Immunologic mast cell-mediated responses and histamine release are attenuated by heparin.

Heparin has been shown to act as a competitive inhibitor of inositol 1,4,5-triphosphate (InsP3) receptors in various cell types. Because InsP3 is one of the second messengers involved in stimulus-secretion coupling in mast cells, it is possible that heparin may inhibit mast cell-mediated reactions. Therefore, in allergic sheep, we tested this hypothesis in two mast cell-mediated reactions induced by immunologic and nonimmunologic stimuli: immediate cutaneous reaction (ICR) and acute bronchoconstrictor response (ABR). In 12 sheep allergic to Ascaris suum antigen, the surface area of the skin wheal was determined 20 min after intradermal injection (0.05 ml) of increasing concentrations of specific antigen, compound 48/80, and histamine, without and after pretreatment with heparin (100, 300, or 1,000 U/kg i.v.). Antigen, compound 48/80, and histamine produced concentration-dependent increases in ICR. Heparin "partially" inhibited the ICR to antigen and compound 48/80 in a dose-dependent manner without modifying the ICR to histamine. The heparin preservative benzyl alcohol was ineffective. In 11 additional sheep, specific lung resistance was measured before and after inhalation challenges with antigen, compound 48/80, and histamine without and with aerosol heparin pretreatment (1,000 U/kg). Heparin blocked the antigen- and compound 48/80-induced bronchoconstriction without modifying the airway effects of histamine. In isolated human uterine mast cells, heparin inhibited the anti-immunoglobulin E- but not the calcium ionophore- (A23187) induced histamine release. These data suggest that heparin inhibits the ICR and ABR induced by stimuli that produce immunologic and nonimmunologic mast cell degranulation without attenuating the effects of histamine.(ABSTRACT TRUNCATED AT 250 WORDS)     

A possible role for PAF in allergen-induced late responses: modification by a selective antagonist

We determined whether platelet-activating factor (PAF) plays a role in allergen-induced airway responses by studying the effects of a selective PAF antagonist WEB-2086 on antigen-induced early and late airway responses in allergic sheep. In seven sheep, inhaled Ascaris suum produced significant early (282%) and late (176%) increases in specific lung resistance (sRL). WEB-2086 (1 mg/kg iv) given 20 min before antigen challenge did not affect the early response, but the peak late increase in sRL was only 37% over base line (P less than 0.05 vs. control). To study the mechanism by which PAF contributes to antigen-induced responses, we evaluated the effects of pharmacological probes on PAF-induced bronchoconstriction. Inhaled PAF (dose range 75-700 micrograms) caused reproducible (r = 0.781, P less than 0.05) increases in sRL in eight sheep. The PAF-induced bronchoconstriction was blocked by WEB-2086 (1 mg/kg iv) and by the leukotriene antagonist FPL-55712 (30 mg by aerosol); however, neither the cyclooxygenase blocker indomethacin (2 mg/kg iv) nor the histamine H1-antagonist chlorpheniramine (2 mg/kg iv) blocked the PAF response. WEB-2086, however, did not block bronchoconstriction induced by aerosol leukotriene D4, indicating that PAF acts indirectly through leukotrienes. Finally, we determined whether PAF could induce late airway responses. Inhaled PAF produced an immediate increase in sRL in all seven sheep tested, but late airway responses were observed in only three of the seven sheep.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of early and late pulmonary responses with inhaled leukotriene D4 in allergic sheep

Some allergic sheep respond to inhalation of Ascaris suum antigen with both immediate and late increases in airflow resistance (late response). The mechanism of the late response is unknown but recent evidence suggests that the initial generation of slow-reacting substance of anaphylaxis (SRS-A) immediately after antigen challenge is a necessary pre-requisite for the physiologic expression of this late response. Based on this evidence we hypothesized that airway challenge with leukotriene D4 (LTD4), an active component of SRS-A would produce acute and late airway responses in allergic sheep similar to those observed with antigen. In five allergic sheep with documented early and late pulmonary responses to Ascaris suum antigen, inhalation of leukotriene D4 aerosol (delivered dose (mean +/- SE) 0.55 +/- 0.08 ug) resulted in significant early and late increases in specific lung resistance (SRL). In three allergic sheep which only demonstrated acute responses to antigen, LTD4 aerosol (delivered dose 0.59 +/- 0.09 ug) only produced an acute increase in SRL. In the late responders pretreatment with aerosol cromolyn sodium (1 mg/kg) did not affect the acute response but blunted the late increase in SRL. Pretreatment with aerosol FPL-57231 (1% w/v solution) completely blocked both the acute and late responses. These data support the hypothesis that initial release of LTD4 in the airways of sensitive animals is important for the physiologic expression of the late response.     

Nedocromil sodium in allergen-induced bronchial responses and airway hyperresponsiveness in allergic sheep

We examined the effects of nedocromil sodium, a new drug developed for the treatment of reversible obstructive airway disease, on allergen-induced early and late bronchial responses and the development of airway hyperresponsiveness 24 h after challenge in nine allergic sheep. On occasions greater than 2 wk apart the sheep were treated with 1) placebo aerosol (buffered saline) before and 3 h after antigen challenge, 2) an aerosol of nedocromil sodium (1 mg/kg in 3 ml buffered saline) before antigen challenge and placebo 3 h after challenge, and 3) placebo aerosol before and nedocromil sodium aerosol 3 h after challenge. Early and late bronchial responses were determined by measuring specific lung resistance (sRL) before and periodically after challenge. Airway responsiveness was assessed by determining from dose-response curves the carbachol concentration (in % wt/vol) that increased sRL to 5 cmH2O/s. In the placebo trial, antigen challenge resulted in early and late increases in sRL over a base line of 353 +/- 32 and 131 +/- 17% (SE), respectively. Both early and late increases in sRL were blocked (P less than 0.05) when the sheep were pretreated with nedocromil sodium. When nedocromil was given after the early response, the late response was reduced significantly. Eight of nine sheep developed airway hyperresponsiveness 24 h after antigen challenge. In these eight sheep, carbachol concentration before antigen challenge was 2.6 +/- 0.3%, 24 h later carbachol concentration was significantly lower (1.8 +/- 0.3%). Both nedocromil sodium treatments blocked (P less than 0.05) this antigen-induced airway hyperresponsiveness.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production of early and late pulmonary responses with inhaled leukotriene D4 in allergic sheep

Some allergic sheep respond to inhalation of antigen with both immediate and late increases in airflow resistance (late response). The mechanism of the late response is unknown but recent evidence suggests that the initial generation of slow-reacting substance of anaphylaxis (SRS-A) immediately after antigen challenge is a necessary pre-requisite for the physiologic expression of this late response. Based on this evidence we hypothesized that airway challenge with leukotriene D₄ (LTD₄), an active component of SRS-A would produce acute and late airway responses in allergenic sheep similar to those observed with antigen. In five allergic sheep with documented early and late pulmonary responses to antigen, inhalation of leukotriene D₄ aerosol (delivered dose {mean ±SE} 0.55±0.08 ug) resulted in significant early and late increases in specific lung resistance (SR_L). In three allergic sheep which only demonstrated acute responses to antigen, LTD₄ aerosol (delivered dose 0.59±0.09ug) only produced an acute increase in SR_L. In the late responders pretreatment with aerosol cromolyn sodium (1 mg/kg) did not affect the acute response but blunted the late increase in SR_L. Pretreatment with aerosol FPL-57231 (1% w/v solution) completely blocked both the acute and late responses. These data support the hypothesis that initial release of LTD₄ in the airways of sensitive animals is important for the physiologic expression of the late response.     

Modification of bronchial blood flow during allergic airway responses

Ascaris suum antigen effects on mean airflow resistance (RL) and bronchial arterial blood flow (Qbr) were studied in allergic anesthetized sheep with documented airway responses. Qbr was measured with electromagnetic flow probes, and supplemental O2 prevented antigen-induced hypoxemia. Aerosol challenge with this specific antigen increased RL and Qbr significantly. Cromolyn sodium aerosol pretreatment prevented antigen-induced increases in RL but not in Qbr. Intravenous cromolyn, however, prevented increases in Qbr and RL, suggesting a role for mast cell degranulation in both bronchomotor and bronchovascular responses to antigen. Antigen-induced increases in Qbr were not solely attributable to histamine release. Indomethacin pretreatment attenuated the antigen-induced increase in Qbr, thus suggesting that vasodilator cyclooxygenase products contribute to the vascular response. Antigen challenge significantly decreased Qbr after indomethacin and metiamide pretreatment, which suggests that vasoconstrictor substances released after antigen exposure also modulate Qbr; however, released vasodilators overshadow vasoconstrictor effects. Thus antigen challenge affects Qbr by locally releasing histamine and vasodilator prostaglandins as well as vasoconstrictor substances. These effects were independent of antigen-induced changes in systemic and pulmonary hemodynamics.     

Indomethacin and FPL-57231 inhibit antigen-induced airway hyperresponsiveness in sheep

We compared the development of antigen-induced airway hyperresponsiveness (AHR) 24 h after challenge with Ascaris suum antigen in allergic sheep with acute (n = 7) and with dual (n = 7) airway responses and then attempted to modify this AHR. Cholinergic airway responsiveness was determined by measuring the carbachol dose required to increase specific lung resistance (sRL) 150% (i.e., PC150). Subsequently the sheep were challenged with antigen and sRL was measured at predetermined times to document the presence or absence of a late response. PC150 was redetermined 24 h later followed by bronchoalveolar lavage (BAL) to assess inflammation. Only dual responders developed AHR (PC150 decreased, P less than 0.05). There were no significant differences in BAL between the two groups. Six dual responders were then, on separate occasions (greater than or equal to 3 wk), pretreated with placebo, indomethacin (2 mg/kg iv), or a leukotriene antagonist, FPL-57231 (30 mg inhaled). Neither agent significantly affected the acute response to antigen. Only FPL pretreatment blocked the late response, but both agents blocked the antigen-induced AHR 24 h later. BAL at 24 h showed no significant differences. These results indicate that only dual responders develop AHR 24 h after antigen challenge. This AHR appears independent of the late increase in sRL or the severity of pulmonary inflammation. AHR appears to be sensitive to agents that interfere with the early release or actions of cyclooxygenase and lipoxygenase metabolites in dual responders.     

Immunologic and nonimmunologic responsiveness in ragweed-sensitized dogs

The relationship between airway responsiveness to inhaled antigen and histamine, immunologic release of lung histamine, immunologic responsiveness of skin, and specific immunoglobulin E (IgE) antibodies were examined in 11 inbred allergic dogs immunized with extracts of ragweed and grass and 5 nonimmunized control dogs from the same colony. Airway responsiveness to antigen and histamine was characterized by the doses that increased the airflow resistance of the total respiratory system to twice the control values (ED200). Highly significant correlations were found between airway responsiveness and cutaneous responsiveness to antigen and other immunologic characteristics (e.g., IgE and histamine released from lung by inhaled antigen) in all dogs. In ragweed-sensitized dogs, there was an inverse correlation between immunologic responsiveness (reflected by the cutaneous response to antigen and histamine released from lung by inhaled antigen) and nonimmunologic responsiveness of airways (histamine ED200: r = 0.73, P less than 0.05 and r = 0.75, P less than 0.01, respectively). Antigen ED200 was also correlated with histamine release from lung after antigen inhalation (r = 0.74; P less than 0.01). We conclude that airway reactions to inhaled antigen in allergic dogs are dependent not only on immunologic factors but also on the degree of nonimmunologic airway responsiveness to histamine and that these factors are correlated inversely.     

Effect of disodium cromoglycate on mast cell-mediated immediate-type allergic reactions

We investigated the effect of disodium cromoglycate (DSCG) on mast cell-mediated immediate-type hypersensitivity. DSCG inhibited systemic allergic reaction induced by compound 48/80 dose-dependently. Passive cutaneous anaphylaxis was inhibited by 71.6% by oral administration of DSCG (1 g/kg). When DSCG was pretreated at concentration rang from 0.01-1000 g/kg, the serum histamine levels were reduced in a dose dependent manner. DSCG also significantly inhibited histamine release from rat peritoneal mast cell (RPMC) by compound 48/80. We confirmed that DSCG inhibited compound 48/80-induced degranulation of RPMC by alcian blue/nuclear fast red staining. In addition, DSCG showed a significant inhibitory effect on anti-dinitrophenyl IgE-mediated tumor necrosis factor-alpha production. These results indicate that DSCG inhibits mast cell-mediated immediate-type allergic reaction.     

Effects of leukotriene D4 on mucociliary and respiratory function in allergic and nonallergic sheep

We determined the effect of aerosol challenge with leukotriene D4 (LTD4) on specific lung resistance (sRL) and tracheal mucous velocity (TMV) in conscious sheep with (allergic) and without (nonallergic) Ascaris suum hypersensitivity. In allergic sheep LTD4 in concentrations of 50, 100, and 150 micrograms/ml produced dose-dependent increases in mean sRL by 44 (P = NS), 154 (P less than 0.05), and 233% (P less than 0.05), respectively. The increase in sRL produced by 150 micrograms/ml LTD4 was prevented by FPL 55712, an antagonist of slow-reacting substance of anaphylaxis. In nonallergic sheep 150 micrograms/ml LTD4 failed to elicit a significant change in sRL. In contrast to the changes in airway mechanics, concentrations of LTD4 as low as 25 micrograms/ml produced significant decreases in TMV in allergic sheep. The maximum decrease in TMV at this dose occurred 2 h after challenge; with larger doses of LTD4 (100 and 150 micrograms/ml) the maximum effect was observed 3 h after challenge. Furthermore, 150 micrograms/ml LTD4 reduced TMV in nonallergic sheep (mean decrease 43%, P less than 0.05). FPL 55712 only had a minor effect on the LTD4-induced decreases in TMV. We conclude that allergic sheep exhibit greater airway responsiveness to inhaled LTD4 than nonallergic sheep but that this difference is not evident for the concomitant changes in mucociliary transport. This suggests that the allergic state is associated with an increased responsiveness to LTD4 in tissues controlling airway caliber but not in those contributing to mucociliary function.     

Separation of late bronchial responses from airway hyperresponsiveness in allergic sheep

Allergic sheep with antigen-induced early and late responses were used to determine whether airway hyperresponsiveness (AHR) to carbachol is present during the late response and whether blocking the late response with the leukotriene D4 (LTD4) antagonist MK-571 also blocks this AHR. To do this, we first showed that MK-571 blocked the antigen-induced late response, and then, in a separate study, we determined the effect of MK-571 treatment on airway responsiveness 6 h after antigen challenge (at the start of the late response). MK-571 (5 mg, by metered dose inhaler) given 30 min before and 4 h after Ascaris suum challenge had no effect on the acute response to antigen but blocked (P less than 0.05) the late response compared with placebo (n = 7). In the second study (n = 6), the antigen-induced acute increases in mean specific lung resistance (sRL) were also similar in the placebo (249%) and drug trials (247%). By 6 h postchallenge, however, mean sRL in the placebo trial began to increase (54%, P less than 0.05), whereas in the drug trial mean sRL was baseline. Nevertheless, AHR was apparent in both trials as indicated by a mean twofold leftward shift in the dose-response curves to inhaled carbachol (P less than 0.05 vs. prechallenge). Bronchoalveolar lavage at 6 h showed that MK-571 did not prevent the inflammatory cell influx into the lung. These observations suggest that although LTD4 may be a mediator of the late response in sheep, it is not a primary mediator affecting cholinergic AHR during this period.     

Hyperoxia prevents hypoxia-induced bronchial hyperreactivity via a cyclooxygenase-independent mechanism

We tested the hypothesis that prior exposure to alveolar hyperoxia prevents the hypoxia-induced enhancement of bronchial reactivity, possibly via a cyclooxygenase-dependent mechanism. In 15 sheep, specific lung resistance (sRL) was measured before and after 30 min of exposure to either air or a hypoxic gas mixture (13% O2). The sheep then inhaled 50 breaths of aerosolized 5% histamine solution (n = 9) or 10 breaths of 2.5% carbachol solution (n = 9), and measurements of sRL were repeated. On subsequent days the above protocols were repeated after a 30-min exposure to hyperoxia (O2 greater than or equal to 95%), without or after pretreatment with indomethacin (2 mg/kg). After air-sham exposure, carbachol and histamine increased mean sRL to 370 +/- 40 (SE) and 309 +/- 65% of baseline, respectively. Exposure to the hypoxic gas mixture had no effect on baseline sRL but enhanced the airway responsiveness to carbachol and histamine; mean sRL increased to 740 +/- 104 and 544 +/- 76% of baseline, respectively (P less than 0.05). Prior 30-min exposure to hyperoxia prevented the hypoxia-induced enhancement of bronchial reactivity to carbachol (sRL = 416 +/- 66% of baseline) and histamine (sRL = 292 +/- 41% of baseline) without affecting the airway responsiveness to these agents after air. Pretreatment with indomethacin did not reverse the protective effects of hyperoxia or the hypoxia-induced enhancement of bronchial reactivity. We conclude that 1) prior exposure to alveolar hyperoxia prevents the hypoxia-induced enhancement of bronchial reactivity and 2) neither the protective effects of hyperoxia nor the hypoxia-induced enhancement of bronchial reactivity is mediated via a cyclooxygenase-dependent mechanism.     

Monocyte chemoattractant protein-1 mediates cockroach allergen-induced bronchial hyperreactivity in normal but not CCR2-/- mice: the role of mast cells.

Bronchial eosinophil and mononuclear cell infiltrates are a hallmark of the asthmatic lung and are associated with the induction of reversible airway hyperreactivity. In these studies, we have found that monocyte chemotactic protein-1 (MCP-1), a CC (beta) chemokine, mediates airway hyperreactivity in normal and allergic mice. Using a murine model of cockroach Ag-induced allergic airway inflammation, we have demonstrated that anti-MCP-1 Abs inhibit changes in airway resistance and attenuate histamine release into the bronchoalveolar lavage, suggesting a role for MCP-1 in mast cell degranulation. In normal mice, instillation of MCP-1 induced prolonged airway hyperreactivity and histamine release. In addition, MCP-1 directly induced pulmonary mast cell degranulation in vitro. These latter effects would appear to be selective because no changes were observed when macrophage-inflammatory protein-1alpha, eotaxin, or MCP-3 were instilled into the airways of normal mice or when mast cells were treated in vitro. Airway hyperreactivity was mediated by MCP-1 through CCR2 because allergen-induced as well as direct MCP-1 instilled-induced changes in airway hyperreactivity were significantly attenuated in CCR2 -/- mice. The neutralization of MCP-1 in allergic animals and instillation of MCP-1 in normal animals was related to leukotriene C4 levels in the bronchoalveolar lavage and was directly induced in pulmonary mast cells by MCP-1. Thus, these data identify MCP-1 and CCR2 as potentially important therapeutic targets for the treatment of hyperreactive airway disease.     

The Tec family kinase, IL-2-inducible T cell kinase, differentially controls mast cell responses

The Tec family tyrosine kinase, IL-2-inducible T cell kinase (Itk), is expressed in T cells and mast cells. Mice lacking Itk exhibit impaired Th2 cytokine secretion; however, they have increased circulating serum IgE, but exhibit few immunological symptoms of allergic airway responses. We have examined the role of Itk in mast cell function and FcepsilonRI signaling. We report in this study that Itk null mice have reduced allergen/IgE-induced histamine release, as well as early airway hyperresponsiveness in vivo. This is due to the increased levels of IgE in the serum of these mice, because the transfer of Itk null bone marrow-derived cultured mast cells into mast cell-deficient W/W(v) animals is able to fully rescue histamine release in the W/W(v) mice. Further analysis of Itk null bone marrow-derived cultured mast cells in vitro revealed that whereas they have normal degranulation responses, they secrete elevated levels of cytokines, including IL-13 and TNF-alpha, particularly in response to unliganded IgE. Analysis of biochemical events downstream of the FcepsilonRI revealed little difference in overall tyrosine phosphorylation of specific substrates or calcium responses; however, these cells express elevated levels of NFAT, which was largely nuclear. Our results suggest that the reduced mast cell response in vivo in Itk null mice is due to elevated levels of IgE in these mice. Our results also suggest that Itk differentially modulates mast cell degranulation and cytokine production in part by regulating expression and activation of NFAT proteins in these cells.     

Comparison of Airway Responses in Sheep of Different Age in Precision-Cut Lung Slices (PCLS)

Background

Animal models should display important characteristics of the human disease. Sheep have been considered particularly useful to study allergic airway responses to common natural antigens causing human asthma. A rationale of this study was to establish a model of ovine precision-cut lung slices (PCLS) for the in vitro measurement of airway responses in newborn and adult animals. We hypothesized that differences in airway reactivity in sheep are present at different ages.

Methods

Lambs were delivered spontaneously at term (147d) and adult sheep lived till 18 months. Viability of PCLS was confirmed by the MTT-test. To study airway provocations cumulative concentration-response curves were performed with different allergic response mediators and biogenic amines. In addition, electric field stimulation, passive sensitization with house dust mite (HDM) and mast cells staining were evaluated.

Results

PCLS from sheep were viable for at least three days. PCLS of newborn and adult sheep responded equally strong to methacholine and endothelin-1. The responses to serotonin, leukotriene D₄ and U46619 differed with age. No airway contraction was evoked by histamine, except after cimetidine pretreatment. In response to EFS, airways in PCLS from adult and newborn sheep strongly contracted and these contractions were atropine sensitive. Passive sensitization with HDM evoked a weak early allergic response in PCLS from adult and newborn sheep, which notably was prolonged in airways from adult sheep. Only few mast cells were found in the lungs of non-sensitized sheep at both ages.

Conclusion

PCLS from sheep lungs represent a useful tool to study pharmacological airway responses for at least three days. Sheep seem well suited to study mechanisms of cholinergic airway contraction. The notable differences between newborn and adult sheep demonstrate the importance of age in such studies.     

Effect of age on lung mechanics and airway reactivity in lambs

We studied airway reactivity (AR) to aerosolized histamine, carbachol, and citric acid in lambs 1 mo of age to adulthood. Awake lambs were intubated and studied in a plethysmograph that measured dynamic compliance (Cdyn), resistance of the lung (RL), and functional residual capacity (FRC). Pleural pressure was measured using a Silastic balloon in the pleural space, and airway opening pressure (Pao) was measured using a catheter placed 1-2 cm distal to the nasotracheal tube. At the ages of 1, 3, 5, and 7 mo and adulthood, measurements of Cdyn, RL, and FRC were obtained in 46 sheep (22 males, 24 females). AR to carbachol, histamine, and citric acid was measured in each sheep in randomized order on three separate days by giving increasing concentrations of the drug in a noncumulative fashion. The dose that would have caused a 35% reduction in Cdyn (ED65Cdyn), a doubling of RL (ED200RL), or a 50% increase in FRC (ED150FRC) was calculated. In both males and females, base-line Cdyn increased (r = 0.81, P less than 0.01) with age, as did FRC (r = 0.87, P less than 0.01). There was no significant change in RL in either sex with age or in the group as a whole. There was a significant increase in AR to both histamine and carbachol with increasing age as measured by a decrease in ED65Cdyn (P less than 0.01 and P less than 0.05, respectively) with age. There was no significant change in AR with age as measured by RL or FRC for any of the three bronchoconstrictors tested.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibitory Effect of Eriodictyol on IgE/Ag-Induced Type I Hypersensitivity

Mast cells are the principal effector cells involved in the allergic response, through the release of histamine. We investigated the effect of eriodictyol, derived from the painted maple and yerba santa, on mast cell degranulation and on an allergic response in an animal model. We also investigated its effect on the expression of the ceramide kinase (CERK) involved in calcium-dependent degranulation, and on ceramide activation by multiple cytokines. Eriodictyol suppressed the release of beta-hexosaminidase, a marker of degranulation, and the expression of interleukin (IL)-4 mRNA. It inhibited the expression of CERK mRNA, reduced the ceramide concentration in antigen-stimulated mast cells, and suppressed the passive cutaneous anaphylaxis (PCA) reaction in mice in a dose-dependent manner. These results suggest that eriodictyol can inhibit mast cell degranulation through inhibition of ceramide kinase, and that it might potentially serve as an anti-allergic agent.     

Attenuation of platelet-activating factor-induced airway responses with methylprednisolone succinate in sheep.

Intratracheal instillation of platelet-activating factor (PAF) in sheep produces bronchoconstriction and airway hyperresponsiveness (AHR) by a two-stage process that involves the initial stimulation of PAF receptors followed by the secondary generation of proinflammatory mediators. Because the biological effects of PAF may be mediated by these proinflammatory metabolites, it is possible that a steroidal anti-inflammatory agent would modify the airway responses of PAF. We measured specific lung resistance (sRL) in sheep (n = 11) before, immediately after, and serially for up to 2 h after intratracheal instillation of PAF (30 micrograms/kg). Airway responsiveness was measured 2 h post-PAF when sRL had returned to baseline and was expressed as the cumulative provocating dose of carbachol that increased sRL to 4 l.cmH2O.l-1.s (PD4). PD4 was determined on a control day and on different experiment days without or after treatment with intravenous methylprednisolone (MPS; 1 mg/kg) administered 3 h before (n = 6), 20 min before PAF (n = 7), or 20 min after PAF challenge (n = 7). PAF increased sRL by 222 +/- 44% (SE) above baseline and decreased PD4 of carbachol by 44 +/- 5% (P less than 0.05). Pretreatment (both 3 h and 20 min) with MPS attenuated the PAF-induced increases in sRL, whereas its administration 20 min post-PAF had no effect. Irrespective of the effects on sRL, MPS administration inhibited the PAF-induced AHR.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibitory effect of eriodictyol on IgE/Ag-induced type i hypersensitivity

Mast cells are the principal effector cells involved in the allergic response, through the release of histamine. We investigated the effect of eriodictyol, derived from the painted maple and yerba santa, on mast cell degranulation and on an allergic response in an animal model. We also investigated its effect on the expression of the ceramide kinase (CERK) involved in calcium-dependent degranulation, and on ceramide activation by multiple cytokines. Eriodictyol suppressed the release of beta-hexosaminidase, a marker of degranulation, and the expression of interleukin (IL)-4 mRNA. It inhibited the expression of CERK mRNA, reduced the ceramide concentration in antigen-stimulated mast cells, and suppressed the passive cutaneous anaphylaxis (PCA) reaction in mice in a dose-dependent manner. These results suggest that eriodictyol can inhibit mast cell degranulation through inhibition of ceramide kinase, and that it might potentially serve as an anti-allergic agent.     

Inhaled porcine pancreatic elastase causes bronchoconstriction via a bradykinin-mediated mechanism.

Neutrophil elastase has been linked to inflammatory lung diseases such as chronic obstructive pulmonary disease, adult respiratory distress syndrome, emphysema, and cystic fibrosis. In guinea pigs, aerosol challenge with human neutrophil elastase causes bronchoconstriction, but the mechanism by which this occurs is not completely understood. Our laboratory previously showed that human neutrophil elastase releases tissue kallikrein (TK) from cultured tracheal gland cells. TK has been identified as the major kininogenase of the airway and cleaves both high- and low-molecular weight kininogen to yield lysyl-bradykinin. Because inhaled bradykinin causes bronchoconstriction and airway hyperresponsiveness in asthmatic patients and allergic sheep, we hypothesized that elastase-induced bronchoconstriction could be mediated by bradykinin. To test this hypothesis, we measured lung resistance (RL) in sheep before and after inhalation of porcine pancreatic elastase (PPE) alone and after pretreatment with a bradykinin B(2) antagonist (NPC-567), the specific human elastase inhibitor ICI 200,355, the histamine H(1)-antagonist diphenhydramine hydrochloride, the cysteinyl leukotriene 1 receptor antagonist montelukast, or the cyclooxygenase inhibitor indomethacin. Inhaled PPE (125-1,000 microg) caused a dose-dependent increase in RL. Aerosol challenge with a single 500 microg dose of PPE increased RL by 132 +/- 8% over baseline. This response was blocked by pretreatment with NPC-567 and ICI-200,355 (n = 6; P < 0.001), whereas treatment with diphenhydramine hydrochloride, montelukast, or indomethacin failed to block the PPE-induced bronchoconstriction. Consistent with pharmacological data, TK activity in bronchial lavage fluid increased 134 +/- 57% over baseline (n = 5; P < 0.02). We conclude that, in sheep, PPE-induced bronchoconstriction is in part mediated by the generation of bradykinin. Our findings suggest that elastase-kinin interactions may contribute to changes in bronchial tone during inflammatory diseases of the airways.