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Kinases are believed to play a crucial role in the expression and activation of inflammatory mediators in the airway, in T-cell function and airway remodelling. Important kinases such as Inhibitor of kappaB kinase (IKK)2, mitogen activated protein (MAP) kinases and phsopho-inositol (PI)3 kinase regulate inflammation either through activation of pro-inflammatory transcription factors such as activating protein-1 (AP-1) and nuclear factor kappaB (NF-kappaB), which are activated in airway disease, or through regulation of mRNA half-life. Selective kinase inhibitors have been developed which reduce inflammation and some characteristics of disease in animal models. Targeting specific kinases that are overexpressed or over active in disease should allow for selective treatment of respiratory diseases. Interest in this area has intensified due to the success of the specific Abelson murine leukaemia viral oncogene (Abl) kinase inhibitor imatinib mesylate (Gleevec) in the treatment of chronic myelogenous leukaemia. Encouraging data from animal models and primary cells and early Phase I and II studies in other diseases suggest that inhibitors of p38 MAP kinase and IKK2 may prove to be useful novel therapies in the treatment of severe asthma, chronic obstructive pulmonary disease (COPD), cystic fibrosis and other inflammatory airway diseases.

Airway mucosal changes have been reported in chronic cough. In cough variant asthma and in eosinophilic bronchitis, there is evidence of eosinophil infiltration and sub-basement membrane thickening. In non-asthmatic cough, an increase of bronchoalveolar mast cells, mucosal mononuclear cells, and epithelial shedding have been reported. In a more recent study, evidence of airway wall remodelling has been observed in both asthmatic and non-asthmatic cough, such as an increase in sub-basement membrane thickness, goblet cell area, vascularity and vessel size. Smooth muscle area was increased in non-asthmatic coughers. Heightened cough sensitivity in non-asthmatic coughers was related to the degree of goblet cell hyperplasia and epithelial shedding. Cough reflex may be heightened by increased production of growth factors that might be further enhanced by the physical effects of cough on the airways. Mast cells may participate in the cough pathophysiology through release of growth factors as well as tussive mediators. Changes in the airway wall mucosa and epithelium may be important in the pathogenesis of cough receptor sensitization.

Neuropeptides such as neuropeptide Y (NPY) have long been proposed to play a role in the pathogenesis of inflammatory diseases. NPY is a 36 amino acid neuropeptide which participates in the regulation of a large number of physiological and pathophysiological processes in the cardiorespiratory system, immune system, nervous system and endocrine system. Serum levels of NPY are increased during exacerbations of asthma, whereas the number of NPY-immunoreactive nerves in the airways remains constant in the airways of patients with inflammatory airway diseases such asthma or rhinitis. Next to a role in the regulation of glandular activity, NPY exerts a major influence on humoral and cellular immune functions. In this respect, NPY is known to modulate potent immunological effects such as immune cell distribution, T helper cell differentiation, mediator release, or natural killer cell activation. In addition to these direct effects, NPY also acts as an immunomodulator by influencing the effects of a variety of other neurotransmitters. Whereas the peptide has been focused for therapeutic options in the central nervous system, a potential use in the treatment of pulmonary inflammatory disorders has not been revealed so far due to the complex pulmonary effects of NPY. However, since selective antagonists and agonists and gene-depleted animals for the different receptors are now available, NPY may be of value for future strategies in airway nerve modulation.

Protease-activated receptors are G protein-coupled receptors activated by serine-proteases. Protease-activated receptor 2 is involved in the regulation of airway smooth muscle tone but its effects vary according to species and experimental conditions. We determined the effects of protease-activated receptor 2 activation on smooth muscle tone and airway reactivity to histamine in guinea pigs and smoking or non-smoking humans. The effects of trypsin and protease-activated receptor activating peptide on the isometric tension and response to histamine of guinea pig tracheal and human bronchial rings were studied. Human tissues were obtained from 6 smokers and 4 non-smokers. We assessed the effects of epithelial removal, inhibitors of cyclooxygenases, nitric oxide synthases, neutral endopeptidase and antagonists of acetylcholine, histamine, bradykinin and tachykinin receptors. Bronchomotor responses to protease-activated receptor 2 activation were variable in guinea pig, in half of animals PAR2 activation induced smooth muscle relaxation through the epithelial release of prostanoids but not of nitric oxide. In human airways, protease-activated receptor 2 activation reduced responsiveness to histamine in bronchial rings from smokers but increased responsiveness in bronchi from non-smokers. This study demonstrates an influence of tobacco smoking on the effect of protease-activated receptor 2 activation on airway responsiveness in humans, with an increased protection against histamine-induced contractions, probably through an increased epithelial release of prostanoids. The role of airway protease-activated receptor 2 may be to maintain smooth muscle tone homeostasis.

Background: Protease-activated receptors 1 and 2 (PAR-1 and PAR-2) are 7-transmembrane G protein-coupled receptors activated by serine proteases in many cell types, including monocytes-macrophages, leading to the production of pro-inflammatory mediators, cytokines, and growth factors. Objective: We determined the influence of chronic smoking and asthma on the expression of PAR-1 and PAR-2 receptors on alveolar macrophages (AMs). Methods: We used RT-PCR and immunocytochemistry with confocal microscopy to determine mRNA and protein expression of PAR-1 and PAR-2 in AMs obtained from healthy smokers, asthmatic patients, and healthy subjects. In addition, we examined the effect of IL-1β and LPS. Results: PAR1 mRNA was decreased, whereas PAR2 mRNA was increased in 24-hour cultured AMs from smokers when compared with values in AMs from healthy subjects. Paradoxically, there was a higher degree of PAR-1 protein staining in AMs from smokers, whereas PAR-2 staining was similar in smokers and healthy subjects. PAR-1 and PAR-2 mRNA and protein expression were similar in asthmatic patients and control subjects. IL-1β and LPS had no effect on PAR1 and PAR2 gene expression by AMs. Conclusions: There is a dissociation between gene and protein expression of PAR-1 and PAR-2. PAR-1 protein overexpression in AMs from smokers might be important in the pathophysiology of chronic airways disease. (J Allergy Clin Immunol 2003;111:367-73.)

Background Asthma is associated with remodeling of the extracellular matrix (ECM) and increased airway obstruction, and the mechanisms of this process are unknown. Matrix metalloproteinases (MMPs) are a group of enzymes capable of degrading the ECM. They are released along with their inhibitors, tissue inhibitor of MMP (TIMP). Study objectives To determine whether severe, persistent asthma is associated with increased levels of MMP-9 in the airway compared with mild asthma, and to assess the effect of both allergen exposure and steroid treatment on MMP-9 and TIMP-1 levels. Design Prospective analysis of levels and activity of MMP-9 and TIMP-1 in BAL fluid (BALF) and induced sputum obtained from asthmatics of differing disease severity. In patients with mild asthma, MMP-9 and TIMP-1 levels were studied in induced sputum following allergen challenge and in BALF after inhaled steroid therapy. Patients Eighteen patients with mild asthma, 10 patients with severe asthma, and 10 nonsmoking, atopic subjects had their sputum studied. Fourteen of the patients with mild asthma underwent allergen challenge. BAL was collected from 16 patients with mild asthma before and after 4 weeks treatment with inhaled budesonide, 800 μg bid, or placebo. Results Patients with severe asthma had increased levels and activity of sputum MMP-9 in their sputum compared with patients with mild asthma and normal subjects. Allergen challenge increased the MMP-9/TIMP-1 ratio and MMP-9 activity. Inhaled budesonide had no effect on MMP-9 or TIMP-1 in patients with mild asthma. Conclusions MMP-9 may play a role in chronic airway inflammation and remodeling in asthma, as concentrations are increased in severe, persistent asthma and following allergen challenge. Inhaled steroids may not affect MMP-9 and TIMP in patients with mild asthma, and additional studies in patients with more severe asthma are needed.

The role of nitric oxide (NO) in allergic inflammation and bronchial hyperresponsiveness is unclear. We studied a selective prodrug nitric oxide synthase (NOS)-2 inhibitor, L-N(6)-(1-iminoethyl)lysine 5-tetrazole amide (SC-51). In ovalbumin-sensitized and challenged rats, exhaled NO levels increased by 3 h following challenge (3.73 +/- 0.74 ppb; P0.05), peaking at 9 h (11.0 +/- 2.75; P0.01) compared to saline controls (1.87 +/- 0.26; P0.05 and 2.81 +/- 0.18; P0.01). Immunoreactive lung NOS2 expression was increased in ovalbumin-challenged rats compared with ovalbumin-sensitized, saline-challenged rats at 8 h post-challenge. SC-51 (10 mg/kg; p.o.) inhibited allergen-induced increase in exhaled NO levels to 1.3 +/- 0.17 ppb. SC-51 inhibited bronchial hyperresponsiveness in ovalbumin-sensitized and challenged rats (P0.05). In sensitized non-exposed rats, SC-51 increased bronchial responsiveness (P0.05). SC-51 reduced the allergen-induced increase in bronchoalveolar lavage neutrophils, but caused a nonsignificant reduction in bronchial mucosal eosinophil numbers. NO generated through NOS2 contributes to allergen-induced bronchial hyperresponsiveness but not to bronchial eosinophilia, indicating that these are independently expressed.

The two kinin receptors, B(1) and B(2), are upregulated in inflammation and may play a role in diseases such as asthma. In pulmonary A549 cells, TNF-alpha or interleukin-1 beta dramatically increased bradykinin B(1) and B(2) receptor mRNA expression and this response was prevented by dexamethasone. In primary human bronchial epithelial cells, bradykinin B(1) receptor mRNA expression showed a similar trend, whereas bradykinin B(2) receptor showed almost constitutive expression. Radioligand-binding studies revealed significant increases in bradykinin B(2) receptor protein expression following both interleukin-1 beta and TNF-alpha treatment of A549 cells; however, no evidence was found for bradykinin B(1) receptor. Functionally, the bradykinin B(2) receptor ligand, bradykinin, but not the B(1) ligand, des-Arg(10)-kallidin, produced a marked increase in prostaglandin E(2) release when administered following interleukin-1 beta treatment. Arachidonic acid release in response to bradykinin was markedly enhanced by prior incubation with interleukin-1 beta and this was prevented by the prior addition of dexamethasone.

In human airway smooth muscle cells, the levels of RANTES were increased upon stimulation with interleukin-1beta together with tumour necrosis factor-alpha (TNF-alpha) (10 ng ml(-1) for each). In this study, we have assessed the effects of prostaglandin E(2) and the beta(2)-adrenoceptor agonist, fenoterol on RANTES (regulated upon activation, normal T cell expressed and secreted) release by these cells. The levels of RANTES released by human airway smooth muscle cells were measured after 24 h of treatment. Prostaglandin E(2) and fenoterol, only in presence of a cyclo-oxygenase inhibitor indomethacin (10(-6) M), provoked a concentration-dependent reduction in RANTES release. These data suggest that, in settings where cyclo-oxygenase activity is low, both drugs may relieve the symptoms of airway diseases by reducing RANTES production.

Background Interleukin-5 (IL-5) is essential for the formation of eosinophils, which are thought to have a major role in the pathogenesis of asthma and other allergic diseases. We aimed to assess the effects of monoclonal antibody to IL-5 on blood and sputum eosinophils, airway hyper-responsiveness, and the late asthmatic reaction to inhaled allergen in patients with mild asthma. Methods We did a double-blind randomised placebo-controlled trial, in which a single intravenous infusion of humanised (IgG-K) monoclonal antibody to IL-5 (SB-240563) was given at doses of 2.5 mg/kg (n=8) or 10.0 mg/kg (n=8). The effects of treatment on responses to inhaled allergen challenge, sputum eosinophils, and airway hyper-responsiveness to histamine were measured at weeks 1 and 4 with monitoring of blood eosinophil counts for up to 16 weeks. Findings Monoclonal antibody against IL-5 lowered the mean blood eosinophil count at day 29 from 0.25x10(9)/L (95% CI 0.16-0.34) in the placebo group to 0.04x10(9)/L (0.00-0.07) in the 10 mg/kg group (p Interpretation A single dose of monoclonal antibody to IL-5 decreased blood eosinophils for up to 16 weeks and sputum eosinophils at 4 weeks, which has considerable therapeutic potential for asthma and allergy. However, our findings question the role of eosinophils in mediating the late asthmatic response and causing airway hyper-responsiveness.

Vitamin A binds to retinoic acid receptors, which in turn may interact with other transcription factors. We determined its effect (2500 and 5000 IU/kg) on nuclear factor-kappaB binding activity in the lung, airway inflammation and bronchial hyperresponsiveness in rats exposed to ozone. Ozone (3 ppm, 3 h) caused neutrophil influx into bronchoalveolar lavage fluid (16.2+/-0.8 x 10(5) cells/ml, p < 0.01) and bronchial hyperresponsiveness (-logPC200ACh = 2.54+/-0.19, p < 0.05, compared to control animals, respectively). Vitamin A inhibited this neutrophilia dose-dependently together with the increased DNA-binding activity of nuclear factor-KB in lung extracts. Vitamin A did not affect bronchial hyperresponsiveness at both doses. Vitamin A inhibits ozone-induced neutrophilic inflammation through a reduction in nuclear factor-kappaB DNA binding activity.

We have studied the role for 5-hydroxytryptamine (5-HT) in the modulation of the cough reflex by examining the effect of a selective 5-HT1A receptor agonist, 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT) on cough and respiratory rate in conscious guinea pigs. Animals were placed in a box and exposed to the tussive agent citric acid (0.5 M) for 10 min, 3 min after terbutaline (0.05 mg/kg i.p.) was administered to prevent bronchoconstriction. 8-OH-DPAT inhibited at low doses (0.008 and 0.016 mg/kg) but potentiated at high doses (0.25 and 0.5 mg/kg) the citric acid-induced number of coughs, but dose-dependently increased respiratory rate. Methysergide (0.05–5 mg/kg), a 5HT1 and 5HT2 receptor antagonist, and ketanserin (0.05 mg/kg), a 5HT2 receptor antagonist, had no effects on cough or respiratory rate. Methysergide inhibited the increased cough responses and respiratory rate induced by 8-OH-DPAT at high doses, while ketanserin was without effect. These results suggest that 8-OH-DPAT may induce both an inhibition and activation of the cough reflex, the latter involving central 5HT1-receptor activation.

The human cough reflex is still poorly understood, although it is known to occur independently of bronchoconstriction. Sensitization of the cough reflex is a unifying hypothesis for chronic dry cough in several conditions, including gastroesophageal acid reflux, angiotensin-converting enzyme inhibitor cough, and cough-variant asthma. The most common cause of chronic dry cough is a group of related conditions of chronic rhinitis, sinusitis, and postnasal drip. In these cases the cough reflex may be sensitized through an action of inflammatory mediators from the nasal mucosa on the airways or a reflex sensitization of airway sensory nerves. The association of cough with gastroesophageal reflux may occur through a local esophageal-tracheobronchial reflex. Angiotensin-converting enzyme inhibitor cough is a side effect of treatment in about 10% of patients; it probably results from inhibition of the degradation of kinins, particularly bradykinin, in the airway. Why some patients with asthma have cough as the principal feature of their disease is unclear. Tachykinins are probably involved in the mechanism of sensitization of the cough reflex, and the development of neuropeptide antagonists may open new research opportunities. A study that used ambulatory recording of cough in a group of subjects with asthma confirmed the presence of significant cough, the frequency of which did not correlate with lung function or diurnal variation in peak flow. This finding highlights the problem of cough in patients with asthma, a problem that probably has been underestimated in the past.

Trimellitic anhydride is a cause of occupational asthma in humans. We have previously found that tracheal instillation of trimellitic anhydride conjugated to guinea pig serum albumin induces acute bronchoconstriction and airway plasma exudation in sensitised animals, responses mediated primarily via histamine release. In the present study, neural mechanisms mediating bronchoconstriction and goblet cell secretion were determined in trimellitic anhydride-sensitised guinea pigs using the ganglionic blocker hexamethonium to eliminate efferent reflex mechanisms, pretreatment with capsaicin to eliminate afferent mechanisms, or cimetidine and mepyramine to eliminate histamine-mediated mechanisms. The magnitude of secretion of intracellular mucus from tracheal goblet cells was quantified morphometrically as a mucus score which is inversely related to the degree of discharge. Guinea pigs were injected intradermally either with 0.1 ml 0.3% trimellitic anhydride in corn oil or with corn oil alone as control. Fourteen to eighteen days later all sensitised animals had developed specific immunoglobulin (Ig) G1 antibodies whereas the controls had not. Tracheal instillation of conjugated trimellitic anhydride in anaesthitised animals significantly increased airway lung resistance ( R L ) 24-fold in sensitised guinea pigs (34.3 ± 7.9 cm H 2 O · ml −1 · s) compared with controls (1.4 ± 0.1 cm H 2 O · ml −1 · s). Mucus score was significantly reduced by 51% (indicating goblet cell secretion) in sensitised guinea pigs (183 ± 22 mucus score units) compared with controls (372 ± 41 mucus score units). The antihistamines significantly inhibited conjugated trimellitic anhydride-induced bronchoconstriction by 89%, but did not significantly affect goblet cell discharge. Hexamethonium alone did not significantly affect conjugated trimellitic anhydride-induced bronchoconstriction or goblet cell secretion. Capsaicin pretreatment (in combination with hexamethonium) significantly inhibited golet cell discharge (by 80%) but had no significant effect on bronchoconstriction. We conclude that conjugated trimellitic anhydride challenge of trimellitic anhydride-sensitised guinea pigs induces goblet cell discharge and bronchoconstriction via different mechanisms with activation of capsaicin-sensitive sensory nerves responsible for secretion and histamine release responsible for airway constriction. The guinea pig model of trimellitic anhydride-induced occupational asthma may prove useful in examination of mechanisms of goblet cell secretion in allergic diseases.

We have investigated the effect of inhaled HOE 140, a novel bradykinin B2 receptor antagonist, against bradykinin- and vagal stimulation-induced airway microvascular leakage and bronchoconstriction in anesthetized guinea-pigs. Lung resistance was measured for 6 min after challenge, followed by measurement of extravasation of Evans blue dye into airway tissues, used as an index of airway microvascular leakage. Bradykinin was given by inhalation (1 mM, 45 breaths) and bilateral vagus nerves were stimulated electrically during a 5-min period (3 and 10 Hz, 5 V, pulse width of 5 ms), both of which caused a significant increase in lung resistance and leakage of dye in the airway. HOE 140 (20 and 200 microM, 60 breaths) completely abolished both the airway effects induced by bradykinin, whereas even the higher dose of HOE 140 had no effect against those induced by electrical vagal stimulation. In conclusion, airway microvascular leakage and bronchoconstriction induced by inhaled bradykinin are mediated by activation of bradykinin B2 receptors in the guinea-pig. In contrast, mechanisms via bradykinin B2 receptors do not play an important role in the acute airway responses induced by vagal stimulation.

We have investigated the effect of a new bradykinin receptor antagonist, Hoe 140 (D-Arg- Hyp3,Thi5,D-Tic7,Oic8]-bradykinin), on bradykinin- and platelet-activating factor (PAF)-induced bronchoconstriction and airway microvascular leakage in anesthetized guinea pigs. Extravasation of Evans blue dye and lung resistance were measured simultaneously. Both i.v. (15 nmol/kg) and inhaled bradykinin (1 mM, 45 breaths) caused a significant increase in lung resistance and leakage of dye at all airway levels. Hoe 140 (100 nmol/kg i.v.) almost completely inhibited these airway responses induced by bradykinin except for dye extravasation in trachea induced by inhaled bradykinin. Inhaled PAF (3 mM, 30 breaths) significantly increased lung resistance and leakage of due at all airway levels, but Hoe 140 had no effect on these responses. Bradykinin-induced bronchoconstriction and airway microvascular leakage are predominantly mediated by activation of B2 receptor, since Hoe 140 is a B2 receptor antagonist. Bradykinin receptor-mediated mechanisms do not play an important role on inhaled PAF-induced bronchoconstriction and microvascular leakage.

Bronchial responsiveness to inhaled acetylcholine (ACh) and inflammatory cell recruitment in bronchoalveolar lavage fluid (BALF) were studied in inbred Brown-Norway rats actively sensitized to, and later exposed to, ovalbumin (OA). We examined animals 21 days after initial sensitization at 18 to 24 hours, or 5 days after a single challenge, or after the last of seven repeated exposures administered every 3 days. BALF was examined as an index of inflammatory changes within the lung. Animals repeatedly exposed to OA aerosols had an increased baseline lung resistance and a significant increase in bronchial responsiveness to inhaled ACh compared to control animals at both 18 to 24 hours and 5 days after the last OA exposure. Sensitized animals receiving a single OA aerosol also demonstrated bronchial hyperresponsiveness (BHR) to inhaled ACh (p less than 0.01) at 18 to 24 hours of a similar order as the multiple-exposed group. There was a significant increase in eosinophils, lymphocytes, and neutrophils in BALF at 18 to 24 hours but not at 5 days after single or multiple exposure to OA aerosol in the sensitized groups. Control animals demonstrated no changes in bronchial responsiveness, although a small but significant increase in inflammatory cells was observed compared to saline-only treated animals. There was a significant correlation between bronchial responsiveness and eosinophil counts in the BALF in the single allergen-exposed group (Rs = 0.68; p less than 0.05). We conclude that (1) BHR after allergen exposure in sensitized rats is associated with the presence of pulmonary inflammation but persists despite the regression of inflammatory cells in BALF after multiple OA exposures, and (2) this rat model has many characteristics of human allergen-induced BHR.

Bradykinin can be inactivated by the peptidases angiotensin-converting enzyme (ACE) and neutral endopeptidase (NEP), both of which are present in the airways. We evaluated the role of these enzymes in bradykinin-induced airway microvascular leakage and lung resistance in anesthetized and mechanically ventilated guinea pigs. We studied the effects of captopril (inhaled; 350 nmol), a specific ACE inhibitor, and phosphoramidon (inhaled; 7.5 nmol), a specific NEP inhibitor. Airway microvascular leakage was measured with the albumin marker Evans Blue dye (20 mg/kg i.v.), and airflow obstruction was measured as lung resistance (RL). Bradykinin was given by inhalation (0.1, 0.3 and 1 mM; 45 breaths), and caused a dose-dependent increase in both RL and airway microvascular leakage. Inhibition of NEP or ACE potentiated the bradykinin-induced microvascular leakage in main bronchi and proximal and distal intrapulmonary airways. However, only NEP inhibition significantly potentiated the extravasation of Evans Blue dye into the tracheal wall and lumen. The combined inhibition of NEP and ACE significantly potentiated plasma leakage at all airway levels, as well as the increase in RL induced by inhaled bradykinin. Recovery RL after one lung inflation significantly correlated with the extravasation of Evans Blue dye in the tissue at all airway levels, indicating that airway edema may have contributed to airway narrowing. We conclude that in the guinea pig, both NEP and ACE modulate bradykinin-induced airway microvascular leakage.

Furosemide has been shown recently to protect asthmatic patients against certain bronchoconstrictor challenges. We investigated the effect of furosemide on eosinophil function. Since furosemide may be exerting its inhibitory effect on the eosinophil by inhibiting anion transport, we also assessed the effects of the anion transport inhibitors 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid (DIDS). Furosemide, NPPB and DIDS inhibited the eosinophil respiratory burst in response to leukotriene B 4 (LTB 4 ) and, to a smaller extent, inhibited the response to opsonized zymosan (OZ). To assess whether the anion transport inhibitors were achieving their inhibitory effect by inhibiting an influx of Cl − ions into the eosinophil, the effect of removing extracellular Cl − on eosinophil function was determined. OZ-induced H 2 O 2 production was inhibited by removing extracellular Cl − whereas the LTB 4 response was not affected by the concentration of extracellular Cl − .

Many different mediators have been implicated in allergic responses and allergic diseases.14 It is becoming increasingly apparent that no single mediator can account for allergic diseases, such as asthma, rhinitis, eczema, or anaphylaxis, and that the clinical manifestation depends on a complex interaction of mediators that are released from a variety of cells. Although previous research concentrated on the mast cell as the key cell in allergic reactions, it now appears likely that other inflammatory cells, such as macrophages, eosinophils , neutrophils, platelets, and lymphocytes, may also contribute to the inflammatory response. The recognition that cells, other than mast cells, such as eosinophils,5 platelets,6 and macrophages,7 may be activated by IgE-dependent mechanisms has focused attention on these other cells as sources of mediators. Histamine,8 prostaglandinsg and leukotrienes3 have been previously proposed as playing a role in mediating allergic responses. Another mediator that is receiving increasing attention is PAF, a highly potent inflammatory mediator formed, like prostaglandins and leukotrienes, by the action of phospholipase A, on membrane phospholipids. The purpose of this article is to review the various effecs of PAF that may be relevant to allergic diseases and to speculate on its possible involvement in allergy. With the introduction of specific PAF antagonists during the next few years, the role of this inflammatory mediator will be clarified. had previously been demonstrated in the rabbit that platelets provided the major source of histamine among the blood elements,” but it was not known whether histamine release during allergen challenge represented a direct or indirect consequence of the recognition of allergen by antibody. Histamine release in sensitized rabbits undergoing antigen challenge was demonstrated to be the consequence of IgE-dependent activation of basophils that, in turn, released a soluble product capable of eliciting platelet activation.““S ‘* This product was termed PAF and was subsequently demonstrated to be a phospholipid.‘3 The chemical structure was then identified in 1979 by three independent groups as 1-o-alkyl-2-acetyl-sn-glyceryl-3phosphorylcholine (Fig. 1). 14-16 This phospholipid has been variously referred to as PAF-acether,14 acetyl glyceryl ether phosphorylcholine,‘5 or antihypertensive polar renomedullary lipid.16 The original nomenclature (PAF) will be used throughout this article, although it is now apparent that PAF has many other actions in addition to activation of platelets.

Because mast cells (MCs) in skin of atopic dogs are heterogeneous with respect to tissue fixation and staining properties, we determined the effect of antigen on each type of MC in vivo. Skin biopsies were done in anesthetized, ragweed-sensitized dogs before and at 1, 3, 6, and 24 hours after intradermal injections of ragweed antigen (n = 5) or glycerin diluent (n = 4). In each case, one biopsy specimen was fixed with formalin, and a second specimen from an adjacent abdominal site was fixed with basic lead acetate. In sections stained with Alcian blue, 49.7% more MCs (p less than 0.05) were detected in tissue fixed with basic lead acetate ("typical" plus "atypical" MCs: 2916 +/- 581/mm3; mean +/- SEM) than in tissue fixed with formalin ("typical" MCs: 1955 +/- 537/mm3). After antigen, the number of "typical" MCs detectable in tissue sections progressively decreased during the 24-hour period, whereas the number of "atypical" MCs was lowest at 1 hour and had increased at 24 hours. After diluent, MC numbers did not change significantly over time. A late-phase response (LPR), detected clinically as induration and edema, was present 6 hours after antigen in four of five dogs, but LPR was not detected after diluent. The size of LPR was correlated (r = 0.85; p less than 0.05) with the decrease in the number of "typical" MCs at 6 hours. We conclude that the response of the "typical" and "atypical" MC to antigen in vivo differs markedly. The "atypical" MCs participate in the early, acute response to antigen, and the "typical" MCs may be associated with the development of the LPR.

We studied the effect of a selective thromboxane-synthetase inhibitor, sodium (E)-3-[4-(1-imidazolymethyl)-phenyl]-2-propanoate) (OKY-046) on the late-phase response to antigen in ragweed-sensitized dogs. Skin biopsies were performed before and 1, 6, and 24 hours after ragweed injection. OKY-046 was infused (100 micrograms.kg-1.min) from 1 hour before until 6 hours after intracutaneous ragweed in five dogs. The early clinical response to ragweed (wheal at 20 minutes) was not changed by OKY-046. A late-phase response (induration at 6 hours) was not observed in any of the OKY-046-treated dogs but was present at 6 hours in 4/5 dogs without OKY-046. Typical mast cells responded similarly in both groups with progressive degranulation during 24 hours. Maximal degranulation of atypical mast cells was delayed to 6 hours with OKY-046, whereas these cells responded completely at 1 hour without OKY-046. The inflammatory response to ragweed followed the same pattern in both groups, but the numbers of each cell type were decreased with OKY-046. With OKY-046, the cutaneous response to histamine was not changed significantly from baseline at 6 hours but was increased (p less than 0.05) at 24 hours, whereas without OKY-046, histamine response was significantly increased at 6 hours (p less than 0.001) and 24 hours (p less than 0.01). We conclude that OKY-046 alters the antigen-induced response of atypical mast cells, the subsequent cellular and clinical late-phase response, and prevents the increase in histamine response.

We studied the cutaneous response to intradermal antigen using clinical, histologic, and physiologic criteria in ragweed-sensitized dogs. The clinical response was measured early (the wheal at 20 minutes) and late (induration at 6 hours). We assessed cutaneous responsiveness to histamine before and 6 hours after injection (intradermally) of ragweed (n = 5, antigen group) and diluent (n = 4, 10% glycerin in 0.9% NaCl, sham group); we measured the wheal in response to histamine (1.0 ng to 0.1 mg, intradermally), constructed a dose-response curve, and interpolated the provocative dose (milligrams) of histamine required to create a wheal 10 mm larger than the response to saline control. Skin biopsy specimens were obtained before and after injection of either ragweed or diluent. Consistent with the human late-phase response, neutrophils and eosinophils were present in the dermis at 1 hour, maximal at 6 hours, and decreased at 24 hours. Mononuclear cells increased significantly at 6 hours and were the predominant cells present at 24 hours after antigen. The late clinical response correlated only with influx of eosinophils (rs = 0.85; p less than 0.005). Histamine responsiveness increased markedly after antigen (p less than 0.0001), did not change after glycerin diluent (sham), and was correlated with the intensity of neutrophil influx at 6 hours (rs = 0.69; p less than 0.05), and to a much greater degree with mononuclear cell influx at 6 hours (rs = 0.85; p less than 0.005).