Mast Cell Stabilizer Research Articles

Epidermal Mechanical Scratching-Induced ROS Exacerbates the Itch-Scratch Cycle via TRPA1 Activation on Mast Cells in Atopic Dermatitis.

Atopic dermatitis (AD) is a chronic inflammatory skin disease characterized by the itch-scratch cycle. Itching, induced by irritants or allergens that stimulate pruriceptive neurons, triggers uncontrollable mechanical scratching, leading to epidermal barrier disruption, immune response activation, inflammatory mediator release, and further stimulation of pruritus conduction. Although oxidative stress and immune cells can exacerbate this cycle, the correlation between mechanical scratching, epidermal oxidative stress, and dermal mast cell activation in AD remains unclear. Here, by examining clinical specimens of AD, establishing a three-dimensional co-culture system of HaCaT and LAD2 cells, and utilizing a mechanical scratching mouse model of AD, we found that reactive oxygen species produced by mechanically stimulated HaCaT can activate TRPA1 on mast cells presenting tryptase (TPS). Implementing a free radical scavenger and TRPA1 inhibitor can inhibit mast cell activation and type II inflammatory response, thereby alleviating itching and skin lesions in AD. These results indicate that active oxygen scavenging combined with TRPA1 inhibition can inhibit the itch-scratch cycle, which may present a potential approach for the treatment of AD.

Silibinin, a PLC-β3 inhibitor, inhibits mast cell activation and alleviates OVA-induced asthma.

The immunoglobulin E (IgE) receptor FcεRI (Fc epsilon RI) plays a crucial role in allergic reactions. Recent studies have indicated that the interaction between FcεRIβ and the downstream protein phospholipase C beta 3 (PLCβ3) leads to the production of inflammatory cytokines. The aim of this study was to develop small molecules that inhibit the protein-protein interactions between FcεRIβ and PLCβ3 to treat allergic inflammation. Additionally, PLCβ3 has emerged as a potential target protein for treating allergic inflammation. In this study, we employed a virtual screening technique to search the Taiwan Traditional Chinese Medicine Database, followed by a second screening using absorption, distribution, metabolism, excretion, and toxicity (ADMET). Among the compounds screened, silibinin exhibited the best performance, forming strong hydrogen bond interactions with residues of PLCβ3, with a binding free energy of -119.277 kcal/mol. Therefore, silibinin effectively blocked the interaction between FcεRIβ and PLCβ3. Silibinin reduced the production of allergic inflammatory cytokines, including cytokine-induced neutrophil chemoattractant 2a (CINC-2a), interleukin-2 (IL-2), cytokine-induced neutrophil chemoattractant 1 (CINC-1), interleukin 1α (IL-1α), macrophage inflammatory protein 3 alpha (MIP3α), interferon γ (IFN-γ), activin A, granulocyte macrophage colony stimulating factor (GM-CSF), intercellular adhesion molecule-1 (ICAM-1), interleukin 4 (IL-4), interleukin 13 (IL-13), Fas ligand (FasL) and tumor necrosis factor alpha (TNF-α), without inducing cytotoxicity. Furthermore, in studies of IgE-mediated allergic responses, silibinin also decreased the expression of surface IgE receptors (FcεRIs). Moreover, silibinin effectively alleviated allergen-induced asthma responses and reduced the infiltration of inflammatory immune cells into the lungs of an OVA-induced allergic airway inflammation mouse model. Taken together, these results demonstrate the potential antiallergic mechanism of silibinin both in vitro and in vivo, making it a promising candidate for the development of asthma therapeutics.

Casticin reduces rosacea-related inflammation by inhibiting mast cell activation via Mas-related G protein-coupled receptor X2.

Rosacea is a chronic inflammatory disease characterized by persistent erythema, papules, and pustules, mainly on the skin of the face. Rosacea is difficult to treat; therefore, identifying new treatments is crucial. Mas-related G protein-coupled receptor X2 (MRGPRX2)-mediated mast cell (MC) activation is essential in the pathogenesis of rosacea. Casticin has been shown to exert anti-inflammatory effects; however, it remains unclear whether it can inhibit MRGPRX2 in treating rosacea. This study determined the therapeutic efficacy of casticin against rosacea by inhibiting MRGPRX2-mediated MC activation. A mouse model of LL37-induced rosacea-like dermatitis was employed. The pathological changes were evaluated using hematoxylin and eosin (H&E) staining, and MCs and CD4+ T cells were observed. Inflammatory mediators were analyzed using ELISA. Mouse skin lesions were collected for transcriptomic sequencing. We used an MRGPRX2-mediated MC degranulation model to evaluate the inhibitory effects of casticin in vitro. Molecular docking analysis, molecular dynamics simulations, and surface plasmon resonance evaluated the binding between casticin and MRGPRX2. Casticin attenuated the LL37-induced inflammatory phenotype and reactions in rosacea-like dermatitis. RNA-seq data showed that casticin inhibited MC activation in a mouse model of rosacea. Furthermore, casticin significantly reduced CD4 + T-cell infiltration. Moreover, casticin inhibited MC activation as an MRGPRX2 antagonist in vitro and in vivo by influencing the NF-κB signaling pathway. Our study demonstrated that casticin exhibits therapeutic efficacy against rosacea by inhibiting MC activation via MRGPRX2.

KIRA6 is an Effective and Versatile Mast Cell Inhibitor of IgE-mediated Activation.

Mast cell (MC)-driven allergic diseases are constantly expanding and require the development of novel pharmacological MC stabilizers. Allergen/antigen (Ag)-triggered activation via crosslinking of the high-affinity receptor for IgE (FcεRI) is fundamentally regulated by SRC family kinases, for example, LYN and FYN, exhibiting positive and negative functions. We report that KIRA6, an inhibitor for the endoplasmic reticulum stress sensor IRE1α, suppresses IgE-mediated MC activation by inhibiting both LYN and FYN. KIRA6 attenuates Ag-stimulated early signaling and effector functions such as degranulation and proinflammatory cytokine production/secretion in murine bone marrow-derived MCs. Moreover, Ag-triggered bronchoconstriction in an ex vivo model and IgE-mediated stimulation of human MCs were repressed by KIRA6. The interaction of KIRA6 with three MC-relevant tyrosine kinases, LYN, FYN, and KIT, and the potential of KIRA6 structure as a pharmacophore for the development of respective single-, dual-, or triple-specificity inhibitors, was evaluated by homology modeling and molecular dynamics simulations. We found that KIRA6 particularly strongly binds the inactive state of LYN, FYN, and KIT with comparable affinities. In conclusion, our data suggest that the chemical structure of KIRA6 as a pharmacophore can be further developed to obtain an effective MC stabilizer.

Agonism of the glutamate receptor GluK2 suppresses dermal mast cell activation and cutaneous inflammation.

Activation of dermal mast cells through the Mas-related G protein-coupled receptor B2 receptor (MrgprB2 in mice and MrgprX2 in humans) is a key component of numerous inflammatory skin diseases, including dermatitis and rosacea. Sensory neurons actively suppress mast cell activation through the regulated release of glutamate, resulting in reduced expression of Mrgprb2 as well as genes associated with proteins found in mast cell granules. To determine whether exogenous glutamate receptor agonism could suppress mast cell function, we determined that mast cells have relatively selective expression of the glutamate receptor ionotropic, kainate 2 (GluK2). A GluK2-specific agonist, SYM2081, effectively inhibited mast cell degranulation in response to MrgprB2 agonism in both murine mast cells and human skin explants in vitro as well as in vivo after both intradermal and topical administration of SYM2081 to mice. Analyses of transcriptomic datasets from SYM2081-treated mast cells using standard differential expression approaches and an interpretable machine learning technique revealed a previously unrecognized cellular program coordinately regulated by GluK2 agonism. GluK2 agonism suppressed the expression of Mrgprb2 and genes associated with mast cell proliferation. Suppression of mast cell proliferation by SYM2081 exposure was confirmed on the basis of reduced Ki-67 expression and BrdU incorporation in vitro and in vivo. Last, pretreatment with SYM2081 reduced skin inflammation in murine models of dermatitis and rosacea. Thus, agonism of GluK2 represents a promising approach to suppress mast cell activation and may prove beneficial as therapy for inflammatory diseases in which mast cell activation is pathogenic.

Acupuncture at Weizhong (BL40) attenuates acetic acid-induced overactive bladder in rats by regulating brain neural activity through the modulation of mast cells and tibial nerves

ObjectiveThe present study evaluated the effects of deep acupuncture at Weizhong acupoint (BL40) on bladder function and brain activity in a rat model of overactive bladder (OAB), and investigated the possible mechanisms around the acupuncture area that initiate the effects of acupuncture. MethodsAdult female Sprague–Dawley rats were randomly divided into six groups, comprising a control group, model group, group treated with deep acupuncture at BL40, group treated with shallow acupuncture at BL40, group treated with acupuncture at non-acupoint next to BL40, and group treated with acupuncture at Xuanzhong (GB39). Urodynamic evaluation was used to observe the urination, and functional magnetic resonance imaging was used to observe the brain activation. The mechanism of acupuncture at BL40 in regulating bladder function was explored by toluidine blue staining and enzyme-linked immunosorbent assay, and the mechanism was verified by stabilizing mast cells (MCs) or blocking tibial nerve. ResultsDeep acupuncture at BL40 significantly increased the intercontraction interval in OAB rats and enhanced the mean amplitude of low frequency fluctuation of primary motor cortex (M1), periaquaductal gray matter (PAG), and pontine micturition center (PMC). It also increased the zero-lag functional connectivity between M1 and PAG and between PAG and PMC. Shallow acupuncture at BL40 and acupuncture at non-acupoint or GB39 had no effect on these indexes. Further studies suggested that deep acupuncture at BL40 increased the number and degranulation rate of MCs as well as the contents of 5-hydroxytryptamine, substance P, and histamine in the tissues around BL40. Blocking the tibial nerve by lidocaine injection or inhibiting MC degranulation by sodium cromoglycate injection obstructed the effects of acupuncture on restoring urinary function and modulating brain activation in OAB rats. ConclusionDeep acupuncture at BL40 may be more effective for inhibiting OAB by promoting degranulation of MCs around the acupoint and stimulating tibial nerve, thereby regulating the activation of the brain area that controls the lower urinary tract.

Successful novel use of dupilumab for gastrointestinal involvement of idiopathic hypereosinophilic syndrome: case report and review of the literature.

Hypereosinophilic syndrome (HES) is characterized by blood and tissue hypereosinophilia leading to organ damage. Gastrointestinal involvement is the third most common manifestation. We present a patient with idiopathic HES with secondary eosinophilic esophagitis (EoE), gastritis, and enteritis, corticosteroids-dependent, azathioprine- and mepolizumab-refractory. The patient achieved clinical and histopathologic remission following dupilumab treatment. A 28year-old female presented with chronic episodic nausea and emesis since childhood and initial diagnosis of primary eosinophilic gastrointestinal disease (EGID), improved with corticosteroids, refractory to azathioprine. She was found to have peripheral eosinophilia and multifactorial anemia, with iron, B12, and folate deficiencies. Esophageal, gastric, duodenal, and terminal ileum biopsies showed significant eosinophilic infiltrate. Bone marrow biopsy at age 31 confirmed HES diagnosis. By age 32, she became total parental nutrition (TPN)-dependent. She failed trials of benralizumab and mepolizumab [anti-interleukin (IL)-5 inhibitors], and cromolyn (mast-cell stabilizer). After developing new esophageal stricture, we initiated dupilumab (IL-4/13 inhibitor), recently FDA-approved for EoE. After 9weeks, esophageal stricture, gut tissue eosinophilia, and prior intestinal ulcerations resolved. She ceased TPN and is tolerating a non-restricted diet, with complete symptom resolution. Our patient's complete remission with dupilumab shows promise for broadening its use in treating GI involvement in HES, along with primary EGIDs.

Aloe-emodin relieves allergic contact dermatitis pruritus by inhibiting mast cell degranulation

Urushiol-induced allergic contact dermatitis (ACD) is a chronic inflammatory skin disease in which skin barrier dysfunction leads to pruritus and eczematous lesions. ACD is triggered by immune imbalance. Aloe emodin is an anthraquinone derivative extracted from rhubarb, aloe and other traditional Chinese medicines. It has a wide range of pharmacological effects, including anti-inflammatory, anti-tumor, and anti-allergic effects. The purpose of our study was to demonstrate the effectiveness of aloe-emodin on urushiol-induced acute pruritus and allergic contact dermatitis. The results showed that urushiol could stimulate keratinocytes to release chemokines CXCL1, CXCL2, CCL2, TSLP, and TNF-α, which recruit or activate mast cells. Aloe-emodin treatment inhibited inflammatory-response-induced mast cell degranulation in skin lesions and suppressed the expression of inflammatory cytokines, such as interleukin-4, and interleukin-6. Therefore, the results indicate that aloe-emodin can improve urushiol-induced acute pruritus and allergic contact dermatitis in mice by inhibiting mast cell degranulation.

“Relative symmetry with electronegativity of different key-groups” strategy for MRGPRX2 antagonist design and its effect on antigen-induced pulmonary inflammation

MRGPRX2 antagonists possess the potential for the treatment of allergic rhinitis, atopic dermatitis, and chronic urticaria. Previously, we identified a class of diaryl urea (DPU) MRGPRX2 antagonists with sub-micromolar IC50 values in vitro. However, the structure–activity relationship remains unclear. Herein, we adopted a “relative symmetry with electronegativity of different key-groups” strategy for further modification of DPUs to achieve a promising MRGPRX2 antagonist with higher activity and safety. Electrostatic potential energy analysis and biological evaluation revealed that B-1023 and B-5023, that possess relatively symmetric electron-withdrawing substituents, remarkable inhibited mast cell degranulation at a sub-micromolar IC50in vitro and alleviated anaphylactic symptoms. Furthermore, B-1023, mitigated antigen-induced pulmonary inflammation (AIPI) in mice and competitively bonded to MRGPRX2. In summary, the “relative symmetry with electronegativity of different key-groups” strategy provided a drug design pattern for MRGPRX2 antagonists and identified promising antiallergic precursors for AIPI treatment.

Mast Cells Contribute to Pressure Overload-Induced Myocardial Hypertrophy by Upregulating TRPV4 via Histamine: Role of Ca2+/ CnA/NFATc3 Signaling Pathway.

To investigate whether cardiac mast cells (MCs) participate in pressure overload-induced myocardial hypertrophy through the regulation of transient receptor potential vanilloid 4 (TRPV4). Pressure overload-induced myocardial hypertrophy was induced via abdominal aortic constriction (AAC). Myocardial hypertrophy was evaluated by measuring the heart weight index (HW/BW), lung weight index (LW/BW), ratio of heart weight to tibia length (HW/TL), ratio of lung weight to tibia length (LW/TL), and cross-sectional area of myocardial cells. qRT-PCR was used to detect the mRNA expression of TRPV4. Western blotting was used to detect the protein expression of TRPV4, mast cell tryptase, myosin heavy chain beta (β-MHC), calcineurin A (CnA), and nuclear factor of activated T-cell c3 (NFATc3). ELISA was used to measure the levels of brain natriuretic peptide (BNP) and histamine. Fluo4 AM was used to detect the calcium signal in H9c2 myocardial cells. Compared with those of the sham rats, the myocardial mast cells, tryptase, HW/BW, LW/BW, HW/TL, and LW/TL, the cross-sectional area of the myocardial cells, and the expression of β-MHC, TRPV4, CnA, and NFATc3 in the myocardial tissue and the serum BNP of the AAC-treated rats increased significantly, whereas the MC stabilizer cromolyn sodium (CS) reversed these indicators. In H9c2 cardiomyocytes, treatment with histamine and the TRPV4 agonist GSK1016790A upregulated the expression of TRPV4, β-MHC, BNP, CnA and NFATc3 and increased calcium ion influx, whereas these effects were inhibited by the H2 receptor inhibitor famotidine and the TRPV4 inhibitor HC067047. Cardiac MCs participate in pressure overload-induced myocardial hypertrophy through the upregulation of TRPV4 via its mediator histamine, and the Ca2+/CnA/NFATc3 signaling pathway is involved in this process.

Biotinylated Heptapeptides with D-amino Acids Suppress Allergic Reactions by Inhibiting Mast Cell Activation and Antagonizing the Histamine Receptor

Biotinylated Heptapeptides with D-amino Acids Suppress Allergic Reactions by Inhibiting Mast Cell Activation and Antagonizing the Histamine Receptor

Anti-Pruritic and Immunomodulatory Effects of Coix [Coix lacryma-jobi L. var. ma-yuen (Rom. Caill.) Stapf.] Sprouts Extract.

This study explored the anti-pruritic and immunomodulatory effects of Coix sprouts extract, focusing on histamine release and IL-31 cytokine production in HMC-1 cells. The extract significantly inhibited both factors, indicating its potential for pruritus relief. In a pruritus induction mouse model, Coix sprouts extract outperformed prednisolone in anti-pruritus effectiveness, also improving skin lesions and inhibiting mast cell infiltration. The extract suppressed tryptase expression, reduced release, inhibited mast cell proliferation, and lowered nitric oxide production, suggesting anti-inflammatory properties. Coix sprouts extract shows promise in suppressing inflammation and pruritus, making it a valuable candidate for clinical use. Additionally, the analysis of coixol content in Coix sprouts revealed variations in growth time, indicating their potential as functional materials with anti-pruritus and immune-enhancing applications.

Involvement of Molecular Mechanism and Biological Activities of Pemirolast: A Therapeutic Review

: This review paper aims to investigate the therapeutic benefits of pemirolast across different medical conditions, including asthma, allergic rhinitis, cancer, conjunctivitis, etc. The prevalence of allergic conditions has risen in recent decades, primarily attributed to environmental factors. This paper explores the role of pemirolast in managing and mitigating these diseases, shedding light on its potential applications in the context of evolving environmental influences. : This review paper investigates the diverse biological activities exhibited by pemirolast, encompassing its roles as an antimicrobial, analgesic, antiviral, anti-inflammatory, antioxidant, antihistamine, mast cell stabilizer, anti-tubercular, anticancer, anti-asthmatic, anti-malarial, diuretic, antianxiety, and antifungal agent. The positive outcomes of pemirolast application in various diseases are highlighted, showcasing its potential across a spectrum of medical conditions. : The pharmacological impact of pemirolast extends to diverse inflammatory mediators, enzymes, and hormones associated with various diseases. Pemirolast demonstrates inhibitory effects on key elements such as eosinophil activation, histamine, leukotriene, IgE, mast cells, basophils, prostaglandin, interleukin, T-helper cells, macrophage T-cells, neutrophils, tryptase, T-lymphocytes, interferons I-III, Amyloid β (Aβ) peptide, dsRNA transcription, GABA, dopamine, serotonin, and norepinephrine. This comprehensive exploration underscores pemirolast inhibitory actions across disorders, emphasizing its potential therapeutic relevance in diverse pathological conditions. : This review paper illuminates pemirolast potassium's versatile biological and therapeutic applications across various diseases. The potential synergies of combining pemirolast with buspirone, ritanserin, theophylline, and capreomycin are explored, showcasing its ability to elicit beneficial responses in addressing diverse ailments.

Plasma-Activated AVC Hydrogel for Reactive Oxygen and Nitrogen Species Delivery to Treat Allergic Contact Dermatitis.

Allergic contact dermatitis (ACD) is a common inflammatory skin disease that accounts for approximately 20% of all occupational skin diseases. As an adverse and recurrent inflammatory dermatological agent, ACD shows insufficient response to current therapies largely owing to abnormal inflammatory activation and accompanying bacterial infection in lesions. Cold atmospheric plasma is a noninvasive fledgling reactive oxygen and nitrogen species (RONS)-based therapeutic technique for ACD treatment; however, its clinical adoption has been hindered due to the risk of electrical burns and insufficient delivery of the plasma-generated RONS. To address these limitations, we constructed plasma-activated AVC (PA-AVC) hydrogels loaded with plasma-generated RONS for ACD treatment as an alternative to the common direct plasma irradiation treatment. The proposed PA-AVC hydrogels were produced on a biodegradable acryloyldimethylammonium taurate/VP copolymer (AVC) with the aid of a novel air discharge plasma without the involvement of any catalyst. In vitro data showed that abundant RONS were produced and incorporated into the PA-AVC hydrogels via complex gas-liquid reactions between the air discharge plasma and hydrosolvent; additionally, the PA-AVC hydrogels exhibited excellent storage, slow release and transdermal delivery of RONS as well as good antibacterial effects. Moreover, in vivo experiments demonstrated that PA-AVC hydrogels effectively alleviated the ACD symptoms, such as skin redness and swelling, reduced epidermal thickening and inhibited mast cell infiltration and IL-9, TNF-α, and TSLP expression with no evident systemic toxicity. Our results revealed that long-acting plasma-activated AVC hydrogels could be effective therapeutic agents for local ACD treatment.

Intestinal Epithelial Cell Brush Border Membrane Cl:HCO3 Exchanger Regulation by Mast Cells in Chronic Ileitis.

Malabsorption of NaCl is the primary cause of diarrhea in inflammatory bowel disease (IBD). Coupled NaCl absorption occurs via the dual operation of Na:H and Cl:HCO3 exchange in the brush border membrane (BBM) of villus cells. Cl:HCO3 exchange is mediated by BBM transporters DRA (downregulated in adenoma) and PAT1 (putative anion transporter 1) in the mammalian small intestine. DRA/PAT1-mediated Cl:HCO3 exchange was significantly downregulated in the BBM of villus cells in a rabbit model of chronic ileitis, while Na:H exchange was unaffected. The inhibition of Cl:HCO3 exchange was restored in the rabbits when treated with a broad-spectrum immunomodulator, i.e. a glucocorticoid, indicating that the downregulation of DRA/PAT1 is likely to be immune-mediated during chronic enteritis. Mucosal mast cells are one type of key immune cells that are known to proliferate and release immune inflammatory mediators, thus playing a significant role in the pathogenesis of IBD. However, how mast cells may regulate DRA- and PAT1-mediated Cl:HCO3 exchange in a rabbit model of chronic ileitis is unknown. In this study, treatment of rabbits with chronic intestinal inflammation with the mast cell stabilizer ketotifen did not affect the mucosal architecture of the inflamed intestine. However, ketotifen treatment reversed the inhibition of Cl:HCO3 activity in the BBM of villus cells. This restoration of Cl:HCO3 activity to normal levels by ketotifen was found to be secondary to restoring the affinity of the exchangers for its substrate chloride. This observation was consistent with molecular studies, where the mRNA and BBM protein expressions of DRA and PAT1 remained unaffected in the villus cells under all experimental conditions. Thus, this study indicates that mast cells mediated the inhibition of coupled NaCl absorption by inhibiting Cl:HCO3 exchange in a rabbit model of chronic enteritis.

Mast Cells Mediate Acute Inflammatory Responses After Glenoid Labral Tears and Can Be Inhibited With Cromolyn in a Rat Model

Background: Injuries to the glenoid labrum have been recognized as a source of joint pain and discomfort, which may be associated with the inflammatory responses that lead to the deterioration of labral tissue. However, it is unclear whether the torn labrum prompts mast cell (MC) activation, resulting in synovial inflammatory responses that lead to labral tissue degeneration. Purpose: To determine the potential influence of activated MC on synovial inflammatory responses and subsequent labral tissue degeneration and shoulder function deterioration in a rat model by monitoring MC behavior and sequential inflammatory responses within the synovial tissue and labral tissue after injury, suture repair, and MC stabilizer administration. Study Design: Controlled laboratory study. Methods: Anteroinferior glenoid labral tears were generated in the right shoulder of rats (n = 20) and repaired using a tunneled suture technique. Synovial tissue inflammatory responses were modulated in some rats with intraperitoneal administration of an MC stabilizer—cromolyn (n = 10). At weeks 1 and 3, MC activation, synovial inflammatory responses, and labral degeneration were histologically evaluated. Simultaneously, gait analysis was performed before and after surgical repair to assess the worsening of the shoulder function after the injury and treatment. Results: Resident MC degranulation after labral injury (50.48% ± 8.23% activated at week 1) contributed to the initiation of synovial tissue inflammatory cell recruitment, inflammatory product release, matrix metalloproteinase-13, and subsequent labral tissue extracellular matrix degeneration. The administration of cromolyn, an MC stabilizer, was found to significantly diminish injury-mediated inflammatory responses (inflammatory cell infiltration and subsequent proinflammatory product secretion) and improve shoulder functional recovery. Conclusion: MC activation is responsible for labral tear–associated synovial inflammation and labral degeneration. The administration of cromolyn can significantly diminish the cascade of inflammatory reactions after labral injury. Clinical Relevance: Our findings support the concept that MC stabilizers may be used as a complementary therapeutic option in the treatment and repair of labral tears.

GPR35 agonists inhibit TRPA1-mediated colonic nociception through suppression of substance P release.

The development of nonopioid analgesics for the treatment of abdominal pain is a pressing clinical problem. To address this, we examined the expression of Gi/o-coupled receptors, which typically inhibit nociceptor activation, in colonic sensory neurons. This led to the identification of the orphan receptor GPR35 as a visceral analgesic drug target because of its marked coexpression with transient receptor potential ankyrin 1 (TRPA1), a mediator of noxious mechanotransduction in the bowel. Building on in silico docking simulations, we confirmed that the mast cell stabiliser, cromolyn (CS), and phosphodiesterase inhibitor, zaprinast, are agonists at mouse GPR35, promoting the activation of different Gi/o subunits. Pretreatment with either CS or zaprinast significantly attenuated TRPA1-mediated colonic nociceptor activation and prevented TRPA1-mediated mechanosensitisation. These effects were lost in tissue from GPR35-/- mice and were shown to be mediated by inhibition of TRPA1-evoked substance P (SP) release. This observation highlights the pronociceptive effect of SP and its contribution to TRPA1-mediated colonic nociceptor activation and sensitisation. Consistent with this mechanism of action, we confirmed that TRPA1-mediated colonic contractions evoked by SP release were abolished by CS pretreatment in a GPR35-dependent manner. Our data demonstrate that GPR35 agonists prevent the activation and sensitisation of colonic nociceptors through the inhibition of TRPA1-mediated SP release. These findings highlight the potential of GPR35 agonists to deliver nonopioid analgesia for the treatment of abdominal pain.

Mast cells promote choroidal neovascularization in a model of age-related macular degeneration

‘Wet’ age-related macular degeneration (AMD) is characterized by pathologic choroidal neovascularization (CNV) that destroys central vision. Abundant evidence points to inflammation and immune cell dysfunction in the progression of CNV in AMD. Mast cells are resident immune cells that control the inflammatory response. Mast cells accumulate and degranulate in the choroid of patients with AMD, suggesting they play a role in CNV. Activated mast cells secrete various biologically active mediators, including inflammatory cytokines and proteolytic enzymes such as tryptase. We investigated the role of mast cells in AMD using a model of CNV. Conditioned media from activated mast cells exerts proangiogenic effects on choroidal endothelial cells and choroidal explants. Laser-induced CNV in vivo was markedly attenuated in mice genetically depleted of mast cells (KitW−sh/W−sh) and in wild-type mice treated with mast cell stabilizer, ketotifen fumarate. Tryptase was found to elicit pronounced choroidal endothelial cell sprouting, migration and tubulogenesis; while tryptase inhibition diminished CNV. Transcriptomic analysis of laser-treated RPE/choroid complex revealed collagen catabolism and extracellular matrix (ECM) reorganization as significant events correlated in clusters of mast cell activation. Consistent with these analyses, compared to wildtype mice choroids of laser-treated mast cell-deficient mice displayed less ECM remodelling evaluated using collagen hybridizing peptide tissue binding. Findings herein provide strong support for mast cells as key players in the progression of pathologic choroidal angiogenesis and as potential therapeutic targets to prevent pathological neovascularization in ‘wet’ AMD.

Cromolyn sodium and masitinib combination inhibits fibroblast-myofibroblast transition and exerts additive cell-protective and antioxidant effects on a bleomycin-induced invitro fibrosis model.

Idiopathic pulmonary fibrosis (IPF) is a progressive and fatal fibrotic lung disease. While recent studies have suggested the potential efficacy of tyrosine kinase inhibitors in managing IPF, masitinib, a clinically used tyrosine kinase inhibitor, has not yet been investigated for its efficacy in fibrotic lung diseases. In a previous study on an invitro neurodegenerative model, we demonstrated the synergistic antitoxic and antioxidant effects of masitinib combined with cromolyn sodium, an FDA-approved mast cell stabilizer. This study aims to investigate the anti-fibrotic and antioxidant effects of the masitinib-cromolyn sodium combination in an invitro model of pulmonary fibrosis. Fibroblast cell cultures treated with bleomycin and/or hydrogen peroxide (H2O2) were subjected to masitinib and/or cromolyn sodium, followed by assessments of cell viability, morphological and apoptotic nuclear changes, triple-immunofluorescence labeling, and total oxidant/antioxidant capacities, besides ratio of Bax and Bcl-2 mRNA expressions as an indication of apoptosis. The combined treatment of masitinib and cromolyn sodium effectively prevented the fibroblast myofibroblast transition, a hallmark of fibrosis, and significantly reduced bleomycin / H2O2-induced apoptosis and oxidative stress. This study is the first to demonstrate the additive anti-fibrotic, cell-protective, and antioxidant effects of the masitinib-cromolyn sodium combination in an invitro fibrosis model, suggesting its potential as an innovative therapeutic approach for pulmonary fibrosis. Combination therapy may be more advantageous in that both drugs could be administered in lower doses, exerting less side effects, and at the same time providing diverse mechanisms of action simultaneously.

Targeting mast cell activation alleviates anti-MHC I antibody and LPS-induced TRALI in mice by pharmacologically blocking the TLR3 and MAPK pathway

Transfusion-related lung injury (TRALI) poses a significant risk following blood transfusion and remains the primary cause of transfusion-related morbidity and mortality, primarily driven by the activation of immune cells through anti-major histocompatibility complex class I (anti-MHC I) antibody. However, it remains to be defined how immune microenvironmental cue contributes to TRALI. Here, we uncover that activated mast cells within the immune microenvironment promote lung inflammation and injury in antibody-mediated TRALI, both in vitro and in vivo. This was demonstrated by co-culturing lipopolysaccharide (LPS)-pretreated mast cell line with anti-MHC I antibody and establishing a “two-hit” TRALI mouse model through intratracheal injection of LPS followed by tail-vein injection of anti-MHC I antibody. Importantly, mast cell-deficient KitW-sh/W-sh mice exhibited markedly reduced lung inflammation and injury responses in antibody-mediated TRALI compared with wild-type mice. Mechanistically, activation of toll-like receptor 3 (TLR3)/mitogen-activated protein kinase (MAPK) signaling pathway in mast cells contributes to the enhanced production of proinflammatory factors. These excessive proinflammatory factors produced by activated mast cells contribute to lung inflammation and injury in antibody-mediated TRALI. Pharmacologically targeting the TLR3/MAPK pathway to inhibit mast cell activation normalizes the proinflammatory microenvironment and alleviates lung inflammation and injury in the preclinical TRALI mouse model. Overall, we find that activation of mast cells via the TLR3/MAPK pathway contributes to lung inflammation and injury in antibody-mediated TRALI, providing novel insights into its underlying mechanisms. Furthermore, targeting activated mast cells and the associated pathway offers potential therapeutic strategies for antibody-mediated TRALI.