Mast cells from human gastric mucosa: a comparative study with lung and colonic mast cells.

Mast cells isolated from human gastric mucosa released histamine on challenge with IgE-directed ligands and calcium ionophores but were essentially unresponsive to a variety of non-immunological stimuli. Moreover, immunologically induced histamine secretion from these cells was inhibited by a number of anti-allergic agents including anti-asthmatic chromones, beta-adrenoceptor agonists and phosphodiesterase inhibitors. In total, these data indicate that mast cells from the human gastric mucosa are in many respects functionally similar to their lung and colonic counterparts.     

Mast cells in tissue healing: from skin to the gastrointestinal tract

Mast cells are largely found at interfaces between the environment and the internal milieu. Early knowledge of the mast cell suggested a role predominantly associated with allergy and pathologic response to antigens, but more recent research has shown a myriad of functions is likely. Wound healing is a complex process of lysis and reconstitution controlled by a series of cell signalling proteins. Mast cells have been shown to play a significant role in the early inflammatory stage of wound healing and also influence proliferation and tissue remodelling in skin. Emerging work implicates the mast cell as a modulator of intestinal healing particularly following surgical anastomosis. The study of mast cells and wound healing involves the use of cell studies and animal models through the use of mast cell inhibitors, promoters and mast cell deficient rodent strains. This review addresses wound healing in skin and the gastrointestinal tract and specifically identifies data pertaining to the role of the mast cell in the process of cell breakdown, repair and regeneration.     

Effect of enalapril on the skin response to bradykinin in man.

We tested the effect of oral enalapril on intradermal bradykinin to determine if kininase II inhibition occurs with therapeutic doses in vivo. Six normal male volunteers took either 5 mg enalapril orally or placebo on 2 days. Three hours later bradykinin was injected into the skin of the back in doses increasing from 10(-11) to 10(-9) M. Enalapril increased the bradykinin-induced wheal. Inhibition of kininase II may cause accumulation of endogenous bradykinin. This could be an important mechanism in the occasionally reported side effect of angioedema with the angiotensin converting enzyme (ACE) inhibiting group of drugs.     

Mast cells from human colonic mucosa and submucosa/muscle: a comparison with human lung mast cells.

The functional properties of enzymically dispersed mast cells from human lung parenchyma, colonic mucosa and colonic submucosa/muscle were compared. In general, the cells responded in a similar fashion to the histamine releasing action of a variety of immunological and non-immunological stimuli. However, some differences were observed in their responses to anti-allergic compounds. In total, these data indicate that there are subtle variations between human colonic and pulmonary mast cells but that these differences are much less sharply defined than in the rodent.     

Prostaglandin D2 release from human skin mast cells in response to ionophore A23187

1. Cells were dispersed from human foreskin by proteolytic digestion and enriched or depleted in mast cell content by density gradient flotation on discontinuous gradients of Percoll. 2. Cells were harvested at six interfaces on the density gradient. Mast cell purity ranged from 0.6-85.0%, compared to 5.5% in the unfractionated cells. 3. Challenge of the cells with the calcium ionophore A23187 resulted in release of both histamine and prostaglandin D2 (PGD2). In fractions depleted of mast cells, histamine release and net PGD2 generation were low, but increasing amounts of these mediators were released as mast cell purity was increased up to 59%. 4. Overall, there was a significant correlation between the net generation of PGD2 and histamine (r = 0.9234, P less than 0.001) and also between PGD2 release and mast cell number (r = 0.7475, P less than 0.001). 5. These data provide the first direct evidence of the capacity of the human cutaneous mast cell to synthesize and release PGD2.     

Differential release of histamine and eicosanoids from human skin mast cells activated by IgE-dependent and non-immunological stimuli.

1. Cells were dispersed from human foreskin using a mixture of collagenase and hyaluronidase and separated into mast cell-depleted (less than 1%) or enriched (greater than 75%) preparations by density-gradient centrifugation. 2. Challenge of gradient fractions with epsilon-chain-specific anti-human IgE stimulated the release of histamine, prostaglandin D2 (PGD2) and leukotriene C4 (LTC4). The release of eicosanoids was significantly correlated with that of histamine, suggesting that they are derived from the mast cell population of the dispersate. In highly purified (76.2 +/- 4.2%) mast cell preparations, maximum net release of histamine, PGD2 and LTC4 was 3432 +/- 725, 84.9 +/- 10.8 and 6.6 +/- 1.2 pmol/10(6) nucleated cells. 3. The non-immunological stimuli substance P, vasoactive intestinal peptide (VIP), somatostatin, compound 48/80, morphine and poly-L-lysine released similar amounts of histamine to anti-IgE, but 12 to 21 fold less PGD2 and LTC4. 4. These studies suggest that IgE-dependent and non-immunological stimuli activate human skin mast cells by different secretory mechanisms, a hypothesis supported by our previous findings of differences in Ca2+ requirements and time-course of histamine release. Activation by the non-immunological mechanism may be of importance in vivo due to the close anatomical association between skin mast cells and dermal nerve-terminals containing neuropeptides.     

Mast cell histamine secretion in response to somatostatin analogues: structural considerations

Comparative studies of the activity of somatostatin and of several of its analogues in releasing histamine from rat mast cells suggest that the integrity of the positively charged amino terminus and of lysines at positions four and nine of somatostatin may be necessary to preserve its histamine releasing activity. D-Lys4 substitution reduced activity by 80% while D-Lys9 substitution increased it four fold. Replacement of the amino terminal Ala by Tyr or simultaneous removal of Ala1,Gly2 and the amino portion of the now terminal Cys3 inhibited the activity by about 95%. Finally, dihydrosomatostatin retained 33% activity while an analogue where both Cys sulfur atoms were permanently blocked by acetamidomethyl groups retained only about 13% activity. Using information from these studies and from the literature, two-dimensional and space-filling models approximating the conformation of somatostatin were constructed and compared with a plausible corresponding model of the hexameric form of 48/80, the most active congener of this classic mast cell secretagogue. By such modelling it was possible to show that the orientation of the cationic moieties in the two molecules could be similarly arranged thereby perhaps explaining the ability of these compounds to induce mast cell secretion.     

Tissue response as a functional discriminator of receptor heterogeneity: effects of mixed receptor populations on Schild regressions.

A model is described that predicts the behavior of competitive antagonists in tissues with more than one receptor mediating response. The receptor stimuli for two receptor types are summed and processed, via a cellular stimulus-response mechanism, into tissue response. The primary receptor is described by a standard Langmurian isotherm, and a secondary receptor input with a variable maximal strength, for which the agonist has variable sensitivity, is added. The prediction of drug effects in this system does not depend on the way in which the two stimuli are combined or on the absolute magnitudes of the parameters used to make the calculations. The model is maximally flexible, in that no pharmacological significance is put on the magnitudes of the inputs from the secondary receptor system (i.e., they can vary with either agonist intrinsic efficacy, receptor number, or efficiency of stimulus-response coupling). The theoretical Schild regressions for selective antagonists in two-receptor systems are calculated for various secondary receptor inputs. These regressions generally are curvilinear whenever the secondary receptor significantly contributes to agonist response. These calculated data also indicate that minor variations in biological input from secondary receptor systems would obscure curvature in the Schild regression and result in a seemingly linear regression with a slope of less than unity. However, further calculations indicate three possible ways to use Schild analysis to detect receptor heterogeneity in tissues. One indicator of receptor heterogeneity is a change in the slope of the dose-response curve for the agonist in the presence of a selective antagonist. A second indicator would be a marked heteroscedasticity of errors in the Schild regression, i.e., the magnitude of the standard errors in the ordinate values would depend upon the concentration of the antagonist. A third, and most experimentally accessible, aspect of heterogeneous receptor systems predicts that changes in the overall sensitivity of organ response mechanisms will differentially alter the relative strength of two receptor inputs. This would be observed as a change in the potency of an antagonist. Under these circumstances, differences in the stimulus-response characteristics of a tissue would result in a change in the Schild regression for a selective antagonist. These concepts are discussed in terms of the use of Schild analysis in functional systems for the detection of physiologically relevant mixtures of receptors and the possible advantages over biochemical binding data.     

The mast cells of the newborn rat diaphragm and their response to histamine liberators.

The mast cell population of rat diaphragm was estimated between birth and adulthood and found to rise with an increase in the age of rat studied. Degranulation of these cells was observed in rats from all age groups, following treatment with compound 48/80 and dextran. The association of mast cells with the blood vessel wall in adult rat diaphragm was not observed in the comparable tissues of newborn rats. These findings are discussed in relationship to the poor vascular permeability reactions exhibited by newborn and young rats.     

Plasma membrane adenosine triphosphatases in rat peritoneal mast cells and macrophages--the relation of the mast cell enzyme to histamine release.

Adenosine triphosphatases activated by calcium or magnesium have been demonstrated on the outer surface of rat peritoneal mast cells and macrophages. The plasma membrane ATPases in the two types of cells have similar but not identical properties. Mg²⁺ is somewhat more effective than Ca²⁺ in stimulating both the enzymes. They are not influenced by sodium and potassium and not inhibited by ouabain and oligomycin. Ethacrynic acid inhibits both, but the mast cell enzyme is more sensitive to it. The enzyme on the macrophage has five to thirty-seven times higher activity (average seventeen times) than that on the mast cell. The apparent K_m of the enzymes in intact cells, incubated with adenosine triphosphate for 5 min, is estimated to be 36 μM for mast cells and 30 μM for macrophages. The optimal pH for the mast cell and the macrophage enzymes is 6.7 and 7.1 respectively. The activities of the two enzymes rise similarly with temperature up to 37° but differ at 47°, the macrophage enzyme being less active at this temperature than at 37°. Phosphatidyl serine, which stimulates anaphylactic and dextran-induced histamine release, causes about 40 per cent stimulation of the plasma membrane ATPase of mast cells in the absence of Ca²⁺ and Mg²⁺ but has no appreciable effect in their presence. No change in the mast cell enzyme could, however, be observed in relation to histamine release induced by dextran, compound $\text{48}\text{80}$ and ATP. But ethacrynic acid, which in 1 mM concentration inhibits 50 per cent of the mast cell enzyme activity, also causes pronounced inhibition of histamine release induced by all the three agents in the same concentration. The inhibition is not influenced by the presence of glucose, suggesting that ethacrynic acid does not inhibit histamine release by blocking energy metabolism. Ethacrynic acid apparently acts at another site. The site of action could very well be plasma membrane ATPase. There is also a correlation between the inhibition of the mast cell enzyme by sodium fluoride and lack of calcium and their inhibitory effect on histamine release. The possible involvement of the plasma membrane ATPase of mast cells in the process of exocytosis leading to histamine release is discussed.     

Long-term cultures of mast cells: a new model for studying the allergic response

Mast cells are the key cells of allergic reactions and probably play also a role in chronic inflammatory reactions resulting in fibrosis. Although their biochemical and functional properties have been extensively investigated, several studies have been hampered by the absence of a tissue culture system to keep these cells alive and functionally active for long periods of time. Recently we have developed an in vitro system in which rat peritoneal mast cells are cocultured with 3T3 mouse skin derived fibroblasts (MC/3T3). Under these tissue culture conditions, mast cells do not proliferate and maintain their viability and functional activity for more than a month. This system allowed us to carry out long-term studies on the functional properties of mast cells. We have found that mast cells activated both by IgE-dependent and IgE-independent stimuli survive, and slowly replenish their histamine content. After a non-immunological challenge mast cells retain their full potential to release histamine upon a repeated similar challenge. In contrast, immunologically challenged mast cells become partially unresponsive to a similar activation event for up to 3 weeks. By exploiting this long-term culture system we described a novel type of mast cell activation induced by cytokines. The onset of this activation is slow and its course appears to be chronic-continuous, very different from the classical anaphylactic type activation that is completed within few minutes. Since MC/3T3 release higher amounts of histamine, this system is a sensitive tool to investigate the antiallergic properties of various drugs. By employing MC/3T3 cultures we were able to show that the gold salt auranofin inhibits histamine release from mast cells stimulated by different secretagogues. In addition, salbutamol inhibited histamine release from repeatedly challenged mast cells; and nedocromil sodium was effective in preventing mast cell activation when incubated for a week with MC/3T3.     

Design, syntheses and properties of nucleic Acid switch in response to external stimuli

The control of molecular properties using external stimuli is an attractive research area that offers the potential for regulation of various biological phenomena. This review summarizes the concept, design, syntheses and properties of nucleic acid switches in response to external stimuli, namely, "external-stimuli-responsive bridged nucleic acids monomer". From (1)H-NMR experiments, every external-stimuli-responsive bridged nucleic acid monomer was found to have an N-conformation, while an S-conformation was predominantly observed after exposure to a proper stimulus. Each bridged nucleic acid monomer was effectively introduced into oligodeoxynucleotides using an automated DNA synthesizer. Moreover, oligonucleotides modified with these bridged nucleic acid monomer were changed in their hybridization property and tolerance to enzymatic digestion in response to each stimulus. These results clearly showed that external-stimuli-responsive bridged nucleic acids monomers should work as a nucleic acid switch, and have the potential for regulation of various biological phenomena.     

Allergy or inflammation? From neuropeptide stimulation of human skin mast cells to studies on the mechanism of the late asthmatic response

This short review examines two examples of studies into the mechanisms of allergic responses which have particular relevance to inflammation research. The first is the ability of human skin mast cells, but not those derived from lung, adenoids, tonsils or intestine, to release histamine in response to stimulation by neuropeptides including substance P, vasoactive intestinal polypeptide (VIP) and somatostatin. The neuropeptide activation site does not appear to be a classical tachykinin receptor but rather a binding site of low affinity and low specificity capable of interacting with neuropeptides and compounds with similar physicochemical characteristics. In contrast to IgE-dependent activation, neuropeptide stimulation of skin mast cells induces a rapid release of histamine with minimal generation of PGD2 and LTC4. This pseudo-allergic reaction is thought to underlie the weal and flare response in the skin and may have a role in urticaria. The second example describes studies to elucidate the mechanisms of the late asthmatic response by use of a guinea-pig model. As in man, both early and late phase responses in the guinea-pig are inhibited by sodium cromoglycate whereas only the early response is inhibited by the beta-adrenoceptor stimulant drug salbutamol. Examination of bronchoalveolar fluid has shown a temporal relationship between an airways neutrophilia and the late response. However, pharmacological manipulation and the use of an anti-neutrophil serum has shown that these events are not interdependent. The role of the airways eosinophilia requires further investigation.     

Progress in allergy signal research on mast cells: signal regulation of multiple mast cell responses through FcepsilonRI

The crosslinking of FcepsilonRI by IgE and antigen (Ag) on mast cells initiates activation cascades that lead to allergic responses. Although it was thought that IgE binding to FcepsilonRI is a passive "sensitization", recent reports suggest that IgE actively promotes mast cell survival in the absence of Ag. However, it is largely unknown how these distinct responses are delivered through the same receptor, FcepsilonRI, depending on the types of stimli. As an underlying molecular mechanism for the generation of diverse responses through FcepsilonRI, we found that the quantity and the duration of the signal through the FcepsilonRI gamma chain (FcRgamma) determine different mast cell responses. Furthermore, FcRgamma-mediated sustained Erk activation is critical for IgE-induced mast cell survival through autocrine production of IL-3. Transmembrane adaptors LAT and NTAL contribute to the maintenance of prolonged Erk activation through membrane retention of the Ras-activating complex, Grb2-Sos. In this review, the signal regulation for the distinct responses of mast cells through FcepsilonRI are discussed.