Guinea Pig Lung Mast Cells Research Articles

20(S)-Protopanaxatriol inhibits release of inflammatory mediators in immunoglobulin E-mediated mast cell activation

BackgroundAntiallergic effect of 20(S)-protopanaxatriol (PPT), an intestinal metabolite of ginseng saponins, was investigated in guinea pig lung mast cells and mouse bone marrow-derived mast cells activated by a specific antigen/antibody reaction. MethodsIncreasing concentrations of PPT were pretreated 5min prior to antigen stimulation, and various inflammatory mediator releases and their relevant cellular signaling events were measured in those cells. ResultsPPT dose-dependently reduced the release of histamine and leukotrienes in both types of mast cells. Especially, in activated bone marrow-derived mast cells, PPT inhibited the expression of Syk protein, cytokine mRNA, cyclooxygenase-1/2, and phospholipase A2 (PLA2), as well as the activities of various protein kinase C isoforms, mitogen-activated protein kinases, PLA2, and transcription factors (nuclear factor-κB and activator protein-1). ConclusionPPT reduces the release of inflammatory mediators via inhibiting multiple cellular signaling pathways comprising the Ca2+ influx, protein kinase C, and PLA2, which are propagated by Syk activation upon allergic stimulation of mast cells.

20(S)-Protopanaxadiol 및 20(S)-Protopanaxatriol이 활성화된 비만세포로부터의 염증 매개체 유리에 미치는 영향

인삼 사포닌은 면역계에 다양한 약리 효과를 발휘한다. 20(S)-프로토파낙사다이올 (PPD) 및 20(S)- 프로토파낙사트리올 (PPT)은 장내 세균에 의하여 생성되는 인삼 대사체의 일종이며 생체 내 투여 시 순환계에서 탐지된다. 활성화된 비만 세포로부터의 염증 매개체 유리에 미치는 20(S)-프로토파낙사다이올 (PPD) 및 20(S)-프로토파낙사트리올 (PPT)의 영향을 평가하였다. 인삼 사포닌 대사체를 처치 후, 히스타민 유리는 활성화된 해명 폐 비만세포에서 평가하였으며, 인터루킨-4, 인터루킨-8, 및 종양괴사인자-알파 유리는 HMC-1 비만세포에서 평가하였다. 결과는 다음과 같다. PPT는 최고 <TEX>$100\;{\mu}M$</TEX> 농도에서 PMA에 의하여 자극된 HMC-1 세포로부터의 인터루킨-4 유리를 완전히 차단하였다. 또한, 이는 HMC-1 세포로부터의 인터루킨-8의 유리를, PMA와 DMSO동시 처치 시얻어진 수치를 기준으로 대략 40-50% 정도 억제하였다. PPD는 최고 <TEX>$100\;{\mu}M$</TEX> 농도에서 해명 폐 비만세포로부터의 히스타민 유리를 초래하였으나 통계적 유의성은 없었다. PPD는 HMC-1 세포에 PMA와 DMSO 동시 처치 시 얻어진 수치를 기준으로 할 때, 인터루킨-4의 유리를 대략 89% 정도 억제하였으나, 인터루킨-8의 유리에는 유의적인 효과를 초래하지 않았다. 그러나 PPD 및 PPT 모두, PMA에 의하여 자극된 HMC-1 세포로부터의 종양괴사 인자-알파의 유리에는 전혀 효과를 나타내지 않았다. 그러므로 본 연구 결과는 PPD와 PPT가 경구로 투여된 인삼 추출물의 면역조절 작용을 담당하는 장내 인삼 대사체 중의 한 종류임을 제시한다. Ginseng saponins have various pharmacological effects on the immune system. 20(S)-protopanaxadiol (PPD) and 20(S)-protopanaxatriol (PPT) are the species of ginseng saponin metabolites that are formed by human intestinal bacteria and detected in circulation. The effects of PPD and PPT on the inflammatory mediator release from the activated mast cells were tested. Histamine release was evaluated in activated guinea pig lung mast cells, and the secretion of interleukin-4 (IL-4), interleukin-8 (IL-8), and the tumor necrosis factor-<TEX>${\alpha}$</TEX> (TNF-<TEX>${\alpha}$</TEX>) was assessed in an HMC-1 cell after treating it with ginseng saponin metabolites. The results are as follows. PPT, at its maximum concentration of <TEX>$100\;{\mu}M$</TEX>, completely abolished the secretion of IL-4 from the PMA-stimulated HMC-1 cell. It also inhibited IL-8 secretion from the same cells by about 40-50% of the PMA-treated DMSO control. PPD, at its maximum concentration of <TEX>$100\;{\mu}M$</TEX>, showed a tendency to induce histamine release from the guinea pig lung mast cells. It inhibited the secretion of IL-4 (by 89% of the PMA-treated DMSO control) in the PMA-stimulated HMC-1 cell, but did have a significant effect on the IL-8 release from the same cell. Both PPD and PPT showed no effects, however, on the release of TNF-<TEX>${\alpha}$</TEX> from the PMA-stimulated HMC-1 cell. These results suggest that PPD and PPT are from the ginseng metabolites that are responsible for the immunomodulating activity of ginseng extracts when they are taken orally.

Eupatilin Blocks Mediator Release Via Tyrosine Kinase Inhibition in Activated Guinea Pig Lung Mast Cells

Eupatilin, an extract from Artemisia asiatica Nakai, is known to exert anti-gastric ulcer, anticancer, and anti-inflammatory effects. The aim of this study was to elucidate whether eupatilin has antiallergic reactions in activated guinea pig lung mast cells compared to apigenin and genistein. Mast cells were purified from guinea pig lung tissues by using enzyme digestion and rough and discontinuous density Percoll gradient. The purified mast cells were sensitized with immunoglobulin (Ig) G1 (anti-OVA antibody) and challenged with ovalbumin (OVA). Histamine was assayed using an automated fluorometric analyzer, leukotrienes by radioimmunoassay, and tyrosine phosphorylation by immunoblotting. Intracellular Ca2+ was analyzed by confocal laser scanning microscopy, protein kinase C (PKC) activity using protein phosphorylated with [γ-32P]ATP, and phopholipase D activity (PLD) and phosphatidic acid by using labeled phosphatidyl alcohol. Eupatilin, apigenin, or genistein reduced histamine release and leukotriene synthesis in a does-dependent manner. Eupatilin inhibited mediators to a greater extent than apigenin or genistein. Eupatilin, apigenin, and genistein initially blocked phosphorylation of Syk tyrosine and Ca2+ influx, PLD activity, phosphatidic acid, and Ca2+-dependent PKC α/βII activities during mast cell activation in a dose-dependent manner. Our data suggest that eupatilin initially inhibits Syk kinase, and then blocks downstream multisignal pathways and Ca2+ influx during mast cell activation triggered by a specific antigen–antibody reaction. Thus, eupatilin may have use clinically as a treatment for inflammatory disorders associated with allergic diseases including asthma.

Peroxynitrite Modulates Release of Inflammatory Mediators from Guinea Pig Lung Mast Cells Activated by Antigen-Antibody Reaction

Background: Peroxynitrite (ONOO^–), the product of the reaction between the superoxide anion (·O₂^–) and nitric oxide (NO), is produced during inflammatory disease and may be a major cytotoxic agent. No reports are available as to whether ONOO^– generates or modulates inflammatory mediator release from activated guinea pig lung mast cells. In this study, we explored the modulatory role of intracellular ONOO^– on inflammatory mediator release (histamine and leukotrienes) from activated mast cells. Methods: Guinea pig lung mast cells were purified by the enzyme digestion, and by using the rough and discontinuous Percoll density gradients. Mast cells were sensitized with IgG1 (anti-ovalbumin) antibody and challenged with ovalbumin (OVA). The intracellular ROS formation was determined by following the oxidative production of 2′, 7′-dichlorofluorescein diacetate (DCFH-DA), dihydrorhodamine 123 (DHR), and anti-nitrotyrosine antibody immunofluorescence. Histamine was assayed using a fluorometric analyzer, leukotrienes by radioimmunoassay, intracellular Ca²⁺ levels by confocal scanning microscopy, and PLA₂ activity using prelabeling of [³H]arachidonic acid. Results: ROS detected by DCFH-DA weakly increased in mast cells activated with OVA (1.0 g/ml), and the ROS so generated was inhibited by ebselen (50 µM). However, the ROS detected by DHR increased 3-fold under the same conditions. Peroxynitrite scavengers sL-MT, DMTU, and inhibitor FeTPPS inhibited ROS formation but the NADPH oxidase inhibitor diphenyleneiodonium (DPI) only partially inhibited this formation. Dimethyl thiourea (DMTU) and seleno-L-methionine (sL-MT) inhibited the tyrosine nitration of cytosolic proteins, the release of histamine and leukotrienes, Ca²⁺ influx, and the PLA₂ activity evoked by mast cell activation. Conclusion: The data obtained suggests that the ROS generated by the antigen/antibody reaction activated mast cells is ONOO^–, and that this modulates the release of inflammatory mediators via Ca²⁺-dependent PLA₂ activity.

Eupatilin blocks mediator release via tyrosine kinase inhibition of initial stage in activated guinea pig lung mast cells

Eupatilin blocks mediator release via tyrosine kinase inhibition of initial stage in activated guinea pig lung mast cells

The Inhibitory Mechanism of Rebamipide on the Mediator Release in the Guinea Pig Lung Mast Cells Activated with Specific Antigen-Antibody Reactions

This study aim was to assess the effects of rebamipide on the mechanism of histamine release and biosynthesis and release of leukotrienes caused by mast cell activation. We purified mast cells from guinea pig lung tissues by the use of enzyme digestion, the rough and the discontinuous density percoll gradient method. Mast cells were sensitized with IgG1 (anti-OVA) antibody and challenged with ovalbumin. Mast cells were also stimulated with A23187 and the intracellular Ca²⁺ level was measured. Histamine and leukotrienes were measured by automated fluorometric analyzer and radioimmunoassay, respectively. The intracellular Ca²⁺ level was analyzed using a confocal laser scanning microscope. Protein kinase C (PKC) activity was determined by protein phosphorylated with [γ-³²P]ATP. The phospholipase D activity was assessed by the labeled phosphatidylalcohol. Mass 1,2-diacylglycerol (DAG) was measured by the [³H]DAG produced when prelabeled with [³H]myristic acid. PLA₂ activity was determined by measuring the arachidonic acid released from the labeled phospholipids. Rebamipide decreased the releases of histamine and leukotrienes, and completely blocked Ca²⁺ influx during mast cell activation by antigen-antibody reactions. It also decreased the release of histamine and leukotrienes during mast cell activation by A23187. The PKC and PLD activities were also decreased by rebamipide in a dose-dependent manner. Rebamipide inhibited the mass DAG production and PLA₂ activity during mast cell activation. The data suggest that rebamipide inhibits intracellular signals and blocks Ca²⁺ influx in mast cells activated by specific antigen-antibody reactions, which in turn inhibits histamine release and leukotriene generation.

THE POTENTIATION OF THE HISTAMINE RELEASE INDUCED BY ADENOSINE IN MAST CELLS FROM GUINEA PIG LUNG AND HEART: SHARP DEPENDENCE ON THE TIME OF PREINCUBATION

We studied here the effect of a wide range of adenosine concentration and time of preincubation, on the histamine release induced in the guinea pig mast cells by different stimulus. Adenosine (10−5–10−3m) potentiated the histamine release induced by antigen in the guinea pig heart (isolated and dispersed tissue) and lung mast cells but not induced by ionophore A23197. The potentiation caused by adenosine (10−4m) was maximum after 1–3 min of preincubation and is probably an extracellular effect since it was not avoided by dipyridamol (3×10−7–10−6m) that inhibit the uptake of adenosine. Similar potentiation was also produced by the adenosine mimetic 2-chloroadenosine (10−5m) and both effects were inhibited by 8-phenyltheophylline indicating an effect on the type A receptors. It is suggested that the adenosine potentiation may not be related to changes on the cyclic AMP levels. 2000 Academic Press@p$hr

Effects of ethanol, acetaldehyde, and acetic acid on histamine secretion in guinea pig lung mast cells

We studied the direct effects of ethanol and its metabolites on the guinea pig lung mast cell, and the alterations caused in the histamine release induced by different stimuli. Guinea pig lungs cells dispersed by collagenase were used throughout. High concentrations of ethanol (100 mg/ml), acetaldehyde (0.3–3 mg/ml) and acetic acid (3 mg/ml) induced histamine release that was not inhibited by sodium cyanide (0.3 mM). Lower concentration of ethanol (10 mg/ml) and acetic acid (0.3 mg/ml), but not acetaldehyde, inhibited the histamine release induced by antigen and ionophore A23187. The histamine release induced by phorbol 12-miristate 13-acetate (1 μM) was also inhibited by ethanol (10 mg/ml). Changes in the levels of calcium, glucose and phosphatidic acid did not influence the effect of ethanol. We conclude that high doses of ethanol, acetaldehyde, and acetic acid cause a cytotoxic histamine release by independent mechanisms. Low concentrations of acetic acid inhibit the histamine release by pH reduction. Ethanol acts by a generalized effect that is independent of calcium and glucose suggesting a nonspecific effect that, nevertheless, is not cytotoxic since it can be reversed by washing the cells.

Inhibitory effect of ginsenoside on the mediator release in the guinea pig lung mast cells activated by specific antigen-antibody reactions

We reported that some components of ginsenosides decreased mediator release which was evoked by the activation of mast cells caused by specific antigen-antibody reactions. This study aimed to assess the effects of ginsenoside, Rb 1, which belongs to the protopanaxadiol, on the mechanism of mediator release during mast cell activation. Pretreatment of Rb 1 (100 μg) significantly decreased histamine and leukotriene in a dose-dependent manner during mast cell activation. The PLD activity during mast cell activation decreased in the pretreatment of Rb 1 (300 μg). The amount of DAG produced by PLC activity decreased because of Rb 1 pretreatment. The amount of mass DAG decreased due to Rb 1 pretreatment during mast cell activation. Rb 1 (300 μg) pretreatment strongly inhibited the incorporation of the [ 3H]methyl moiety into phospholipids. The data suggest that Rb 1, purified from Korean Red Ginseng Radix, inhibits an increase of DAG production during mast cell activation caused by antigen-antibody reactions, which is mediated via phosphatidylcholine-PLD and phosphatidylinositol-PLC systems. This is then followed by the inhibition of histamine releases. Furthermore, Rb 1 reduces the phosphatidylcholine production by inhibiting the methyltransferase I and II, and the reduction of phosphatidylcholine production inhibits leukotriene release.

The effects of cromakalim on the mediator releases from guinea pig lung mast cell activated by specific antigen-antibody reactions.

The inhibitory effect of cromakalim on the mediator release from mast cells caused by antigenantibody reactions was in controversy with the specific antigen used. However, it has recently been observed that cromakalim inhibits the release of mediators from superfused tracheal and parenchymal strips or lung mast cells after passive sensitization with the IgG1 antibody. An attempt, therefore, was made to determine the inhibitory mechanisms of cromakalim on the release of mediators such as histamine and leukotriene released by the activation of enzymes during mast cell activation. Guinea pig lung mast cells were purified through enzyme digestion, rough percoll and continuous percoll density gradients. The purified mast cells were prelabeled with [3H]palmitic acid. PLD activity was assessed more directly by the production of labeled phosphatidylethanol by PLD-mediated transphosphatidylation in the presence of ethanol. In the cells labelled with [3H]myristic acid, [3H] DAG production was measured. The methyltransferase activity was assessed by measuring the incorporation of [3H]methyl moiety into phospholipids in sensitized mast cells labelled with L-[3H] methylmethionine. cAMP level was measured by radioimmunoassay. Cromakalim resulted in a decrease in the amount of histamine and leukotrienes releases by 30% in the ovalumin-induced mast cell. Cromakalim had little effect on phospholipase D activity enhanced by the activated mast cell. Cromakalim inhibited the initial increase of diacylglycerol production during mast cell activations. Cromakalim inhibited the phospholipid methylation increased in the activated mast cell. These results show that cromakalim decreases histamine release by inhibiting the initial increase of 1,2-diacylglycerol during the mast cell activation, which is mediated via the phosphatidylinositide-phospholipase C system rather than the phosphatidylcholine-phospholipase D system. Furthermore, cromakalim reduces phosphatidylcholine production by inhibiting the methyltransferase, which decreases the conversion of phosphatidylcholine into arachidonic acid and inhibits the production of leukotrienes.

Peptidoleukotriene (pLT) release from guinea pig lung mast cells.

Guinea pig lung mast cells can be isolated and purified to high purity. This has given us the opportunity to study in greater detail mediator release from these cells. Both immunologic (ovalbumin sensitized) and nonimmunologic (calcium ionophore, CaI) stimuli caused a dose-dependent release of histamine and pLT from monodispersed lung cells and highly purified lung mast cells. Examination of the time release curve for pLT revealed a 5-min lag in the release of this mediator and a peak release at 60 min after challenge with antigen. Verification of pLT release was obtained by use of the 5-lipoxygenase inhibitor A64077 (Zileuton). Pretreatment of lung mast cells with the 5-lipoxygenase inhibitor prevented release of pLT by either antigen or CaI but had no appreciable effect on histamine release (HR). The pulmonary mast cell appears to be an important contributor to pLT release in the guinea pig lung.

Passive sensitization of guinea-pig lung mast cells and alveolar macrophages with anti-ovalbumin IgG1 and IgG2 antibodies

In the present study, we performed experiments to explore the role of IgG immunoglobulins in type I hypersensitivity reactions in the guinea-pig. Our results suggest that IgG immunoglobulins may be involved in allergic reactions, both through histamine release from mast cells and thromboxane A2 release from macrophages after challenge of passively sensitized cells.

Microfilament-associated degranulation of sensitized guinea-pig lung mast cells

When sensitized guinea-pig lung mast cells were exposed to antigen, granules were pushed out on to the cell surface. Subsequently, thin filaments, some extending as long as 15 microns, were projected radially with the extruded granules. The latter became swollen in the extracellular medium and the elongated filaments became shorter, until, within 7-8 min, the granules were reincorporated into the cytoplasm. The time course of morphological changes corresponded approximately to that of changes in the intracellular Ca2+ concentrations. The filaments connecting the extruded granules to the cell surface were stained with rhodamine-phalloidin, indicating that they consisted mainly of actin.

The Effect of Parainfluenza 3 Infection on Guinea Pig Basophil and Lung Mast Cell Histamine Release

The guinea pig infected with parainfluenza 3 (P-3) has provided an animal model to study mechanisms of virus-induced asthma and airway hyperreactivity. Evidence to identify the mechanisms by which P-3 infection enhances airway hyperreactivity, however, has not been established. The present study evaluated the effect of P-3 infection on histamine release (HR) from isolated peripheral blood guinea pig basophils and lung mast cells. Basophil HR was determined in animals made basophilic with 13 daily intraperitoneal injections of sheep blood. On Day 10 of the sensitization, blood was obtained from all animals, and leukocyte HR was determined to sheep gammaglobulin (SG), which served as the secretagogue. After these baseline studies, the animals were separated into two groups; one was insufflated with P-3 virus and the other with growth medium that did not contain virus. Four days later, animals were killed, basophils and lung mast cells were isolated, HR was determined, and lung tissue was cultured for the presence of virus. In basophils isolated from animals proven to be infected with P-3 virus, we found significantly more histamine released in response to the lower doses of SG and a significant shift in the -log EC50 dose of this antigen causing HR in these cells. Experiments conducted with the calcium ionophore A23187 as the basophil secretagogue in the same experimental design did not demonstrate similar HR findings. Furthermore, a defect in calcium disposition did not account for the enhanced HR from basophils isolated from infected animals. HR from pulmonary mast cells was similar in cells isolated from P-3-infected and control insufflated animals.(ABSTRACT TRUNCATED AT 250 WORDS)

ACE‐Inhibitor‐Induced Enhancement of Spontaneous and IgE‐Mediated Histamine Release from Mast Cells and Basophilic Leukocytes and the Modulatory Effect of Capsaicin Sensitive Nerves

Frequently reported adverse inflammatory skin and airway reactions have been reported in subjects being medicated with angiotensin converting enzyme (ACE)-inhibitors. Intradermally evoked wheal and flare reactions to ovalbumin, capsaicin and bradykinin, in ovalbumin sensitized guinea pigs, was previously demonstrated to be enhanced by pretreatment with the ACE-inhibitor MK 422 (the active parent diacid of enalapril). In vitro results from this study demonstrate that the ACE-inhibitor MK 422 degranulated guinea pig lung and skin mast cells as well as human basophils, and enhanced allergen-evoked histamine release. Local capsaicin pretreatment in vivo of guinea pig skin decreased spontaneous and allergen-triggered release of histamine in vitro from skin mast cells. No clear enhancing effect of MK 422 was seen on the allergen-triggered histamine release in vitro from capsaicin pretreated skin, and the spontaneous release was unaffected by the ACE-inhibitor. The allergen-triggered wheal and flare reaction in ovalbumin sensitized guinea pigs was potentiated by MK 422 and the late phase reaction of the inflammatory response was especially augmented. Capsaicin pretreatment of the guinea pigs abolished this late phase reaction as well as the inflammatory enhancing effect of MK 422. Our in vitro results from capsaicin pretreated skin indicate that the reduced inflammatory response in vivo in capsaicin pretreated skin is due not only to capsaicin induced depletion of neuropeptides from sensory nerves, but also to secondary degranulation of mast cells by one or more of these peptides.

Production and properties of histamine-releasing factor of guinea pigs.

The production of histamine-releasing factor (HRF) by human mononuclear cells has previously been reported. In this paper we describe the production of HRF by guinea pig spleen cells, thymocytes, and PBMC. Guinea pig lymphoid cells were cultured either alone or in the presence of mitogens (PHA and Con A) or specific Ag(OVA and keyhole limpet hemocyanin) and the dialyzed cell-free supernatant was tested for histamine-releasing activity on guinea pig lung mast cells and blood basophils. Lung mast cells were isolated by enzymatic digestion and partially purified by countercurrent elutriation and discontinuous Percoll gradient centrifugation. Guinea pig spleen cells, thymocytes, and PBMC spontaneously produced significant amounts of HRF. The production was enhanced upon stimulation with PHA or specific Ag in animals immunized with Ag in CFA. Two distinct species of HRF were identified with m.w. of 50,000 to 70,000 and 5000 to 8000 by gel chromatography. HRF is a trypsin- and chymotrypsin-sensitive heat-stable protein. It does not bind to Con A-Sepharose and its production is not inhibited by tunicamycin. HRF-induced histamine release from lung mast cells is a temperature-dependent process and is complete in 10 min at 37 degrees C. Intradermal injection of HRF caused an immediate ear-swelling reaction in guinea pigs. The most severe ear-swelling reactions did not resolve within 1 h, but instead evolved over a period of 12 to 24 h.

Activation of guinea pig lymphocytes and mast cells by grain-dust extract

Previous studies have demonstrated that grain dust can stimulate lymphocyte proliferation and the production of interleukin-1 by macrophages. This study was undertaken to investigate whether grain dust could stimulate the production of histamine-releasing factor (HRF) by guinea pig spleen cells. We also studied the direct action of grain dust on guinea pig lung mast cells and basophils. Plastic nonadherent cells from immunized animals were cultured for 24 hours in the presence of grain dust or lipopolysaccharide, and the cell-free supernatants were assayed for HRF activity in the mast cell and basophil histamine release test. Lung mast cells were isolated by enzymatic digestion and discontinuous Percoll gradient centrifugation. It has been demonstrated that grain dust stimulated the production of HRF by spleen cells from the immunized animals but not from the control animals. Stimulation of spleen cells with lipopolysaccharide did not enhance the HRF production. Gel chromatography of grain dust-stimulated supernatant revealed that HRF has a molecular weight in the range of 50 to 70 kd and 5 to 8 kd. We also found that grain dust directly released significant amount of histamine from both mast cells and basophils. The results of this study suggest that grain dust contains some potent active substances that can activate lymphocytes, mast cells, and basophils.

Monoclonal anti-histamine antibody: Preparation, characterization and application to enzyme immunoassay of histamine

An enzyme immunoassay to measure histamine has been developed. A histamine-bovine serum albumin conjugate was prepared using 1,4-benzoquinone as the coupling agent and was employed to immunize mice for the preparation of monoclonal antibodies against histamine. After an initial screening to identify antigen-binding monoclonal antibodies the clones were isolated by limiting dilution cloning, grown in ascites and antibodies which had been secreted into the ascitic fluid were precipitated by ammonium sulphate at 50% saturation. A systematic approach for the determination of epitope specificities of monoclonal antibodies was performed. It was found that for the most specific antibody the main epitope encompassed the 2-histaminyl-1,4-benzoquinone moeity and that the K D value determined by indirect ELISA was 1.5 × 10 −8 M for the hapten part of the immunogen and 4.6 × 10 −10 M for a histamine-Bq-ovalbumin conjugate. The selected monoclonal antibody could not recognized histidine or methyl-histamine. Using this antibody, we developed an enzyme immunoassay for histamine and pg amounts could be detected. The same assay was used to quantify the allergic release of histamine from guinea pig lung mast cells. Results obtained either by the present enzyme immunoassay or by a fluorometric assay were closely correlated (correlation coefficient r = 0.9702, n = 37).

A procedure for isolation and partial purification of guinea pig lung mast cells

Pulmonary mast cells were obtained from guinea pig lung using a combination of enzymatic digestion of tissue, centrifugal elutriation, and density gradient centrifugation on Percoll. In the initial procedure, lung tissue was enzymatically digested with collagenase and elastase in four 30 min incubations. Typically, monodispersed cell suspensions contained 4% mast cells. Further purification of these lung mast cells using elutriation and Percoll gradients consistently yielded mast cells of 40–78% (mean 51%) purity. These cells were morphologically intact, viable and found to be functional as determined by histamine release evoked by antigen and anti-guinea pig IgG 1 antibody.

Modulation of histamine release by fatty acids. A new in vitro model investigating adverse drug reactions in various species

Histamine release caused by drugs and/or their solvents is a well known phenomenon. In this study, both in vivo (anaesthetized and conscious dogs) and in vitro (isolated rat peritoneal, human and guinea-pig lung mast cells) models were used. Cremophor E1 and six derivatives of 12-hydroxystearic acid were compared for their histamine releasing abilities. Although the three types of isolated mast cells responded similarly, histamine release being observed with DH (the diester of 12-hydroxystearic acid with polyethylene glycol), TN (12-hydroxystearic acid polymerized with ethylene oxide) and ME (the monoester of 12-hydroxystearic acid esterified with polyethylene glycol), the anaesthetized dog exhibited elevated blood histamine levels and clinical symptoms after administration of all the solubilizing agents, except TN and ME. The reasons for this discrepancy are not known. The addition of the drugs (Althesin or propanidid) to their solubilizing agents caused histamine release, which was not observed with the solubilizing agent alone. This is the first demonstration of an in vitro model, which copies the clinical situation, i.e. solvent does nothing but the solvent plus drug combination causes histamine release and hence adverse reactions.