Phorbol esters induce a slow, non-cytotoxic release of histamine from rat peritoneal mast cells

The phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) and the synthetic diacylglycerol, 1-oleoyl-2-acetylglycerol (OAG) were employed to investigate the consequences of protein kinase C activation in rat peritoneal mast cells. TPA (10 ng/ml) induced a biphasic release of histamine from mast cells. At short periods of incubation histamine release was low; about 10% at 10 min. At 15 min a second phase of release commenced, achieving a maximum at 90 min incubation by which time about 70% of cell histamine was released. Histamine release by TPA was dependent on glycolytic and oxidative metabolism, temperature-dependent and occurred in purified mast cells. In contrast, histamine release induced by OAG 50 micrograms/ml was maximal after 20 min. Studies using RHC 80267, a diacylglycerol (DAG) lipase inhibitor, indicated that metabolism of OAG by DAG lipase as well as spontaneous degradation accounts for the comparatively short-lived response to OAG.     

The roles of calmodulin and protein kinase C in histamine secretion from mast cells.

The effect of a new calmodulin-antagonist, 5-iodo-1-C8, with a high selectivity for calmodulin in comparison to protein kinase C, has been investigated on histamine secretion from mast cells. It has been found to be much more sensitive for the inhibition of histamine secretion than the earlier calmodulin-antagonists, trifluoperazine and W7. The effect of four inhibitors of protein kinase C, viz. staurosporine, K252a, tamoxifen and sphingosine, has also been studied on histamine secretion from mast cells. All of them caused dose-dependent inhibition of histamine secretion induced by the three secretagogues used: antigen, compound 48/80 and the calcium ionophore A23187. K252a was tested against histamine release, induced by the stimulation of protein kinase C alone with the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA) or the synthetic diacylglycerol, 1-oleoyl-2-acetyl-rac-glycerol (OAG). In both the cases K252a caused dose-dependent inhibition of histamine release. Staurosporine was also tested against TPA and was found to inhibit the release induced by it. Potentiation and inhibition (modulation) of secretagogue-induced histamine release by simultaneous protein kinase C stimulation with TPA or OAG have been demonstrated before. The potentiation and inhibition are shown to be antagonized by staurosporine. The observations point to the involvement of both calmodulin and protein kinase C in the histamine secretion process from mast cells.     

The actions of retinal and retinoic acid on histamine release from rat peritoneal mast cells

Histamine release from rat peritoneal mast cells induced by anti-IgE was potentiated by the retinoids: retinoic acid 2-10 microM and retinal 1-5 microM. Retinal also produced a concentration-dependent increase in anti-IgE-stimulated 45Ca uptake by these mast cells. A similar potentiating action of both retinoids was observed on histamine release induced by 12-O-tetradecanoylphorbol-13-acetate (TPA). For both anti-IgE- and TPA-induced histamine release, the potentiating effect of the two retinoids was only observed in the presence of extracellular calcium. In contrast, histamine release induced by compound 48/80 was inhibited by retinal 1-5 microM and by retinoic acid 10-50 microM and the inhibition was the same in the presence as in the absence of extracellular calcium 1 mM. Histamine release induced by calcium and the calcium ionophore A 23187 was inhibited by retinoid acid 2-10 microM and by retinal 10 microM. Inhibitions of compounds 48/80-induced histamine release by cromoglycate and by retinal were additive. It is concluded that while the actions of retinoids on rat peritoneal mast cells are consistent with the inhibition of protein kinase C, another action of these compounds, unrelated to this enzyme, may explain the data.     

The actions of inhibitors of diacylglycerol kinase and diacylglycerol lipase on histamine release from rat peritoneal mast cells.

1. RHC 80267 and R59 022 are selective inhibitors of diacylglycerol (DAG) lipase and DAG kinase enzymes respectively. These inhibitors were examined with regard to their effects on oleoylacetylglycerol (OAG)-and anti-IgE- induced histamine secretion in rat peritoneal mast cells. 2. RHC 80267, 10 microM and R59 022, 50 microM both enhanced OAG-induced histamine release by 30% and 40% respectively. 3. In the concentration range 3-30 microM, R59 022 enhanced anti-IgE-induced histamine release by up to about 40%, whereas RHC 80267 was without effect. 4. The enhancement of anti-IgE-induced histamine release by R59 022 is consistent with a role for protein kinase C in transducing immunological signals to rat peritoneal mast cells. 5. The lack of effect of RHC 80267 in this situation may indicate that in the mast cell, DAG kinase is more active than DAG lipase in degrading physiological levels of DAG.     

Characteristics of the effect of phorbol ester on rat mast cells: interaction with anti-IgE

The phorbol ester 12-0-tetradecanoyl-13-acetate (TPA) induced a dose-dependent release of histamine from rat peritoneal mast cells at concentrations from 3 to 100 ng/ml. The release is biphasic: an early phase being complete in 15 min and being followed by a second phase extending for more than 50 min. Concentrations of TPA greater than 100 ng/ml produced decreasing releases of histamine. Synergistic interaction in the induction of histamine secretion was observed between TPA and A23187 and between TPA and anti-IgE. Such synergism with anti-IgE was only manifest at low concentrations of TPA and with incubations of the cells with TPA for 5 mins or less. At higher concentrations of TPA and longer incubations, TPA inhibited the response of rat peritoneal mast cells to anti-IgE stimulation. Synergism between A23187 and TPA was observed only at low levels of histamine release induced by calcium plus A23187: at higher levels of release TPA was inhibitory.     

Differentiation of second messenger systems in mast cell activation

Pretreatment of rat peritoneal mast cells with either Staurosporine or an analog K-252a, lead to a dose-related inhibition of histamine release when stimulated with Anti-IgE (IC50: Staurosporine = 110 nM; K-252a = 100 nM). In contrast, the two PKC inhibitors (1-1000 nM) failed to inhibit histamine release induced by compound 48/80 (0.5-1 micrograms/ml). Exposure of Anti-Asc-IgE sensitized mouse bone marrow derived mast cells to Asc-BSA lead to the release of both histamine (510 ng +/- 12.6 ng/10(6) cells) and immunoreactive Leukotriene C4 (27.0 +/- 12.6 ng/10(6) cells). LTC4 release was inhibited by Staurosporine and K-252a with an IC50 of 75 nM for both compounds. Pretreatment of rat peritoneal mast cells with PMA 100 nM lead to a small but significant release of histamine (18.3 +/- 3.6%). Pretreatment of these cells with K-252a or Staurosporine lead to a dose related inhibition of histamine release with an ED50 of 10 nM for Staurosporine and 60 nM for K-252a. Treatment of rat peritoneal mast cells with the calcium ionophore A23187 lead to a significant release of histamine which was not inhibited by either of the two kinase inhibitors (0.1-1000 nM). The two kinase inhibitors also inhibited mouse bone marrow derived mast cell proliferation in response to IL-3 with IC50 of 80 nM for Staurosporine and 270 nM for K-252a.     

The influence of tetradecanoyl-phorbol-acetate (TPA) on histamine release from isolated rat mast cells

A detailed investigation of the influence of tetradecanoyl-phorbol-acetate (TPA) on isolated rat mast cells was undertaken in order to explore the possible involvement of protein kinase C in histamine release. TPA alone could induce histamine release in a medium without calcium, whereas 1 mM CaCl2 suppressed the release. TPA in combination with a low concentration of the ionophore A23187 induced a considerable histamine release. Preincubation with TPA followed by incubation with the ionophore induced a similar release at low concentrations of TPA (less than or equal to 2.5 nM) whereas the response was reduced at higher concentrations of TPA. The inhibition after preincubation with TPA was almost at a maximum within 2 min and was due to a decreased rate of release. TPA could also increase antigen-induced histamine release. After preincubation the potency of low concentrations of TPA increased, whereas higher concentrations (50 nM) became inhibitory. The effects of preincubation were almost fully expressed after 2 min and were not due to altered kinetics of the release. The interaction of oleoylacetylglycerol (OAG) with the ionophore A23187 and with antigen resembled that of TPA, but OAG was considerably less potent. Preincubation with TPA was inhibitory to the histamine release induced by compound 48/80, particularly in the absence of calcium. The release induced by TPA and the ionophore or antigen was calcium-dependent and energy-requiring, and the effects of TPA persisted after washing the cells before exposure to antigen or the ionophore. Preincubation with the protein kinase C inhibitor isoquinolinesulfonyl-methylpiperazine (H7) slightly enhanced the histamine release induced by the combination of TPA and the ionophore.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of ryanodine on histamine release from rat peritoneal mast cells induced by anti-IgE.

Ryanodine strongly inhibited histamine release from rat peritoneal mast cells induced by anti-IgE. Ryanodine also inhibited Ca(2+)-mobilization from the intracellular Ca(2+)-store as well as histamine release in mast cells activated by anti-IgE. These results suggest that the effect of ryanodine on histamine release from rat mast cells might be due to the inhibition of Ca2+ release from the intracellular Ca2+ store.     

Differential inhibition of histamine release from mast cells by protein kinase C inhibitors: staurosporine and K-252a.

Pretreatment of rat peritoneal mast cells with either staurosporine or an analog K-252a [(8R*,9S*,11S*)-(-)-9-hydroxyl-9-methoxycarbonyl-8-methyl-2,3, 9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11-atrizadibenzo- [a,g]cycloocta[cde]trinden-1-one] led to a concentration-related inhibition of histamine release when the cells were stimulated with anti-IgE (IC50: staurosporine = 110 nM; K-252a = 100 nM). In contrast, the two protein kinase C (PKC) inhibitors (1-1000 nM) partially (less than 15%) inhibited histamine release induced by compound 48/80 (0.5 to 1 micrograms/mL). Furthermore, prostaglandin E2 (PGE2) synthesis mediated by anti-IgE from rat peritoneal mast cells was also inhibited by staurosporine and K-252a (IC50 = 100 nM). Exposure of anti-arsenate IgE (anti-Ars-IgE) sensitized mouse bone marrow derived mast cells to arsenate-bovine serum albumin (Ars-BSA) led to the release of both histamine (510 +/- 12.6 ng/10(6) cells) and immunoreactive leukotriene C4 (LTC4) (27.0 +/- 2.6 ng/10(6) cells). Both histamine and LTC4 release was inhibited by staurosporine and K-252a with an IC50 of 50 nM for both compounds. We also characterized a 45K molecular weight protein which is phosphorylated by PKC after Ars-BSA or phorbol, 12-myristate, 13-acetate (PMA) stimulation. This protein is phosphorylated in a broken cell preparation in which PKC is activated by phosphatidylserine/Diolein and Ca2+. Peptide mapping by V8 protease of the phosphorylated 45K protein revealed that the 45K protein phosphorylation patterns induced by IgE or PMA or in the broken cell preparation are identical. Pretreatment of 32P-labeled mouse bone marrow derived mast cells with either staurosporine or K-252a led to a concentration-related inhibition of 45K protein phosphorylation induced by PMA or Ars-BSA. This inhibition of protein phosphorylation correlated well with the inhibition of histamine and leukotriene release in bone marrow derived mast cells.     

Interaction of neurotensin with the substance P receptor mediating histamine release from rat mast cells and the flare in human skin.

1 Substance P induced histamine release from rat peritoneal mast cells in a dose-dependent manner over the concentration range 1 to 10 microM. 2 At concentrations in the range 2.5 to 1 0 microM, neurotensin produced only about 5% release of histamine, which was substantially less than the maximum effect obtained with substance P. 3 Neurotensin, 2.5 to 10 microM produced graded inhibition of histamine release induced by substance P. The inhibitory effect of neurotensin was not seen when histamine release was induced by an antigen-antibody effect of neurotensin was not seen when histamine release was induced by an antigen-antibody reaction or by the ionophore, A 23187. Some evidence was obtained to suggest that compound 48/80 may interact with the same receptor as substance P and neurotensin. 4 [D-Arg8]neurotensin, [D-Arg9]neurotensin, xenopsin and the C-terminal octapeptide of substance P (SP4-11) all inhibited histamine release by substance P, but physalaemin did not. 5 Neurotensin inhibited the wheal and flare reactions induced by substance P in human skin. 6 [D-Trp7,9]substance P released histamine from rat mast cells and was about 12 times more potent than substance P itself. [D-Trp7,9]SP1-11 also produced wheal and flare responses in human skin, being 1.8 times more potent than substance P in the production of flare.     

The effect of prednisolone on substance P-induced vascular permeability in mice.

The effect of prednisolone on the substance P (SP)-induced vascular permeability increase in male ddY, WBB6 F1(-)+/+ (control) and WBB6 F1-W/WV (no mast cell in skin or internal organs) mice was investigated. 1) SP (1-10,000 pg/site) increased vascular permeability in ddY, WBB6 F1(-)+/+ and WBB6 F1-W/WV mice ears. 2) SP (100 pg/site)-induced vascular permeability was inhibited by prednisolone (10 mg/kg) administered intraperitoneally 3 to 12 hours prior to the elicitation of the reaction in ddY mice. When dexamethasone at a dose of 1 mg/kg was administered intraperitoneally 2 to 24 hours prior to the elicitation of the reaction, significant inhibition was observed. When prednisolone was administered intraperitoneally 8 hours prior to the elicitation of the reaction, the SP-induced capillary permeability increase in both ddY and WBB6 F1-W/WV mice was clearly inhibited by the drug at doses of 5 and 10 mg/kg. 3) Diphenhydramine (1 and 10 mg/kg) inhibited SP-induced vascular reaction in ddY mice but not in WBB6 F1-W/WV mice. 4) Atropine (10 mg/kg) inhibited SP-induced vascular reaction in both ddY and WBB6 F1-W/WV mice. But acetylcholine did not cause an increase of vascular permeability in ddY and WBB6 F1-W/WV mice ears. 5) Prednisolone (5 mg/kg) inhibited histamine- and serotonin-induced vascular permeability in ddY and WBB6 F1-W/WV mice ears. 6) Prednisolone (5 and 10 mg/kg) inhibited the SP-induced histamine release from ddY mice peritoneal mast cells. These results suggest that the vascular effect of SP is mediated by both mast cell dependent (release of histamine from mast cells) and mast cell independent mechanisms. Prednisolone inhibits the SP-induced vascular permeability mediated by both mechanisms in mice.     

Inhibition of protein kinase activity by apomorphine

The effect of apomorphine (1-20 microM) on protein kinase activity was studied in extracts from rat peritoneal mast cells and brain tissue. Apomorphine inhibited the cyclic AMP-dependent and the calcium-plus phosphatidylserine-dependent protein kinase activity with an IC50 between 1 and 6 microM, depending on the tissue and on the protein kinase involved. This effect might explain previous results on the apomorphine-induced inhibition of histamine release in rat peritoneal mast cells.     

Inhibitory effect of anti-allergic agent NCO-650 on histamine release induced by various secretagogues

Histamine release from rat peritoneal mast cells induced by antigen and anti-IgE was essentially complete within 2 min and 3 min, respectively, but that due to Concanavalin A (Con A) was complete only within 9 min. An anti-allergic agent NCO-650 [trans-4-Guanidinomethylcyclohexanecarboxylic acid p-tert-butylphenyl ester hydrochloride], which is a strong inhibitor of trypsin, dose-dependently inhibited anti-IgE-induced histamine release from rat peritoneal mast cells. Moreover, the rate and extent of histamine release from rat peritoneal mast cells induced by various histamine liberators such as antigen, concanavalin A, ionophore A 23187 and compound 48/80 are significantly diminished in samples incubated with NCO-650. The IC50 values of NCO-650 on histamine release induced by antigen, anti-IgE, Concanavalin A, A23187 and compound 48/80 were in the order of micromolar range, i.e. 1.9, 3.6, 4.6, 2.9 and 6.1 microM, respectively. On a molecular basis, NCO-650 is 1000-fold more potent than DSCG, an anti-allergic drug, in inhibiting the antigen-induced histamine release. The present results suggest that the effect of NCO-650 might be due to the inhibition of a common process underlying the release of histamine by various histamine liberators.     

Histamine release induced by dextran: the nature of the dextran receptor

Dextrans of molecular weight 10(4) to 2 x 10(6) induced histamine release from rat peritoneal mast cells in the presence of calcium (1 mM) and phosphatidyl serine (10 micrograms/ml). Glucose and low molecular weight dextrans inhibited the histamine release induced by high molecular weight dextrans but the inhibition could not be explained in terms of a simple competitive model. Structure-activity relationships for the inhibition of dextran-induced histamine release by a number of saccharides demonstrated that substitution at the C-3, C-4, and C-6 positions of glucose were most important for activity. Inhibition of histamine release by glucose was specific for the dextran stimulus. Soluble IgE and IgG antibodies failed to interfere with histamine release induced by dextran. Phlorizin specifically inhibited the histamine release induced by dextran. Purification of mast cells on albumin or ficoll gradients produced a selective loss of response to dextran which was not due to the removal of non-mast cells but to some change in the mast cells themselves. The possible nature of the dextran receptor is discussed.     

Effect of dantrolene on histamine release from rat peritoneal mast cells.

Dantrolene strongly and dose-dependently inhibited histamine release from rat peritoneal mast cells induced by anti-IgE. Dantrolene inhibited Ca(2+)-mobilization from intracellular Ca(2+)-store as well as histamine release in mast cells activated by anti-IgE, the effect on both these phenomena being closely correlated. These results suggested that the effect of dantrolene on histamine release from rat mast cell might be due to the inhibition of Ca(2+)-release from intracellular Ca(2+)-store.     

Cyclic AMP independent inhibition by papaverine of histamine release induced by compound 48/80.

Compound 48/80-induced histamine release from isolated rat peritoneal mast cells was inhibited in a dose-dependent manner by papaverine (ic₅₀ approx 20 μM). This effect of papaverine was not influenced by PGE₁ (14–140 μM), even though PGE₁ markedly increased must cell cAMP levels. Papaverine (0.5 mM) completely inhibited histamine release without causing any change in cAMP levels. Theophylline (0.1 and 0.5 mM) potentiated histamine release induced by submaximal concentrations of compound 48/80, while cAMP levels were increased. IBM X was as potent as papaverine in causing inhibition of mast cell phosphodiesterase. IBM X (0.14–0.7 mM) had no effect on histamine release but caused a 6–20 fold increase in mast cell cyclic AMP. Papaverine inhibition of histamine release was gradual at the onset and was parallelled by a depletion of mast cell ATP content. The inhibition of 48/80-induced histamine release and depletion of mast cell ATP levels was reversed by glucose. It is concluded that papaverine induced inhibition of 48/80-induced histamine release is independent of cAMP, is unrelated to phosphodiesterase inhibition but is dependent upon inhibition of energy production.     

Effects of phorbol ester, serine phospholipids, and calcium on histamine release from mast cells and basophils

The effects of calcium, serine phospholipids and the phorbol ester tetradecanoylphorbol acetate (TPA), a known activator of protein kinase C, have been examined on mastocytes from different sources. A complex and differential interaction between these agents and histamine secretion was observed in human basophil leucocytes and in peritoneal mast cells from the rat, mouse and hamster. On this basis, the possible role of kinase C in histamine release is discussed.     

Substance P-induced histamine release from rat peritoneal mast cells and its inhibition by antiallergic agents and calmodulin inhibitors.

Substance P-induced histamine release and Ca2+ release from the intracellular Ca store of rat peritoneal mast cells were inhibited by both antiallergic drugs and microtubule inhibiting agents. It was found that in the case of antiallergic compounds, histamine release inhibition may be intimately related to the inhibition of Ca2+ release from the intracellular store in which the microtubules play an important role. When mast cells were pretreated with either theophylline or dibutyryl cAMP, the inhibition of histamine release was closely related to the inhibition of Ca2+ release from the intracellular Ca store. Calmodulin inhibitors were also effective in inhibiting histamine release from mast cells induced by substance P. The inhibitory potencies of calmodulin inhibitors on histamine release from mast cells were closely correlated with those exerted on calmodulin activity.     

On the actions of substance P,somatostatin, and vasoactive intestinal polypeptide on rat peritoneal mast cells and in human skin

Substance P (SP), somatostatin (Som), and vasoactive intestinal polypeptide (VIP) induced a concentration-dependent release of histamine from isolated rat peritoneal mast cells. The release of histamine induced by these neuropeptides was inhibited by preincubation of the cells with the SP analogue [D-Pro4,D-Trp7,9,10]-SP4-11 (SP-A) (10 microM), and also by benzalkonium chloride (10 microM). In addition, SP-A inhibited histamine release induced by compound 48/80, whilst that induced by goat anti-(rat-IgE) was unaffected. In human skin, intradermal injection of SP, Som, or VIP produced flare and wheal responses. The flares to all three peptides were inhibited by preinjection of the skin with SP-A (25 pmol), whilst the wheal responses were unaffected. It is concluded that the receptors mediating histamine release and the flare response are similar, and that SP, Som, and VIP are acting at a similar receptor to produce these effects. It is probable that this receptor is also the site of action of compound 48/80.     

Chondroitin sulphate inhibits connective tissue mast cells

1. Mast cells derive from the bone marrow and are responsible for the development of allergic and possibly inflammatory reactions. Mast cells are stimulated by immunoglobulin E (IgE) and specific antigen, but also by a number of neuropeptides such as neurotensin (NT), somatostatin or substance P (SP), to secrete numerous pro-inflammatory molecules that include histamine, cytokines and proteolytic enzymes. 2. Chondroitin sulphate, a major constituent of connective tissues and of mast cell secretory granules, had a dose-dependent inhibitory effect on rat peritoneal mast cell release of histamine induced by the mast cell secretagogue compound 48/80 (48/80). This inhibition was stronger than that of the clinically available mast cell 'stabilizer' disodium cromoglycate (cromolyn). Inhibition by chondroitin sulphate increased with the length of preincubation and persisted after the drug was washed off, while the effect of cromolyn was limited by rapid tachyphylaxis. 3. Immunologic stimulation of histamine secretion from rat connective tissue mast cells (CTMC) was also inhibited, but this effect was weaker in umbilical cord-derived human mast cells and was absent in rat basophilic leukemia (RBL) cells which are considered homologous to mucosal mast cells (MMC). Oligo- and monosaccharides were not as effective as the polysaccharides. 4. Inhibition, documented by light and electron microscopy, involved a decrease of intracellular calcium ion levels shown by confocal microscopy and image analysis. Autoradiography at the ultrastructural level showed that chondroitin sulphate was mostly associated with plasma and perigranular membranes. 5. Chondroitin sulphate appears to be a potent mast cell inhibitor of allergic and nonimmune stimulation with potential clinical implications.