imuno TF mitigated the allergic asthma due to the restoration of balance between the responses of Th1/Th2 as well as Treg cells, and their respective transcription factors the T-bet/STAT6 and Foxp3. [ABSTRACT FROM AUTHOR]

The high-affinity IgE receptor, FcεRI, plays a key role in the antigen-induced mast cell activation. Regulations for FcεRI are not yet well understood. TAFA4 is a molecule derived from neuron tissues, and has immune regulation functions. This study aims to clarify the role of TAFA4 in the regulation of FcεRI expression in mast cells. Nasal secretions were collected from patients with allergic rhinitis (AR) and healthy control (HC) subjects. TAFA4 levels of nasal secretions were evaluated by ELISA. A mouse model AR was developed using ovalbumin as the specific antigen. Negative correlation between TAFA4 and tryptase levels in nasal secretions was observed. TAFA4 could suppress the antigen-related mast cell activation. TAFA4 modulated the transcription of Fcer1g (FcεRI γ gene) in mast cells. Signals from the TAFA4-PTEN-PU.1 axis restricted FcεRI expression in mast cells. Administration of TAFA4 attenuated experimental AR. TAFA4 suppressed the expression of FcεRI in mast cells of airway tissues. TAFA4 can down regulate the expression of FcεRI in mast cells to suppress experimental AR. The data suggest that TAFA4 has translation potential to be developed as an anti-allergy therapy. TAFA4 suppressed the expression of FceRI in mast cells of airway tissues. Signals from the TAFA4-PTEN-PU.1 axis restricted FceRI expression in mast cells. Administration of TAFA4 attenuated experimental AR. [ABSTRACT FROM AUTHOR]

Objective: To characterize demographically and clinically the patients with anaphylaxis treated in a third level health institution in Medellin, Colombia. Methods: A cross-sectional descriptive observational study was carried out, which includedpatients were diagnosed with anaphylaxis between 2009 and 2019. Information was retrieved from medical records through a collection instrument. Subsequently, a descriptive statistical analysis of proportions and measures of central tendency of the variables of interest was performed. Results: A total of 1820 records were reviewed and data from 253 patients were included. Among the reported comorbidities, drug allergy was the most prevalent (28%). The most frequent manifestations of anaphylaxis were cutaneous and respiratory. Most of the cases presented basal tryptase values < 11.4 ng/mL (94.7%). Different etiological agents (food, drugs, insects and latex) were reported, and their frequency varied according to age. Adrenaline, steroids, and antihistamines were the treatments of choice in 39.9, 34.3, and 39.9% of cases, respectively. Conclusions: The characteristics of anaphylaxis in a medical center in Colombia coincide with those reported in Latin American. The treatment of anaphylaxis is not standardized, which makes it necessary to educate the health personnel and develop national guidelines. [ABSTRACT FROM AUTHOR]

Background: Evaluation of perioperative hypersensitivity (POH) is challenging, and accurate screening tools are needed to optimize the diagnostic process. We aimed to assess and validate the diagnostic value of a published algorithm (using tryptase and clinical presentation) to identify appropriate individuals for further testing for IgE‐mediated POH. Methods: We analysed the clinical presentation (tryptase elevation, cardiovascular, respiratory, skin involvement) of patients proceeding to testing for possible IgE‐mediated POH at a single tertiary referral centre, relative to subsequent skin testing and specific IgE results. Clinical presentations by drug class were also determined. Results: In 293 consecutive patients, the use of a published algorithm based on one or more of; (i) defined increase in serum tryptase, (ii) involvement of at least two‐organ systems, or (iii) presentation with new urticaria and/or angioedema; was highly sensitive [98.8% (CI95: 95.7–99.9%)] but less specific [34.6% (CI95: 25.7–44.4%)] in identifying patients testing positive on skin testing and/or specific IgE. Presentation with cardiovascular symptoms was also sensitive [89.8%(CI95: 84.2–94.0%)], while the combination of respiratory symptoms and increased tryptase was most specific [85.9%(CI95:76.6–92.5%)]. Respiratory involvement was more common in neuromuscular blocking agent allergy, while urticaria/angioedema was more common in antibiotic allergy. Conclusion: The published algorithm (of tryptase rise, two‐organ involvement or new urticaria/angioedema) is highly sensitive, and appropriate as a screening tool to identify patients suitable for testing for IgE‐mediated POH. [ABSTRACT FROM AUTHOR]

Background: Anaphylaxis is the most acute and life‐threatening manifestation of allergic disorders. Currently, there is a need to improve its medical management and increase the understanding of its molecular mechanisms. This study aimed to quantify the extravasation underlying human anaphylactic reactions and propose new theragnostic approaches. Methods: Molecular determinations were performed in paired serum samples obtained during the acute phase and at baseline from patients presenting with hypersensitivity reactions. These were classified according to their severity as Grades 1, 2 and 3, the two latter being considered anaphylaxis. Tryptase levels were measured by ImmunoCAP, and serum protein concentration was quantified by Bradford assay. Human serum albumin (HSA) and haemoglobin beta subunit (HBB) levels were determined by Western blot and polyacrylamide gel electrophoresis, respectively. Results: A total of 150 patients were included in the study. Of them, 112 had experienced anaphylaxis (83 and 29 with Grade 2 and 3 reactions, respectively). Tryptase diagnostic efficiency substantially improved when considering patients' baseline values (33%–54%) instead of the acute value threshold (21%). Serum protein concentration and HSA significantly decreased in anaphylaxis (p <.0001). HSA levels dropped with the severity of the reaction (6% and 15% for Grade 2 and 3 reactions, respectively). Furthermore, HBB levels increased during the acute phase of all hypersensitivity reactions (p <.0001). Conclusions: For the first time, the extravasation underlying human anaphylaxis has been evaluated based on the severity of the reaction using HSA and protein concentration measurements. Additionally, our findings propose new diagnostic and potential therapeutic approaches for this pathological event. [ABSTRACT FROM AUTHOR]

Mast cell (MC) plays a central role in intestinal permeability; however, few MC-targeting drugs are currently available for protection of the intestinal barrier in clinical practice. A nonfluorinated Lidocaine analog 2-diethylamino-N-2,5-dimethylphenyl acetamide (JM25-1) displays anti-allergic effect, but its impact on MC remains elusive. In this study, we explored whether JM25-1 has therapeutic potential on intestinal barrier defect through stabilizing MC. JM25-1 alleviated release of β-hexosaminidase and cytokine production of MC. The paracellular permeability was redressed by JM25-1 in intestinal epithelial cell monolayers co-cultured with activated MC. In vivo, JM25-1 diminished intestinal mucosal MC amount and cytokine production, especially downregulating the expression of CRHR1, accompanied by an increase of CRHR2. Protective effects appeared in JM25-1-treated stress rats with a recovery of weight and intestinal barrier integrity. Through network pharmacology analysis, JM25-1 showed a therapeutic possibility for irritable bowel syndrome (IBS) with predictive targeting on PI3K/AKT/mTOR signaling. As expected, JM25-1 reinforced p-PI3K, p-AKT, p-mTOR signaling in MC, while the mTOR inhibitor Rapamycin reversed the action of JM25-1 on the expression of CRHR1 and CRHR2. Moreover, JM25-1 successfully remedied intestinal defect and declined MC and CRHR1 expression in rat colon caused by colonic mucus of IBS patients. Our data implied that JM25-1 possessed therapeutic capacity against intestinal barrier defects by targeting the CRH receptors of MC through PI3K/AKT/mTOR signaling. [ABSTRACT FROM AUTHOR]

Thymic stromal lymphopoietin (TSLP) has been shown to play a central role in the initiation and persistence of allergic responses. We evaluated whether tezepelumab, a human monoclonal anti-TSLP antibody, improved the efficacy of subcutaneous allergen immunotherapy (SCIT) and promoted the development of tolerance in patients with allergic rhinitis. We conducted a double-blind parallel design trial in patients with cat allergy. A total of 121 patients were randomized to receive either intravenous tezepelumab plus subcutaneous cat SCIT, cat SCIT alone, tezepelumab alone, or placebo for 52 weeks, followed by 52 weeks of observation. Nasal allergen challenge (NAC), skin testing, and blood and nasal samples were obtained throughout the study. At week 52, the NAC-induced total nasal symptom scores (TNSS) (calculated as area under the curve [AUC 0-1h ] and as peak score [Peak 0-1h ] during the first hour after NAC) were significantly reduced in patients receiving tezepelumab/SCIT compared to SCIT alone. At week 104, one year after stopping treatment, the primary end point TNSS AUC 0-1h was not significantly different in the tezepelumab/SCIT group compared to SCIT alone, while TNSS Peak 0-1h was significantly lower in those receiving combination treatment versus SCIT. Transcriptomic analysis of nasal epithelial samples demonstrated that treatment with the combination of SCIT/tezepelumab, but neither monotherapy, caused persistent downregulation of a gene network related to type 2 inflammation that was associated with improvement in NAC responses. Inhibition of TSLP augments the efficacy of SCIT during therapy and may promote tolerance after a 1-year course of treatment. (ClinicalTrials.gov NCT02237196). [Display omitted] [ABSTRACT FROM AUTHOR]

Mast cells are granulated immune sentinels responsible for allergic inflammation. Allergen‐induced FcεRI‐signaling leads to rapid degranulation in the early‐phase and sustained production and release of pro‐inflammatory mediators in the late phase. Glycogen synthase kinase 3 (GSK3) is a constitutively active serine/threonine kinase and a central molecular convergence point for several pro‐inflammatory pathways. GSK3 inhibition has been shown to reduce inflammation but has not yet been fully characterized in mast cell activation. Therefore, the objective of this study was to evaluate GSK3 as a putative therapeutic target in allergic inflammation using the GSK3 inhibitor, CHIR99021. Here, we found that GSK3 inhibition impaired ROS production and degranulation. Through modulation of MKK4‐JNK, c‐jun, and NF‐κB signaling, GSK3 inhibition reduced the production/release of IL‐6, IL‐13, TNF, and CCL1, while only the release of CCL2 and CCL3 was impaired. Furthermore, CHIR99021‐mediated GSK3 inhibition altered the pro‐inflammatory phenotype of mast cells, reducing c‐kit receptor levels. This implicated GSK3 in FcεRI signaling, reducing release of IL‐6, TNF, and CCL1 when stimulated through FcεRI, while CCL2 and CCL3 remained unaffected, and were increased when stimulated with SCF only. These results identify GSK3 as a potential therapeutic target of utility warranting further consideration in contexts of pathological mast cell activation. [ABSTRACT FROM AUTHOR]

Epicutaneous exposure to protein allergens, such as papain, house dust mite (HDM), and ovalbumin (OVA), represents an important mode of sensitization for skin diseases including protein contact dermatitis, immunologic contact urticaria, and atopic dermatitis. These diseases are inducible by re-exposure to an allergen at both original skin sensitization and distant skin sites. In this study, we examined the serum IgE/IgG1 response, differentiation of T-helper (TH) cells, and epicutaneous TH recall response in mice pre-sensitized with protein allergens through the back skin and subsequently challenged on the ear skin. Repeated epicutaneous sensitization with allergenic proteins including papain, HDM, OVA, and protease inhibitor-treated papain, but not bovine serum albumin, induced serum allergen-specific antibody production, passive cutaneous anaphylaxis responses, and TH2 differentiation in the skin draining lymph node (DLN) cells. Sensitization with papain or HDM, which have protease activity, resulted in the differentiation of TH17 as well as TH2. In papain- or HDM-sensitized mice, a subsequent single challenge on the ear skin induced the expression of TH2 and TH17/TH22 cytokines. These results suggest that allergenic proteins induce the differentiation of TH2 in skin DLN cells and an antibody response. These findings may be useful for identifying proteins of high and low allergenic potential. Moreover, allergenic proteins containing protease activity may also differentiate TH17 and induce TH2 and TH17/TH22 recall responses at epicutaneous challenge sites. This suggests that allergen protease activity accelerates the onset of skin diseases caused by protein allergens. [ABSTRACT FROM AUTHOR]

Mast cell activation plays a key role in various allergic diseases and anaphylaxis. Several methods/techniques can be used for detection of mast cell activation. However, there was no previous systematic evaluation to compare the efficacy of each method/technique. The present study thus systematically compared various markers for mast cell activation induced by IgE cross-linking. The widely used RBL-2H3 mast cells were sensitized with anti-DNP (dinitrophenyl) IgE overnight and activated with DNP-BSA (bovine serum albumin) for up to 4 h. The untreated cells and those with anti-DNP IgE sensitization but without DNP-BSA activation served as the controls. Intracellular calcium level gradually increased to ~2-fold at 1 h, reached its peak (~5-fold) at 2 h, and returned to the basal level at 3-h post-activation. The increases in cellular tryptase level (by Western blotting) (~0.3- to 0.4-fold) and average cell size (~2.5-fold) and decrease of nucleus/cytoplasm ratio (~0.4- to 0.5-fold) were marginal at all time-points. By contrast, β-hexosaminidase release and CD63 expression (by both flow cytometry and immunofluorescence detection/localization), secreted tryptase level (by Western blotting), and tryptase expression (by immunofluorescence detection/localization) stably and obviously increased (~10-fold as compared with the untreated control and sensitized-only cells or detectable only after activation). Based on these data, the stably obvious increases (by ≥ 10-fold) in β-hexosaminidase release, CD63 expression (by both flow cytometry and immunofluorescence staining), secreted tryptase level (by Western blotting), and tryptase expression (by immunofluorescence staining) are recommended as the markers of choice for the in vitro study of mast cell activation using RBL-2H3 cells. [ABSTRACT FROM AUTHOR]

Granzymes comprise a group of proteases involved in the killing of infected or cancerous cells by the immune system. Although best studied in T cells and natural killer (NK) cells, they are also expressed in some innate immune cells. Granzymes B and C are encoded in the mouse chymase locus that also encodes a number of mast cell-specific proteases. In line with this, mast cells can express granzyme B, although how this is regulated and their ability to express other granzymes is less well studied. We therefore examined how IL-33, a cytokine able to activate mast cells but not induce degranulation, regulated granzyme B and C levels in mast cells. Granzyme C, but not B, mRNA was strongly up-regulated in bone marrow-derived mast cells following IL-33 stimulation and there was a corresponding increase in granzyme C protein. These increases in both granzyme C mRNA and protein were blocked by a combination of the p38α/β MAPK inhibitor VX745 and the MEK1/2 inhibitor PD184352, which blocks the activation of ERK1/2. ERK1/2 and p38α activate the downstream kinases, mitogen and stress-activated kinases (MSK) 1 and 2, and IL-33 stimulated the phosphorylation of MSK1 and its substrate CREB in an ERK1/2 and p38-dependent manner. The promoter for granzyme C contains a potential CREB-binding site. Bone marrow-derived mast cells from either MSK1/2 double knockout or CREB Ser133Ala knockin mice were unable to up-regulate granzyme C. Together these results indicate that IL-33-induced granzyme C expression in mast cells is regulated by an MSK1/2-CREB-dependent pathway. [ABSTRACT FROM AUTHOR]

Purpose: The causes of anaphylaxis in a general hospital may differ from those occurring in a community setting. Underlying diseases in admitted patients and vague presenting symptoms can make the diagnosis of anaphylaxis difficult. Serum tryptase measurements may provide valuable evidence for diagnosing anaphylaxis in admitted patients. Materials and Methods: This study was designed as a retrospective study of 53 patients with an anaphylaxis episode at a Korean tertiary care general hospital. Tryptase levels were measured at baseline and different time points from the onset of anaphylaxis. Results: Drugs (42 cases; 79.2%) and foods (10 cases; 18.9%) were the most common causes of anaphylaxis. In drug-induced anaphylaxis, antibiotics (24.5%), anticancer medications, which included monoclonal antibodies (22.6%), and contrast agents (11.3%) were the most frequent causes. The muscle relaxant eperisone (5.7%), neuromuscular blocking agent rocuronium (5.7%), and its antagonist sugammadex (3.8%) were other frequent triggering agents. Wheat-dependent exercise-induced anaphylaxis was the most common entity in food-induced anaphylaxis. Tryptase concentrations were higher in patients with higher grades of anaphylaxis, as well as in accidental anaphylaxis, compared to meticulously provoked anaphylaxis. Overall diagnostic sensitivity was higher for tryptase algorithm criteria (=[1.2×baseline+2] µg/L: 71.4%) than for abnormal tryptase level criteria (=11.4 µg/L: 52.8%). Conclusion: The triggers of anaphylaxis in a Korean tertiary care hospital were diverse, including beta-lactam antibiotics, anticancer medications, contrast medias, eperisone, nonsteroidal anti-inflammatory drugs, rocuronium, sugammadex, and wheat. Tryptase measurements provided valuable evidence for diagnosis, and the sensitivity of algorithm criteria was superior to that of the abnormal value criteria. [ABSTRACT FROM AUTHOR]