Antagonism of airway reactivity induced by ovalbumin antigen in guinea pigs by 5-amino-4-imidazolecarboxamide riboside Dale I?. Bergren, PhD,* Robert G. Townley, MD,** Diana L. Marquardt, MD,**** Harry E. Gruber, MD,**** Virginia A. Bergren, BS* Omaha, Neb., and San Diego,

and

Calif.

The effect of 5-amino-4-imidazolecarboxamide riboside (AICA riboside), a modulator of purine metabolism, was studied on antigen-induced bronchospasm in ovalbumin (OA)-sensitized guinea pigs. In separate experiments, sodium cromoglycate (SCG) and terbutaline were used to compare their effectiveness with that of AICA riboside (wtlvol). AICA riboside and SCG were administered as an aerosol daily for a minimum of 2 weeks before OA aerosol challenge. Terbutaline, as an aerosol, was administered once 5 minutes before OA challenge. Airway reactivity was determined through the use of a whole-body plethysmograph by monitoring specific airway resistance (SR,,). OA aerosol challenge of 0.05%, O.l%, and 0.25% (wtlvol), administered for a period of 1 minute, increased SR,,. Each of the three agents attenuated the effect of OA on SR.,,, although terbutaline demonstrated more consistency and potency as compared to either AICA riboside or SCG. However, at moderate degrees of OA challenge, AICA riboside appeared to be as effective as either agent. Although the mechanism of action of AICA riboside remains uncertain, it may have therapeutic benefit in the treatment of asthma or allergic diseases. (J ALLERGYCLIN IMMUNOL1991;88:604-12) Key words: Guinea pigs, ovalbumin antigen, AICA riboside, spec$c airway resistance, terbutaline, sodium cromoglycate, whole-body plethysmograph

Purine metabolismmay have appreciableeffects on airway tone and pulmonary inflammation during allergic reactions or episodesof asthma.Adenosine has been demonstratedto induce bronchospasmwhen it is inhaled by individuals with asthmabut not by individuals with normal airway reactivity. ’ Severalstudies tend to underscorethe role of preformed mast cell mediatorreleasein the mechanismof action of inhaled

From the Creighton University School of Medicine, *Department of Biomedical Sciences,Division of Physiology, and **Department of Medicine, Division of Allergy, Omaha,Neb.; University of California-San Diego, University of California Medical Center, ***Division of Allergic and Immunologic Diseases,School of Medicine, San Diego, and ****Gensia Pharmaceuticals,Inc., Diego, Calif. Supportedby Gensia Pharmaceuticals,Inc., San Diego, Calif. Received for publication April 24, 1990. Revised March 12, 1991. Accepted for publication May 22, 1991. Reprint requests: Dale R. Bergren, PhD, Associate Professorof Physiology, Creighton University School of Medicine, Department of Biomedical Sciences,Division of Physiology, 2500 California St., Omaha, NE 68178. l/1/31234

604

Abbreviations used AICA riboside: 5-Amino-4-imidazolecarboxamide OA: Ovalbumin SCG: Sodium cromoglycate SR,,: Specific airway resistance Ag: Antigen ANOVA: Analysis of variance AUC: Area under the curve

adenosineas a bronchoconstrictor.2AFurthermore, exogenousadenosinehasbeen reported to potentiate the release of granule-associatedmediators from stimulated mastcells obtained from rat peritoneum,* mouse bone marrow,5 guinea pig lung,6 or human lung,7-9 suggesting a role for adenosine in augmenting the allergic response. Studiesof the effect of adenosineon airway smooth muscletone report varying results. In somelaboratory animals, such as the guinea pig, adenosinerelaxes tracheal smooth muscle,‘@12 although this effect appears to depend on how the tracheal muscle is cut.‘* In other models, adenosinecausescontraction of air-

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way smooth muscle.4*‘**I3 In addition to its possible direct eff;ectson airway smooth muscle, adenosine may also causereflex bronchoconstriction and tachypnea. It increasesthe activity of rapidly adapting receptors and C fiber endings.4,” Adenosine also enhancesthe contractile responseto transmural stimulation of isolated bronchial smoothmuscle,16possibly by cholinergic neurotransmitterrelease,since this potentiation is blocked by tetrodotoxin. Modulation of purine metabolism pharmacologically may be of value in the treatment of asthmaand allergic diseases. AICA riboside is a purine analog that may itself act as a substratein purine metabolism. Mouse bone marrow-derived mast cells cultured in the presenceof micromolar concentrations of AICA riboside for a few days exhibit a marked decreasein the releaseof preformedor newly generatedmediators after challenge with specific Ag or the calcium ionophore, A23 187.5 Although AICA riboside has been demonstratedto produce adenosinein ischemic myocardial tissue, the mast cell effects appear not to be related to adenosine.l7The precisetype of modulation of purine metabolism by AICA riboside that results in mast cell stabilization is not known. AICA riboside is currently in human clinical trials for cardioprotective applications. Because AICA riboside appearsto “stabilize” cultured mast cells without altering cell viability or mediator concentrations, we studied whether AICA riboside would demonstrateantibronchoconstrictor effects in sensitized guinea pigs challenged with aerosolsof a specific Ag.

ential pressure transducers (model DP 45-14, Validyne, Northridge, Calif.) sensedchangesin nasaland thoracic box pressures.The signals from thesetransducerswere relayed to a noninvasive respiratory analyzer (Buxco Electronics, Inc.). The respiratory analyzer was connectedto an on-line person computer with scrolling monitor and data-printing capabilities. Parametersmonitored included thoracic box pressure,nasalbox pressure,SR,, tidal volume, respiratory rate, and minute volume. BaselevelSR,, was determined after the guinea pig was accustomedto being placedwithin the plethysmographwhen the parametersbeing monitored had stabilized. SR,, was again measured1 minute after OA aerosol challenge for a total of 10 additional minutes. The guinea pigs were then removed form the whole-body plethysmograph. OA administration Challenge of OA aerosol to the respiratory system was performed while the guinea pigs were being housed in the whole-body plethysmograph. Three different dosesof OA were chosen for use in this study from preliminary experiments with various doses (O.OS%,O.l%, and 0.25% dissolved in distilled water). The nature of the OA challenge was a 50 to 60-secondaerosol exposure generatedby ultrasonic nebulizers (model 65, DeVilbiss Co., Sommerset, Pa.) producing particle diametersof 0.5 to 5 pmol/L into the nasal chamberof the whole-body plethysmograph.Immediately after the aerosol challenge, the air in the nasal chamber was flushed with room air generatedfrom an air pump with an adjustable air valve. No guinea pigs with obvious airway reactivity to the OA challenge received another OA challenge within a 4-day period of time to avoid possible OA tachyphylaxis. Drug groups

The guinea pigs were separatedinto four drug-treatment groups after the lbday inoculation period to OA. The four drug-treatment groups were no pretreatmentand pretreatHartley guinea pigs (300 to 900 gm, Sasco, Omaha, ment with AICA riboside, SCG, or terbutaline. For AICA Neb.) were usedfor this study.Theguineapigswerehoused riboside and SCG,thedrugadministration consistedof aeroin stainlesssteelmeshcages,watered,andfedwith standard sol exposure of 15 minutes in duration generatedby two guineapig chow ad libitum. Theguineapigsweresensitized ultrasonic nebulizers for a minimum of 14 continuous days to OA (Sigma Chemical Co., St. Louis, MO.) according to in an aerosol chamber (14 cm deep, 32 cm wide, and 51 the methodsof Andersson.‘* Each guinea pig was injected cm long). The resulting aerosol generatedduring the drug intraperitoneally with 0.5 ml of 0.9% saline solution contreatmentsaturatedthe air in the chamberso that visibility taining 10 pg of OA and 100 mg of AI(O Ten days into the chamber was obscured. Doses of AICA riboside later., the guinea pigs were injected intraperitoneally with and SCG used were 0. l%, l%, and 5% (wt/vol in 10 p,gof CIA in 0.5 ml isotonic saline. Two weeks after the distilled water). The 5% solution of AICA riboside is near initial injection, sensitization to OA was demonstrable.The the highest concentration we could achieve. This dose of sensitization to OA lasts for several months. This method AICA riboside was chosenalong with 1%and 0.1% asdoses of OA sensitization producesbronchial reactivity mediated that would be expected to demonstrateantagonismof Ag primarily via IgE-like antibodies. challengeif suchpropertiesof the agentexists. Similar doses of SCG and terbutaline were chosen to match the concenMeasurement of SR., in conscious trations of AICA riboside (wt/vol) that were used in this guinea pigs study. No physiologic or behavioral abnormalities were observed in either group of daily treatment. In other studies Pulmonary function was measuredin consciousand nonnot reported here, intravenous injeciion of AICA riboside sedatedgdnea pigs by the use of a whole-body plethyshad no adverseeffects and appearsto attenuatethe cardiomograph(model P, Buxco Electronics Inc., Sharon, Conn.) vascular effects of intravenous Ag challenge. patterned after that developed by Pennock et a1.19DifferMATERIAL Animal6

AND METHODS

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TIME (minutes) FIG. 1. Percent increase above baselevel SR, in sensitized guinea pigs in response to l-minute OA aerosol challenge versus time; 0.05% (0) (N = 17). 0.1% (0) (N = 19), and 0.25% (A) (N = 19). All points for each dose are increased over baselevel SR., (p < 0.05) except minute 1 at 0.1% (+).

Terbutaline was administered as an aerosol (0.1% and 1%) to the nasal chamber for a l-minute period 5 minutes before the Ag challenge while the guinea pig was being housed in the plethsmograph. Terbutaline solution of 5% was not used becausethe demonstrableeffectivenessof the 1% solution, evenat the highest concentrationof OA aerosol used. Terbutaline was also not testedagainstthe lowest dose of OA aerosol challenge (0.05%).

Data analysis A data analyzer (Data Logger, Buxco Electronics, Inc.) in line with the respiratory analyzer was programmedto averageand then print the values for each parametermonitored at j-second intervals. After OA challenge, the pulmonary functions were monitored for 10 minutes. The SR., at each minute was checkedwith those data ? 10 seconds from the actual minute value. If the SR, value was aberrant because of animal movement within the plethysmograph, for example, then an SR, measurementwas substituted from either side of the actual minute value with that which was without such interference. Percentchangesin SR, from the baselevelSR, was used for statistical analysis becauseof variations betweenguinea pigs in baselevelSR,. This procedureis routinely used in analysis of SR,.*’ SR,, was analyzed within each of the drug groups (against time and doses) and between groups (a similar dose against time only) by meansof a one-way ANOVA. The derivation of the AUC was similar to that describedby Richards et a12’ and was calculated for each experiment by subtracting the baselevelSR, from the SR,, at each minute after the Ag challenge. Statistical analysis was again by means of one-way ANOVA. Should the ANOVA reject the nulrhypothesis, the Newman-Keuls’ test

was usedto determinestatistical separationbetweengroups. The lowest level of significance used for the one-tailed test was equal to 0.05. These statistical methodsare described by Zar.22

RESULTS OA challenge by aerosol to the airways of sensitized guinea pigs increased SR,, in a time- and dosedependent manner compared to the vehicle challenge (Fig. 1). SR,, was increased for all doses above baselevel except for minute 1 at 0.1% aerosol challenge of OA. Although guinea pig airway reactivity exhibited considerable variability, when reactivity was compared between subjects, individual reactivity to repeated OA challenge on different days was reasonably consistent. OA challenge increased the AUC of SR,, for all three doses versus that of the vehicle water (Fig. 2). The AUC was greater after 0.25% OA challenge compared to 0.05% OA challenge. Daily 5% AICA riboside treatment by aerosol inhalation reduced the effect of 0.05% OA aerosol challenge on SR,, (Figs. 3 and 4). Temporally, statistical separation between the nontreated and 5% AICA riboside-treated guinea pigs occurred at minutes 4 to 10 (Fig. 3), and the AUC difference had reached statistical significance (p < 0.05, Fig. 4). Although 1% AICA riboside treatment decreased the AUC approximately 50% versus the nontreated group, interanimal variation did not allow statistical separation (Fig. 4). Treatment with SCG also demonstrated a trend to attenuate the increase in SR,, in response to 0.05% OA

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* -I-

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FIG. 2. Percent increase in the AUC above baselevel aerosol challenge and to its vehicle water in sensitized **p < 0.05 verses OA 0.05%.

OA 0.25%

OA 0.1%

OA 0.05%

SR, for 10 minutes after a l-minute OA guinea pigs; *p < 0.05 versus the vehicle;

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I

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* I

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TIME (minutes) FIG. 3. Antagonism of OA challenge by AICA riboside aerosol pretreatment. Percent change in SR, versus time; *p < 0.05 versus nontreated group. OA 0.05% versus no pretreatment (0) (N = 17). AICA riboside aerosol pretreatment of 0.1% (El) (N = 81, 1.0% (~1 (N = 6) and 5.0% I+) (N = 6).

challenge but did not attain statistical separationwhen the AUC was analyzed (Fig. 4). However, versus time, 5% SCG attenuated that effect of 0.05% OA aerosolchallenge on SR, during minutes4 to 10 after aerosol challenge. OA aerosol challenge of 0.1% increasedSR,, from minutes 2 to minutes 10 in guinea pigs receiving no pretreatment. Only terbutaline statistically attenuated the effect of OA on SR,, at this dosage(Fig. 5). Both dosesof te:rbutalineattenuatedthe effect of 0.1% OA

aerosol challenge from minutes 1 through 10. Both AICA riboside and SCG had heterogenouseffects. Heterogeneousaction hasbeenreportedelsewherefor SCG.13, 23, 24 OA aerosolchallengeof 0.25% increasedSR,, from minutes 1 through 10 in the untreatedgroup of guinea pigs (Fig. 1). The AUC of the 0.25% OA challenge versus the vehicle and 0.1% OA aerosol challenge was also elevated (Fig. 2). AICA riboside of 0.1% and 1% reducedthe effect of 0.25% OA aerosolchal-

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I

gf;vetreatment OSCG FIG. 4. Percent increase in the AUC above the baselevel SR., for 10 minutes after a l-minute OA aerosol challenge of 0.05% in sensitized guinea pigs. Hatched column represents no pretreatment (N = 17). shaded column represents AICA riboside pretreatment (N = 8 for O.l%, 6 for 1.0%. and 6 for 5-O%), and open columns represents SCG pretreatment (N = 5 for O.l%, 6 for 1.0%. and 6 for 5.0%). See methods for details of pretreatment; *p < 0.05 versus no pretreatment.

0.1% ShIretreatment

0.1%

1 .O%

5.0%

OSCG CXITERB FIG. 5. Percent increase in the AUC above the baselevel SR,, for 10 minutes after a l-minute OA aerosol challenge of 0.1% in sensitized guinea pigs. Hatched columns represents no pretreatment (N = 19), shaded columns represents AICA riboside pretreatment (N = 8 for O.l%, 6 for 1.0%. and 6 for 5.0%). open columns represents SCG pretreatment (N = 5 for 0.1%. 6 for l.O%, and 6 for 5.0%), cross-hatched columns represents terbutaline pretreatment (N = 10 for 0.1% and 10 for 1.0%). See methods for details of pretreatment; *p < 0.05 versus no pretreatment.

lenge on SR,, (Fig. 6, p < 0.05). Separationbetween 0.1% and 1% AICA riboside pretreatment and OA challenge with no pretreatmentoccurred from minute 2 through minute 10 and from minute 3 through 10,

respectively. In guinea pigs treated with 5% AICA riboside, SR,, was lower at minutes 3 and 4 versus SR,, in guinea pigs that received no pretreatment. Terbutaline solutions of 0.1% and 1% also afforded

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0 SCG ECITERB FIG. 6. Percent increase in the AUC above the baselevel SR, for 10 minutes after a l-minute OA aerosol challenge of 0.25% in sensitized guinea pigs. Hatched columns represents no pretreatment (N = 191, shaded columns represents AICA riboside pretreatment (N = 8 for O.l%, 6 for l.O%, and 6 for 5.0%), open columns represents SCG pretreatment (N = 5 for O.l%, 6 for 1.0%. and 6 for 5.0%) cross-hatched columns represent terbutaline pretreatment (N = 10 for 0.1% and 10 for 1.0%). See methods for details of pretreatment; *p < 0.05 versus no pretreatment.

protection from the effect of 0.25% OA challenge (Fig. 6). Temporally, separation between the nontreated group and the group that received 0.1% and 1% terbutaline treatment occurred from minute 2 through 10. The AUC for all dosesof the SCG-treated guinea pigs was not statistically separablefrom AUC of the nontreatedguinea pigs (Fig. 6). However, with temporal analysis, 5% SCG significantly attenuated the effect of 0.25% OA challenge at mintues 3, 4, and 5 versus that of the nontreated group. DISCUSSION In the conscious and nonsedatedguinea pig, daily administration of AICA riboside aerosolattenuatedthe effect of OA aerosol challenge on SR,. The exceptions to this finding tended to be at doses of AICA riboside or OA that were intermediate in nature and were malt-elikely to be attributed to the large interexperimental variability of guinea pig airway responsivenessrather than to discrete differences in airway constriction or relaxation induced by theseagents.For example, 0.1% and 1% AICA riboside were more effective.than 5% in antagonizingthe effectsof 0.25% OA aerosol (Fig. 6), whereas5% AICA riboside was more effective than either 0.1% or 1% in antagonizing the effects of 0.05% OA aerosol (Figs. 3 and 4). Terbutaline and SCGwere testedasantigenic agents to compare the magnitude of their protective effect with that of AICA riboside at similar dosages

(wt/vol). The acute effect of &adrenergic stimulation of the airways by terbutaline markedly attenuated the rise in SR,, induced by OA challenge. This effect was evident at both concentrationstested(Figs. 5 and 6). The failure to demonstrateany dose-responserelationship with terbutaline suggeststhat even lower concentrationscould be equally useful. SCG was chosen as an agent that exerts its antiasthmatic effects acutely in some situations or at times requires more long-term administration to achieve the desired effects. SCG produced a variable decreasein the rise in SR,, causedby 0.1% OA aerosol challenge (Fig. 5), with the most consistent protection observed at the highest concentration of SCG and the lowest dose of OA (Fig. 3). In a comparison of AICA riboside, terbutaline, and SCG, terbutaline was nearly always as good or better than either of the other two agents, although at the highest concentrationof OA challenge, AICA riboside but not SCG performed equally well (Fig. 6). The heterogeneity of the dose-responsecurve to SCG is not an unexpectedfinding. Other investigators have reported13. 23,24and commentedthat subjectswith asthma could be heterogeneousin their responseto SCG. The effect of SCG antagonismof increasedSR,, by sulphur dioxide in patients was dose dependent, although individual variability is apparentwith perusal of thesedata. However, of the two dosesof SCGtested in that study, reactivity to methacholine increasedas

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a result of the administration of the higher dose.” The effect of SCG to antagonize 5’-N-ethylcarboxanide adenosine,an adenosineA,-receptor agonist in rats,13 was heterogeneouswith respectto dosagesof SCG as well. It hasbeenreported in rats that the effectiveness of SCG vary with respect to immunization conditions.2s SR,, was measuredin awakeand nonsedatedguinea pigs by the use of a whole-body plethysmograph. We have usedthis methodin the study of airway reflexes.26 The total resistanceof air movement into and out of the lung (SR,,) is the sum of the upper and lower airway resistances,that is, nasal and/or mouth, phatyngeal, laryngeal, and extrathoracic and intrathoracic airways. Numerous studies demonstrate that in patients with asthma, the upper airways also contribute to increased airway resistance.27-29 Narrowing of the upper airways results from local mast cell degranulation and by reflex constriction of the larynx30 and glottis. Both mechanisms occur in the lowe?’ airways as well. Therefore, therapeutic agents that stabilize mast cells or pharmacologically antagonize smoothmuscle constriction in the lower airways could be expected to attenuate the action of Ag challenge in the upper airways by similar mechanisms. Airway reactivity of guinea pigs to mediators, such as histamine, has been studied with whole-body plethysmography. Reliable dose-responsereactivity to histamine is reportedby Thorne and KaroP* using this method. Furthermore, intersubject variability in this model is reported to be similar to that in humanswhen FEV, in responseto a provocative concentrationcausing a 20% fall in FEV, is measured. Histamine antagonism was not reported in that study, however. Although the effect of Ag challenge by aerosoland its pharmacologic antagonism in guinea pigs has not been reported to our knowledge in the whole-body plethysmograph, Stewart et a1.33have monitored insufflation pressurein responseto aerosolsof OA Ag challenge in anesthetizedguinea pigs. Administration of the P,-adrenergic agonist, fenoterol (2.5 mg/kg, intravenously), 5 minutes before OA, abolishedthe Ag-induced increaseof insufflation pressure. Other antagonists were not studied. Andersson and Bergstrand testedthe long-term effects of SCG challenge and aminophylline on OA Ag-induced bronchoconstriction in sensitized and anesthetizedguinea pigs. Long-term treatmentof either agent for 3 weeks decreasedbut did not abolish bronchial reactivity to Ag challenge as measuredby monitoring lung resistance and dynamic lung compliance. However, SCG was administered as a daily intraperitoneal injection rather than by aerosol administation in that study and with only one dosage(10 mg/kg).

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Therefore, the effect of Ag challenge by aerosolon SR, in the presentstudy is similar to results reported in other studies with several different techniques. In addition, the marked effectiveness of &adrenergic agentswas observed in the present study was similar to that reportedin earlier studiesin guineapigs.33SCG had also beendemonstratedto reducebut not abolish, bronchoconstriction caused by OA Ag challenge in guinea pigs,*’ an observation also reportedin the present study. The choice of long-term daily administration of AICA riboside was made becauseof the previously reported finding that at least 2 days of exposure to AICA riboside in culture was required to inhibit the releaseof granule-associatedor newly generatedmediators from mousebonemarrow-derived mastcells.’ Because other dosing schedules for AICA riboside were not tested, it remains unknown whether the ability of AICA riboside to antagonizethe effects of OA aerosol requires more than one dose or whether one nebulization is an optimal frequency. Considering the encouraging results from these experiments, these questions will be addressedin expandedstudies. The sametype of concernscould be expressedfor the SCG dosing schedule as to whether the doses and times chosenconstitute a fair comparisonof the two putative antiasthmaticagents. Again, theseinitial efforts were to ascertainwhether AICA riboside possessedany potential as an antiasthmatic agent. Therefore, identical doses and time schedules were used for SCG and AICA riboside with some efficacy demonstratedfor each agent under these experimental conditions. Whether the apparent superiority of AICA riboside efficacy over SCG is valid will require more extensive experimentation, including human studies. AICA is currently in clinical trials for cardioprotective application. The mechanism of action of AICA riboside to attenuatethe responseof OA aerosolchallenge on SR,, is not known. AICA riboside apparently has significant intracellular actions. It readily entersintracellular compartments and has been reported to prevent ischemic injury and to enhanceadenosineproduction in myocardial tissue.” AICA riboside is reported to increase intracellular pools of adenosine triphosphate and guanosine triphosphate in postischemic myocardium34and adenosineconcentrationin blood.” In cultured mouse bone marrow-derived mast cells, AICA riboside reduced by 50% the release of P-hexosaminidaseinduced by either specific antigen or A23 187.5An inhibition of leukotriene C, generation from the mast cells was also demonstrableafter severaldays incubation with AICA riboside. Acute or simultaneousadministration of AICA riboside had no

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effect on the quantity of P-hexosaminidasereleasein responseto Ag or A23187 challenge. In addition, this acute treatment had no effect on the action of adenosine to cause the release of P-hexosaminidase,implying that AICA riboside did not act through blockade of adenosinereceptors.Therefore, AICA riboside may possesspharmacologic properties of mast cell stabilization. Furthermore, 5-amino-4-imidazolecarboxamide-1-P-D-ribofuranosyl-5’-triphosphatewas determined to have been produced in the cells exposedto AICA riboside.5 Inhibition of mast cell mediator release may be explained by 5-amino-4-imidazolecarboxamide-.l-P-D-ribofuranosyl-5’-triphosphate competing with adenosinetriphosphate or guanosine triphosphateat an intracellular site that would ultimately alter mediator release, perhapsthe intracellular P receptor. P receptorsmay inhibit enzyme systems,such as adenyl cyclase.35Whether the protective effects of AICA riboside on airway hyperresponsivenessare limited to the mast cell is still unclear. In the present study, a purine analog, AICA riboside, demonstrated antiallergic actions against OA challenge by aerosol with a whole-body plethysmograph to monitor SR,, in a sensitized guinea pig model. T!heultimate use of this agent in the treatment of airway reactivity will be determinedonly by human clinical trials. REFERENCES 1. Cushley MJ, Tattersfield AE, Holgate ST. Inhaled adenosine and guanosine on airway resistance in normal and asthmatic subjects. Br .I Clin Phannacol 1983;15:161-5. 2. Marquardt DL, Parker CW, Sullivan TJ. Potentiation of mast cell mediator release by adenosine. J Immunol 1978; 120(3):871-g. 3. Nishibori J. Shimamura K, Yokoyama H, Tsutsumi K, Saeki K. Differential effects of adenosine on histamine secretion induced by antigen and chemical stimuli. Arch Int Pharmacodyn Ther 1983;265:17-28. 4. Pauwcls R, van der Straeten M. The bronchial effects of adenosine in the rat. Arch Int Pharmacodyn Ther 1986;280 (supp1)2:229-39. 5. Marquardt DL, Gruber HE. Inhibition of mast cell mediator releasmeby 5-amino-4-imidazolecarboxamide riboside. Biothem Pharmacol 1986;35(24):4415-21. 6. Wehon AF, Simko BA. Regulatory role of adenosine in antigen-induced histamine release from the lung tissue of actively sensitized guinea pigs. Biochem Pharmacol 1980: 29: 1085-92. 7. Church MK, Hughes PJ, Holgate ST. Adenosine modulation of histamine release from human basophils and mast cells [Abstract]. Feb Proc 1983;42:6152. 8. Hughes PJ, Holgate ST, Church MK. Adenosine inhibits and potentiates IgE-dependent histamine release from human lung mast cells by an A,-purinoceptor-mediated mechanism. Biothem Pharmacol 1984;33(23):3847-52. 9. Peachell PT, Columbo M, Kagey-Sobotka A, Lichtenstein LM, Marone G. Adenosine potentiates mediator release from human lung mast cells. Am Rev Respir Dis 1988;138:1143-51,

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10. Brown CM, Collis MG. Evidence for an Al/R, adenosine receptor in the guinea pig trachea. Br J Pharmacol 1982;76: 381-8. II. Coleman RA. Effects of some purine derivatives on the guinea pig trachea and their interaction with drugs that block adenosine uptake. Br J Pharmacol 1976;57:51-7. 12. Satchel1 D, Smith R. Adenosine causes contractions in spiral strips and relaxations in transverse strips of guinea pig trachea: studies on mechanism of action. Em J Pharmacol 1984;101:243-7. 13. Pauwels RA, van der Straeten ME. An animal model for adenosine-induced bronchoconstriction. Am Rev Respir Dis 1987;136:374-8. 14. Chemiack NS, Runold M, Prabhakar NR, Mitra H. Effect of adenosine on vagal sensory pulmonary afferents. Fed Proc 1987;43(3):825. 15. Runold M, Prabhakar NR, Mitra H, Chemiack NS. Adenosine stimulates respiration by acting on vagal receptors. Fed Proc 1987;43(3):825. 16. Gustafsson LE. Wiklund NP, Cederqvist BO. Apparent enhancement of cholinergic transmission in rabbit bronchi via adenosine A, receptors. Eur J Pharmacol 1986;120:179-85. 17. Gruber HE, Hoffer ME, McAllister DR, et al. Increased adenosine concentration in blood from ischemic myocardium by AICA riboside. Circulation 1989;SO:1400-l 1. 18. Andersson P. Antigen-induced bronchial anaphylaxis in actively sensitized guinea pigs: pattern of response in relation to immunization regimen. Allergy 1980;35:65-71, 19. Pennock BE, Cox CP, Rogers RM, Cain WA. Wells JH. A noninvasive technique for measurement of changes in specific airway resistance. J Appl Physiol: Respir Environ Exert Physiol 1979;46(2):399-406. 20. Abraham WM. Pelejunt JC, Yerger L, Marchette B. Characterization of a late-phase pulmonary response after antigen challenge in allergic sheep. Am Rev Respir Dis 1983;128:83944. 21. Richards R, Simpson SF, Renwick AG, Holgate ST. Inhalation rate of sodium cromoglycate determines plasma pharmacokinetics and protection against AMP-induced bronchoconstriction in asthma. Eur Respir J 1988;1(10):896-901. 22. Zar JH. Biostatistical analysis. Englewood Cliffs, N.J.: Prentice-Hall, 1974. 23. Ben-Dov I, Bar-Yishay E, Godfrey S. Heterogeneity in the response of asthmatic patients to pre-exercise treatment with cromolyn sodium. Am Rev Respir Dis 1983;127:113-6. 24. Myers DJ, Bigby BG, Boushey HA. The inhibition of sulfur dioxide-induced bronchoconstriction in asthmatic subjects by cromolyn is dose dependent. Am Rev Respir Dis 1986;133:1150-3. 25. Andersson P, Bergstrand H. Antigen-induced bronchial anaphylaxis in actively sensitized guinea-pigs: effect of long-term treatment with sodium cromoglycate and aminophylline. Br J Pharmacol 1981;74:601-9. 26. Bergren DR. Capsaicin challenge, reflex bronchoconstriction, and local action of substance P. Am J Physiol 1988;254:R845R52. 27. Campbell AH, Imberger H, Jones B. Increased upper airway resistance in patients with airway narrowing. Br J Dis Chest 1976;70:58-65. 28. Higenbottam T, Payne J. Glottis narrowing in lung disease. Am Rev Respir Dis 1982;125:746-50. 29. Nacleiro RM, Meier HL, Kagey-Sobotka A. Mediator release after nasal airway challenge with allergen. Am Rev Respir Dis 1983;128:597-602. 30. Nadel JA. Autonomic regulation of airway smooth muscle. In:

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Nadel JA, ed. Physiology and pharmacology of the airways. New York: Marcel Dekker, 1980:217-57. 31. Lisboa C. Jardim J, Angus E, Macklem PT. Is extrathoracic airway obstruction important in asthma?Am Rev Respir Dis 1980;122:115-21. 32. Thome PS, Karol MH. Assessmentof airway reactivity in guinea pigs: comparison of methods employing whole body plethysmography.Toxicology 1988;52(1-2):141-63. 33. Stewart AG, FennessyMR. Lewis SJ. Attenuation of antigen-

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induced bronchospasmby fenoterol in the guinea pig. Agents Actions 1984;14(1):318. 34. Swam JL, Hines JJ, Sabina RL, Holmes EW. Accelerated repletion of ATP and GTP pools in postischemiccanine myocardium using a precursor of purine de novo synthesis. Circ Res 1982;51:102-5. 35. Londos C, Wolff J. Two distinct adenosine-sensitivesites on adenylate cyclase. Pros Nat1Acad Sci USA 1977;74:54826.

lmmunoglobulin concentrations in nasal secretions differ between patients with an IgE-mediated rhinopathy and a non-IgE-mediated rhinopathy Siebe J. Swart, MD,* Siebren van der Baan, MD, PhD,** Joke J. E. Steenbergen, Jos J. P. Nauta,** Gerard J. van Kamp, PhD,*** and Jeike Biewenga, PhD* Amsterdam, The Netherlands Nasal secretions from patients with an &E-mediated rhinopathy, patients with a non-&E-mediated rhinopathy, and healthy control subjects were collected with a newly developed direct aspiration system. Total protein, albumin, secretory IgA (sIgA), IgE, IgG, and Igki concentrations were measured in the nasal secretions to detect whether the nasal pathology is reJected in nasal secretion protein concentrations. It was found that protein and immunoglobulin concentrations in nasal secretions were inversely related to amount of secretion in the nasal cavity. Both patients’ groups had a significantly higher sIgA to protein ratio than the healthy control subjects. Furthermore, patients with an &E-mediated rhinopathy had significantly higher sIgA and IgM to total protein ratios in their nasal secretions than patients with a non-IgE-mediated rhinopathy. Probably these dtyerences are due to changes in immunoregulation. (J ALLERGYCUN IMMUNOL1991;88:612-19.) Key words: IgE-mediated rhinopathy, non-IgE-mediated immunoglobulins

From the *Department of Cell Biology, Division of Histology, Medical Faculty, and Departments of **Otorhinolatyngology/Head and Neck Surgery and ***Clinical Chemistry, Academic Hospital, Vrije Universiteit, Amsterdam, The Netherlands. Supported in part by Astra PharmaceuticsBV, Rijswijk, and by Glaxo BV, Nieuwegein, The Netherlands. Received for publication Oct. 19, 1990. Revised May 22, 1991. Accepted for publication May 22, 1991. Reprint requests: Jeike Biewenga, PhD, Department of Cell Biology, Division of Histology, Medical Faculty, Vrije Universiteit, Van der Boechorststraat7, 1081 BT Amsterdam, The Netherlands. l/1/31235

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rhinopathy, nasal secretion,

Abbreviations used

SFT: sIgA: Ab: AR: HRF’:

Immune respiratory noglobulin the results

Skin prick test Secretory IgA Antibody Allergic rhinitis Horseradishperoxidase

and inflammatory reactions in the upper tract are reflected in protein and immucomposition of nasal secretions. ’ However, of nasal secretion analysis in patients with