Regulatory role of extracellular Na<Superscript>+</Superscript> and Ca<Superscript>2+</Superscript> ions in nerve growth factor induced histamine secretion from rat mast cells

OBJECTIVE AND DESIGN: This study was aimed to investigate effects of extracellular Na+ and Ca2+ ions on nerve growth factor (NGF) induced histamine release from mast cells. MATERIAL: Isolated peritoneal mast cells were obtained from male Wistar rats. METHODS: Cells were suspended in solution with different concentration of Na+ and Ca2+ ions and stimulated with NGF. Histamine release was assayed spectrofluorometrically. RESULTS: NGF (0.001-1 microg/ml) dose-dependently releases histamine from mast cell at physiological extracellular Na+ (134 mM) and Ca2+ (1 mM) conditions. Lowering extracellular Ca2+ ions to 0.1 mM reduced histamine response to nearly basal level. However, the removal of extracellular Na+ ions significantly enhanced the secretion provoked by NGF (0.6 microg/ml) in low Ca2+ medium. Amiloride, an inhibitor of Na+/Ca2+ and Na/H+ exchangers inhibited the potentiating effect of sodium free conditions. CONCLUSIONS: Our results suggest that the activity of Na+/Ca2+ and/or Na+/H+ exchange mechanisms could be of particular importance in the secretory process of mast cells induced by NGF.     

Release of histamine and tryptase during continuous and interrupted cutaneous challenge with allergen in humans.

To help in understanding the patterns of in vivo mediator release in human allergic skin reactions, we have used a skin chamber model to challenge the denuded bases of skin blisters of 11 sensitive subjects with pollen antigens (Ags) and codeine (C), a mast cell degranulator. Challenges were performed either (1) continuously for 6 hours or (2) in an intermittent fashion that is, Ag or C for the first hour, buffer for the next 4 hours, and then Ag or C during the sixth hour. Fluids in the overlying chamber were assayed for levels of the mast cell components, histamine and tryptase. There was peak release of both histamine and tryptase during the first hour of Ag incubation (89 +/- 11 ng/ml and 1428 +/- 260 ng/ml, respectively). At continuous Ag-challenge sites, there was a plateau of histamine levels (8.0 to 9.5 ng/ml) during the next 4 hours, whereas tryptase levels decreased progressively to baseline levels. Challenge of continuous Ag-incubation sites with C, a mast cell activator, led to another peak release of both histamine and tryptase. At interrupted Ag-challenge sites, histamine levels decreased abruptly, and tryptase levels decreased progressively after the first hour. Rechallenge of such sites with Ag during the sixth hour induced a peak release of histamine but no increase in tryptase levels. Continuous challenge with C for up to 5 hours in other sites induced an initial peak histamine release without a subsequent plateau. However, such a plateau of histamine (but not tryptase) release occurred after an initial C challenge if Ag was subsequently incubated in a continuous fashion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibitory effect of traxanox sodium on IgE-mediated histamine release from passively-sensitized mast cells of the rat in vitro

Traxanox sodium, a benzopyranopyridine derivative showing a potent oral antiallergic activity in the rat, was compared with disodium cromoglycate (DSCG) for ability to block the release of histamine from the rat mast cell in vitro. Traxanox sodium showed dose-, antigen- and time-dependent inhibiton of the IgE-mediated release of histamine. The 50% inhibitory concentration was 0.04 microM for traxanox sodium, 1 microM for DSCG and 660 microM for theophylline. All these drugs blocked the release of histamine potentiated by preincubation of the mast cell with 10 micro M adenosine at lower concentrations than those which could inhibit the IgE-mediated histamine release. In addition, traxanox sodium at concentration of 1-100 microM inhibited the histamine release caused by 0.25 microgram/ml compound 48/80 in the presence and absence of calcium, and the drug at 100 micro M slightly inhibited the release caused by 0.2 microgram/ml ionophore A23187. These results suggest that traxanox sodium is a more potent inhibitor than DSCG on the histamine release from the mast cells of the rat, and a part of its antiallergic action is due to a selective inhibition of the immunological release of allergic mediators from the mast cell.     

Effects of anti-CD11a, anti-CD11b and anti-CD18 on histamine release from human basophils primed with IL-3.

This study was carried out to investigate the possible role of adhesion molecules in the histamine release from human basophils primed with recombinant IL-3 (rIL-3). Anti-IgE monoclonal antibody (0.1 microgram/ml) did not induce apparent histamine release by itself, however, remarkable histamine release was induced from rIL-3-primed human basophils triggered by anti-IgE. Bear-1 (anti-CD11b monoclonal antibody) inhibited the histamine release enhanced by rIL-3, however SPV-L7 (anti-CD11a monoclonal antibody) or CLB-LFA1/1 (anti-CD18 monoclonal antibody) did not inhibit the histamine release enhanced by rIL-3 at all. The present study suggests that alpha-chain of Mac-1 is involved in the priming effect of IL-3 to increase histamine release from human basophils.     

Regulation of histamine release from human bronchoalveolar lavage mast cells by stem cell factor in several respiratory diseases

We investigated the effects of stem cell factor (SCF) on histamine release (HR) from human bronchoalveolar lavage (BAL) mast cells. BAL cells were recovered from lavage performed in patients undergoing clinical bronchoscopy. SCF (0.02–20 ng/ml), which is by itself a poor secretagogue (mean ± SEM HR: 3.7 ± 0.9%; n = 27), strongly enhanced HR induced by anti-IgE in a concentration-related manner. Significant potentiation began at 0.2 ng/ml (30 ± 10°0; p <0.05; n = 12) and reached a plateau at 2 ng/ml (40 ± 10%; P <0.01 at 2 ng/ml and 45 ± 10%; P <0.01 at 20 ng/ml; n = 12). In contrast, SCF failed to enhance HR induced by calcium ionophore A23187. Among the BAL cell samples initially unresponsive to anti-IgE (55° of samples), 36% (10/28) were converted to responders if the cells were shortly preincubated with SCF. In 25% of samples (7/27), SCF (20 ng/ml) caused direct HR of 10 ± 2.1 %. The mast cells which released histamine when challenged with SCF also secreted higher levels of histamine in response to anti-IgE and calcium ionophore than those nonresponsive to SCF. While interleukin (IL)-3 and IL-5 (20 ng/ml) were unable to modulate immunologic HR. GM-CSF (20 ng/ml) produced significant potentiation ( P <0.05), which was, however, smaller than that observed with SCF. The rate of responders to anti-IgE in atopic asthma (47 %) was greater than that in control (9%) and intrinsic asthma (10%) but not different from that in some other respiratory diseases such as chronic bronchitis (44%), lung cancer (47%), or interstitial disease (68%,). The potentiation of HR afforded by SCF did not differ significantly among the several disease groups. We conclude that, whatever the underlying respiratory disease, SCF selectively enhances IgE-mediated HR from human BAL mast cells. Furthermore, this cytokine is sometimes necessary to render mast cells able to release histamine in response to anti-IgE.     

Evidence of ongoing mast cell and eosinophil degranulation in symptomatic asthma airway.

To assess whether mast cell and eosinophil (EOS) degranulation occurs in the airway of subjects with moderately symptomatic asthma, we have measured levels of preformed mast cell-derived mediators (histamine and tryptase) and EOS-derived mediators (major basic protein and EOS-derived neurotoxin) in bronchoalveolar lavage fluid (BALF) obtained from patients with symptomatic (N = 14) and asymptomatic asthma (N = 9) and patients without asthma (N = 6). Both the FEV1 (1.52 +/- 0.33 L:55% +/- 15% of predicted FEV1) and the forced expiratory flow at 50% (FEF50) (1.11 +/- 0.62 L/sec:26% +/- 14% of predicted FEF50) in the patients with symptomatic asthma were significantly lower than the corresponding values for FEV1 (3.16 +/- 0.45 L:86% +/- 10% of predicted FEV1) and the FEF50 (4.04 +/- 1.54 L/sec:71% +/- 25% of predicted FEF50) in the patients with asymptomatic asthma. Levels of histamine (4.8 +/- 5.0 ng/ml versus 0.2 +/- 0.2 ng/ml) (p = 0.05), EOS-derived neurotoxin (420.6 +/- 959.4 ng/ml versus 12.6 +/- 7.7 ng/ml) (p = 0.05), major basic protein (31.4 +/- 46.6 ng/ml versus less than 9 ng/ml) (p = 0.05), and percent EOSs (10.6% +/- 7.0% versus 1.1% +/- 0.9% of BAL cells) (p = 0.0006) were all significantly elevated in BALF from symptomatic compared to asymptomatic patients with asthma. The elevated levels of tryptase (13.2 +/- 14.8 ng/ml versus 3.9 +/- 3.9 ng/ml) in BALF from symptomatic compared to asymptomatic patients with asthma approximated, but did not reach, statistical significance. Spontaneous histamine release from BAL mast cells of symptomatic patients with asthma was 46% +/- 5% compared to 5% +/- 2% in asymptomatic patients with asthma. In response to antihuman IgE, histamine release from BAL mast cells recovered from asymptomatic patients with asthma increased to 25% +/- 10%, whereas in BAL mast cells of symptomatic patients with asthma, no anti-IgE potentiation of histamine release occurred. This study suggests that mast cell and EOS degranulation is ongoing in the airway of patients with moderately symptomatic asthma.     

Release of Mediators from Purified Rat Mast Cells during Phagocytosis

Purified mature rat peritoneal mast cells, on exposure to zymosan or latex beads, phagocytize these particles, although less efficiently than macrophages. During phagocytosis, histamine, beta-glucuronidase, and eosinophil chemotactic factor are released from mast cells in a time-, temperature- and dose-dependent fashion. Complement components, cytochalasin B (5 microgram/ml), and indomethacin (10-6M), enhanced mediator release, whereas compound BW 755C (20 microgram/ml), a cyclooxygenase and lipoxygenase inhibitor of arachidonate metabolism, totally abolished this process. Phagocytosis of mast cell thus activates intracellular mechanisms that closely resemble those observed with other phagocytic cells. These observations add a new perspective to the role of mast cells in inflammatory events.     

Naturally occurring aliphatic polyamines-induced histamine release from rat peritoneal mast cells

Rat peritoneal mast cells were incubated with different concentrations of naturally occurring aliphatic polyamines, spermine and spermidine, at 0.1-10 mM and the amount of histamine release into the supernatant solutions was measured. The addition of each polyamine to the suspensions of the mast cells caused a histamine release in a dose-dependent manner. The effect of 10 mM spermine and spermidine was as much as that of 0.5 microgram/ml compound 48/80. The histamine release from the cells incubated with each polyamine was rapid and the amount of histamine release into the supernatant solutions reached a maximum at 1 min with the incubations. 0.1 mM spermine, which in itself could not cause a significant histamine release, showed a tendency to enhance anti-IgE-induced histamine release from the mast cells.     

Mouse IL-3 induces histamine release from mouse peritoneal mast cells.

In this study, we have attempted to determine whether mouse peritoneal mast cells released histamine in response to IL-3. Recombinant mouse (m)IL-3 induced histamine release from mouse peritoneal mast cells in a dose-dependent fashion. Histamine release did not occur in the absence of phosphatidyl serine (PS), and was dependent on PS concentrations. The release was 14.3 +/- 3.8 and 43.5 +/- 11.5% (mean +/- SEM, n = 5) at 1 nM IL-3 in the presence of 10 and 20 micrograms/ml of PS. Calcium was required for the response, and in the absence of calcium, significant histamine release was not observed. The kinetics were slower than those of anti-IgE-induced response. IL-3-induced histamine release reached a peak within 15 min, while that by anti-IgE reached 80% of the maximum in 3 min. Lower concentrations of IL-3, which failed to directly induce histamine release, did not enhance anti-IgE-induced histamine release. Other cytokines, including mIL-4, mIL-5, m-granulocyte-macrophage colony-stimulating factor, human (h)IL-1 alpha, hIL-1 beta and hIL-8, neither induced histamine release nor enhanced anti-IgE induced histamine release. IL-4 had no capacity to enhance IL-3-induced histamine release. These results suggest that locally produced IL-3 might modulate mast cell-related inflammation through histamine release from mast cells.     

Effect of sensitization on spontaneous and phosphatidylserine-induced histamine release and on cyclic AMP and GMP levels in isolated rat mast cells.

Sprague Dawley rats were sensitized with 20 microgram or 100 mg egg albumin (using pertussis vaccine as adjuvant). Mast cells isolated from the former group of animals showed a higher degree of histamine release upon challenge in vitro with egg albumin than those from the latter group. Using the lower amount of antigen for immunization mast cells from Hooded Lister rats showed an even higher degree of histamine release induced by antigen. An increased antigen-induced histamine release was associated with an increased spontaneous and phosphatidylserine-induced histamine release. Histamine release induced by phosphatidylserine was found to be specific in so far as it was calcium dependent and theophylline-inhibited. The basal level of cyclic AMP in mast cells was significantly depressed by sensitization. There was a relationship between the cyclic AMP/cyclic GMP ratio and the degree of spontaneous, phosphatidylserine-induced and anaphylactic histamine release. The results suggest that sensitization induces an increased release of histamine not only to the specific antigenic stimulus but also to more unspecific stimuli. Concomitantly there is a fall in the cyclic AMP/cyclic GMP ratio. The relationship between these two phenomena is discussed.     

Palytoxin both induces and inhibits the release of histamine from rat mast cells

Palytoxin (PTX) is a potent releaser of histamine from rat mast cells. The concentration response curve is bell-shaped with its range between 0.05 ng/ml and 5 micrograms/ml and its maximum at 50 ng/ml. The release of histamine by PTX is specific because (a) heat-inactivated (50 degrees C) mast cells are insensitive to PTX, and (b) extracellular calcium is essential. Removal of extracellular potassium as well as the presence of borate shifts the curve to the left by factor of 10. At higher concentrations (5 micrograms/ml), PTX inhibits the release of histamine due to compound 48/80, MCD-peptide, Con-A, ionophore A-23187, and PTX itself.     

Effect of cytokines on mediator release from human dispersed lung mast cells

To evaluate the contribution of human lung mast cells (HLMC) to allergic inflammation, we investigated whether or not cytokines, including stem-cell factor (SCF), monocyte chemotactic and activating factor (MCAF), and RANTES, activate HLMC. SCF induced histamine release from dispersed HLMC in a dose-dependent fashion (P<0.01). The release was 7.8 ± 1.0% at 500 ng/ml SCF (n= 9). This response was also observed in chopped lung tissue. HLMC from which surface IgE molecules had been removed by treatment with lactic acid responded to SCF, while these cells lost their response to anti-IgE. The process was relatively rapid and reached a maximum in 5 min. This response required extracellular calcium, and it was observed at 37°C, but not at 4°C or 20°. A brief preincubation (10 min) with lower concentrations of SCF, which were ineffective in releasing histamine, enhanced anti-IgE-induced histamine release (P<0.05), while its enhancing effect was lost by the longer preincubation (30 min). SCF did not prime basophils to enhance stimulated-histamine release. Interleukin (IL)-1α, IL-1β, IL-3, IL-4, IL-5, granulocyte/macrophage-colony stimulating factor (GM-CSF), MCAF, and RANTES neither induced histamine release nor enhanced the release stimulated by anti-IgE after a 10- or 30-min preincubation. The combination of IL-3 and IL-4 showed no effect on histamine release from HLMC. Leukotriene (LT)C₄/D₄/E₄ production by SCF was negligible, as compared with anti-IgE-induced LT production. SCF at 1.5 ng/ml augmented anti-IgE-induced LT generation significantly (536+ 117 pg/10⁵ mast cells and 1569 ± 258 pg/10⁵ mast cells; P<0.01). These results provide further evidence that numerous aspects of the phenotype of mast cells and basophils are heterogeneous, including structure, relevant secretagogues, and pharmacologic control.     

Histamine suppression of in vivo eosinophil accumulation and histamine release in human allergic reactions

Although it has been shown that histamine inhibits antigen-induced in vitro histamine release from basophils, it is unclear whether histamine inhibits in vivo mediator release in human allergic reactions. We report effects of exogenous histamine on histamine release and inflammatory cell responses in antigen-challenged skin sites in eight ragweed-sensitive individuals. Four heat-suction blisters in each subject were unroofed, and a collection chamber was appended to each blister base. Chamber A contained 1000 PNU/ml ragweed extract; chamber B contained buffered saline (control fluid); chamber C contained 1000 PNU/ml ragweed and 50 ng (5 x 10(-7) M) of histamine; and chamber D contained histamine alone (50 ng). Comparative analyses of chamber histamine levels in individual subjects showed that (1) histamine levels in chamber A were significantly greater than those in chamber B (p less than 0.01) and that histamine levels in chamber C were not significantly different than those in chamber D (p less than 0.5). Likewise, comparison of eosinophils attaching to membrane filters appended to the chamber bases for 2 hr showed that there were significantly more eosinophils in chamber A than in chamber B (p less than 0.01) and that there was no significant difference in eosinophil numbers on filters appended to chamber C vs chamber D. In three of four subjects studied, addition of exogenous histamine (50 ng/ml) to ragweed before intradermal injection inhibited the ultrastructural mast cell alterations seen within 10 min after injection of ragweed alone. In the one subject in which mast cell alterations were not prevented, exogenous histamine also did not inhibit antigen-induced histamine release or subsequent eosinophil accumulation in the skin chambers.     

Interferon-alpha/beta inhibits IgE-dependent histamine release from rat mast cells.

Although mast cells and interferons are both involved in numerous immune and inflammatory responses, little is known about how microenvironmental factors such as interferons (IFNs) influence mast cell function. To study this question, sensitized peritoneal mast cells (greater than 98% purity) obtained from rats infected 4 weeks earlier with the parasite Nippostrongylus brasiliensis were preincubated for 24 hr with rat IFN-alpha/beta in RPMI-1640, then stimulated to degranulate with worm antigens. In the absence of antigen, IFN-alpha/beta had no noticeable effect on histamine release. However, in the presence of antigen, IFN-alpha/beta (150-1500 U/ml) inhibited histamine release in a dose-dependent manner (22.2 +/- 7.5% to 56.3 +/- 6.9%, n = 10). This inhibitory effect was neither heat (56 degrees for 1 hr) nor acid (pH 2 for 18 hr) labile, but was completely blocked by anti-IFN antibodies. In the presence of compound 48/80 (1 microgram/ml) or substance P (5 X 10(-5) M), IFN-alpha/beta was ineffective at modulating histamine release. Histamine release induced by antigen in the presence of the membrane phospholipid phosphatidyl-serine (30 micrograms/ml) was inhibited by IFN in a dose-dependent manner, but maximal inhibition (25.3 +/- 2.7%, n = 10) was reached at a lower concentration of IFN (750 U/ml) than when antigen was used alone. Therefore, rat IFN-alpha/beta appears to inhibit histamine release from rat mast cells in a dose- and stimulus-dependent manner and may do so by reducing the fluidity of the cell membrane.     

The role of mast cells in inflammatory processes: evidence for nerve/mast cell interactions

In the rat, there is considerable evidence of mast cell/nerve interaction both in the normal and infected intestine. Between 67 and 87% of all mast cells in the intestinal lamina propria of rats infected 22-35 days earlier with Nippostrongylus brasiliensis were touching nerves. These membrane contacts were between subepithelial mast cells and nonmyelinated nerves containing substance P, calcitonin gene-related peptide and neurone specific enolase. 2.5S nerve growth factor (NGF) has a significant enhancement effect on antigen-induced histamine release without addition of phosphatidylserine, and the in vivo administration of NGF to rats causes both connective tissue and mucosal mast cells to dramatically increase in number. All of these effects are both dose dependent and NGF specific, as evidenced by inhibition with anti-NGF. 2.5S NGF also causes in vitro increase of colonies in methylcellulose cultures of human peripheral blood. The effects of NGF in this system are synergistic with other T cell-derived growth factors and relatively specific for metachromatic cell growth. These observations support the conclusions that nerves and mast cells may constantly communicate and provide a structural and conceptual framework whereby the central nervous system may communicate with inflammatory events.     

Comparison of histamine release from peritoneal mast cells derived from diabetic and control rats

Clinical and experimental diabetes are associated with an increased number of mast cells and elevated tissue histamine concentrations. This study compared histamine release from peritoneal mast cells derived from diabetic and control rats. Experimental diabetes was induced by a single i.v. injection of streptozotocin (50 mg/kg body weight). Measurement of plasma glucose levels confimed the diabetic state. Peritoneal mast cells were stimulated for 10 min with the lectin concanavalin A (0.5–100 g/ml) in the presence or absence of phosphatidylserine, clinical dextran (0.6–1200 g/ml) in the presence of phosphatidylserine, the calcium ionophore A23187 (0.1–1 M) or the basic releasing agent compound 48/80 (0.1–10 g/ml). Histamine release induced by these agents was similar in both populations. Further studies will compare the differences in histamine release from mast cells isolated from different tissues, e.g. heart and lung. In addition, physiological stimuli which are altered in the diabetic state (e.g. hyperosmolalar solutions and free radical generating systems) are under investigation.     

Histamine release from dispersed human intestinal mast cells

To study the human intestinal mast cell of children and adults, we combined a sensitive glassfibre-based histamine assay with the enzymatic and mechanical dispersion of surgical specimens or mucosal biopsies. The method yields between 1.2 x 10(3) to 4.6 x 10(3) mast cells/mg tissue constituting 1.2% to 5.3% of total cell count. The mast cell yield, however, depends on the intestinal tissue specimen used for dispersion. Aliquots containing 1500 mast cells per sample are sufficient for measuring significant amounts of histamine (greater than or equal to 0.15 ng histamine per sample), thus making it possible, to carry out approximately 75 tests for four mucosal biopsies of 10 mg each. The intestinal mast cell releases histamine in a dose-dependent manner on challenge with anti-IgE (6-600 U/ml), ionophore A23187 (0.25-1.0 microM), and Concanavalin A (0.7-25.0 micrograms/ml). The histamine release shows interindividual variation with a net histamine release between 0 to 2.5 ng/samples dependent on the secretatogue. In general, it is not necessary to passively sensitize the mast cells to obtain a sufficient histamine release response to anti-IgE challenge, indicating the presence of intact and functional cell-bound IgE. However, it is shown that four of 10 non-atopic intestinal mast cell samples could be passively sensitized with human plasma containing either mite- or grass-specific IgE without stripping off the IgE first. This indicates the presence of free and preserved Fc-receptors on the dispersed mast cells in some subjects. In addition, it is found that the phorbolester TPA increases the histamine release response to A23187 and turns anti-IgE non-responding mast cells into responding mast cells, but TPA alone at 2 to 16 ng/ml has no histamine releasing effect. In patients with anti-IgE responding mast cells no additional effect of TPA is seen. Finally, no substantial differences between mast cells of children and adults are demonstrated.     

Characterization of purification-associated reduction in IgE-dependent histamine release from rat peritoneal mast cells

Histamine release from purified rat peritoneal mast cells (PMC) was examined and compared to that from a non-purified preparation (PEC). Both PEC and PMC released similar amounts of histamine upon stimulation with compound 48/80, calcium ionophore A23187 and substance P. In contrast, IgE-dependent histamine release from PMC caused by antigen, anti-IgE and concanavalin A was very low compared to that of PEC. The reduced IgE-dependent histamine release from PMC, however, was recovered when PMC was reconstituted with non-mast cells (NMC) present in the peritoneal cavity. The effect was time-dependent and reached a plateau in 30 min. NMC from both sensitized and non-sensitized rats recovered the reduced histamine release from PMC dose-dependently. The potentiating effect of NMC was observed even in the presence of excess amount of phosphatidylserine. Supernatants of NMC and a mixture of PMC and NMC incubated for 1 hr at 37°C, however, failed to potentiate the histamine release. These results demonstrate that IgE-dependent histamine release from rat peritoneal mast cells is upregulated by other cells present in the peritoneal cavity, and that the mechanism involved is distinct from that of phosphatidylserine.