Mucosal Mast Cells Research Articles

Giardia excretory-secretory proteins modulate the enzymatic activities of mast cell chymase and tryptase

BackgroundMast cells are involved in the host immune response controlling infection with the non-invasive intestinal protozoan parasite Giardia intestinalis. Experimental infections in rodents with G. intestinalis showed increased intestinal expression of mucosal and connective mast cell specific proteases suggesting that both mucosal and connective tissue mast cells are recruited and activated during infection. During infection Giardia excretory-secretory proteins (ESPs) with immunomodulatory capacity are released. However, studies investigating potential interactions between Giardia ESPs and the connective tissue mast cell specific serine proteases, i.e. human chymase and mouse mast cell protease (mMCP)-4 and, human and mouse tryptase (mMCP-6) remain scarce. ResultsWe first investigated if soluble Giardia proteins (sGPs), which over-lap extensively in protein content with ESP fractions, from the isolates GS, WB and H3, could induce mast cell activation. sGPs induced a minor activation of bone marrow derived mucosal-like mast cells, as indicated by increased IL-6 secretion and no degranulation. Furthermore, sGPs were highly resistant to degradation by human tryptase while human chymase degraded a 65 kDa sGP and, wild-type mouse ear tissue extracts degraded several protein bands in the 10 to 75 kDa range. In striking contrast, sGPs and ESPs were found to increase the enzymatic activity of human and mouse tryptase and to reduce the activity of human and mouse chymase. ConclusionOur finding suggests that Giardia ssp. via enhancement or reduction of mast cell protease activity may modulate mast cell-driven intestinal immune responses. ESP-mediated modulation of the mast cell specific proteases may also increase degradation of tight junctions, which may be beneficial for Giardia ssp. during infection.

Enterocolic increase of cannabinoid receptor type 1 and type 2 and clinical improvement after probiotic administration in dogs with chronic signs of colonic dysmotility without mucosal inflammatory changes.

Colonic dysmotility in dogs can cause different GI signs. Sometimes, histology of enterocolic biopsies does not reveal inflammatory infiltrates or mucosal lesions that are typically associated with clinical disease activity. It is speculated that, similarly to humans, colonic dysmotility may be anxiety-based, although recent data demonstrate that irritable bowel syndrome (IBS) could result from acute infectious enteritis. Specific Lactobacillus spp. strains administered orally in humans induced the expression of μ-opioid and cannabinoid receptors in mucosal enterocytes, modulating intestinal morphine-like analgesic functions. We investigated the potential association of GI signs caused by colonic dysmotility and mucosal expression of cannabinoid receptors in intestinal epithelial cells and the number of mucosal mast cells. Ten to 15 endoscopic biopsies were collected from colonic mucosa of 20 dogs diagnosed with dysmotility disturbances before and after probiotic (Slab51 bacterial blend; Sivoy® ) administration (3-month period). Number and distribution of mast cells (MCs), and cannabinoid receptor type 1 (CB1) and type 2 (CB2) were evaluated by immunohistochemistry and PCR. Results were compared to data obtained from five clinically healthy dogs (archive samples). Decreased numbers of MCs (P<.0001) and increased CB1- and CB2-positive epithelial cells (P<.0001) in diseased dogs were positively associated with post-treatment CCECAI scores (P<.0001). Our results suggest that probiotic administration can reduce signs of colonic dysmotility, possibly due to microbiota modulation and epithelial cell receptor-mediated signaling in intestinal mucosa.

Potential effects of xylopic acid on acetic acid-induced ulcerative colitis in rats.

Xylopic acid (XA) has been reported to exhibit analgesic activity, alleviate neuropathic pain in rodents, and demonstrate anti-inflammatory effects. Intrarectal challenge of rats with acetic acid induces colitis that bears resemblance in terms of its pathogenesis, histopathology, and inflammatory profile to that in humans. Reactive oxygen species are implicated as the main driving force in this inflammatory bowel disease. This study aimed to investigate the anti-colitic potential of XA. We investigated the effect of XA on body weight, disease activity, inflammatory cell infiltration, and generation of reactive oxygen species. Rats were treated with XA or sulphasalazine, challenged intrarectally with acetic acid with macroscopic and microscopic findings made after eight days. Administration of XA to rats with colitis resulted in an increase in body weight with significant (p<0.05) improvement of the disease profile macroscopically. We observed decreased gross mucosal injury, reduced inflammation, and cellular proliferation with XA administration. Treatment with XA also resulted in decreased colonic epithelial expression of argyrophylic nucleolar organizer regions (AgNORs) with significant decrease (p<0.0001) in the quantitative expression of AgNORs/nucleus ratio to levels comparable with non-colitic control. We also observed reduced proliferation of mucosal mast cell in the colonic segment of the rats treated with XA. Treatment with XA also significantly (p<0.0001) increased the activity of SOD, CAT, and APx while it decreased the activity of MPO and MDA levels. Xylopic acid possesses anti-colitic activity in rats induced with acetic acid colitis.

Deletion of ΔdblGata motif leads to increased predisposition and severity of IgE-mediated food-induced anaphylaxis response.

BackgroundPrevious studies have revealed an important role for the transcription factor GATA-1 in mast cell maturation and degranulation. However, there have been conflicting reports with respect to the requirement of GATA-1 function in mast cell dependent inflammatory processes. Herein, we examine the requirement of GATA-1 signaling in mast cell effector function and IgE-mast cell-dependent anaphylaxis.ObjectiveTo study the requirement of GATA-1 dependent signaling in the development and severity of IgE-mast cell-dependent anaphylaxis in mice.MethodsWild type (Balb/c) and mutant ΔdblGata (Balb/c) mice were employed to study the role of GATA-1 signaling in in vitro IgE-mediated activation of bone marrow derived mast cells (BMMCs). Murine models of passive IgE-mediated and oral antigen-induced IgE-mediated anaphylaxis were employed in mice. Frequency of steady state mast cells in various tissues (duodenum, ear, and tongue), peritoneal cavity, and clinical symptoms (diarrhea, shock, and mast cell activation) and intestinal Type 2 immune cell analysis including CD4+ Th2 cells, type 2 innate lymphoid cells (ILC2), and IL-9 secreting mucosal mast cells (MMC9) were assessedResultsIn vitro analysis revealed that ΔdblGata BMMCs exhibit a reduced maturation rate, decreased expression of FcεRIα, and degranulation capacity when compared to their wildtype (WT) counterparts. These in vitro differences did not impact tissue resident mast cell numbers, total IgE, and susceptibility to or severity of IgE-mediated passive anaphylaxis. Surprisingly, ΔdblGata mice were more susceptible to IgE-mast cell-mediated oral antigen induced anaphylaxis. The increased allergic response was associated with increased Type 2 immunity (antigen-specific IgE, and CD4+ TH2 cells), MMC9 cells and small intestine (SI) mast cell load.ConclusionDiminished GATA-1 activity results in reduced in vitro mast cell FcεRIα expression, proliferation, and degranulation activity. However, in vivo, diminished GATA-1 activity results in normal homeostatic tissue mast cell levels and increased antigen-induced CD4+ Th2 and iMMC9 cell levels and heightened IgE-mast cell mediated reactions.

Possible Involvement of Intracellular Calcium-Independent Phospholipase A2 in the Release of Secretory Phospholipases from Mast Cells-Increased Expression in Ileal Mast Cells of Crohn's Disease.

Increased activity of secretory phospholipases A2 (sPLA2) type-II was previously observed in ileum of Crohn’s disease (CD). Our aims were to explore the involvement of calcium-independent (i)PLA2β in the release of sPLA2s from the human mast cell (MC) line (HMC-1) and investigate expressions of cytosolic (c)PLA2α, iPLA2β, sPLA2-IIA and sPLA2-V in MCs of CD ileum. The release of sPLA2 was investigated in HMC-1 by immunocytochemistry and ELISA. The expression intensities of PLA2s in mucosal MCs, and the proportion of PLA2-positive MCs, were investigated in normal ileum and in ileum from patients with CD by immunohistochemistry. The calcium ionophore-stimulated release of sPLA2-IIA and sPLA2-V from HMC-1 was reduced by the iPLA2-inhibitor bromoenol lactone. All four PLA2s were detectable in mucosal MCs, both in normal ileum and in CD, but the proportion of iPLA2β-containing mucosal MCs and the expression intensity of sPLA2-IIA was increased in CD. Results indicate that iPLA2β is involved in the secretion of sPLA2s from HMC-1, and suggest that iPLA2β-mediated release of sPLA2 from intestinal MCs may contribute to CD pathophysiology. Ex vivo studies on isolated mucosal mast cells are however needed to clarify the precise role of MC PLA2s in the inflammatory processes of CD.

IL-13–induced intestinal secretory epithelial cell antigen passages are required for IgE-mediated food-induced anaphylaxis

Food-induced anaphylaxis (FIA) is an IgE-dependent immune response that can affect multiple organs and lead to life-threatening complications. The processes by which food allergens cross the mucosal surface and are delivered to the subepithelial immune compartment to promote the clinical manifestations associated with food-triggered anaphylaxis are largely unexplored. We sought to define the processes involved in the translocation of food allergens across the mucosal epithelial surface to the subepithelial immune compartment in FIA. Two-photon confocal and immunofluorescence microscopy was used to visualize and trace food allergen passage in a murine model of FIA. Ahuman colon cancer cell line, RNA silencing, and pharmacologic approaches were used to identify the molecular regulation of intestinal epithelial allergen uptake and translocation. Human intestinal organoid transplants were used to demonstrate the conservation of these molecular processes in human tissues. Food allergens are sampled by using small intestine (SI) epithelial secretory cells (termed secretory antigen passages [SAPs]) that are localized to the SI villous and crypt region. SAPs channel food allergens to lamina propria mucosal mast cells through an IL-13-CD38-cyclic adenosine diphosphate ribose (cADPR)-dependent process. Blockade of IL-13-induced CD38/cADPR-dependent SAP antigen passaging in mice inhibited induction of clinical manifestations of FIA. IL-13-CD38-cADPR-dependent SAP sampling of food allergens was conserved in human intestinal organoids. We identify that SAPs are a mechanism by which food allergens are channeled across the SI epithelium mediated by the IL-13/CD38/cADPR pathway, regulate the onset of FIA reactions, and are conserved in human intestine.

Scratching Beneath the Surface: Linking Skin Pathology with Food Allergy

A link between atopic dermatitis and food allergy has long been suspected but remains elusive. In this issue, Leyva-Castillo etal. (2019) show how mechanical injury of the skin initiates a cascade of events that stimulate the expansion of mucosal mast cells and promote food anaphylaxis.

Transient Suppression and Functional Modulation of Mucosal Mast Cells for the Treatment of Allergic Disease

Abstract Food allergy is estimated to affect about 5 percent in children and adults and causes life-threatening reactions. Especially food allergy in infant and child shows severe gastrointestinal symptoms, including diarrhea, vomiting, and abdominal pain. To control the food allergy, allergen desensitization by oral immunotherapy (OIT), has been shown to an effective therapeutic approach; however, limited information is currently available for basic understandings of OIT induced immunological events at intestinal mucosa. In this study, we found that orally desensitized intestinal mast cells acquired the regulatory function for the inhibition of food allergic response. Indeed, the gene analysis showed the increase of regulatory cytokine IL-2 and IL-10 expressions in orally-desensitized intestinal mast cells; on the contrary, decreased Th2 cytokine expressions were suppressed by epigenetic control. Thereby, mice depleting these regulatory mast cells during OIT led to the reduction of functional regulatory T cell-mediated inhibition of allergic responses. These findings newly uncovered characteristics desensitized mast cells to acquire regulatory suppressive milieu in the allergic disease mucosa and indicated the novel roles of mast cells in the effective anti-allergic therapy.

The Role of Myofibroblasts and Mast Cells in Oral Mucosa Repair After Fractional Laser Treatment

The aimof this study was to clarify the features of the reparative process of the rat's oral mucosa in the later periods after fractional laser treatment with an analysis of the possible involvement of mast cells and myofibroblasts in this process.Material and methods.The samples of the oral mucosa of male Wistar rats (n=9) were used as a material for this study. Fractional laser treatment was carried out using stLase (DentalPhotonics, USA) with power P=7–10 W and pulse duration tp=100–200 ms (wavelength 980 nm). Histological sections of the oral mucosa from the control zones (unaffected) and zones treated with laser radiation were stained with hematoxylin and eosin and with Masson’s aniline blue. To identify mast cells, toluidine blue was used. For the detection of blood vessels and myofibroblasts, immunohistochemical reaction to smooth-muscle α-actin was performed.Results.On the 28th day after fractional laser treatment in rat oral mucosa the signs of incomplete repair were present. At this period in laser treated areas within the lamina propria rows of densely adjacent myofibroblasts were found. In the rows of myofibroblasts the mast cells are not visualized, while in the similar areas of the intact mucosa mast cells are present in large numbers.Conclusion.Fractional laser treatment stimulates the prolongation of regeneration process of rat oral mucosa while formation of myofibroblasts occurs in the laser treated zone. During this process mast cells may be involved. The ability to degranulate under laser radiation was shown.

Mast cell-mediated splanchnic cholestatic inflammation

Splanchnic mast cells increase in chronic liver and in acute-on-chronic liver diseases. We administered Ketotifen, a mast cell stabilizer, and measured the mast cells in the splanchnic organs of cholestatic rats. These groups were studied: sham-operated rats (S; n = 15), untreated microsurgical cholestasic rats (C; n = 20) and rats treated with Ketotifen: early (SK-e; n = 20 and CKe; n = 18), and late (SK-l; n = 15 and CK-l; n = 14). The cholestatic rats showed systemic and splanchnic impairments, such as ascites, portal hypertension, and biliary proliferation and fibrosis. The rats also showed a splanchnic increase of TNF-α, IL-1β and MCP-1, and a reduction of IL-4, IL-10 and antioxidants. An increase of VEGF in the ileum and mesenteric lymphatic complex was associated with a liver reduction of TGF-β1. Ketotifen reduces the degree of hepatic insufficiency and the splanchnic inflammatory mediators, as well as VEGF and TGF-ß1 levels. Ketotifen also reduces the connective tissue mast cells in the mesenteric lymphatic complex of cholestatic rats, while increases the hepatic mucosal mast cells. In cholestatic rats, Ketotifen improves liver function and ascites, and also reduces pro-inflammatory mediators in the splanchnic area. The decrease in connective tissue mast cells in the mesenteric lymphatic complex due to the administration of Ketotifen would lead to the improvement of the inflammatory splanchnic response, and consequently the abovementioned complications.

Inhibition of mast cell degranulation by melanin

Melanin is a dark naturally occurring pigment produced in nature and in many organisms. Although several reports have demonstrated applications for melanins in various therapeutic treatments, to date, no research has examined the anti-allergic effect of melanin. In this study, we for the first time found that solubilized or synthesized soluble melanin acts as a potent inhibitor of the degranulation of mast cells. We found that squid-ink-derived melanin significantly inhibited antigen-IgE-FcεRI-mediated degranulation of the mucosal mast cell line RBL-2H3. A homogenized melanin nanoparticle prepared by laser ablation also clearly suppressed antigen-induced mast cell degranulation. We also successfully solubilized synthetic melanin in a neutral biochemical buffer and found that it also significantly inhibited IgE-sensitized mast cells. The anti-degranulation activity of synthesized melanin was abolished in the melanin fraction below 50-kD molecular weight. All melanins used in this study did not exert significant cell death. Signal transduction analysis revealed that melanin suppressed antigen-triggered phosphorylation of signaling molecules as well as calcium influx. Transmission electron microscopy revealed that homogenized melanin nanoparticles partially attached to the cell surface and some nanoparticles were internalized to the cell. Flow cytometry revealed that the number of FcεRI-bound IgE molecules was decreased by melanin. Fluorescence recovery after photobleaching analysis indicated that melanin attenuated both plasma membrane and cytoplasmic fluidity, implying that melanin increased their viscosities. In vivo experiments using passive systemic anaphylaxis (PSA) and passive cutaneous anaphylaxis (PCA) mouse models demonstrated that oral administration of melanin accelerated the recovery of decreased body temperature after antigen infection in PSA, and combination sensitization of IgE with melanin attenuated antigen-induced extravasation in PCA. These findings indicated that melanin exhibits preventative effects against IgE-mast cell-mediated anaphylaxis. This study provides the first evidence that homogenized melanin may be a potential therapeutic agent for diseases involving mast cells.

Germ-Free Mice Exhibit Mast Cells With Impaired Functionality and Gut Homing and Do Not Develop Food Allergy.

Background: Mucosal mast cells (MC) are key players in IgE-mediated food allergy (FA). The evidence on the interaction between gut microbiota, MC and susceptibility to FA is contradictory.Objective: We tested the hypothesis that commensal bacteria are essential for MC migration to the gut and their maturation impacting the susceptibility to FA.Methods: The development and severity of FA symptoms was studied in sensitized germ-free (GF), conventional (CV), and mice mono-colonized with L. plantarum WCFS1 or co-housed with CV mice. MC were phenotypically and functionally characterized.Results: Systemic sensitization and oral challenge of GF mice with ovalbumin led to increased levels of specific IgE in serum compared to CV mice. Remarkably, despite the high levels of sensitization, GF mice did not develop diarrhea or anaphylactic hypothermia, common symptoms of FA. In the gut, GF mice expressed low levels of the MC tissue-homing markers CXCL1 and CXCL2, and harbored fewer MC which exhibited lower levels of MC protease-1 after challenge. Additionally, MC in GF mice were less mature as confirmed by flow-cytometry and their functionality was impaired as shown by reduced edema formation after injection of degranulation-provoking compound 48/80. Co-housing of GF mice with CV mice fully restored their susceptibility to develop FA. However, this did not occur when mice were mono-colonized with L. plantarum.Conclusion: Our results demonstrate that microbiota-induced maturation and gut-homing of MC is a critical step for the development of symptoms of experimental FA. This new mechanistic insight into microbiota-MC-FA axis can be exploited in the prevention and treatment of FA in humans.

Identification of Soluble Mediators in IgG-Mediated Anaphylaxis via Fcγ Receptor: A Meta-Analysis.

Background: Anaphylaxis is an acute and life-threatening allergic response. Classically and most commonly, it can be mediated by the crosslinking of allergens to immunoglobulin E (IgE)- high affinity IgE receptor (FcεRI) complex found mostly on mast cells. However, there is another pathway of anaphylaxis that is less well-studied. This pathway known as the alternative pathway is mediated by IgG and its Fc gamma receptor (Fcγ). Though it was not documented in human anaphylaxis, a few studies have found that IgG-mediated anaphylaxis can happen as demonstrated in rodent models of anaphylaxis. In these studies, a variety of soluble mediators were being evaluated and they differ from each study which causes confusion in the suitability, and reliability of choice of soluble mediators to be analyzed for diagnosis or therapeutic purposes. Hence, the objective of this meta-analysis is to identify the potential soluble mediators that are involved in an IgG-mediated anaphylaxis reaction.Methods: Studies related to IgG-mediated anaphylaxis were sourced from five search engines namely PubMed, Scopus, Ovid, Cochrane Library, and Center for Agricultural Bioscience International (CABI) regardless of publication year. Relevant studies were then reviewed based on specific inclusion factors. The means and standard deviations of each soluble mediator studied were then extracted using ImageJ or Get Data Graph Digitiser software and the data were subjected to meta-analysis.Results: From our findings, we found that histamine, serotonin, platelet activating factor (PAF), β-hexosaminidase, leukotriene C4 (LTC4), mucosal mast cell protease-1 (MMCP-1), interleukins (IL)-4,−6, and−13; tumor necrosis factor alpha (TNF-α), and macrophage inflammatory protein-1α (MIP-1α) were often being analyzed. Out of these soluble mediators, histamine, PAF, β-hexosaminidase, IL-6, and−13, MIP-1α and TNF-α were more significant with positive effect size and p < 0.001. As study effect was relatively small, we performed publication bias and found that there was publication bias and this could be due to the small sample size studied.Conclusion: As such, we proposed that through meta-analysis, the potential soluble mediators involved in rodent IgG-mediated anaphylaxis to be histamine, PAF, β-hexosaminidase, IL-6 and−13 and MIP-1α, and TNF-α but will require further studies with larger sample size.

Intestinal Mucosal Mast Cells: Key Modulators of Barrier Function and Homeostasis.

The gastrointestinal tract harbours the largest population of mast cells in the body; this highly specialised leukocyte cell type is able to adapt its phenotype and function to the microenvironment in which it resides. Mast cells react to external and internal stimuli thanks to the variety of receptors they express, and carry out effector and regulatory tasks by means of the mediators of different natures they produce. Mast cells are fundamental elements of the intestinal barrier as they regulate epithelial function and integrity, modulate both innate and adaptive mucosal immunity, and maintain neuro-immune interactions, which are key to functioning of the gut. Disruption of the intestinal barrier is associated with increased passage of luminal antigens into the mucosa, which further facilitates mucosal mast cell activation, inflammatory responses, and altered mast cell–enteric nerve interaction. Despite intensive research showing gut dysfunction to be associated with increased intestinal permeability and mucosal mast cell activation, the specific mechanisms linking mast cell activity with altered intestinal barrier in human disease remain unclear. This review describes the role played by mast cells in control of the intestinal mucosal barrier and their contribution to digestive diseases.

Expression and clinical significance of mast cells and related immune factors in interstitial cystitis/ bladder pain syndrome

Objective To investigate the significance of mast cells activation, and IL-6, IL-10, sialic acid-binding immunoglobulin-like lectins 8 (Siglec-8) expression in interstitial cystitis/bladder pain syndrome. Methods Bladder mucosal biopsy tissues of 21 patients with IC/BPS admitted to our hospital from March 2016 to October 2017 were taken as the IC/BPS group, and normal bladder mucosa biopsy tissues of 9 patients who underwent ureteroscopy were taken as the control group. In the IC/BPS group, there were 3 males and 18 females aged (56.2±3.4) years, and the patients' pain symptom score (PUF) was (24.6±3.6). Four males and five females in the control group were aged (63±5.1 )years. The infiltration of bladder mucosa mast cells and plasma cells in IC/BPS group and control group was observed by transmission electron microscopy. The expression of IL-6, IL-10 and Siglec-8 were detected by immunohistochemical staining. The relationship between the expression level of the immune index and the PUF score was analyzed. Results In the IC/BPS group, mast cells and plasma cells was observed in 18 of the 21 cases, but no mast cells or plasma cells were observed in the control group. Expression of IL-6 in IC/BPS group 1 case (-), 11 cases(+ ) and 9 cases (+ + ). IL-6 expression in the control group 9 cases were all (-). The difference of IL-6 expression between the IC/BPS group and the control group was statistically significant (P=0.001). Expression of IL-10 were 3 cases (-), 7 cases (+ + ), 11 cases (+ + ) in IC/BPS group, while 7 cases (-), 2 cases (+ + ), and 0 cases (+ + )in the control group, and there was significant difference between the two group (P=0.001). In the IC/BPS group, expression of Siglec-8 were 12 cases (-), 5 (+ ), 4 (+ + ), while in control group 7 cases (-), 2 (+ ), and 0 (+ + ), no statistically significant difference was found between the two group (P=0.214). The PUF score of the patients with IL-10 expression (-) was 19.7±2.1, lower than those with IL-10 (+ + ) (27.1+ 2.5, P<0.001). PUF of patients with IL-10 (+ ) was 22.7±1.8, lower than those with (+ + ) (P=0.001). The PUF score of IL-6 expression (-) was 21. The PUF score of IL-6 expression (+ ) was (23.2 + 3.2), lower than (+ + ) (26.7+ 3.1, P=0.025). The PUF score of (+ + ) IC/BPS patients was (26.6 ±2.4), higher than that of (+ ) IC/BPS patients (21.5±2.1, P<0.01). The PUF score of Siglec-8 expression (+ + ) and (+ ) in IC/BPS group was (21.3±2.0), lower than that of Siglec-8 (-) ( 27.0±2.3, P<0.01). Conclusions Mast cells and plasma cells were expressed in IC/BPS tissues. The expression of IL-6 and IL-10 was positively correlated with clinical symptoms, while the expression of Siglec-8 was negatively correlated with clinical symptoms. Key words: Interstitial cystitis/ bladder pain syndrome; Mast cell; Interleukin-6; Interleukin-10; Sialic acid-binding immunoglobulin-like lectins 8

IL-9 Integrates the Host-Candida Cross-Talk in Vulvovaginal Candidiasis to Balance Inflammation and Tolerance.

Vulvovaginal candidiasis (VVC) is a common fungal infection caused by Candida albicans. The antifungal therapy represents the standard of care but due to the high costs of treatment and to the inability to prevent recurrences, the development of alternative therapeutic approaches is much-awaited. Recently, we have shown that the pathogenesis of C. albicans in the gut is modulated by IL-9, a pleiotropic cytokine able to promote both inflammation and tolerance during C. albicans infection. Herein, by using a mouse model of VVC, we similarly demonstrated that IL-9 might exert a dual role in VVC by contributing to inflammation during the initial immune activation and promoting resolution thereafter. Specifically, IL-9 has a pro-inflammatory activity at the onset of VVC by promoting NLRP3 inflammasome activity and mucosal mast cells expansion but a tolerogenic role in the resolution phase by promoting IL-1Ra production and connective tissue mast cells activation. We further show that a timely IL-9 neutralization at the onset of the inflammatory response ameliorated symptoms and vaginal pathology. Given that vaginal fluids from patients with recurrent VVC had higher levels of IL-9, these findings, by providing novel insights into the pathogenesis of VVC, may pave the way for alternative therapeutic strategies based on IL-9 neutralization.

Upregulation of intestinal mucosal mast cells expressing VPAC1 in close proximity to vasoactive intestinal polypeptide in inflammatory bowel disease and murine colitis.

Mast cells (MCs) and vasoactive intestinal polypeptide (VIP) have been proposed as regulators of the intestinal barrier and inflammation. Our aim was to map the distribution in inflammatory bowel disease (IBD) and murine colitis. MCs, VIP, and VIP-receptors (VPACs) were quantified by immunofluorescence and enzyme-immunoassay (EIA) in ileal tissues (villus epithelium (VE) and adjacent VE, ie, VE next to the follicle-associated epithelium, (FAE)) from Crohn's disease (CD; n=16) and non-IBD patients, and in colonic specimens of ulcerative colitis (UC; n=12) and healthy controls (HCs). In addition, VIP levels were measured in plasma from HCs, non-IBD, and IBD in remission (CD n=30; UC n=30). Colon, ileum, and plasma from mice with dextran sulfate sodium (DSS)-induced colitis and control mice were analyzed likewise. FAE-adjacent VE in ileum of CD possessed more MCs (P<0.05) and MCs expressing VPAC1 (P<0.05), but not VPAC2, compared to controls. Both adjacent and regular VE of CD had more MCs co-localizing/in close proximity to VIP (P<0.05). In UC colon, more MCs (P<0.0005), MCs close to VIP (P<0.0005), and MCs expressing VPAC1 (P<0.05) were found compared to controls. VIP levels were elevated in plasma from CD and UC compared to controls (P<0.0005). Colon of DSS mice showed more MCs and MCs close to VIP (P<0.05) compared to control mice. In vitro experiments revealed MCs expressing VPACs and internalized VIP after 120minutes of VIP-stimulation. Communication between MCs and VIP is upregulated during IBD and mice colitis. In CD patients, the epithelium next to FAE seems to be more involved than the surrounding VE, suggesting increased MC-VIP-interactions in this intestinal region.

The Combination Therapy of Dietary Galacto-Oligosaccharides With Budesonide Reduces Pulmonary Th2 Driving Mediators and Mast Cell Degranulation in a Murine Model of House Dust Mite Induced Asthma.

Background: Dietary non-digestible galacto-oligosaccharides (GOS) suppress allergic responses in mice sensitized and challenged with house dust mite (HDM). Budesonide is the standard therapy for allergic asthma in humans but is not always completely effective.Aim: To compare the efficacy of budesonide or different doses of GOS alone or with a combination therapy of budesonide and GOS on HDM-allergic responses in mice.Methods:BALB/c mice were sensitized and challenged with HDM, while fed a control diet or a diet supplemented with 1 or 2.5 w/w% GOS, and either or not oropharyngeally instilled with budesonide. Systemic and local inflammatory markers, such as mucosal mast cell protease-1 (mMCP-1) in serum, pulmonary CCL17, CCL22, and IL-33 concentrations and inflammatory cell influx in the bronchoalveolar lavage fluid (BALF) were determined.Results: Budesonide or GOS alone suppressed the number of eosinophils in the BALF of HDM allergic mice whereas budesonide either or not combined with GOS lowered both eosinophil and lymphocyte numbers in the BALF of HDM-allergic mice. Both 1 w/w% and 2.5 w/w% GOS and/or budesonide suppressed serum mMCP-1 concentrations. However, budesonide nor GOS alone was capable of reducing Th2 driving chemokines CCL17, CCL22 and IL-33 protein levels in supernatants of lung homogenates of HDM allergic mice, whereas the combination therapy did. Moreover, IL-13 concentrations were only significantly suppressed in mice treated with budesonide while fed GOS. A similar tendency was observed for the frequency of GATA3+CD4+ Th2 and CD4+RORγt+ Th17 cells in the lungs of the allergic mice.Conclusion: Dietary intervention using GOS may be a novel way to further improve the efficacy of anti-inflammatory drug therapy in allergic asthma by lowering Th2 driving mediators and mast cell degranulation.

Genetic diversity between mouse strains allows identification of the CC027/GeniUnc strain as an orally reactive model of peanut allergy

Improved animal models are needed to understand the genetic and environmental factors that contribute to food allergy. We sought to assess food allergy phenotypes in a genetically diverse collection of mice. We selected 16 Collaborative Cross (CC) mouse strains, as well as the classic inbred C57BL/6J, C3H/HeJ, and BALB/cJ strains, for screening. Female mice were sensitized to peanut intragastrically with or without cholera toxin and then challenged with peanut by means of oral gavage or intraperitoneal injection and assessed for anaphylaxis. Peanut-specific immunoglobulins, T-cell cytokines, regulatory T cells, mast cells, and basophils were quantified. Eleven of the 16 CC strains had allergic reactions to intraperitoneal peanut challenge, whereas only CC027/GeniUnc mice reproducibly experienced severe symptoms after oral food challenge (OFC). CC027/GeniUnc, C3H/HeJ, and C57BL/6J mice all mounted a TH2 response against peanut, leading to production of IL-4 and IgE, but only the CC027/GeniUnc mice reacted to OFC. Orally induced anaphylaxis in CC027/GeniUnc mice was correlated with serum levels of Ara h 2 in circulation but not with allergen-specific IgE or mucosal mast cell protease 1 levels, indicating systemic allergen absorption is important for anaphylaxis through the gastrointestinal tract. Furthermore, CC027/GeniUnc, but not C3H/HeJ or BALB/cJ, mice can be sensitized in the absence of cholera toxin and react on OFC to peanut. We have identified and characterized CC027/GeniUnc mice as a strain that is genetically susceptible to peanut allergy and prone to severe reactions after OFC. More broadly, these findings demonstrate the untapped potential of the CC population in developing novel models for allergy research.

Short-term in vitro culture of purity and highly functional rat bone marrow-derived mast cells.

Mast cells (MCs) are responsible for the innate immune response. Rat MCs are more suitable than mouse MCs as models of specific parasite infection processes and ovalbumin-induced asthma. Rat peritoneum-derived MCs and RBL-2H3 cells (an MC cell line) are widely used in disease studies. However, the application of rat bone marrow-derived MCs (BMMCs) are poorly documented in terms of the methodology of rat BMMC isolation. Here, we describe a relatively rapid, efficient, and simple method for the cultivation of rat BMMCs. As compared to previous protocols, rat BMMCs produced with the proposed protocol exhibited advantages in differentiation, proliferation, lifespan, and functionality, which should prove useful for studies of mucosal MC diseases in specific rat models.