Induced Mast Cell Degranulation Research Articles

Topiramate Inhibits Capsaicin-Induced Mast Cell Degranulation and CGRP Release in Rat Dura Mater.

Migraine is a disease that stands out for its high prevalence and socioeconomic costs. It involves the entire trigeminovascular system, the signaling substances, and their targets. However, the role of meningeal mast cells in migraine is still unclear. To better understand one of the components of neurogenic inflammation underlying migraine pathophysiology, we developed an in vivo rat model in which the dura mater was exposed bilaterally to investigate the influence of topiramate on capsaicin-induced mast cell degranulation and CGRP release from dura mater. On the day of the experiment, rats were anesthetized, and a craniectomy was performed on each parietal bone. Test substances were applied in situ over the dura mater using the right and left sides of the dura mater for the test and control, respectively. After exposure, the dura mater was processed for mast cell staining and counting. Using this setup, the effect of capsaicin (10-3 M) was evaluated in rats of both sexes, and subsequently the effect of in situ (10-3 M, 20 µL) and (20 mg/kg/day for 10 days) topiramate treatment on mast cell degranulation and CGRP release were evaluated. In both female and male rats, there was a greater amount of degranulated mast cells in the side stimulated by capsaicin compared to the control side in both females (18 ± 3% vs. 74 ± 3%; p = 0.016) and males (28 ± 2% vs. 74 ± 3%, p = 0.016). In the group treated with topiramate for 10 days prior to the experiments, capsaicin did not induce mast cell degranulation (control 20 ± 1% vs. capsaicin 22 ± 1%, p = 0.375) in contrast to animals treated for 10 days with gavage control (control 25 ± 1% vs. capsaicin 76 ± 1%, p = 0.016). Topiramate applied in situ concomitant with capsaicin did not protect the mast cells from degranulation in response to capsaicin (38 ± 2% vs. 44 ± 1%, p = 0.016). There was a significant reduction in CGRP release from the dura mater in the group treated with topiramate for 10 days compared to the control. This study demonstrates a novel experimental model wherein systemic administration of topiramate is observed to modulate the impact of capsaicin on meningeal mast cell degranulation.

Comparative analyses of various IgE-mediated and non-IgE-mediated inducers of mast cell degranulation for in vitro study.

In vitro investigations of mast cell (MC) degranulation are essential for studying many diseases, particularly allergy and urticaria. Many MC-degranulation inducers are currently available. However, there is no previous systematic comparative analysis of these available inducers in term of their efficacies to induce MC degranulation. Herein, we performed systematic comparisons of efficacies of five well-known and commonly used MC-degranulation inducers. RBL-2H3 cells were sensitized with 50ng/ml anti-DNP IgE or biotinylated IgE followed by stimulation with 100ng/ml DNP-BSA or streptavidin, respectively. For non-IgE-mediated inducers, the cells were treated with 5µg/ml substance P, compound 48/80, or A23187. At 15-, 30-, 45- and 60-min post-induction, several common MC-degranulation markers (including intracellular [Ca2+], β-hexosaminidase release, tryptase expression by immunofluorescence staining, cellular tryptase level by immunoblotting, secretory tryptase level by immunoblotting, CD63 expression by immunofluorescence staining, and CD63 expression by flow cytometry) were evaluated. The data showed that all these markers significantly increased after activation by all inducers. Among them, A23187 provided the greatest degrees of increases in intracellular [Ca2+] and β-hexosaminidase release at all time-points and upregulation of CD63 at one time-point. These data indicate that all these IgE-mediated (anti-DNP IgE/DNP-BSA and biotinylated IgE/streptavidin) and non-IgE-mediated (substance P, compound 48/80, and A23187) inducers effectively induce MC degranulation, while A23187 seems to be the most effective inducer for MC degranulation.

Characterization of the Molecular Diversity and Degranulation Activity of Mastoparan Family Peptides from Wasp Venoms.

Wasp stings have become an increasingly serious public health problem because of their high incidence and mortality rates in various countries and regions. Mastoparan family peptides are the most abundant natural peptides in hornet venoms and solitary wasp venom. However, there is a lack of systematic and comprehensive studies on mastoparan family peptides from wasp venoms. In our study, for the first time, we evaluated the molecular diversity of 55 wasp mastoparan family peptides from wasp venoms and divided them into four major subfamilies. Then, we established a wasp peptide library containing all 55 known mastoparan family peptides by chemical synthesis and C-terminal amidation modification, and we systematically evaluated their degranulation activities in two mast cell lines, namely the RBL-2H3 and P815 cell lines. The results showed that among the 55 mastoparans, 35 mastoparans could significantly induce mast cell degranulation, 7 mastoparans had modest mast cell degranulation activity, and 13 mastoparans had little mast cell degranulation activity, suggesting functional variation in mastoparan family peptides from wasp venoms. Structure-function relationship studies found that the composition of amino acids in the hydrophobic face and amidation in the C-terminal region are critical for the degranulation activity of mastoparan family peptides from wasp venoms. Our research will lay a theoretical foundation for studying the mechanism underlying the degranulation activity of wasp mastoparans and provide new evidence to support the molecular design and molecular optimization of natural mastoparan peptides from wasp venoms in the future.

Abstract B018: The development of macrocyclic peptide Ras inhibitors

Abstract While there has been tremendous progress developing small molecule covalent K-Ras G12C inhibitors that bind Ras-off (GDP) and inhibit nucleotide exchange, there remains a need for additional K-Ras inhibitors with different Ras isoform/mutant selectivity profiles and Ras-on (GTP) mechanisms of inhibition. Of particular interest are compounds with a pan-K-Ras profile that inhibit multiple oncogenic mutants and wild type K-Ras, but achieve a therapeutic window by sparing H and N-Ras. In addition, addressing cancers driven by K-Ras oncogenic mutants beyond G12C, K-Ras amplification, or upstream dysregulation where K-Ras is maintained in the GTP-bound state will require “Ras-on” compounds that prevent effector binding. We were interested in the potential of peptide macrocycles that bind the Switch II pocket of K-Ras, in a different conformation than the small molecules, to inhibit Ras-on by allosterically blocking Raf and other effector binding. KRpep-2d is a 19-residue disulfide bridged peptide macrocycle discovered by phage display against K-Ras G12D bound to GDP. Due to its high molecular weight (2561 Da), polarity and metabolic instability, KRpep2d is inactive in in vitro cell assays. And while biochemical experiments have shown that it binds the Ras-off GDP-bound conformation of K-Ras with high affinity to inhibit nucleotide exchange, it has low affinity for K-Ras GTP and does not allosterically inhibit Raf binding. Using parallel macrocyclic peptide chemistry, we made 180 analogs of KRpep-2d to explore truncation of the exocyclic residues and replacement of the labile disulfide bond. UNP-6752, the best compound from this exploratory library, had K-Ras affinities similar to KRpep2d with a 50-fold K-Ras GDP/GTP preference, but was only 11 residues and cyclized with a non-reducible linker. Critically this compound lacked the eight arginine residues of KrPep-2d, which both preclude passive-cell permeability and induce mast cell degranulation. A second iteration of 350 compounds designed to explore single or multiple amino acid substitutions in the core of UNP-6752 led to compound UNP-7572 with sub-micromolar affinity for K-Ras GTP and an IC50 in a K-Ras-Raf TR-FRET assay of ~800 nM. Working from the UNP-7572 we are tuning selectivity for K-Ras and our progress further improving allosteric Ras-Raf inhibition, cell permeability and activity will be presented. Citation Format: Kristopher Josephson, Anthony Silvestri, Jennifer L. Fribourgh, Walter M. Bray, Brian David, Cameron C. Hanna, Sai Kumar Chakka, Alex Hyatt, Monique Porcella, Dio Dieppa-Matos, Cameron R. Pye, Joshua Schwochert. The development of macrocyclic peptide Ras inhibitors [abstract]. In: Proceedings of the AACR Special Conference: Targeting RAS; 2023 Mar 5-8; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Res 2023;21(5_Suppl):Abstract nr B018.

Brucella abortus induces mast cell activation through TLR-2 and TLR-4

The Gram-negative bacteria Brucella abortus is a major cause of brucellosis in animals and humans. The host innate immune response to B. abortus is mainly associated with phagocytic cells such as dendritic cells, neutrophils, and macrophages. However, as mast cells naturally reside in the main bacterial entry sites they may be involved in bacterial recognition. At present, little is known about the role of mast cells during B. abortus infection. The role of the innate immune receptors TLR2 and TLR4 in activation of mast cells by B. abortus (strain RB51) infection was analyzed in this study. The results showed that B. abortus did not induce mast cell degranulation, but did induce the synthesis of the cytokines IL-1β, IL-6, TNF-α, CCL3, CCL4, and CCL5. Furthermore, B. abortus stimulated key cell signaling molecules involved in mast cell activation such as p38 and NF-κB. Blockade of the receptors TLR2 and TLR4 decreased TNF-α and IL-6 release by mast cells in response to B. abortus. Taken together, our results demonstrate that mast cells are activated by B. abortus and may play a role in inducing an inflammatory response during the initial phase of the infection.

SARS-CoV-2 vaccine excipients polyethylene glycol and trometamol do not induce mast cell degranulation, in an in vitro model for non-IgE-mediated hypersensitivity.

The development of vaccines against SARS-CoV2 brought about several challenges, including the management of hypersensitivity reactions to these formulations. The search for underlying mechanisms involved in these adverse events initially focused on excipients which may trigger mast cell activation responses via non-IgE pathways: polyethylene glycol and trometamol. We sought to determine whether these components, in their pure form, were capable of stimulating mast cells directly. To test this hypothesis, we used an in vitro model for non-IgE-mediated activation that has previously shown degranulation responses induced via MRGPRX2 with known drug agonists of the receptor. Human LAD2 mast cells were incubated with different concentrations (1, 10, 50 mg/ml) of trometamol and of purified polyethylene glycol/Macrogol (molecular weights: 2,000, 3,350, 4,000, and 6,000). Mast cell degranulation was assessed using a beta-hexosaminidase read-out. Interestingly, degranulation responses for all reagents tested showed no significant differences from those obtained from the negative control (basal degranulation). Receptor-silencing assays were therefore not conducted. In summary, purified PEG and trometamol did not induce mast cell degranulation in this in vitro model for the study of non-IgE mechanisms of drug hypersensitivity, previously shown to be useful in the investigation of MRGPRX2 ligands. Studies using complete vaccine formulations, lipid conjugates, and receptor gene variants are needed to further clarify mechanisms of vaccine hypersensitivity.

IgE-Binding and Immunostimulating Properties of Enzymatic Crosslinked Milk Proteins as Influenced by Food Matrix and Digestibility.

Dairy foods are essential in the diet, although in some susceptible individuals they may cause allergy to cow's milk proteins. Therefore, alternative methods are sought to reduce their allergenicity. Transglutaminase (TG) is widely used in dairy products mainly to improve texture. Although it has been claimed that TG can be used to modify the digestibility and allergenicity of foods, its impact within a real matrix has been rarely studied. The aim of this work was to assess the allergenic potential of crosslinked skim milk (SM), milk casein fraction (CN), and whey protein (WP). To this purpose, inhibition ELISA with sera from milk allergic patients, in vitro activation tests of mouse mast cells and splenocytes, and simulated gastrointestinal digestion assays were performed. The results showed that cross-linking increased the binding of IgE to WP, but decreased IgE-binding to SM and CN. However, no differences were observed in the ability of cross-linked proteins to induce mast cell degranulation compared to native proteins. The cross-linking of SM and CN reduced Th2 cytokine release from the splenocytes of sensitized mice. All TG-treated samples exhibited more resistance to in vitro digestion than the untreated proteins and the human IgE binding capacity after digestion was higher. In conclusion, TG treatment of milk proteins does not reduce the risk of eliciting allergic symptoms in cow's milk allergic patients.

Assessment of the safety of the cationic arginine-rich peptides (CARPs) poly-arginine-18 (R18 and R18D) in ex vivo models of mast cell degranulation and red blood cell hemolysis

Our laboratory focuses on the development of novel neuroprotective cationic peptides, such poly-arginine-18 (R18: 18-mer of l-arginine; net charge +18) and its d-enantiomer R18D in stroke and other brain injuries. In the clinical development of R18/R18D, their cationic property raises potential safety concerns on their non-specific effects to induce mast cell degranulation and hemolysis. To address this, we first utilised primary human cultured mast cells (HCMCs) to examine anaphylactoid effects. We also included as controls, the well-characterised neuroprotective TAT-NR2B9c peptide and the widely used heparin reversal peptide, protamine. Degranulation assay based on β-hexosaminidase release demonstrated that R18 and R18D did not induce significant mast cell degranulation in both untreated (naïve) and IgE-sensitised HCMCs in a dose-response study to a maximum peptide concentration of 16 μM. Similarly, TAT-NR2B9c and protamine did not induce significant mast cell degranulation. To examine hemolytic effects, red blood cells (RBCs), were incubated with the peptides at a concentration range of 1–16 μM in the absence or presence of 2% plasma. Measurement of hemoglobin absorbance revealed that only R18 induced a modest, but significant degree of hemolysis at the 16 μM concentration, and only in the absence of plasma. This study addressed the potential safety concern of the application of the cationic neuroprotective peptides, especially, R18D, on anaphylactoid responses and hemolysis. The findings indicate that R18, R18D, TAT-NR2B9c and protamine are unlikely to induce histamine mediated anaphylactoid reactions or RBC hemolysis when administered intravenously to patients.

Effect of low-intensity pulsed ultrasound on mast cell degranulation and fibroblast expression on type 2 diabetes mellitus rats wound healing process

Impaired wound healing is one of the Diabetes mellitus complications. Low-intensity Pulsed Ultrasound (LIPUS) therapy may accelerate the impaired wound healing. The use of LIPUS therapy in the early inflammatory phase can induce mast cell degranulation, and in the proliferative phase it can increase collagen synthesis by fibroblasts. The purpose of this study was to determine the effects of LIPUS therapy on mast cell degranulation and fibroblastexpression in the healing process of punch biopsy wound in rats with type 2 diabetes mellitus. Twenty-four Sprague dawley (n=24) were designed into type 2 diabetes mellitus by injecting Nicotinamide and Streptozotocin, then divided into six groups: diabetes mellitus without LIPUS (DM3, DM7, DM14) and diabetes mellitus with LIPUS (DML3, DML7, DML14), 4 each, and punch biopsy wounds were made on the dorsal skin. The DML group received LIPUS therapy in the wound area (frequency 3 MHz, intensity 0.5 W/cm2, duty cycle 20%, duration 3 minutes every day for 3 days (DML3), 7 days (DML7), and 14 days (DML14). The wounded tissue area was stained with toluidine blue to observe mast cell degranulation and immunohistochemical type HSP-47 to observe fibroblast expression. Two-Way ANOVA and Post Hoc LSD tests were used to determine the differences in mast cell degranulation and fibroblast expression. The results showed that mast cell degranulation and fibroblast expression in the DML group were higher than in the DM group (table 1). Pearson test showed a correlation between mast cell degranulation and fibroblast expression (p=0.00; r= 0.839). LIPUS therapy increases mast cell degranulation and fibroblast expression in type 2 diabetes mellitus rat model. The higher the mast cell degranulation, the higher fibroblast expressions.

Role of Mast Cells in HIV-associated Chronic Widespread Pain

Chronic widespread pain (CWP) in people with HIV-1 (PWH) is associated with a high rate of disability and low quality of life, with prevalence estimates ranging from 25%-85%. However, the specific mechanisms that contribute to CWP in HIV are not understood. Previously we demonstrated that PWH with CWP have increased hemolysis, elevated plasma levels of cell-free heme, and attenuated levels of the heme degrading enzyme, heme oxygenase 1 (HO-1). We also found that high cell-free heme and low HO-1 were associated with hypersensitivity in animals. In this study, we hypothesized that high heme and low HO-1 activates mast cells leading to degranulation and release of pain mediators like, histamine and bradykinin in HIV. PWH who self-report CWP were recruited from the University of Alabama at Birmingham HIV clinic. Animal models used in the study included, C57BL/6 mice subjected to hemolysis by intraperitoneal injection of phenylhydrazine hydrochloride (PHZ), HO-1 knockout mice, and HIV-1 transgenic rats overexpressing HIV-1 proteins. Cellular studies were performed in MC/9 mast cells. Human findings show that PWH with CWP have elevated plasma levels of histamine and bradykinin. HIV-1 transgenic rats also have elevated plasma levels of histamine and bradykinin and are hypersensitive as estimated by a decrease in withdrawal threshold to von Frey stimulation. In addition, data in mice with elevated plasma levels of free-heme, or decreased expression of HO-1, show significantly elevated levels of the pain mediators, histamine and bradykinin. Cellular data shows that high heme or low HO-1 induces mast cell degranulation and the release of histamine and bradykinin. Together, these data suggest that heme-dependent mast cell degranulation and the release of pain mediators may be contributing CWP in HIV. Future studies will focus on determining strategies to stabilize mast cell membrane in HIV and reduce hypersensitivity. Grant Number: RO1DA049657 Grant Name: Chronic Widespread Pain in HIV: Novel Mechanisms and Therapeutics.

Fulminant Ehrlichia chaffeensis Infection in a Patient Discovered to Have Indolent Systemic Mastocytosis

Abstract Patients with systemic mastocytosis (SM) are at increased risk for anaphylaxis from vespid and honeybee stings. The response of mastocytosis patients to tick-borne infections has not been reported. A 77-year-old woman hospitalized for progressive symptoms of fever, chills, night sweats, and abdominal pain became obtunded and required intubation, support with pressors, treatment for disseminated intravascular coagulation, and resuscitation from 3 episodes of asystole. During her evaluation, bone marrow biopsy revealed heretofore undiagnosed SM. A history of a tick bite 3 weeks previously was elicited when Ehrlichia chaffeensis infection was diagnosed. Institution of treatment with doxycycline led to gradual recovery. Ehrlichia chaffeensis–induced mast cell degranulation was felt to be responsible for many of her symptoms. This case is a cautionary report of the potential danger of tick-borne infection in patients with SM. How widespread this problem occurs is unknown and will depend on future surveillance in endemic areas.

In Vitro Antioxidant and Mast Cell Stabilizing Activity of Different Extracts of Vanda Spathulata Flowers

Objective: The objective of the present work is to evaluate the petroleum ether, ethyl acetate and methanolic extracts of vanda spathulata on experimental models for in vitro antioxidant activity and mast cell stabilizing activity. Methods: Mast cell stabilization effect was assessed using compound 48/80 induced mast cell degranulation in rat peritoneal mast cell. The antioxidant activity was evaluated using 2, 2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging assay. Results and Discussion: Vanda spathulata methanolic extract (VSME) at doses of 200 and 400mg/kg exhibited significant protection (p<0.01) and petroleum ether (VSPE) and ethylacetate (VSEA) extracts at the dose of 400mg/kg showed significant protection (p<0.05) against mast cell degranulation. VSME revealed better DPPH radical scavenging activity (IC50 value 38.39 µg/ml) which was closely resembled to standard ascorbic acid (IC50 value 33.98 µg/ml) when compared to other extracts. Phytochemical study revealed the presence of alkaloids, saponins, flavonoids, tannins, steroids and glycosides. Conclusion: From the results Vanda spathulata shows mast cell stabilizing activity mainly due to phytochemical constituents and strong antioxidant property of plant extracts.

Immunodominant conformational and linear IgE epitopes lie in a single segment of Ara h 2

Contribution of conformational epitopes to the IgE reactivity of peanut allergens Ara h 2 and Ara h 6 is at least as important as that of the linear epitopes. However, little is known about these conformational IgE-binding epitopes. We investigated the distribution of conformational epitopes on chimeric 2S-albumins. Recombinant chimeras were generated by exchanging structural segments between Ara h 2 and Ara h 6. Well-refolded chimeras, as verified by circular dichroism analysis, were then used to determine the epitope specificity of mAbs by performing competitive inhibition of IgG binding. Furthermore, we delineated the contribution of each segment to the overall IgE reactivity of both 2S-albumins by measuring the chimeras' IgE-binding capacity with sera from 21 patients allergic to peanut. We finally assessed chimeras' capacity to trigger mast cell degranulation. Configuration of the conformational epitopes was preserved in the chimeras. Mouse IgG mAbs, raised against natural Ara h 6, and polyclonal human IgE antibodies recognized different conformational epitopes distributed all along Ara h 6. In contrast, we identified human IgG mAbs specific to different Ara h 2 linear or conformational epitopes located in all segments except the C-terminal one. The major conformational IgE-binding epitope of Ara h 2 was located in a segment located between residues 33 and 81 that also contains the major linear hydroxyproline-containing epitope. Accordingly, this segment is critical for the capacity of Ara h 2 to induce mast cell degranulation. Chimeric 2S-albumins provide new insights on the conformational IgE-binding epitopes of Ara h 2 and Ara h 6. Proximity of the immunodominant linear and conformational IgE-binding epitopes probably contributes to the high allergenicpotency of Ara h 2.

Pain and Cellular Migration Induced by Bothrops jararaca Venom in Mice Selected for an Acute Inflammatory Response: Involvement of Mast Cells.

Bothrops jararaca venom (BjV) can induce mast cell degranulation. In order to investigate the role of mast cells and the interference of the host genetic background in the inflammation induced by BjV, we have used mouse strains selected for maximal (AIRmax) or minimal (AIRmin) acute inflammatory response (AIR). Mice were pretreated with an inhibitor of mast cell degranulation, cromolyn (CROM), and injected in footpads or intraperitoneally (i.p.) with BjV. Pain was measured with von Frey hairs, cell migration in the peritoneum by flow cytometry, and reactive oxygen species (ROS) production by chemiluminescence assays. The nociceptive response to BjV was higher in AIRmax than AIRmin mice; however, this difference was abolished by pretreatment with CROM. BjV induced peritoneal neutrophil (CD11b+ GR-1+) infiltration and ROS secretion in AIRmax mice only, which were partially inhibited by CROM. Our findings evidence a role for mast cells in pain, neutrophil migration, and ROS production triggered by BjV in AIRmax mice that are more susceptible to the action of BjV.

SARS-CoV-2-triggered mast cell rapid degranulation induces alveolar epithelial inflammation and lung injury

SARS-CoV-2 infection-induced hyper-inflammation links to the acute lung injury and COVID-19 severity. Identifying the primary mediators that initiate the uncontrolled hypercytokinemia is essential for treatments. Mast cells (MCs) are strategically located at the mucosa and beneficially or detrimentally regulate immune inflammations. In this study, we showed that SARS-CoV-2-triggered MC degranulation initiated alveolar epithelial inflammation and lung injury. SARS-CoV-2 challenge induced MC degranulation in ACE-2 humanized mice and rhesus macaques, and a rapid MC degranulation could be recapitulated with Spike-RBD binding to ACE2 in cells; MC degranulation altered various signaling pathways in alveolar epithelial cells, particularly, the induction of pro-inflammatory factors and consequential disruption of tight junctions. Importantly, the administration of clinical MC stabilizers for blocking degranulation dampened SARS-CoV-2-induced production of pro-inflammatory factors and prevented lung injury. These findings uncover a novel mechanism for SARS-CoV-2 initiating lung inflammation, and suggest an off-label use of MC stabilizer as immunomodulators for COVID-19 treatments.

Mast Cell Stabilizing, Antianaphylactic and Bronchodilatory activity of Methanolic extract of Averrhoa carambola Fruit

This work was mainly aimed to study the mast cell stabilizing, anti-anaphylactic and bronchodilatory activities of methanolic extract of Averrhoa carambola (ACME). Mast cell stabilization activity was investigated by Compound 48/80 induced mast cell degranulation in rats and antianaphylactic activity was performed by determining the mortality rate of mice upon exposure to compound 48/80. The bronchodilatory effect of ACME was studied on histamine aerosol-induced bronchospasm using guinea pigs, in which occurrence of preconvulsive dyspnea (PCD) was noted as end point. Treatment with ACME (100, 200 and 400mg/kg) showed significant (p<0.05) protection of rat peritoneal mast cells and significantly (p<0.05) reduced the mortality of mice in a dose dependent manner. ACME significantly (p<0.05) increased the time of preconvulsive dyspnea (PCD) in a dose dependent manner that suggestive of bronchodilating activity. Phytochemical studies observed presence of saponins, tannins, steroids, alkaloids, flavonoids and glycosides. From these finding, we concluded that ACME possesses mast cell stabilizing; anti anaphylactic and bronchodilatory activity which might be used in treatment of asthma.

Use of capsaicin as a model in the study of migraine: a literature review

Capsaicin is able to induce mast cell degranulation, an event probably related to the pathophysiologyof a migraine attack. The present review study aimed to address the mechanisms of action of capsaicin and other chemical inducers in mast cell degranulation and an interaction of nerves and events that happen in the dura mater with the activation of mast cells. A survey was carried out in the literature, from 1980 to 2019, in different databases, using the following terms: capsaicin, mast cell and dura mater. 36 articles were selected for this review. Studies indicate that the main mechanisms of action of capsaicin are chemical induction through the activation of TRPV1 channels,allowing calcium influx into neurons in the trigeminal ganglion of the dura mater, activating mast cell degranulation, releasing pro-inflammatory (e.g., histamine, oxide nitric) and vasoactive (e.g., CGRP and substance P) substances. Therefore, the use of capsaicin may be a tool to be used in an animal model to better understand the pathophysiology of migraine.

PD01-07 MEDICAL-GRADE MANUKA HONEY (MEDIHONEY®) POTENTIAL EFFICACY IN THE MANAGEMENT OF INTERSTITIAL CYSTITIS/BLADDER PAIN SYNDROME: AN IN VITRO MODEL OF UROTHELIAL INFLAMMATION

You have accessJournal of UrologyInfections/Inflammation/Cystic Disease of the Genitourinary Tract: Interstitial Cystitis (PD01)1 Sep 2021PD01-07 MEDICAL-GRADE MANUKA HONEY (MEDIHONEY®) POTENTIAL EFFICACY IN THE MANAGEMENT OF INTERSTITIAL CYSTITIS/BLADDER PAIN SYNDROME: AN IN VITRO MODEL OF UROTHELIAL INFLAMMATION Muhammadbakhoree Yusuh, Laurie Lau, Omar Abdelwahab, Kamaluddeen Garba, Prapussara Sirikhansaeng, Brian Birch, and Bashir Lwaleed Muhammadbakhoree YusuhMuhammadbakhoree Yusuh More articles by this author , Laurie LauLaurie Lau More articles by this author , Omar AbdelwahabOmar Abdelwahab More articles by this author , Kamaluddeen GarbaKamaluddeen Garba More articles by this author , Prapussara SirikhansaengPrapussara Sirikhansaeng More articles by this author , Brian BirchBrian Birch More articles by this author , and Bashir LwaleedBashir Lwaleed More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000001965.07AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Interstitial Cystitis/Bladder Pain Syndrome (IC/BPS) refers to “spontaneous” conditions of varied aetiologies and defined as chronic pelvic pain, pressure or discomfort, urge to void, frequency and nocturia. It is characterised by bladder wall inflammation, defective bladder urothelium, and increased levels of the pro-inflammatory cytokine TNF-α in the bladder tissue. The release of TNF-α by the urothelial cells further aggravates the inflammatory response in patients with IC/BPS. Commonly, Medical grade Manuka honey (Medihoney® (MH)) possesses strong pro-angiogenic and anti-microbial properties. Our recent findings have highlighted its anti-inflammatory effects through inhibition of induced mast cell degranulation. In addition, we demonstrated that MH is well tolerated by the rat urothelial cells where it was cytoprotective against acid damage. In the present study, we investigated the cytotoxic and anti-inflammatory effects of MH using an in vitro model of primary human urothelial cells (HUCs). METHODS: Cytotoxic effect of various concentrations of MH on HUCs at 1, 24, 48, and 72 hours was evaluated using MTT assay. In addition, the release of the pro-inflammatory cytokines IL-8 and IL-6 in the HUCs after 24-hour incubation with TNF-α (10 ng/ml) with/without pre-treatment with 4% MH was assessed using ELISA assays. RESULTS: One hour incubation with MH (2%, 4%, 6% and 8% w/v) did not induce significant cytotoxic effects on the HUCs. However, when the cells were incubated with MH for a longer periods, we observed an incremental cytotoxic effect, especially at 6% and 8% (Figure 1). In addition, 4% MH significantly decreased the TNF-α induced release of IL-6 and IL-8 in the urothelial cells (Figure 2). CONCLUSIONS: Our results indicate that MH is not cytotoxic to urothelial cells and has potential use as an intravesical therapeutic agent for the management for bladder inflammatory conditions such as IC/BPS. Source of Funding: N/A © 2021 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 206Issue Supplement 3September 2021Page: e33-e33 Advertisement Copyright & Permissions© 2021 by American Urological Association Education and Research, Inc.MetricsAuthor Information Muhammadbakhoree Yusuh More articles by this author Laurie Lau More articles by this author Omar Abdelwahab More articles by this author Kamaluddeen Garba More articles by this author Prapussara Sirikhansaeng More articles by this author Brian Birch More articles by this author Bashir Lwaleed More articles by this author Expand All Advertisement Loading ...

Itch in Hymenoptera Sting Reactions.

Insect stings and the resulting itch are a ubiquitous problem. Stings by members of the insect order Hymenoptera, which includes sawflies, wasps, bees and ants, and especially by bees and wasps are extremely common, with 56–94% of the population being stung at least once in their lifetime. The complex process of venom activity and inflammation causes local reactions with pain and pruritus, sometimes anaphylactic reactions and more seldomly, as in case of numerous stings, systemic intoxication. We reviewed the literature regarding itch experienced after Hymenoptera stings, but found no study that placed a specific focus on this topic. Hymenoptera venoms are composed of many biologically active substances, including peptide toxins and proteinaceous toxins. Peptide toxins from bee venom cause cell lysis and ion channel modulation in the peripheral and central nervous systems, while toxins from wasp venom induce mast cell degranulation and chemotaxis of polymorphonuclear leukocytes in the skin. The proteinaceous toxins cause a disruption of the cell membranes and necrotic cell death, degradation of hyaluronan (an extracellular matrix glycosaminoglycan), increased vascular permeability, hemolysis, as well as activated platelet aggregation. Mediators which could be directly involved in the venom-induced pruritus include histamine and tryptase released from mast cells, interleukin-4 and interleukin-13 from Th2 lymphocytes, as well as leukotriene C4. We postulate that a pruriceptive itch is induced due to the pharmacological properties of Hymenoptera venoms.

Corrigendum] Cryptotanshinone inhibits IgE‑mediated degranulation through inhibition of spleen tyrosine kinase and tyrosine‑protein kinase phosphorylation in mast cells

Following the publication of the above article, an interested reader drew to the authors' attention that they had mentioned that activated PKCδ phosphorylates IKKβ in order that IKKβ is relocated to the plasma membrane, resulting in the induction of mast cell degranulation; however, four references the authors had included did not seem to support this statement. The authors have re-examined their paper, and realized that the four references the reader mentioned were indeed cited incorrectly, and wish to rectify this error through revising the third paragraph in the Discussion section, the References section, and an associated figure (Fig.6C) in order to avoid any further misunderstandings on the part of the readership. First, the authors wish to revise the wording of the third and fourth paragraphs of the Discussion, as featured on pp.1101-1102, to the following (changed text is indicated in bold): 'We showed that CRT exerts anti-AD effect through inhibition of the mast cell degranulation in mast cells. Upon IgE/antigen stimulation, the immunoreceptor tyrosine-based activation motif (ITAM) region of FcεRI receptor which is on the mast cell surface is phosphorylated and the initial signalling protein kinases Lyn and Syk are recruited to the ITAM (28,29). Then, the activated Lyn and Syk leads to phosphorylation of the transmembrane adaptor linker for activation of T cells (LAT). Phosphorylated LAT which is a scaffold for multimolecular signalling complexes and activates PLCγ through phosphorylation. The activated PLCγ hydrolyses phosphatidylinositol biphosphate (PIP2) to generate second signalling molecules IP3 and DAG, which activate PKCs including PKCδ to induce the mast cell degranulation (30,31). On the other hand, cross-linking of FcεRI also activates IKKβ, which moves to the lipid raft fractions and phosphorylates synaptosomal-associated protein23 (SNAP-23) leading to degranulation (7). Since PKCδ phosphorylates IKKα, but not IKKβ (32), it is not likely that two signalling pathways are directly connected. In this study, novel function of CRT on phosphorylations of Lyn/Syk kinases in mast cells is elucidated for the first time. Furthermore, it is likely that this inhibitory effect of CRT on Lyn/Syk kinases negatively affected activities of their downstream signalling molecules including PLCγ, PKCδ, and IKKβ, which leads to decrease in mast cell degranulation by CRT treatment. Besides the inhibitory effect of CRT on mast cell degranulation, here we provide additional evidence that CRT exerts anti-AD effects through inactivation of MAPK and NF‑κB. It has been reported that CRT regulates the activities of MAPK and NF‑κB in various cell types. In rhabdomyosarcoma, hepatoma, and breast carcinoma, CRT activates MAPK p38/JNK and suppresses ERK1/2, followed by caspase-independent apoptosis (10,33,34). In chronic myeloid leukaemia cells, CRT enhances TNF‑α-induced apoptosis through the activation of MAPK p38 (35). In smooth muscle cells, CRT exerts anti-migration/invasion effect as it inhibits TNF‑α/NF‑κB signalling pathway (36).' Secondly, the authors wish to make the following changes to the Reference list: New references 30-32 have been inserted to the list, as follows: 30. Ozawa K, Szallasi Z, Kazanietz MG, BlumbergPM, MischakH, Mushinski JF and Beaven MA: Ca2+-dependent and Ca2+-independent isozymes of protein kinase C mediate exocytosis in antigen-stimulated rat basophilic RBL-2H3 cell. J Biol Chem268: 1749-1756, 1993. 31. Cho SH, Woo CH, Yoon SB and Kim JH: Protein kinaseCδ functions downstream of Ca2+ mobilization in FcεRI signaling to degranulation in mast cells. J Allergy Clin Immunol114: 1085-1092, 2004. 32. Yamaguchi T, Miki Y and Yoshida K: Protein kinaseCδ activates IκB-kinaseα to induce the p53 tumor suppressor in response to oxidative stress. Cell Signal19: 2088-2097, 2007. The addition of these new references means that the former references 30-33 have been accordingly renumbered to references 33-36. Finally, the authors have revised Fig.6C, as it appeared on p.1102, in order to assist the understanding of the readers, and the corrected version of Fig.6 appears on the next page. All these corrections have been approved by all the authors, with the exception of the first author, Sumiyasuren Buyanravjikh, who is no longer uncontactable. The authors regret that these errors were included in the paper, even though they did not substantially alter any of the major conclusions reported in the study, are grateful to the Editor for allowing them this opportunity to publish a Corrigendum, and apologize to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 18: 1095‑1193, 2018; DOI: 10.3892/mmr.2018.9042].