Mucosal Mast Cell Degranulation Research Articles

Ultrastructural evidence for piecemeal and anaphylactic degranulation of human gut mucosal mast cells in vivo.

One hundred and seventeen coded intestinal biopsies were examined by electron microscopy and evaluated for morphological evidence of mast cell and basophil secretion in situ. Sixty percent of the biopsies had evidence of secretion. Mast cell secretion was evident in control biopsies, many of which were obtained from uninvolved tissues of patients with inflammatory bowel disease. Biopsies of inflamed continent pouches from ulcerative colitis (UC) patients showed more mast cell secretion than noninflamed UC pouch biopsies. This evidence of mast cell secretion supports recent work that documents high constitutive levels of histamine in jejunal fluids of Crohn's disease patients and suggests a proinflammatory role for mast cells in inflammation associated with pouchitis.

Relationship between mast cell degranulation and jejunal myoelectric alterations in intestinal anaphylaxis in rats

The effects of two degranulators of mast cells and intestinal anaphylaxis on jejunal myoelectric activity were compared in rats fasted for 15 hours. Attempts to antagonize the motility changes were performed using antagonists of histamine and serotonin and a cyclooxygenase and lipoxygenase inhibitor. Hooded Lister rats were chronically fitted with electrodes implanted in the jejunal wall. A group of rats was sensitized to egg albumin and challenged 14 days later by intraduodenal infusion of antigen. Sensitized animals had serum titers ≥ 1:64. The other group was administered with mast cells degranulators. Both 48/80 (1 mg/kg), a degranulator of connective mast cells, and bromolasalocid (2 mg/kg), acting on connective and mucosal mast cells, induced a phase of total spiking inhibition followed by a progressive irregular spiking activity until the recovery of migrating myoelectric complex pattern (about 3 hours after injection). In contrast, antigen challenge disrupted the migrating myoelectric complex pattern, which was replaced by a peculiar pattern characterized by propagated spike burst, lasting 98 ± 11.3 minutes. Chlorpheniramine (1 mg/kg) antagonized only the inhibitory phase induced by degranulators and was ineffective on the intestinal anaphylaxis-induced motor changes. Methysergide (1 mg/kg) and indomethacin (5 mg/kg) significantly reduced the degranulator effects as well as the anaphylaxis-induced alterations of intestinal motility. It is concluded that anaphylaxis-induced motor disturbances are relevant to mucosal mast cell degranulation involving 5-hydroxytryptamine and arachidonic acid derivative products, whereas histamine release appears to be a minor component.

Immunologically activated chloride channels involved in degranulation of rat mucosal mast cells.

Crosslinking of type I Fc epsilon receptors (Fc epsilon RI) on the surface of basophils or mast cells initiates a cascade of processes leading to the secretion of inflammatory mediators. We report here a correlation between mediator secretion and the activation of Cl- channels in rat mucosal-type mast cells (line RBL-2H3). Stimulation of RBL cells by either IgE and antigen or by a monoclonal antibody specific for the Fc epsilon RI, resulted in the activation of Cl- ion channels as detected by the patch-clamp technique. Channel activation occurred slowly, within minutes after stimulation. The channel has a slope conductance of 32 pS at potentials between 0 and -100 mV, and an increasing open-state probability with increasing depolarization. Activation of apparently the same Cl- channels could be mimicked without stimulation by isolating inside-out membrane patches in tyrode solution. Parallel inhibition of both Cl- channel activity and mediator secretion, as monitored by serotonin release, was observed by two compounds, the Cl- channel blocker 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) and the anti-allergic drug cromolyn. NPPB inhibited both the antigen-induced Cl- current and the serotonin release, where half-maximal inhibition occurred at similar doses, at 52 microM and 77 microM, respectively. The drug cromolyn, recently found to inhibit immunologically induced mediator secretion from RBL cells upon intracellular application, also blocks Cl- channels (IC50 = 15 microM) when applied to the cytoplasmic side of an inside-out membrane patch. The observed Cl- channel activation upon immunological stimulation and the parallel inhibition of channel current and of serotonin release suggests a functional role for this Cl- channel in mediator secretion from the mast cells studied.

Mast Cell/Nerve InteractionsIn VitroandIn Vivo

In this report, we review the evidence for mast cell/nerve interactions. We believe that the morphologic and functional evidence now strongly support a purposeful and biologically significant interaction between these two cell types. This interaction has physiologic consequences and appears to be able to regulate such local events as chloride ion secretion by epithelial cells of the intestinal and respiratory tracts in experimental models. In this way, the mast cell and nerve may be considered as a functional homeostatic regulatory unit. The extent to which this unit may be involved in maintenance of normal integrity of mucous membranes or other structures, in health and in disease, is not clear at the present and will require considerably more investigation and elucidation. However, the concept of such an interaction is an interesting one and may bring new approaches of a therapeutic and diagnostic nature to bear on some old problems. The observations reviewed in this report that psychologic conditioning may itself cause mucosal mast cell degranulation and mediator release in the rat is significant. We consider that this evidence of central nervous system control of mast cell degranulation is an extension of the idea that mast cells and nerves communicate.

Gastric response to mucosal IgE-mediated reactions.

The effect of immunoglobulin E (IgE)-mediated anaphylaxis has been extensively studied in the small intestine, but little information is available on the response of the stomach to IgE-mediated mucosal reactions to food proteins. The effect of luminal antigenic challenge on gastric acid secretion, gastric emptying, and mucosal mast cell degranulation was examined in rats sensitized to egg albumin or in sham-treated controls. Intraluminal challenge of the stomach with egg albumin in sensitized animals significantly increased gastric acid secretion and delayed gastric emptying. The response was specific for the sensitizing antigen as challenge with bovine serum albumin was without effect. Sham-treated animals showed no response to egg albumin or bovine serum albumin. The increase in gastric acid secretion was reproduced by antigen challenge in naive animals passively transferred with hyperimmune serum. This effect was abolished by prior heat treatment of the serum. In sensitized animals challenged with egg albumin, there was histological evidence of mast cell degranulation in the stomach mucosa, increased intraluminal release of histamine, and increased serum levels of rat mast cell protease II, a marker specific for mucosal mast cell degranulation. The findings indicate that the stomach is a target organ for IgE-mediated reactions to food proteins. Antigen challenge in sensitized animals leads to increased gastric acid secretion and delayed emptying and evidence of mucosal mast cell activation.

The protective mechanism of FPL55712 against stress-induced gastric ulceration in rats.

Cold-restraint stress produced stomach ulceration which was accompanied by a decreased glandular mucosal mast cell count and blood flow. Pretreatment with FPL55712, a leukotriene antagonist, dose-dependently prevented ulcer formation and mast cell depletion; however, it did not affect the reduced mucosal blood flow, nor did it significantly influence acid secretion and pepsin output. The role of the leukotrienes in stress ulceration is discussed in the light of the findings with FPL55712 on gastric glandular mucosal mast cell degranulation.

5 MUCOSAL MAST CELLS IN THE BABY RAT JEJUNUM

Mucosal mast cells (MMC) are found mainly within the jejunal submucosa. MMC increase in intestinal graft-versus-host reaction, and their degranulation is observed In parasitic enteropathy. Weaning is associated with morphological changes resembling those seen in mild cell mediated intestinal injury. We studied MMC in the young rat jejunum in relation lo weaning changes. Jejunal morphology, crypl cell production rate, intraepithelial lymphocytes (IEL) and goblet cells were studied, and MMC were counted in rat mast cell protease II antibody (anti-RMCP II) and toluidine blue stained sections. Serum RMCP II was determined with a direct ELISA method. In the next few days following weaning the jejunal villi shortened by about 30%. crypts elongated by the same amount, crypt cell production rate more than doubled, and the numbers of IEL showed a marked increase. The MMC counts showed a steady increase from birth to the time of weaning, sharp fall on the two days immediately after weaning, and further increase thereafter. Serum RMCP II level displayed a sudden rise at the time of weaning, with gradual decline thereafter. Mucosal mast cell degranulation, obviously associated with the event of weaning, may be responsible for the observed morphological changes through epithelial cell damage. Weaning is a period of temporary loss of oral tolerance in rats; this might be a result of jejunal injury caused by mucosal masl cell protease.

Gastroprotective effect of pentacaine: role of mast cells.

Pentacaine was found to prevent the development of acute haemorrhagic lesions induced by ethanol in rats in a dose-dependent way. Electron microscopy in the untreated group showed extensive disruption of the surface epithelium and deep necrosis of the mucosa after ethanol exposure. Degranulation or even complete destruction of mast cells was observed. The microvasculature exhibited several signs of derangement. After pentacaine treatment, these signs were absent and no degranulation of mucosal mast cells was observed. The mast cell-mediated effect of pentacaine appears to be only one component of its gastroprotective action.

Mast cell protease release and mucosal ultrastructure during intestinal anaphylaxis in the rat

Intestinal anaphylaxis is associated with disturbances in gut function that are antigen-specific and dependent on mast cell degranulation. Using an animal model of intestinal anaphylaxis, we have correlated alterations in water and electrolyte transport, associated with intraluminal challenge, with specific intestinal mucosal mast cell degranulation by following systemic as well as local release of rat mast cell protease II. This protease is specific for intestinal mucosal mast cells and is known to selectively attack type IV collagen, which is found in basement membranes. Intraluminal antigen challenge in sensitized animals dramatically increased serum and intraluminal levels of rat mast cell protease II. Serum levels continued to rise throughout the duration of antigen challenge. Although light microscopy of challenged intestine demonstrated little distortion of mucosal architecture, ultrastructural examination revealed significant disruption to the basement membrane and underlying collagenous matrix of the intestinal mucosa. Our findings indicate that during mucosal immunoglobulin E-mediated reactions, rat mast cell protease II is released and is associated with ultrastructural changes in the intestinal mucosa. The systemic appearance of this specific protease provides a serum marker of intestinal anaphylaxis.

UPTAKE OF INGESTED PROTEINS FROM THE RAT CUT DURING A LOCALLY-INDUCED HYPERSENSITIVITY REACTION EVIDENCED BY RELEASE OF RAT MAST CELL PROTESE II

Hypersensitivity reactions to dietary proteins may lead to an increased permeability of the gut to macromolecular antigen molecules and thereby contribute to further sensitization. Rats sensitized to ovalbumin (OA) were tube-fed 1g of bovine serum albumin (BSA) and challenged one hour later with 100mg OA introduced directly into the jejunum. Serum samples taken after challenge were analysed for BSA by RIA, and for rat mast cell protease II (RMCP II), a specific marker for mucosal mast cell degranulation, by ELISA. One hour after antigen challenge the levels of RMCP II in the sera of challenged rats were significantly higher than both the pre-challenge levels and those of sham-challenged controls (p<0.002). There was no correlation between RMCP II levels and BSA levels in the challenged rats. We conclude that local hypersensitivity reactions in the rat gut may be unequivocally demonstrated by RMCP II release. However, a concomitant increased uptake of “bystander” antigen is not a consistent finding. A role for such an increased uptake in the aetiology of dietary intolerance is not supported by those results.

Intestinal anaphylaxis in the rat: jejunal response to in vitro antigen exposure.

In previous studies we showed that rats sensitized to egg albumin respond to in vivo intraluminal antigen with decreased net absorption of Na+, Cl-, and water. These abnormalities are associated with high serum levels of immunoglobulin E (IgE) antibodies and mucosal mast cell degranulation. In the present in vitro study electrical parameters, unidirectional fluxes of Na+ and Cl-, and levels of cAMP were determined in jejunum from sensitized and control rats during a basal period and after antigen addition. In Ussing chambers potential difference and short-circuit current increased significantly in tissue from sensitized rats after addition of 100 micrograms/ml of egg albumin to both mucosal and serosal surfaces. These changes were accompanied by a reversal of net Cl- absorption to net Cl- secretion. The presence of doxantrazole, a mast cell-stabilizing agent, in the buffer prevented these abnormalities. No changes occurred in response to antigen challenge in tissue from controls. In a further series of experiments the antigen was added only to the mucosal side of the tissue in Ussing chambers. In these studies short-circuit current increased after a lag period of approximately 25 min and was significantly increased (P less than 0.025) at 35 min. cAMP levels increased significantly in jejunal slices from sensitized rats exposed to antigen for 2 min. Our findings suggest that the in vivo transport abnormalities induced by IgE-mediated mucosal reactions to a food protein are related to antigen stimulation of a Cl- secretory process.

Effects of somatostatin on ethanol-induced gastric erosions in the rat: role of mast cells.

Somatostatin (S) inhibits hemorrhagic gastric erosions produced by ethanol. In this study we compared the dose-dependent effects of linear (reduced) and cyclic (oxidized) S with respect to mast cell degranulation. The gastric mucosal injuries were more inhibited by linear S than by cyclic S. But linear S aggravated injury at a certain dose (10(-7) mol/rat). Mucosal mast cell degranulation correlated significantly with the area of hemorrhagic mucosal lesions (r = 0.91). Both cytoprotection as well as aggravation potency of S may be connected to gastric mucosal mast cell activity in the rat.

Transport abnormalities during intestinal anaphylaxis in the rat: Effect of antiallergic agents

Our previous studies demonstrated that rats sensitized to egg albumin had reduced intestinal absorption of water and electrolytes in response to intraluminal antigen. The rapid onset of this effect and reduction in mucosal histamine and numbers of granulated mast cells in the lamina propria suggested a reaginic (IgE) mechanism involving mast cell mediators. In this study we examined the effect of antiallergic agents on the intestinal transport abnormalities in our model. Sensitized rats, 14 days after intraperitoneal injection of 10 μg of egg albumin plus alum had specific IgE serum titers ⩾ 1:64; control rats had no measurable IgE antibodies. Net fluxes of Na +, Cl −, and H 2O were determined by in vivo perfusion during a 1-hour antigen-free period and then a 1-hour antigen period. Sodium cromoglycate, administered intravenously (20 mg/kg) or in the perfusate (5 × 10 −4 mol/L) failed to prevent mucosal mast cell degranulation as evidenced by histamine release or the decrease in absorption of H 2O, Na +, and Cl − induced by antigen exposure. In contrast, 10 −3 mol/L of doxantrazole in the perfusate completely inhibited these changes. Histamine receptor antagonists, H 1, diphenhydramine, or H 2, cimetidine, in perfusates had no effect on the transport abnormalities. Our findings support a role for intestinal mucosal mast cells, but not connective tissue mast cells, in the pathogenesis of the intestinal dysfunction associated with mucosal IgE-mediated reactions to food proteins and suggest that mast cell mediators other than histamine are involved.

In situ degranulation of human nasal mucosal mast cells: Ultrastructural features and cell-cell associations

Mucosal mast cells in human nasal turbinates obtained at surgery were studied by electron microscopy before and after exposure en bloc to a degranulating stimulus. Mast cells occurred as solitary cells and in islets typically containing one mast cell and one to three mononuclear cells. All islets and most solitary mast cells were associated with thin cytoplasmic processes of distinctive, branched stromal cells that also phagocytosed debris from pyknotic mast cells. Mast cells were grouped into three categories based largely on published ultrastructural criteria. Resting (nondegranulating) mast cells possessed secretory granules that typically packed the cytoplasm and contained a densely stained amorphous material and scroll-like crystalline profiles. Granules in some resting cells were larger, more polymorphic, and appeared nearly homogeneous. Secretory granules of degranulating mast cells possessed crystalline, scroll-like, or reticular constituents predominantly with interspersed amorphous material. Granules were fewer in number and were often concentrated in the peripheral cytoplasm. Few degranulating mast cells exhibited labyrinth formation or exocytosis of granule contents. Largely degranulated mast cells had few if any typical granules but possessed small vacuoles containing recognizable granule remnants. Resting and degranulating mast cells possessed very long cell surface projections that, in places, interdigitated to form stacks of parallel folds above the plasmalemma, resulting in a threefold increase in cell-surface area. The surface membrane of the folds appeared to be continuous with that lining intracytoplasmic channels extending from the cell surface to the Golgi zone.

Effector cell heterogeneity in immediate hypersensitivity reactions.

Histologic heterogeneity in the basophil and mast cell populations has been apparent for many years. The advent of tissue culture and cell separation procedures has now made it possible to explore distinct populations of basophils and mast cells, as should be evident from this review. In fact, the logical extension of such technology is the requirement that cell preparations used in exploring basophil and mast cell function be carefully defined to permit comparison of data from one laboratory to another. While this is a practical application of the knowledge of the characteristics of heterogeneity, the implications for future developments in the understanding of basophil and mast cell function are more theoretical. While both basophils and mast cells respond to degranulating stimuli and antiallergic compounds in a similar fashion, as a rule, this review has noted several exceptions including the failure of disodium cromoglycate to prevent rat mucosal mast cell degranulation induced by immunologic stimuli. Such observations suggest that the ability of given drugs to inhibit allergic responses in one target organ and not in another may be due in part to differing mast cell responses. This hypothesis can be extended to variation in response among infants, children, and adults to such drugs as antihistamines. Furthermore, the variety of symptoms seen in individuals may be a reflection of differing responses of those individuals' mast cells from organ to organ. For instance, one subject with ragweed sensitivity might express this sensitivity as asthma, and a second subject with ragweed sensitivity might express this sensitivity as rhinitis. This would depend upon the ability of their pulmonary and upper airway mast cells to bind ragweed-specific IgE, degranulate to IgE-mediated stimuli, or to be regulated by intrinsic control mechanisms. Of a yet more speculative nature is the attempt to discern the basis for basophil, and particularly mast cell, heterogeneity. The function of the mast cell is unknown, but theories include the rejection of parasites; regulation and repair of connective tissue; regulation of the microvasculature; regulation of gastric acid secretion; limitation of delayed hypersensitivity reactions; and detoxification of surrounding tissues. Any or all of these theories may be correct, however, such a diversity of possible biologic roles for the mast cell suggests that mast cell subpopulations may have highly specialized functions reflected in stimuli that lead to their proliferation, their response to degranulating stimuli, and their mediator content.(ABSTRACT TRUNCATED AT 400 WORDS)

Human intestinal mucosal mast cells: expanded population in untreated coeliac disease.

Previous retrospective studies of intestinal mucosal mast cells in coeliac disease have given divergent results, and we have recently reported that inappropriate methodology could account for these discrepancies. In this prospective study, mucosal mast cell counts were performed in Carnoy fixed, peroral jejunal biopsy specimens from patients with coeliac disease, both untreated and treated with a gluten-free diet; and from controls (mainly irritable bowel syndrome). Mean mucosal mast cell count in 27 control subjects was 146/mm2, SD 29. Significantly higher values were obtained in untreated coeliac disease (mean 243, SD 41, p less than 0.001) returning to the normal range in coeliacs treated with a gluten-free diet with normal jejunal biopsy morphology. In seven patients mucosal mast cell counts were performed in multiple jejunal biopsies, and these showed that mucosal mast cell distribution was not patchy. There was no evidence of degranulation of intestinal mucosal mast cells under the conditions of routine biopsy (overnight fast). An increase in mucosal mast cells in untreated coeliac disease may be one explanation for the high number of IgE positive stained cells in the intestinal mucosa that has been reported by some authors.

The effect of disodium cromoglycate on rectal mucosal histamine release, eosinophil exudation and disease activity in active ulcerative colitis.

Disodium cromoglycate has been reported to benefit some patients with active ulcerative colitis. We have investigated the effect of cromoglycate in high dose on rectal mucosal mast cell degranulation (assessed indirectly by measurement of mucosal histamine release, using in vivo rectal dialysis, and the eosinophil content of the rectal inflammatory exudate) and on disease activity in eight patients in relapse. Although cromoglycate (1600 mg by mouth daily for 2 weeks) did not affect histamine release, it reduced (P less than 0.05) the eosinophil content of the exudate towards normal. Stool frequency also fell (P less than 0.05) but other aspects of bowel habit, sigmoidoscopic appearance, rectal mucosal potential difference and electrolyte transport were unaltered. Pre-treatment values of histamine release and eosinophil exudation could not be used to predict the response to cromoglycate. The reduction in eosinophil exudation produced by the drug could be due to inhibition of mast cell release of chemotactic factors other than histamine; the therapeutic effect of cromoglycate was unimpressive.

The influence of cimetidine, a histamine H 2-receptor antagonist, on the gastric effects of reserpine in rats

The effects of the graded doses of cimetidine on both resting and reserpine-evoked gastric acid secretion were examined in relation to their influence on reserpine-induced ulceration, mast cell degranulation and mucosal microcirculatory changes in rat stomachs. Cimetidine 10 mg/kg or above reduced resting or reserpine-provoked gastric acid secretion as well as rumenal and glandular ulceration. However, non-acid-inhibiting doses, 5 mg/kg or below, continued to prevent glandular ulceration. Reserpine-evoked gastric glandular mucosal mast cell degranulation was unaffected by both acid-inhibiting and non-acid-inhibiting doses of cimetidine which dose-dependently blocked the superficial glandular mucosal microcirculatory volume changes. These results suggest that cimetidine prevents reserpine-induced glandular ulceration largely by blocking the ulcerogenic effect of histamine H 2-receptor-mediated mucosal microcirculatory congestion, in contrast to antagonising rumenal lesions through acid inhibition; they also support the idea that reserpine may release histamine mainly from the glandular mucosal mast cells. The possibility of another antiulcer mechanism, due to cytoprotection, is discussed.

Disodium cromoglycate: a novel gastric antiulcer agent?

The influence of oral pretreatment with disodium cromoglycate (2.5 or 5 mg/kg × 9 doses) on the 4-h gatsric effects of i.p. injected reserpine (5 mg/kg) was examined in rats. Disodium cromoglycate markedly prevented reserpine-induced ulceration, mucosal mast cell degranulation and superficial mucosal microcirculatory changes in the glandular portion of the stomach wall. These interesting result point to the possibility that disodium cromoglycate may also have gatsric effects in man.

The effects of zinc sulphate on vagal-induced mast cell changes and ulcers in the rat stomach

The effects of zinc sulphate pretreatment on the formation of gastric ulcers, changes in intragastric pressure and changes in the gastric mucosal mast cell count induced by electrical vagal stimulation were studied in anaesthetized rats. Vagal stimulation produced a high gastric glandular ulcer incidence and ulcer index, increased the intragastric pressure, and reduced the number of granulated mast cells in the gastric mucosa and submucosa. Pretreatment with zinc sulphate (22, 44 ot 88 mg/kg), injected i.p. 48 h beforehand, reversed the changes in these parameters in a dose-related manner. These observations suggest that the inhibitory effects of zinc sulphate on mucosal mast cell degranulation may account for its ability to antagonise vagal-induced gastric glandular ulceration. The mechanisms involved in the aetiology of this type of gastric ulcer are discussed in the light of these results.