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The temporal relationship between increases in airway responsiveness to histamine and late asthmatic responses induced by occupational agents
Moss et al.
J. ALLERGY
42. Pienkowski MM, Norman PS, Lichtenstein LM. Suppression of late-phaseskin reactions by immunotherapy with ragweed extract. J ALLERGYCLIN IMMUNOL 1985;76:729. 43. Peltre G, Lapeyre J, David B. Heterogeneity of grass pollen allergens(Dacrylis glomerura) recognizedby IgE antibodiesin
CLIN. IMMUNOL. FEBRUARY 1987
humanpatients seraby a new nitrocellulose immunoprint technique. Immunol Lett 1982;5:127. 44. Sutton R, Wrigley CW, Baldo BA. Detection of IgE- and IgGbinding proteins after electrophoretic transfer from polyacrylamide gels. J Immunol Methods 1982;52:183.
The temporal relationship between increases in airway responsiveness to histamine and late asthmatic responses induced by occupational agents Stephen R. Durham, M.R.C.P., * Bernard J. Graneek, M.R.C.P., Rosemarie Hawkins, S.R.N., and Anthony J. Newman Taylor, M.Sc., M.R.C.P., M.F.O.M. London, England The temporal relationship between increases in airway responsiveness and the late asthmatic response was assessed in nine patients challenged with occupational agents toluene diisocyanate (one patient), carmine (one patient), maleic anhydride (two patients), colophony (four patients), and trimellitic anhydride (one patient). The provocation concentration of histamine causing a 20% decrease in FEV, (PC,,) was measured before challenge and at approximately 3 hours and 24 hours on control and active-challenge days. Thirteen active challenges provoked eight dejinite late asthmatic responses (maximum fall in FEV, > 15% at 3 to 11 hours). At 3 hours after the challenges that provoked late responses, there was a significant (p < 0.02) decrease in PC,, that was more (p < 0.03) than that observed for the3ve tests provoking early (late FEV, fall 0% to 5%) or equivocal late (FEV, fall 6% to IS%) responses. At 24 hours, PC,, remained decreased (p < 0.05), although it was less so than at 3 hours (p < 0.05) and not signijTcantly when compared with challenge tests causing single early or equivocal late responses. The 3-hour decreases in PC,, were identified when FEV, (jive of seven observations) was MO% of prechallenge values. For the nine independent tests, the j-hour decreases in PC,, correlated (r = 0.72; p < 0.05) with the magnitude of the late falls in FEV,, whereas this was twt observed at 24 hours (r = 0.35; p, not significant). These results demonstrate that in subjects challenged with occupational agents, increases in histamine airway responsiveness precede the late asthmatic response, occur independently of changes in airway caliber, and correlate with the magnitude of the subsequent late response. We suggest that in subjects with occupational asthma, the late asthmatic response depends on the tissue events, possibly airway inJlammation, that underlie the preceding increase in airway responsiveness. (J ALLERGYCLIN hfMUNOL 1987;79:398-406. )
Airway hyperresponsiveness,an increased bronchoconstrictor response to nonspecific provocative I From the Department of Occupational Medicine, Cardiothoracic Institute, Brompton Hospital, London, England. Received for publication Aug. 12, 1985. Accepted for publication Aug. 16, 1986. Reprint requests: A. J. Newman Taylor, M.Sc., Dept. of Occupational Medicine, CardiothoracicInstitute, Brompton Hospital, Fulham Rd., London 3W3 6HP England. *Current address:JohnRadcliffe Hospital, Off Headley Way,Headington, Oxford, U.K.
I 1
Abbreviations used Provocation concentrationof histamine causGl: ing a 20% decreasein FEV, TDI: Toluene diisocyanate
stimuli, is now considered a central feature of bronchial asthma. The association between increases in airway responsivenessand late asthmatic responses has been observed after inhalation of both aller-
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Airway
responsiveness
to histamine
TABLE I. Clinical features of nine subjects studied Age Patient
Ckcupational agent
IV)
Sex
41
M
TDI
18 48
M M
21
M
18 46 40 39 37
M M M F M
Carmine Maleic anhydride Maleic anhydride Colophony Colophony Colophony Colophony Trimellitic anhydride
399
--__-Duration of exposure Ml
o=-
Worker in printing ink factory Batch weigher Storeman
FM,titemlsec I% pm)
PC& ~m%imll
16
3.2 (98)
24
1 17
3.8 ! loo) 3.2 t88,
1.7 X.0
Batch weigher
1
4.3 il II)
3.0
Solderer Solderer Solderer Solderer Maintenance worker
3 0.5 10 3 20
4.5 (105) 2.9 (89) 2.2 (67) 2.1 (100) 4.3 !I 14)
1.1 1.8 0.68 2.3 18.5
Duration of exposure representstime interval between first exposure and onset of work-related respimtory symptoms. S@ects 3. 4, 5, 7, and 9 were atopic (skin prick tests positive to one or more of the following allergens: Derf?m&@#agoi&spterqM#z~. mix& grass dlens, cat fur, or dog hair), and subjects 1, 2, 6, and 8 were nonatopic. Subjects 1, 3, 6. 8, and 9 were exsmokers, and subjects 2, h, 5. and 7 had neversmoked
gens’.’ and occupational agents3 These increases in airway responsiveness were observed at 24 hours after challenge, i.e., after the development of the late response. The consistency of this observation has led to the suggestion4.’ that the increase in airway IXsponsivenessresults from mechanismsthat underlie the late asthmaticresponse,possibly as a consequence of bronchial inflammation. In this study, in subjects challenged with occupational agents, we have sought to clarify the temporal relationship between the late asthmaticresponseand the associatedincreasein air-
way responsiveness.We have measuredairway responsivenessto inhaled histamine before challenge with occupational agents, at 3 hours (before the late asthmatic response), and at 24 hours (after the late asthmatic response), and related the changes in responsivenessto the magnitude of the late response.
T’heclinical featuresof the nine patients (eight male patients and one female patient) studied are presentedin Table 1. The inhalation tests were performed as part of the routine investigation for possible occupational asthma. At the time of the investigations. the subjectswere asymptomatic, and with one exception, patient 7, the baseline FEV, was ~35% of the predicted normal value. Bronchodilators were withheld for at least 24 hours before the challenge tests. With one exception, none of the subjectshad taken disodium cromoplycate or corticosteroids in the week befan the study. The exception, patient 6, had taken regular beclomethasonedipropionate, 250 p,g (one puff from a metered-dose inhaler). daily. until the day before his admission.
The patients were admitkd lo the hospital for a period of 1 to 2 weeks. On day 1, the p8ticnt.swere instmcted in the performanceof nptatpble forced enpiratory maneuvers with a dry wedge-bellows spirometer, (N&graph Ltd., Bucks, U.K.). On day 2 the p&en& had a “control” challenge in which the patients’workexposureswee simulated
in the absenceof theOccupatiOgnl agentsuspcckdto cause asthma.On subsequentdays testswen madewilh increasing exposureslo the suspectedagent. On control and challenge days. the patients’ nonspecific bronchial responsivenesswas measuredby a histamine provocation test. The histamine tests were performed on at least three occasionsduring the day: before the occupatioualexposuretests(generally 10:00 A.M.), at approximately 3 hours after the test (generally 2:00 P.M.), and at 22 to 24 hours (at IO:00 A.M. the following day). In four of the nine p&ents tested, one further histamine test was performed at 6 to 9 hours after the occupational exposure.
The histamine tests were performed by use of a tidal breathing mctl~& and a Wright nebulizer. The nebulizer was calibrated to produce 0.14 mllmin. The duration of inhalation of histamine was stand&i& af 2 minutes. Histamine responsivenesswas defined asPC, and Wasobtained by linear interpolation of the log histamine FEV, dose-response curve. A PC,,,of 8 mg/ml was taken as the cut point between normal and increa.& histamine airway responsiveness.Histamine tests were performed before challenge and at approximately 3 hours and 24 hours after challenge on control days and after exposure to the suspected occupational agent. The precise timings of the histamine tests (when the first dose of histamine was inhaled) are reprcsented in Figs. 1 to 4. The timings of the last dose of
400 Durham et al.
TOLUENE
J. ALLERGY
DIlKXYANATE
FIG. 1. The time course (0 to 24 hours) of changes in FEV, (symbols o, l , A, and n , liters per second) and PC, (solid bars, milligrams per milliliter) in a subject exposed for 30 minutes to increasing concentrations of TDI (0 to 0.02 parts per million). The magnitude and timing of the early (0 to 60 minutes) and late (3 to 11 hours) maximal percent decreases in FEV, are indicated. The hatched bars represent the periods of challenge.
histamine (when the measurementof histamine responsiveness was made) occurred up to 30 minutes after the times indicated. The occupational-typebronchial provocation procedures’ were performed in an isolation cubicle, dimensions 1.5 by 1.3 by 2.2 meters, fitted with an exhaust-extraction fan. The patients wore gowns, caps, gloves, and plastic overshoes. These occupational-type exposures on control and active challenge days were performedin a single-blind fashion in which the patients had no knowledge of the protocol used. The responsewas measuredas FEV, at 5, 10, 15, 30, and 60 minutes, hourly from 1 to 12 hours (or until the patient retired to bed), and on the next day until 22 to 24 hours after challenge. The bestof the threeforced expiratoty maneuversat each time point was used in the subsequent analysis. The incremental exposuresto increasing concentrations (or durations of exposure) of the suspectedagent wereperformedon separatedaysand only when the patients’ histamine responsivenesshad returned to baseline (control day) values. A positive early asthmaticresponsewas defined as a decreasein FEV, that occurred within 1 hour of the test of >15% from the mean of two prechallenge values
CLIN. IMMUNOL. FEBRUARY 1987
and that was >15% from the correspondingtime value on the control day. A positive late asthmatic response was defined as a decreasein FEV, between3 hours and 11 hours after the test of >15% from the prechallenge and corresponding control day values. A late asthmaticresponsewas considered equivocal when a reduction in FEV, of 6% to 15% was observed. Exposure to TDP was performedas follows: The patient first brush painted 100 gm of marine-basevarnish for a period of 30 minutes (control day). On the subsequentchallenge days, he painted varnish to which increasing amounts of TDI, previously determinedto produce reproducible air concentrationsof 0.005 ppm to 0.02 ppm within the cubicle used for provocation, were added (TDI meter, model 7005FR, Sieger Ltd., Poole, Dorset, U. K.). Carmine is a natural brilliant red dye in the form of a powder extractedfrom the dried femaleinsectCoccuscactus and is usedwidely in the cosmeticsindustry.’ On the control day, the patient suspectedof sensitivity to carmine (patient 2) tipped 250 gm of oven-dried lactosepowder mixed with amaranth B.P.C. (Evans Medical, Ltd., Dinistable, Bedford, U. K.), an unrelated dye of similar color to carmine. On the challenge days, he tipped 250 mg of lactose mixed with 0.1% and 0.3% carmine for a period of 60 seconds. The patientssuspectedof sensitivity to maleic anhydride”’ (subjects3 and 4) tipped 250 gm of lactose alone (control day) and mixed with maleic anhydride in the form of crystals ground to a powder and addedin increasing concentrations to 1% by weight for 5 minutes. The patients suspectedof sensitivity to colophony” (subjects5,6,7, and 8) performed manual soldering with solid solder wire (60/40 tin/lead alloy) (control day), and with multicore solderwire containing colophony flux for maximum periods of 15 minutes, 15 minutes, 1 minute, and 3 minutes, respectively. The subject challenged with trimellitic anhydride tipped 250 gm of lactose powder alone (control day) and mixed with trimellitic anhydride 1% for 5 minutes.
Analysis Statistical analysis of the changesin PC, was performed on log-transformeddata. For the subjectswith definite late asthmatic responses(maximal FEV, percent decreaseof > 15% at 3 to 11 hours), the changesin histamine PC, at 3 hours and 24 hours after challenge (log prechallengePC, minus log postchallengePC,) were comparedwith the corresponding values on the control days by use of the Wilcoxon signed-ranktest. The changesin PC,, at 3 hours and 24 hours after challenge for the definite late responderswere aIs0 comparedwith corresponding values in subjectswho developedeither single early (late FEV, fall of 0% to 5%) or equivocal late responses(late FEV, of 6% to 15%) by use of the Mann-Whitney U test. Spearman’srank correlation was used to comparethe changesin histamine P&, at 3 hours and at 24 hours with the magnitude of the late falls in FEV, for the challenge tests. Becausemore than one challenge was performed in three of the patients, we randomly selectedonly one of the results from each of these
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MALEIC
CARMINE
responsiveness
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401
ANHYDRIDE
hours
Pi& 2. The time course (0 to 24 hours) of changes in FEV, (symbols 0, l , and A, titers mr second) and PCnr(solid bars, milligrams per milliliter) in a subject exposed far 1 minute to ti&ng v&h 250 gm of lactose alone and mixed with carmine (0.1% and 0.3%), and in a sub@@ exposed kr 5 minutes to tipping with 250 gm of lactose powder alone and mix8d with meleic an~yd~e (0.2% and 1%). The magnitude and timing of the early (0 to 60 minute91 and late (1 to 12 hm} maximal percent decreases in FEV, a& indicated. The hatched bars represent the periods of challenge. MALEIC
COLOPHONI
ANHYDIUDE
6.0 2.0 0.5 0.125
8.0 2.0 0.5 0.125 Before
0 60 c :: mins
3
5
7
9
11
22
24 *
hours
FIG. 3. The time course (0 to 24 hours) of changes in FEV, (Eymbols o and 0, liters per SIBWI~) and PC, {solid bars, milligrams per milliliter) in a subject exposed for 5 minutes to t&p&g w&h 250 gm of lactose powder alone and mixed with maieic anhydride 1%. and in a subjaa who performed for 5 minutes manual soldering with solid colder wire (tindad altoy, control &allenge) and with multicore solder wire containing c&phony flux. The magnitude end timing of the early (0 to 60 minutes) and late (1 to 12 hours) maximal Percent decreases in FEV, ar. indicated. The hatched bars represent the periods of challenge. three patients in estimating correlation and its significsnce. P values CO.05 were considered statistically sign&ant.
The individual values of FJZV, and PC, for the subjects challenged with occupational agents are rep-
resented in Figs. 1 to 4. On the cm&o1 days for all subjects, the FEV, did no& change by > k 10% of the
prechallenge values. ‘II&teen active chaknges were performed in nine subjects. Three chakqges resulted in a single early asthmatic response. Ten chellenges resulted in an equivocal or definite late response. The responses were arbitrarily divided into thme categories
J. ALLERGY
402 Durham et al. COLOPHON-f
COLOPHONY
Before
0 60 t 3: mlns
3
5
7
9
11
22
24
hours
0 <
60 ::
IllitlS
3
5
7
9
Before
0 60 < :< mins
3
5
7
9
11
22
24
hours
TRIMELLITIC! ANHYDRIDE
COLDPHONY
Before
CLIN. IMMUNOL. FEBRUARY 1987
11
22
24 >
hours
FIG. 4. The time course (0 to 24 hours) of changes in FEV, (symbols o and l , liters per second) and PCs,(solid bars, milligrams per milliliter) in three subjects who performed manual soldering (patient 6, 15 minutes; patient 7, 1 minute; patient 8, 3 minutes) and in one subject (patient 9) exposed for 5 minutes to tipping with 250 gm of lactose powder alone and mixed with trimellitic anhydride 1%. The magnitude and timing of the early (0 to 60 minutes) and late (1 to 12 hours) maximal percent decreases in FEV, are indicated. The hatched bars represent the periods of challenge.
according to the magnitude of the late (3 to 11 hours) falls in PEV, as follows: (1) no late response (late PEV, decrease0% to 5%, asobservedin three subjects with single early responses), (2) equivocal late response(late PEV, decrease6% to 15%, observed as part of a dual responsein two challenges), and (3) definite late response(late PEV, decrease>15%, as observed in eight challenges, either “isolated late” in four or as part of a “dual” responsein four challenges)(Table II). The absolutevalues for PC, before challenge and at 3 hours and 24 hours after challenge (together with the FEV, immediately before the histamine tests in parenthesis)are recorded in Table II. Thirty-nine of the total 43 postchallenge histamine teats were performed when the PEV, was within f 10% of the prechallengevalues. With the marginal exception of subject3, the pre- andpostchallengePC,,
values on the control days were within a twofold histamine concentration difference. Similarly, the prechallenge PC,, values on control and active challenge days were within a twofold concentration difference. There was a wide range of baseline histamine responsiveness. The prechallenge PC, varied from moderately severely increased to within the normal nonasthmatic range (0.68 to 24 mg/ml). There was no significant correlation between the prechallenge PC, and the tendency to develop either an early or late asthmatic response. PC, was measuredat 3 hours after challenge in seven of the eight challenges that provoked definite late responses.PEV, was X0% of the prechallenge value at this time in five of the seventests. After all sevenchallenges, a greater than fourfold reduction in PC, was observed. These 3-hour reductions in PCm
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79 2
TABLE II. PC, in subjects -
challenged
with occupational
agents
-----.
Maximal FE&% deerease postchallenge occupatbnel &W
agent
Control TDI 0.005 TDI 0.01 TDI 0.02 Control Carmine 0.1% Carmine 0.3% Control Maleic anhychide 0.2% Maleic anhydride 1% Control Maleic anhydride 1% Control Colophony 15 min Control Colophony 15 min Control Colophvny 1 min Control Colvphony 3 min Control Trimellitic anhydride 1%
403
.-.________
PC, (mglmi) --.----
TYP of
24.hr’ poet-
Late (3 to 11 hr)
PrecheiiQnge
3-hr* postcheilenge
3 2 8 1 16
1 31 22 30 0 I2
24 (3.17)t >I6 (3.35) >I6 (3.16) 20 (3.37) 1.7 (3.74) I .65(3.53)
24 2.9 1.7 ND 2.h 0.76
39
30
2.6 (4.0)
0.5
1 0
8 31
8.0 (3.20) 3.3 (3.27)
7.9 (3.35) 0.43 (3.201
Dual
30
23
6.4 (3.57)
Control Dual (equivocal late) Control Dual
I 24
3 14
3.0 (4.31) 3.9 (4.42)
6.0 (4.50) 0.53 14.1)
5.2 (4.30) 0.38 (4.4)
4 25
3 16
1. I (4.45) I .25(4.4)
1.4 (4.3) 0.25 (4.1)
1.25 (4.4) 0.6 (4.35)
Control Dual
4 24
9 24
1.8 (2.93) 2.7 (2.70)
1.0 (2.78) 0.61 (2.21)
I .O (2.63) 0.58 12.80)
Control Single early
6 28
6 3
0.68(2.23) 1.3 (2.15)
0.64 (2.17) 0.70 (2.17)
I.3 (2.30) I .29 (2.30)
Control Single early
7 30
5 5
2.3 (2.1) 2.2 (2.35)
1.7 (2.35) 1.5 (2.35)
2.2 (2.35) 1.8 (2.30)
Control Single early
3 44
6 4
18.5 (3.9) 14.5 (3.73)
14.0 (3.99) 9.1 (3.5)
14.5 (3.95) IO. 1 (3.90)
asthmatic reaction
Control Late Late Late Control Dual (equivocal late) Dual Control Late
Early (0 to 60 min) 0
(3.28) (3.26) (1.lSi
chatfetsge 3 ih
(3.35)
13.w (3.22)
.>lh II 0.9 I.5 I .O
(3.15) i3.31) (2.X5) (3.71) (3.62)
(3.30,
0.9
i3.0)
3.8 (3.6) 1.27 (3.09) 1.9 (3.44)
PC, (milligrams per milliliter) before challenge and at 3 hours and 24 hours after challenge are compared with the type of asthmatic responseand the maximum percent decreasein FEV, at 0 to 60 minutes and at 3 to 11 hours after challenge. *The 3-hourpostchallengePC, measurementswere performedat 2% to 3’/2hours(exceptfor subject5 at 4 hvurs). The 24.hour measurements were perfvrmcd at 22 to 24 hours. The precise timings for the individual subjects are representedin Figs. 1 to 4. *The values in parenthesesrepresentthe baseline FEV, (liters per second)recorded immediately before the measurementsof PC,,..
preceded
the development
of the late response and
were significant when they were compared with the control day reductions (p < 0.02) and with the corresponding 3-hour values in the five challenges that provoked equivocal late (two) or single early (three) asthmatic responses (p < 0.03). PC, was measured at 24 hours after challenge after eight challenges provoking definite late responses. A reduction in PC,, was observed after seven of the eight challenges. These 24-hour reductions in PC,, were
significant
when reductions
were compared
with re-
ductions on control day (p -C 0.05), although the magnitude of the changes were less than those observed at 3 hours in the same subjects (p < 0.05) and were not significant when these were compared with the corresponding 24-hour values for the five challenges provoking equivocal late or single early asthmatic responses. The magnitude of the changes in PC?, at 3 hours and at 24 hours (expressed as the ratio prechallenge
404
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J. ALLERGY
correlation was observed at 3 hours (r = -0.38) or at 24 hours (r = -0.05) after challenge.
80
A
10
25
0
DISCUSSION
8
0.5 1 2 4 8 16 Ratio change in histamine PC20 at 3 hours (pre-challenge PC2O/post-challenge PC20) 30@.
Oo
1 25-
0
0
0
0
2015-
0 0 0
lo5-
24 hours .0
CLIN. IMMUNOL. FEBRUARY 1987
r = 0.35 p = NS
0 I I I I I 0.5 1 2 4 8 16 Ratio change in histamine PC20 at 24 hours (pre-challenge PC20/ post-challenge PC20) 0
FIG. 5. A, Ratio change in PCzO(prechallenge PC&/postchallenge PCJ at 3 hours. 8, Ratio’change in PC& (prechallenge PC,/postchallenge PCJ at 24 hours compared with maximum FEV, percent decrease during late period (3 to 11 hours postchallenge). Correlation coefficients were obtained by Spearman’s rank correlation on data from independent challenges (0); data from separate challenges on the same patients (0) were not used for estimation of the correlation coefficients.
PC,,/postchallengePC,) wascomparedwith the magnitude of the late falls in FEV, provoked by the challenge tests (Fig. 5). For the nine independentobservations, there was a significant correlation between the magnitude of the decrease in PC,, at 3 hours and the magnitude of the late response (r = 0.72; p < 0.05), whereas this was not observed for the changes in PC,, at 24 hours (r = 0.35; p, not significant). When the magnitude of the early asthmatic response(maximal percent decreasein FFV, 0 to 60 minutes after challenge) was compared with the changes in histamine responsiveness,no significant
We have found that occupational agents provoked an increasein airway responsivnessto histamine that preceded the development of the late asthmatic response. These early increases in responsivnessappeared to occur independently of changesin airway caliber and were related to the magnitude of the subsequentlate falls in FEV,. Furthermore, in three subjects, increasingintensity of exposureto the provoking agentresulted in an earlier occurrenceof the late asthmatic response. Taken together, these observations suggestthat the underlying tissue events causing the late responsecan begin before the late responsebecomesevident. These studies involved a controlled series of observations in which the changesin histamine responsivenessand FEV, on the active challenge days were comparedwith both prechallengevalues and the corresponding values on a separateday after a control challenge. The criteria we usedfor defining a positive asthmatic responseenabled us to identify significant changesin both FEV, and PC,, without subjecting the patients to undue bronchoconstriction (14% to 44% maximal reductions in FEV, during early and late responses).Thus, although somepatientsexperienced mild symptoms of cough and wheezing during these reactions, at no time was a bronchodilator required to reversetheir responses.The inhalation testswere performed in a single-blind fashion. A double-blind protocol would have been ideal but was not possible becauseof the potential hazards of uncontrolled administration of the occupational agents used. Randomization of the challenges was also consideredinappropriatein view of the necessityto establish baseline observations after a control challenge before the active challenge days. The occupational challenge procedures involved several different agents and methods of administration. The pattern of asthmatic responsesalso varied. Isolated late, dual (early and late), and single early reactions were observed. The pattern of asthmaticresponse and the changesin histamine responsiveness appearedto be unrelatedto the natureof the provoking agent used. However, the changes in histamine responsivenesswere remarkably consistent. Whatever the nature of the test exposure, the developmentof a late asthmatic responsewas precededby a reduction in PCZOat 3 hours after challenge. Interpretation of changesin bronchial responsivenessis complicated when there is a changein airway caliber.‘*, l3 In the present study, the 3-hour increases
VOLUME NUMBER
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79 2
in histamine responsiveness for the subjects with equivocal or definite late responses were identified at a time when they were asymptomatic and their FEV, (in seven of nine observations) was >90% of the prechallenge and corresponding control day values; in two patients (subjects 1 and 3), these 3-hour increases in histamine responsiveness were observed when the FEV, was 100% and 98% of the prechallenge values. [t therefore appears likely that the changes we observed in histamine responsiveness could not be explained by changes in airway geometry. It is possible that the histamine test performed at 3 hours may itself have modified the subsequent late response. However, this appears unlikely for the following reasons: There were no late falls in FEV, after the 3-hour histamine tests performed on control days and after single early asthmatic responses. In six of nine observations in subjects with equivocal or definite late responses, the FEV, returned to pretest values before the subsequent late falls in FEV,; in the subject exposed to increasing concentrations of TDI, the timing of the late response was earlier at higher concentrations of TDI. After the highest concentration (TDI 0.02 ppm), a 3-hour histamine test could not be performed because the late response had already begun. The presence or absence of a histamine test did not appear to modify the late response in this subject. The mechanisms underlying late asthmatic responses are largely unknown. Studies of allergen-induced late asthmatic responses in humans suggest that airway inflammation may be important. Thus, the late asthmatic response has been associated with release of pharmacologic mediators,14.I5 activation of peripheral blood leukocytes,‘6- ” altered T-lymphocyte function.18 peripheral blood, “. ‘” and bronchoalveolar*’ eosinophilia. Airway hyperresponsiveness has similarly been associated with airway inflammation, since many of the agents that increase responsiveness, such as viruses”. ” and ozone,24 also cause bronchial inflammation. Airway responsiveness has previously been demonstrated to be increased after late asthmatic responses induced by allergens” * and by occupational agents.’ Taken together, these observations suggest an important link between late asthmatic responses, airway hypemesponsiveness, and bronchial inflammation. Some authors have speculated that the increased airway responsiveness that has previously been identified after the late response may result from the airway inflammation that accompanies the late response.‘, 4.’ In the present study we have demonstrated that increases in airway responsiveness preceded the late asthmatic response. These early increases in responsiveness appeared to be independent of changes in airway caliber and were identified at a time when the
responsiveness
to histamine
405
subjects were asymptomatic. Our finding that the magnitude of these early increases in airway responsiveness was significantly correlated with the magnitude of the subsequent late response does not necessarily imply a causal relationship. On the other hand, this relationship raises the possibility that the late response depends on the mechanisms that underly the preceding increase in airway responsiveness, presumably as a consequence of airway inflammation Our observations were confined to subjects with occupational asthma. A recent study by Cockroft and Murdock,” published in abstract form, suggested that histamine responsiveness was not increased at 2 hours after allergen challenge in subjects who developed late asthmatic responses. This observation is not inconsistent with our own findings. First. the measurements of histamine responsiveness were performed at 2 hours after challenge and do not exclude a possible increase in histamine responsiveness at 3 hours after allergen inhalation. Second, increases in histamine responsiveness were observed at 2 hours in two of the 12 subjects studied. Third, the magnitude of the Iate responses (mean maximal JYEV, decrease of 17%) was lower when late responses were compared with the responses of the present study group (mean maximal FEV, decrease of 26%). In summary, late asthmatic responses induced by allergens and occupational agents are closely associated with an increase in airway responsiveness to inhaled histamine. We have demonstrated that in subjects challenged with occupational agents. the increase in airway responsiveness precedes the late asthmatic response. In so far as late asthmatic responses correlate closely with occupational3 and seasonal’” asthma, these observations support a central role for increases in airway responsiveness in the induction of clinical asthma. A therapeutic implication of these findings is that pharmacoIogic agents that reverse increased airway responsiveness are likely to inhibit late asthmatic responses and be effeclive in prophylaxis of clinical asthma. REFERENCES 1. Cockroft DW, Ruffin RE, Dolovich J. Hargreavc FE. Allergeninduced increase in nonallergic bronchial reactivity Clin Allergy 1977;7:503. 2. Cartier A. Thomson NC, Frith PA, Robens R, Hargreave FE. Allergen-induced increase in bronchial responsiveness to histamine: relationship to the late asthmatic response and change in auway caliber. J ALLERGY CLIN I,u~riwor. 1982:70: 170. 3. Chan Yeung M. Lam S, Koemer S. Clinical features and natural history of occupational asthma due to western red cedar (Thuju plicata). Am I Med 1982;72:4 Ii. 4. Cockroft DW. Mechanisms of perennial allergic asthma. Lancet 1983;2:253. 5. Kaliner M. Hypothesis on the contribution ot late-phase re-
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et al.
sponsesto the understandingand treatmentof allergic diseases. J ALLERGY CLIN IMMUNOL 1984;73:311.
6. Co&oft DW, Killian DN, Mellon JJA, Hargreave FE. Bronchial reactivity to inhaled histamine: a method and clinical survey. Clin Allergy 1977;7:235. 7. Pepys J, Hutchcroft BJ. Bronchial provocation tests in etiological diagnosis and analysis of asthma. Am Rev Respir Dis 1975;112:829. 8. O’Brien IM, Newman Taylor AJ, Burge PS, Fawcett IW, Pepys J. Toluene diisocyanate-inducedasthma. II. Inhalation challenge tests and bronchial reactivity studies. Clin Allergy 1979;9:7. 9. Burge PS, O’Brien IM, H&es MG, Pepys J. Occupational asthmadue to inhaled carmine. Clin Allergy 1979;9:185. 10. Guerin C, Deschamps0, Guillot YL, Chavaillon JM, Kalb JC. A propos d’un cas d’asthme a l’anhydride maleique. Poumon Coeur 1980;36:393. 11. Burge PS, O’Brien IM, Harries MG, Pepys J. Occupational asthma due to soldering fluxes containing colophony (rosin, pine resin). Eur J Resp [Suppl] 1982;123:65. 12. Benson MK. Bronchial hyperreactivity. Br J Dis Chest 1975; 69:227. 13. Tattersfield AE. Measurementof bronchial reactivity: a question of interpretation. Thorax 1981;36:561. 14. Nagy L, Lee TH, Kay AB. Neutrophil chemotacticactivity in antigen-inducedlate asthmaticreactions. N Engl J Med 1982; 306:497. 15. Durham SR, Lee TH, Cromwell 0, Shaw RJ, Merrett TG, Merrett J, CooperP, Kay AB Immunologic studiesin allergeninduced late-phase asthmatic reactions. J ALLERGY CLIN IMMUNOL 1984;74:49.
16. Durham SR, Carroll M, Walsh GM, Kay AB. Leukocyte activation in allergen-induced late-phase asthmatic reactions. N Engl J Med 1984;311:1398. 17. Venge P, Dahl R, HankanssonL, PettersonC. Generation of
J. ALLERGY
CLIN. IMMUNOL. FEBRUARY 1987
heat-labile chemotactic activity in blood after inhalation challenge and its relationship to neutrophil and monocyteimacrophage turnover and activity. Allergy 1982;37:55. 18. Gerblich AA, Campbell AE, Schuyler MR. Changes in Tlymphocyte subpopulations after antigenic bronchial provocation in asthmatics.N Engl J Med 1984;310:1349. 19. Booij-noord H, de Vries K, Sluiter HJ, Orie NGM. Late bronchial obstructive reaction to experimental inhalation of house dust extract. Clin Allergy 1972;2:43. 20. Durham SR, Kay AB. Eosinophils, bronchial hyperreactivity. and late-phaseasthmaticresponses.Clin Allergy 1985;15:411. 21. de Monchy JGR, Kauffman HF, Venge P, Koeter GH, Jansen HM, Sluiter HJ, de Vries K. Bronchoalveolar eosinophilia during allergen-inducedlate asthmaticreactions.Am Rev Respir Dis 1985;131:373. 22. Empey DW, Laitinen LA, Jacobs L, Gold WM, Nadal JA. Mechanisms of bronchial hyperreactivity in normal subjects after upper respiratory tract infection. Am Rev Respir Dis 1976;113:131. 23. Laitinen LA, Elkin RB, Empey DW, JacobsL, Mills J, Gold WM, Nadel JA. Changesin bronchial reactivity after administration of live attenuatedinfluenza virus. Am Rev Respir Dis [SuppI] 1976;113:A194. 24. Golden JA, Nadel JA, BousheyHA. Bronchial hyperirritability in healthy subjects after exposure to ozone. Am Rev Respir Dis 1978;118:287. 25. .Cockroft DW, Murdock KY. Bronchial responsivenessto inhaled histamine is increased7 and 30 hours but not 2 hours after allergen inhalation in subjects with dual asthmatic responses[Abstract]. Clin Invest Med 1985;8:44. 26, Boulet LP, Carder A, Thomson NC, Roberts RS, Dolovich J, Hargreave FE. Asthma and increasesin nonallergic bronchial responsivenessfrom seasonalpollen exposure.J ALLERGYCLIN IMMUNOL 1983;71:399.
AVAILABLE NOW! The PROCEEDINGS OF THE INTERNATIONAL CONGRESS OF ALLERGOLOGY AND CLINICAL IMMUNOLOGY can be purchasedfrom the Publisher. This collection of “state-of-the-art” presentationsfrom the XII Congressheld October 2025, 1985, in Washington, D.C., brings together the current advancesin basic and applied aspectsof allergy and allergic diseases.It includes 528 pagescovering such topics as IgE, roles of the different cell types and their products, clinical problems, asthma, rhinitis, and reactions to foods and drugs and occupational agents, collected and reviewed by Editor Charles E. Reed, M.D. (U.S.A.) and Associate Editors Joseph Bellanti, M.D. (U.S.A.), Robert J. Davies, M.D. (U.K.), Sidney Friedlaender,M.D. (U.S. A.), Albert Oehling, M.D. (Spain), and Raymond G. Slavin, M.D. (U.S.A.). To purchase, call or write: The C. V. Mosby Company, 11830 Westline Industrial Dr., St. Louis, MO 63146, or telephone FREE l-800-325-4177, Journal Fulfillment, ext. 531 (in Missouri call collect at 314-872-8370, Journal Fulfillment, ext. 531). Prepayment required. Make checkspayable to The C. V. Mosby Company. (All paymentsmust be in U.S. funds drawn on a U.S. bank.) Price: $36.50 in the U.S., $40.50 in Canada, and $41.50 international (surface shipping chargesincluded).
J. ALLERGY
42. Pienkowski MM, Norman PS, Lichtenstein LM. Suppression of late-phaseskin reactions by immunotherapy with ragweed extract. J ALLERGYCLIN IMMUNOL 1985;76:729. 43. Peltre G, Lapeyre J, David B. Heterogeneity of grass pollen allergens(Dacrylis glomerura) recognizedby IgE antibodiesin
CLIN. IMMUNOL. FEBRUARY 1987
humanpatients seraby a new nitrocellulose immunoprint technique. Immunol Lett 1982;5:127. 44. Sutton R, Wrigley CW, Baldo BA. Detection of IgE- and IgGbinding proteins after electrophoretic transfer from polyacrylamide gels. J Immunol Methods 1982;52:183.
The temporal relationship between increases in airway responsiveness to histamine and late asthmatic responses induced by occupational agents Stephen R. Durham, M.R.C.P., * Bernard J. Graneek, M.R.C.P., Rosemarie Hawkins, S.R.N., and Anthony J. Newman Taylor, M.Sc., M.R.C.P., M.F.O.M. London, England The temporal relationship between increases in airway responsiveness and the late asthmatic response was assessed in nine patients challenged with occupational agents toluene diisocyanate (one patient), carmine (one patient), maleic anhydride (two patients), colophony (four patients), and trimellitic anhydride (one patient). The provocation concentration of histamine causing a 20% decrease in FEV, (PC,,) was measured before challenge and at approximately 3 hours and 24 hours on control and active-challenge days. Thirteen active challenges provoked eight dejinite late asthmatic responses (maximum fall in FEV, > 15% at 3 to 11 hours). At 3 hours after the challenges that provoked late responses, there was a significant (p < 0.02) decrease in PC,, that was more (p < 0.03) than that observed for the3ve tests provoking early (late FEV, fall 0% to 5%) or equivocal late (FEV, fall 6% to IS%) responses. At 24 hours, PC,, remained decreased (p < 0.05), although it was less so than at 3 hours (p < 0.05) and not signijTcantly when compared with challenge tests causing single early or equivocal late responses. The 3-hour decreases in PC,, were identified when FEV, (jive of seven observations) was MO% of prechallenge values. For the nine independent tests, the j-hour decreases in PC,, correlated (r = 0.72; p < 0.05) with the magnitude of the late falls in FEV,, whereas this was twt observed at 24 hours (r = 0.35; p, not significant). These results demonstrate that in subjects challenged with occupational agents, increases in histamine airway responsiveness precede the late asthmatic response, occur independently of changes in airway caliber, and correlate with the magnitude of the subsequent late response. We suggest that in subjects with occupational asthma, the late asthmatic response depends on the tissue events, possibly airway inJlammation, that underlie the preceding increase in airway responsiveness. (J ALLERGYCLIN hfMUNOL 1987;79:398-406. )
Airway hyperresponsiveness,an increased bronchoconstrictor response to nonspecific provocative I From the Department of Occupational Medicine, Cardiothoracic Institute, Brompton Hospital, London, England. Received for publication Aug. 12, 1985. Accepted for publication Aug. 16, 1986. Reprint requests: A. J. Newman Taylor, M.Sc., Dept. of Occupational Medicine, CardiothoracicInstitute, Brompton Hospital, Fulham Rd., London 3W3 6HP England. *Current address:JohnRadcliffe Hospital, Off Headley Way,Headington, Oxford, U.K.
I 1
Abbreviations used Provocation concentrationof histamine causGl: ing a 20% decreasein FEV, TDI: Toluene diisocyanate
stimuli, is now considered a central feature of bronchial asthma. The association between increases in airway responsivenessand late asthmatic responses has been observed after inhalation of both aller-
VOLUME NUMBER
79 2
Airway
responsiveness
to histamine
TABLE I. Clinical features of nine subjects studied Age Patient
Ckcupational agent
IV)
Sex
41
M
TDI
18 48
M M
21
M
18 46 40 39 37
M M M F M
Carmine Maleic anhydride Maleic anhydride Colophony Colophony Colophony Colophony Trimellitic anhydride
399
--__-Duration of exposure Ml
o=-
Worker in printing ink factory Batch weigher Storeman
FM,titemlsec I% pm)
PC& ~m%imll
16
3.2 (98)
24
1 17
3.8 ! loo) 3.2 t88,
1.7 X.0
Batch weigher
1
4.3 il II)
3.0
Solderer Solderer Solderer Solderer Maintenance worker
3 0.5 10 3 20
4.5 (105) 2.9 (89) 2.2 (67) 2.1 (100) 4.3 !I 14)
1.1 1.8 0.68 2.3 18.5
Duration of exposure representstime interval between first exposure and onset of work-related respimtory symptoms. S@ects 3. 4, 5, 7, and 9 were atopic (skin prick tests positive to one or more of the following allergens: Derf?m&@#agoi&spterqM#z~. mix& grass dlens, cat fur, or dog hair), and subjects 1, 2, 6, and 8 were nonatopic. Subjects 1, 3, 6. 8, and 9 were exsmokers, and subjects 2, h, 5. and 7 had neversmoked
gens’.’ and occupational agents3 These increases in airway responsiveness were observed at 24 hours after challenge, i.e., after the development of the late response. The consistency of this observation has led to the suggestion4.’ that the increase in airway IXsponsivenessresults from mechanismsthat underlie the late asthmaticresponse,possibly as a consequence of bronchial inflammation. In this study, in subjects challenged with occupational agents, we have sought to clarify the temporal relationship between the late asthmaticresponseand the associatedincreasein air-
way responsiveness.We have measuredairway responsivenessto inhaled histamine before challenge with occupational agents, at 3 hours (before the late asthmatic response), and at 24 hours (after the late asthmatic response), and related the changes in responsivenessto the magnitude of the late response.
T’heclinical featuresof the nine patients (eight male patients and one female patient) studied are presentedin Table 1. The inhalation tests were performed as part of the routine investigation for possible occupational asthma. At the time of the investigations. the subjectswere asymptomatic, and with one exception, patient 7, the baseline FEV, was ~35% of the predicted normal value. Bronchodilators were withheld for at least 24 hours before the challenge tests. With one exception, none of the subjectshad taken disodium cromoplycate or corticosteroids in the week befan the study. The exception, patient 6, had taken regular beclomethasonedipropionate, 250 p,g (one puff from a metered-dose inhaler). daily. until the day before his admission.
The patients were admitkd lo the hospital for a period of 1 to 2 weeks. On day 1, the p8ticnt.swere instmcted in the performanceof nptatpble forced enpiratory maneuvers with a dry wedge-bellows spirometer, (N&graph Ltd., Bucks, U.K.). On day 2 the p&en& had a “control” challenge in which the patients’workexposureswee simulated
in the absenceof theOccupatiOgnl agentsuspcckdto cause asthma.On subsequentdays testswen madewilh increasing exposureslo the suspectedagent. On control and challenge days. the patients’ nonspecific bronchial responsivenesswas measuredby a histamine provocation test. The histamine tests were performed on at least three occasionsduring the day: before the occupatioualexposuretests(generally 10:00 A.M.), at approximately 3 hours after the test (generally 2:00 P.M.), and at 22 to 24 hours (at IO:00 A.M. the following day). In four of the nine p&ents tested, one further histamine test was performed at 6 to 9 hours after the occupational exposure.
The histamine tests were performed by use of a tidal breathing mctl~& and a Wright nebulizer. The nebulizer was calibrated to produce 0.14 mllmin. The duration of inhalation of histamine was stand&i& af 2 minutes. Histamine responsivenesswas defined asPC, and Wasobtained by linear interpolation of the log histamine FEV, dose-response curve. A PC,,,of 8 mg/ml was taken as the cut point between normal and increa.& histamine airway responsiveness.Histamine tests were performed before challenge and at approximately 3 hours and 24 hours after challenge on control days and after exposure to the suspected occupational agent. The precise timings of the histamine tests (when the first dose of histamine was inhaled) are reprcsented in Figs. 1 to 4. The timings of the last dose of
400 Durham et al.
TOLUENE
J. ALLERGY
DIlKXYANATE
FIG. 1. The time course (0 to 24 hours) of changes in FEV, (symbols o, l , A, and n , liters per second) and PC, (solid bars, milligrams per milliliter) in a subject exposed for 30 minutes to increasing concentrations of TDI (0 to 0.02 parts per million). The magnitude and timing of the early (0 to 60 minutes) and late (3 to 11 hours) maximal percent decreases in FEV, are indicated. The hatched bars represent the periods of challenge.
histamine (when the measurementof histamine responsiveness was made) occurred up to 30 minutes after the times indicated. The occupational-typebronchial provocation procedures’ were performed in an isolation cubicle, dimensions 1.5 by 1.3 by 2.2 meters, fitted with an exhaust-extraction fan. The patients wore gowns, caps, gloves, and plastic overshoes. These occupational-type exposures on control and active challenge days were performedin a single-blind fashion in which the patients had no knowledge of the protocol used. The responsewas measuredas FEV, at 5, 10, 15, 30, and 60 minutes, hourly from 1 to 12 hours (or until the patient retired to bed), and on the next day until 22 to 24 hours after challenge. The bestof the threeforced expiratoty maneuversat each time point was used in the subsequent analysis. The incremental exposuresto increasing concentrations (or durations of exposure) of the suspectedagent wereperformedon separatedaysand only when the patients’ histamine responsivenesshad returned to baseline (control day) values. A positive early asthmaticresponsewas defined as a decreasein FEV, that occurred within 1 hour of the test of >15% from the mean of two prechallenge values
CLIN. IMMUNOL. FEBRUARY 1987
and that was >15% from the correspondingtime value on the control day. A positive late asthmatic response was defined as a decreasein FEV, between3 hours and 11 hours after the test of >15% from the prechallenge and corresponding control day values. A late asthmaticresponsewas considered equivocal when a reduction in FEV, of 6% to 15% was observed. Exposure to TDP was performedas follows: The patient first brush painted 100 gm of marine-basevarnish for a period of 30 minutes (control day). On the subsequentchallenge days, he painted varnish to which increasing amounts of TDI, previously determinedto produce reproducible air concentrationsof 0.005 ppm to 0.02 ppm within the cubicle used for provocation, were added (TDI meter, model 7005FR, Sieger Ltd., Poole, Dorset, U. K.). Carmine is a natural brilliant red dye in the form of a powder extractedfrom the dried femaleinsectCoccuscactus and is usedwidely in the cosmeticsindustry.’ On the control day, the patient suspectedof sensitivity to carmine (patient 2) tipped 250 gm of oven-dried lactosepowder mixed with amaranth B.P.C. (Evans Medical, Ltd., Dinistable, Bedford, U. K.), an unrelated dye of similar color to carmine. On the challenge days, he tipped 250 mg of lactose mixed with 0.1% and 0.3% carmine for a period of 60 seconds. The patientssuspectedof sensitivity to maleic anhydride”’ (subjects3 and 4) tipped 250 gm of lactose alone (control day) and mixed with maleic anhydride in the form of crystals ground to a powder and addedin increasing concentrations to 1% by weight for 5 minutes. The patients suspectedof sensitivity to colophony” (subjects5,6,7, and 8) performed manual soldering with solid solder wire (60/40 tin/lead alloy) (control day), and with multicore solderwire containing colophony flux for maximum periods of 15 minutes, 15 minutes, 1 minute, and 3 minutes, respectively. The subject challenged with trimellitic anhydride tipped 250 gm of lactose powder alone (control day) and mixed with trimellitic anhydride 1% for 5 minutes.
Analysis Statistical analysis of the changesin PC, was performed on log-transformeddata. For the subjectswith definite late asthmatic responses(maximal FEV, percent decreaseof > 15% at 3 to 11 hours), the changesin histamine PC, at 3 hours and 24 hours after challenge (log prechallengePC, minus log postchallengePC,) were comparedwith the corresponding values on the control days by use of the Wilcoxon signed-ranktest. The changesin PC,, at 3 hours and 24 hours after challenge for the definite late responderswere aIs0 comparedwith corresponding values in subjectswho developedeither single early (late FEV, fall of 0% to 5%) or equivocal late responses(late FEV, of 6% to 15%) by use of the Mann-Whitney U test. Spearman’srank correlation was used to comparethe changesin histamine P&, at 3 hours and at 24 hours with the magnitude of the late falls in FEV, for the challenge tests. Becausemore than one challenge was performed in three of the patients, we randomly selectedonly one of the results from each of these
VOLUME NUMBER
Airway
79 2
MALEIC
CARMINE
responsiveness
to histamine
401
ANHYDRIDE
hours
Pi& 2. The time course (0 to 24 hours) of changes in FEV, (symbols 0, l , and A, titers mr second) and PCnr(solid bars, milligrams per milliliter) in a subject exposed far 1 minute to ti&ng v&h 250 gm of lactose alone and mixed with carmine (0.1% and 0.3%), and in a sub@@ exposed kr 5 minutes to tipping with 250 gm of lactose powder alone and mix8d with meleic an~yd~e (0.2% and 1%). The magnitude and timing of the early (0 to 60 minute91 and late (1 to 12 hm} maximal percent decreases in FEV, a& indicated. The hatched bars represent the periods of challenge. MALEIC
COLOPHONI
ANHYDIUDE
6.0 2.0 0.5 0.125
8.0 2.0 0.5 0.125 Before
0 60 c :: mins
3
5
7
9
11
22
24 *
hours
FIG. 3. The time course (0 to 24 hours) of changes in FEV, (Eymbols o and 0, liters per SIBWI~) and PC, {solid bars, milligrams per milliliter) in a subject exposed for 5 minutes to t&p&g w&h 250 gm of lactose powder alone and mixed with maieic anhydride 1%. and in a subjaa who performed for 5 minutes manual soldering with solid colder wire (tindad altoy, control &allenge) and with multicore solder wire containing c&phony flux. The magnitude end timing of the early (0 to 60 minutes) and late (1 to 12 hours) maximal Percent decreases in FEV, ar. indicated. The hatched bars represent the periods of challenge. three patients in estimating correlation and its significsnce. P values CO.05 were considered statistically sign&ant.
The individual values of FJZV, and PC, for the subjects challenged with occupational agents are rep-
resented in Figs. 1 to 4. On the cm&o1 days for all subjects, the FEV, did no& change by > k 10% of the
prechallenge values. ‘II&teen active chaknges were performed in nine subjects. Three chakqges resulted in a single early asthmatic response. Ten chellenges resulted in an equivocal or definite late response. The responses were arbitrarily divided into thme categories
J. ALLERGY
402 Durham et al. COLOPHON-f
COLOPHONY
Before
0 60 t 3: mlns
3
5
7
9
11
22
24
hours
0 <
60 ::
IllitlS
3
5
7
9
Before
0 60 < :< mins
3
5
7
9
11
22
24
hours
TRIMELLITIC! ANHYDRIDE
COLDPHONY
Before
CLIN. IMMUNOL. FEBRUARY 1987
11
22
24 >
hours
FIG. 4. The time course (0 to 24 hours) of changes in FEV, (symbols o and l , liters per second) and PCs,(solid bars, milligrams per milliliter) in three subjects who performed manual soldering (patient 6, 15 minutes; patient 7, 1 minute; patient 8, 3 minutes) and in one subject (patient 9) exposed for 5 minutes to tipping with 250 gm of lactose powder alone and mixed with trimellitic anhydride 1%. The magnitude and timing of the early (0 to 60 minutes) and late (1 to 12 hours) maximal percent decreases in FEV, are indicated. The hatched bars represent the periods of challenge.
according to the magnitude of the late (3 to 11 hours) falls in PEV, as follows: (1) no late response (late PEV, decrease0% to 5%, asobservedin three subjects with single early responses), (2) equivocal late response(late PEV, decrease6% to 15%, observed as part of a dual responsein two challenges), and (3) definite late response(late PEV, decrease>15%, as observed in eight challenges, either “isolated late” in four or as part of a “dual” responsein four challenges)(Table II). The absolutevalues for PC, before challenge and at 3 hours and 24 hours after challenge (together with the FEV, immediately before the histamine tests in parenthesis)are recorded in Table II. Thirty-nine of the total 43 postchallenge histamine teats were performed when the PEV, was within f 10% of the prechallengevalues. With the marginal exception of subject3, the pre- andpostchallengePC,,
values on the control days were within a twofold histamine concentration difference. Similarly, the prechallenge PC,, values on control and active challenge days were within a twofold concentration difference. There was a wide range of baseline histamine responsiveness. The prechallenge PC, varied from moderately severely increased to within the normal nonasthmatic range (0.68 to 24 mg/ml). There was no significant correlation between the prechallenge PC, and the tendency to develop either an early or late asthmatic response. PC, was measuredat 3 hours after challenge in seven of the eight challenges that provoked definite late responses.PEV, was X0% of the prechallenge value at this time in five of the seventests. After all sevenchallenges, a greater than fourfold reduction in PC, was observed. These 3-hour reductions in PCm
VOLUME NbMBER
Airway responsiveness to histamine
79 2
TABLE II. PC, in subjects -
challenged
with occupational
agents
-----.
Maximal FE&% deerease postchallenge occupatbnel &W
agent
Control TDI 0.005 TDI 0.01 TDI 0.02 Control Carmine 0.1% Carmine 0.3% Control Maleic anhychide 0.2% Maleic anhydride 1% Control Maleic anhydride 1% Control Colophony 15 min Control Colophony 15 min Control Colophvny 1 min Control Colvphony 3 min Control Trimellitic anhydride 1%
403
.-.________
PC, (mglmi) --.----
TYP of
24.hr’ poet-
Late (3 to 11 hr)
PrecheiiQnge
3-hr* postcheilenge
3 2 8 1 16
1 31 22 30 0 I2
24 (3.17)t >I6 (3.35) >I6 (3.16) 20 (3.37) 1.7 (3.74) I .65(3.53)
24 2.9 1.7 ND 2.h 0.76
39
30
2.6 (4.0)
0.5
1 0
8 31
8.0 (3.20) 3.3 (3.27)
7.9 (3.35) 0.43 (3.201
Dual
30
23
6.4 (3.57)
Control Dual (equivocal late) Control Dual
I 24
3 14
3.0 (4.31) 3.9 (4.42)
6.0 (4.50) 0.53 14.1)
5.2 (4.30) 0.38 (4.4)
4 25
3 16
1. I (4.45) I .25(4.4)
1.4 (4.3) 0.25 (4.1)
1.25 (4.4) 0.6 (4.35)
Control Dual
4 24
9 24
1.8 (2.93) 2.7 (2.70)
1.0 (2.78) 0.61 (2.21)
I .O (2.63) 0.58 12.80)
Control Single early
6 28
6 3
0.68(2.23) 1.3 (2.15)
0.64 (2.17) 0.70 (2.17)
I.3 (2.30) I .29 (2.30)
Control Single early
7 30
5 5
2.3 (2.1) 2.2 (2.35)
1.7 (2.35) 1.5 (2.35)
2.2 (2.35) 1.8 (2.30)
Control Single early
3 44
6 4
18.5 (3.9) 14.5 (3.73)
14.0 (3.99) 9.1 (3.5)
14.5 (3.95) IO. 1 (3.90)
asthmatic reaction
Control Late Late Late Control Dual (equivocal late) Dual Control Late
Early (0 to 60 min) 0
(3.28) (3.26) (1.lSi
chatfetsge 3 ih
(3.35)
13.w (3.22)
.>lh II 0.9 I.5 I .O
(3.15) i3.31) (2.X5) (3.71) (3.62)
(3.30,
0.9
i3.0)
3.8 (3.6) 1.27 (3.09) 1.9 (3.44)
PC, (milligrams per milliliter) before challenge and at 3 hours and 24 hours after challenge are compared with the type of asthmatic responseand the maximum percent decreasein FEV, at 0 to 60 minutes and at 3 to 11 hours after challenge. *The 3-hourpostchallengePC, measurementswere performedat 2% to 3’/2hours(exceptfor subject5 at 4 hvurs). The 24.hour measurements were perfvrmcd at 22 to 24 hours. The precise timings for the individual subjects are representedin Figs. 1 to 4. *The values in parenthesesrepresentthe baseline FEV, (liters per second)recorded immediately before the measurementsof PC,,..
preceded
the development
of the late response and
were significant when they were compared with the control day reductions (p < 0.02) and with the corresponding 3-hour values in the five challenges that provoked equivocal late (two) or single early (three) asthmatic responses (p < 0.03). PC, was measured at 24 hours after challenge after eight challenges provoking definite late responses. A reduction in PC,, was observed after seven of the eight challenges. These 24-hour reductions in PC,, were
significant
when reductions
were compared
with re-
ductions on control day (p -C 0.05), although the magnitude of the changes were less than those observed at 3 hours in the same subjects (p < 0.05) and were not significant when these were compared with the corresponding 24-hour values for the five challenges provoking equivocal late or single early asthmatic responses. The magnitude of the changes in PC?, at 3 hours and at 24 hours (expressed as the ratio prechallenge
404
Durham
3o
et al.
J. ALLERGY
correlation was observed at 3 hours (r = -0.38) or at 24 hours (r = -0.05) after challenge.
80
A
10
25
0
DISCUSSION
8
0.5 1 2 4 8 16 Ratio change in histamine PC20 at 3 hours (pre-challenge PC2O/post-challenge PC20) 30@.
Oo
1 25-
0
0
0
0
2015-
0 0 0
lo5-
24 hours .0
CLIN. IMMUNOL. FEBRUARY 1987
r = 0.35 p = NS
0 I I I I I 0.5 1 2 4 8 16 Ratio change in histamine PC20 at 24 hours (pre-challenge PC20/ post-challenge PC20) 0
FIG. 5. A, Ratio change in PCzO(prechallenge PC&/postchallenge PCJ at 3 hours. 8, Ratio’change in PC& (prechallenge PC,/postchallenge PCJ at 24 hours compared with maximum FEV, percent decrease during late period (3 to 11 hours postchallenge). Correlation coefficients were obtained by Spearman’s rank correlation on data from independent challenges (0); data from separate challenges on the same patients (0) were not used for estimation of the correlation coefficients.
PC,,/postchallengePC,) wascomparedwith the magnitude of the late falls in FEV, provoked by the challenge tests (Fig. 5). For the nine independentobservations, there was a significant correlation between the magnitude of the decrease in PC,, at 3 hours and the magnitude of the late response (r = 0.72; p < 0.05), whereas this was not observed for the changes in PC,, at 24 hours (r = 0.35; p, not significant). When the magnitude of the early asthmatic response(maximal percent decreasein FFV, 0 to 60 minutes after challenge) was compared with the changes in histamine responsiveness,no significant
We have found that occupational agents provoked an increasein airway responsivnessto histamine that preceded the development of the late asthmatic response. These early increases in responsivnessappeared to occur independently of changesin airway caliber and were related to the magnitude of the subsequentlate falls in FEV,. Furthermore, in three subjects, increasingintensity of exposureto the provoking agentresulted in an earlier occurrenceof the late asthmatic response. Taken together, these observations suggestthat the underlying tissue events causing the late responsecan begin before the late responsebecomesevident. These studies involved a controlled series of observations in which the changesin histamine responsivenessand FEV, on the active challenge days were comparedwith both prechallengevalues and the corresponding values on a separateday after a control challenge. The criteria we usedfor defining a positive asthmatic responseenabled us to identify significant changesin both FEV, and PC,, without subjecting the patients to undue bronchoconstriction (14% to 44% maximal reductions in FEV, during early and late responses).Thus, although somepatientsexperienced mild symptoms of cough and wheezing during these reactions, at no time was a bronchodilator required to reversetheir responses.The inhalation testswere performed in a single-blind fashion. A double-blind protocol would have been ideal but was not possible becauseof the potential hazards of uncontrolled administration of the occupational agents used. Randomization of the challenges was also consideredinappropriatein view of the necessityto establish baseline observations after a control challenge before the active challenge days. The occupational challenge procedures involved several different agents and methods of administration. The pattern of asthmatic responsesalso varied. Isolated late, dual (early and late), and single early reactions were observed. The pattern of asthmaticresponse and the changesin histamine responsiveness appearedto be unrelatedto the natureof the provoking agent used. However, the changes in histamine responsivenesswere remarkably consistent. Whatever the nature of the test exposure, the developmentof a late asthmatic responsewas precededby a reduction in PCZOat 3 hours after challenge. Interpretation of changesin bronchial responsivenessis complicated when there is a changein airway caliber.‘*, l3 In the present study, the 3-hour increases
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in histamine responsiveness for the subjects with equivocal or definite late responses were identified at a time when they were asymptomatic and their FEV, (in seven of nine observations) was >90% of the prechallenge and corresponding control day values; in two patients (subjects 1 and 3), these 3-hour increases in histamine responsiveness were observed when the FEV, was 100% and 98% of the prechallenge values. [t therefore appears likely that the changes we observed in histamine responsiveness could not be explained by changes in airway geometry. It is possible that the histamine test performed at 3 hours may itself have modified the subsequent late response. However, this appears unlikely for the following reasons: There were no late falls in FEV, after the 3-hour histamine tests performed on control days and after single early asthmatic responses. In six of nine observations in subjects with equivocal or definite late responses, the FEV, returned to pretest values before the subsequent late falls in FEV,; in the subject exposed to increasing concentrations of TDI, the timing of the late response was earlier at higher concentrations of TDI. After the highest concentration (TDI 0.02 ppm), a 3-hour histamine test could not be performed because the late response had already begun. The presence or absence of a histamine test did not appear to modify the late response in this subject. The mechanisms underlying late asthmatic responses are largely unknown. Studies of allergen-induced late asthmatic responses in humans suggest that airway inflammation may be important. Thus, the late asthmatic response has been associated with release of pharmacologic mediators,14.I5 activation of peripheral blood leukocytes,‘6- ” altered T-lymphocyte function.18 peripheral blood, “. ‘” and bronchoalveolar*’ eosinophilia. Airway hyperresponsiveness has similarly been associated with airway inflammation, since many of the agents that increase responsiveness, such as viruses”. ” and ozone,24 also cause bronchial inflammation. Airway responsiveness has previously been demonstrated to be increased after late asthmatic responses induced by allergens” * and by occupational agents.’ Taken together, these observations suggest an important link between late asthmatic responses, airway hypemesponsiveness, and bronchial inflammation. Some authors have speculated that the increased airway responsiveness that has previously been identified after the late response may result from the airway inflammation that accompanies the late response.‘, 4.’ In the present study we have demonstrated that increases in airway responsiveness preceded the late asthmatic response. These early increases in responsiveness appeared to be independent of changes in airway caliber and were identified at a time when the
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subjects were asymptomatic. Our finding that the magnitude of these early increases in airway responsiveness was significantly correlated with the magnitude of the subsequent late response does not necessarily imply a causal relationship. On the other hand, this relationship raises the possibility that the late response depends on the mechanisms that underly the preceding increase in airway responsiveness, presumably as a consequence of airway inflammation Our observations were confined to subjects with occupational asthma. A recent study by Cockroft and Murdock,” published in abstract form, suggested that histamine responsiveness was not increased at 2 hours after allergen challenge in subjects who developed late asthmatic responses. This observation is not inconsistent with our own findings. First. the measurements of histamine responsiveness were performed at 2 hours after challenge and do not exclude a possible increase in histamine responsiveness at 3 hours after allergen inhalation. Second, increases in histamine responsiveness were observed at 2 hours in two of the 12 subjects studied. Third, the magnitude of the Iate responses (mean maximal JYEV, decrease of 17%) was lower when late responses were compared with the responses of the present study group (mean maximal FEV, decrease of 26%). In summary, late asthmatic responses induced by allergens and occupational agents are closely associated with an increase in airway responsiveness to inhaled histamine. We have demonstrated that in subjects challenged with occupational agents. the increase in airway responsiveness precedes the late asthmatic response. In so far as late asthmatic responses correlate closely with occupational3 and seasonal’” asthma, these observations support a central role for increases in airway responsiveness in the induction of clinical asthma. A therapeutic implication of these findings is that pharmacoIogic agents that reverse increased airway responsiveness are likely to inhibit late asthmatic responses and be effeclive in prophylaxis of clinical asthma. REFERENCES 1. Cockroft DW, Ruffin RE, Dolovich J. Hargreavc FE. Allergeninduced increase in nonallergic bronchial reactivity Clin Allergy 1977;7:503. 2. Cartier A. Thomson NC, Frith PA, Robens R, Hargreave FE. Allergen-induced increase in bronchial responsiveness to histamine: relationship to the late asthmatic response and change in auway caliber. J ALLERGY CLIN I,u~riwor. 1982:70: 170. 3. Chan Yeung M. Lam S, Koemer S. Clinical features and natural history of occupational asthma due to western red cedar (Thuju plicata). Am I Med 1982;72:4 Ii. 4. Cockroft DW. Mechanisms of perennial allergic asthma. Lancet 1983;2:253. 5. Kaliner M. Hypothesis on the contribution ot late-phase re-
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sponsesto the understandingand treatmentof allergic diseases. J ALLERGY CLIN IMMUNOL 1984;73:311.
6. Co&oft DW, Killian DN, Mellon JJA, Hargreave FE. Bronchial reactivity to inhaled histamine: a method and clinical survey. Clin Allergy 1977;7:235. 7. Pepys J, Hutchcroft BJ. Bronchial provocation tests in etiological diagnosis and analysis of asthma. Am Rev Respir Dis 1975;112:829. 8. O’Brien IM, Newman Taylor AJ, Burge PS, Fawcett IW, Pepys J. Toluene diisocyanate-inducedasthma. II. Inhalation challenge tests and bronchial reactivity studies. Clin Allergy 1979;9:7. 9. Burge PS, O’Brien IM, H&es MG, Pepys J. Occupational asthmadue to inhaled carmine. Clin Allergy 1979;9:185. 10. Guerin C, Deschamps0, Guillot YL, Chavaillon JM, Kalb JC. A propos d’un cas d’asthme a l’anhydride maleique. Poumon Coeur 1980;36:393. 11. Burge PS, O’Brien IM, Harries MG, Pepys J. Occupational asthma due to soldering fluxes containing colophony (rosin, pine resin). Eur J Resp [Suppl] 1982;123:65. 12. Benson MK. Bronchial hyperreactivity. Br J Dis Chest 1975; 69:227. 13. Tattersfield AE. Measurementof bronchial reactivity: a question of interpretation. Thorax 1981;36:561. 14. Nagy L, Lee TH, Kay AB. Neutrophil chemotacticactivity in antigen-inducedlate asthmaticreactions. N Engl J Med 1982; 306:497. 15. Durham SR, Lee TH, Cromwell 0, Shaw RJ, Merrett TG, Merrett J, CooperP, Kay AB Immunologic studiesin allergeninduced late-phase asthmatic reactions. J ALLERGY CLIN IMMUNOL 1984;74:49.
16. Durham SR, Carroll M, Walsh GM, Kay AB. Leukocyte activation in allergen-induced late-phase asthmatic reactions. N Engl J Med 1984;311:1398. 17. Venge P, Dahl R, HankanssonL, PettersonC. Generation of
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heat-labile chemotactic activity in blood after inhalation challenge and its relationship to neutrophil and monocyteimacrophage turnover and activity. Allergy 1982;37:55. 18. Gerblich AA, Campbell AE, Schuyler MR. Changes in Tlymphocyte subpopulations after antigenic bronchial provocation in asthmatics.N Engl J Med 1984;310:1349. 19. Booij-noord H, de Vries K, Sluiter HJ, Orie NGM. Late bronchial obstructive reaction to experimental inhalation of house dust extract. Clin Allergy 1972;2:43. 20. Durham SR, Kay AB. Eosinophils, bronchial hyperreactivity. and late-phaseasthmaticresponses.Clin Allergy 1985;15:411. 21. de Monchy JGR, Kauffman HF, Venge P, Koeter GH, Jansen HM, Sluiter HJ, de Vries K. Bronchoalveolar eosinophilia during allergen-inducedlate asthmaticreactions.Am Rev Respir Dis 1985;131:373. 22. Empey DW, Laitinen LA, Jacobs L, Gold WM, Nadal JA. Mechanisms of bronchial hyperreactivity in normal subjects after upper respiratory tract infection. Am Rev Respir Dis 1976;113:131. 23. Laitinen LA, Elkin RB, Empey DW, JacobsL, Mills J, Gold WM, Nadel JA. Changesin bronchial reactivity after administration of live attenuatedinfluenza virus. Am Rev Respir Dis [SuppI] 1976;113:A194. 24. Golden JA, Nadel JA, BousheyHA. Bronchial hyperirritability in healthy subjects after exposure to ozone. Am Rev Respir Dis 1978;118:287. 25. .Cockroft DW, Murdock KY. Bronchial responsivenessto inhaled histamine is increased7 and 30 hours but not 2 hours after allergen inhalation in subjects with dual asthmatic responses[Abstract]. Clin Invest Med 1985;8:44. 26, Boulet LP, Carder A, Thomson NC, Roberts RS, Dolovich J, Hargreave FE. Asthma and increasesin nonallergic bronchial responsivenessfrom seasonalpollen exposure.J ALLERGYCLIN IMMUNOL 1983;71:399.
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