Urinary Nτ-methyihistamine during early and late allergen-induced bronchial-obstructive reactions

Urinary NT-methylhistamine during early and late allergen-induced bronchial-obstructive reactions J. J. Keyzer,

Ph.D.,*

J. J. Keyzer-Udding,

H. F. Kauffman, M.D.,***

Ph.D.,**J.

and K. de Vries,

G. R. de Monchy, M.D.**

Groningen,

M.D.,** The

Netherlands

Urinary N’methylhistamine, a unique histamine metabolite, was determined in 29 patients with bronchial asthma before and after inhalation of a control solution on I day and before and after allergen inhalation on another day. Twenty-eight patients had an early bronchial-obstructive reaction, defined by a decrease in FEV, of 20% or more within I hr after allergen inhalation. A late bronchial-obstructive reaction, 3 to 8 hr after allergen inhalation, was found in 16 patients. The early bronchial-obstructive reaction was attended with a significantly increased NT-methylhistamine excretion. The fall in FEVI was correlated with the degree of bronchial hyperreactivity to histamine, but no significant correlation was found with the increase in urinary NT-methylhistamine excretion. During the late reaction there was no signi$cant difference in the change in NT-methylhistamine excretion when the change was compared with that of the control day. These results suggest that a renewed degranulation of lung mast cells during the late bronchial-obstructive reaction is unlikely. (J ALLERGY CLINIMMVNOL74:240-5, 1984.)

Allergen inhalation in asthmatic subjects may evoke both early and late bronchial-obstructive reactions.’ It is generally assumedthat the early airway obstruction can be attributed to IgE-mediated mast cell degranulation.*a3 About the mechanismunderlying the late bronchial-obstructive reaction, however, lessis known.4-6 As to the pathogenesisof this reaction, the following factors have been suggestedto be involved: deposition of immune complexes in the airways,7 releaseof preformed mast cell-dependent mediators,s’ y cellular infiltration ,6, ’ or arachidonic acid metabolites.lo In several studiess~ *l-l5 plasmahistaminehas been measuredduring allergen-provoked, early bronchialobstructive reactions as a parameter for mast cell deFrom the *Central Laboratory for Clinical Chemistry, and **Department of Allergology, Clinic for Internal Medicine, Univerand ***Department of Chronic sity Hospital, Groningen, Obstructive Lung Diseases (Head Dr. U. Chr. Berg), Beatrixoord, Haren, The Netherlands. Supported by grants 79-15 and 82-20 of The Netherlands Asthma Foundation. Received for publication Aug. 26, 1983. Accepted for publication Feb. 22, 1984. Reprint requests: J. J. Keyzer, Ph.D., Bergschot Centre for Research, P.O.B. 2176, 4800 CD Breda, The Netherlands. 240

I

Abbreviation

FEV,:

Forced

used expiratory

volume

in 1 set

granulation. Although large discrepanciesin histamine levels are reported between different studies,15 the increaseof plasmahistamine observed during the early obstructive reaction and its inhibition by disodium cromoglycate are consistentwith the concept of lung mast cell degranulation. By contrast, studieson plasma histamine during late bronchial-obstructive reactions are scarce and controversials* l*, l6 A very short half-life of plasmahistamine(approximately .l mirP) and artifacts in the plasma-histamine determination causedby leakage from basophilsduring sampling and separation of the plasma1sare the main di,sadvantagesof plasma-histamine measurements. These problems can be overcome by measuring metabolites of histamine in urine. In vitro studiesls, ls demonstrated that human and animal lung tissue have a considerablehistamine methylating capacity. In an in vivo study by use of 14C-histamine administered intrabronchially, it was demonstrated that this histamine was methylated to a higher degree than 14C-.histamine administeredintravenously.” The

VOLUME NUMBER

TABLE

74 3. PART 1

I. Clinical

Urinary

data of 29 patients

with

bronchial

NT-methylhistamine

during

bronchial

reactions

241

asthma Maximal % decrease in FEV,

Patient

1 2 3 4 5 6 I 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29

Age/Sex

17/F 38/M 25/M 23/F 22/M 22/M 30/F 27/M 15/M 33/M 23/M 29/M 31/M 25/M 16/M 26/M 281M 40/M 19/M 33/F 32/M 30/F 45/M 20/M 20/M 16/M 39/M 42/M 69/M

Allergen

House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust mite House dust House dust House dust Grass pollen Grass pollen Grass pollen Cat dander Dog dander Guinea pig dander Aspergillus fumigatus

Histamine threshold (mglml)

FEV, before inhalation (1)

8 4 32 4 4 16 32 4 4 16 2 8 8 8 2 8

2 2 16 4 16 32 64 4 16 64 32 4 32

samestudy demonstratedthat, after administration of “C-histamine intrabronchially, the main part of the labeled NT-methylhistamine was found in the urine collected during the first hour after administration, whereas NT-methylimidazole acetic acid (the main histamine metabolite) for the greater part was excreted in urine during 3 to 6 hr after the histamine administration. In view of theseresults, we have the urinary excretion of NT-methylhistamine chosen as parameter for histamine release by bronchial mast cells in the present study. So far, only one study is reported in which this parameteris usedfor following histamine releaseduring the early bronchial-obstructive reaction provoked by allergen inhalation.21 The present study was intended to assessthe histamine turnover during the early and notably the late allergen-induced, bronchial-obstructive reactions. In a group of 30 patients with bronchial asthma, NTmethylhistamine in the urine was measuredand col-

% Of the predicted value

2.95 2.10 4.60 2.85 4.10 3.05 2.45 2.85 3.90 4.10 3.90 2.35 4.00 4.00 3.40 4.20 3.55 2.90 3.00 1.60 3.25 2.45 3.10 3.30 3.25 4.80 3.10 3.15 2.40

78 62 96 88 101 86 95 72 79 102 116 70 93 95 79 96 83 82 66 58 94 95 116 70 80

101 78 108 103

Early reaction

32 49 29 46 49 38 20 14 35 20 54 20 55 46 22 27 21 41 41 48 51 20 21 43 29 34 29 33 35

Late reaction

62 31
Urine collection on control day

+ + +

-

+ +

+ + + + + + + +

+ + + + + +

+ -

lected during both reactions by use of recently developed techniques.22*23

MATERIAL

AND METHODS

Twenty-nine patients suffering from bronchial asthma (24 men and five women) with either early, late, or combined bronchial-obstructive reactions were studied. Clinical details of these patients are given in Table I. Each patient inhaled a control solution the day before the allergen provocation. None of the patients used corticosteroids regularly. All medication was withheld at least 48 hr before the control day. Bronchial hyperreactivity was measured by serial inhalations of histamine diphosphate solutions in increasing concentrations by use of a Doppel inhaler, Wiesbadener Inhalataren-Vertrieb, Wiesbaden, West Germany, as described previously.6 The lowest concentration at which a fall in FEV, of 20% or more was found was called the histamine-threshold value. Allergen challenge (except for house dust mite) was per-

242

Keyzer et al.

TABLE II. Urinary bronchial-obstructive

J. ALLERGY

NT-methylhistamine reaction

excretion

levels

before

and after an allergen-induced

CLIN. IMMUNOL. SEPTEMBER 1994

early

Whole group (n = 28) Moderate reactors (n = 14) Severe reactors (n = 14) (Decrease in FEV, > 20%) (Decrease in FEV, 20% to 35%) (Decrease in FEV, 2 35%) NT-methylhistamine excretion before allergen inhalation (mean ? SD)* Increase in N7-methylhistamine excretion after inhalation (mean 5 SD)* Significance of the increase Median histamine threshold OwW)

92.4 + 28.0

97.4 r 28.2

87.4 r 28.0

20.3 ? 24.2

19.4 k 28.7

21.2 2x 19.9

p < 0.001

p < 0.05

8

16

p < 0.005

4

*Expressed in pmolimol of creatinine. formed as described for house dust,“ by use of challenges of 1, 4, and 5 min successively at 5-minute intervals. The allergen concentrations were: grass pollen, 1000 p/ml; house dust and Aspergillus fumigatus, 5 mglml; and cat, dog, and guinea pig dander, 2.5 mg/ml. House dust-mite (Dermatophagoides pteronyssinus) extracts were inhaled in four concentrations (100, 500, 2500, and 12500 P./ml) during 1 min successively at 15-minute intervals. All allergen extracts were purchased from Diephuis Laboratories, Groningen, The Netherlands. Allergen challenge was terminated when the FEV, value decreased 20% or more in comparison with the prechallenge value. The percentage of decrease in FEV, during the late bronchial-obstructive reaction (3 to 10 hr after challenge) was calculated from the difference between the FEV, value on the control day and that on the provocation day related to the value on the control day as previously described.‘4 Both the early and the late reaction was considered to be positive when the FEV, value decreased at least 20%. Urine was collected in hourly fractions starting 2 or 3 hr before and continuing up to 9 or 10 hr after inhalation of the control solution or allergen extracts. No urine was collected from nine patients on the control day (Table I). To prevent bacterial growth, urine was voided in polypropylene vials containing 0.5 ml of 20% chlorhexidine solution. Aliquots were stored at -20” C until analysis. During the days that urine was collected, the patients avoided foodstuffs known or suspected to contain histamine such as sauerkraut, cheese, and yogurt. ” When it was necessary, diuresis was enhanced by drinking water. NT-methylhistamine was determined with isotope dilution mass fragmentography as previously described.22s 2.1The samples of an individual person were analyzed in the same series. The within-run coefficient of variation, by use of a pool urine, amounted to 2.3%. In order to permit the measurements of NT-methylhistamine in hourly urines, N7methylhistamine concentrations were expressed in relation to urinary creatinine. The normal range for urinary NTmethylhistamine is 40 to 160 pmol/mol of creatinine.2” Creatinine was determined according to the method of

Chasson et a1.Z6with a coefficient of variation (within-run), by use of a pool urine, of 1.2%.

RESULTS Fig. 1 illustrates an individual example of the relationship bnetweenthe urinary excretion of NT-me& ylhistamirneand the lung-function parameterof FEV, both on the control day and after inhalation of house dust-mite extract. Patient 11, Table I, exhibited a dual reaction after challenge. An increasein NT-methylhistamine excretion was observed only during the early bronchial-obstructive reaction. During the control day urinary NT-methylhistamine excretion demonstrated a tendency to decrease. The changesin NT-methylhistamine excretion calculated from the values of the urines voided before and after inhalation of both the control solution and allergen extracts for 19 patients with an early reaction and 11 patients with a late reaction are illustrated in Fig. 2. The urines after inhalation were those collected afi.er the l-hour period in which the maximal fall in FEVl was registered. As the inhalations started between 8 and 9 A.M., each difference in N7methylhistamine excretion was calculated with respect to the value obtained from the urine collected at 8 A.M. After the early reaction for all except two patients, an increase was observed in NT-methylhistamine excretion in comparisonto the corresponding values on the control day. The meanincrease(n = 19) was 23.8 hmol/mol of creatinine, which was highly significant (paired t test p < 0.001). The decreasein NT-methylhistamine excretion (mean 3.2 pmol/mol) after inhalation

of the control

solution

was not sig-

nificant (paired t testp > 0.1). With respectto the late reaction, both the values on the control day as on the provocation day were significantly lower than those values of the urine collected at 8 A.M. (paired t test

VOLUME NUMBER

74 3. PART 1

Urinary

Control

day

N7-methylhistamine

House

dust

mw

during

bronchial

reactions

243

lnhalatlon

4.0..

6

FIG. 1. Lung function (Table I) on the control of the inhalations.

6

10

12

14

(FEV,) and hourly dav and the house

16

16

urinary dust-mite

p < 0.01). However, NT-methylhistamine excretion on the control day demonstrated no significant difference (paired t testp > 0.5) as compared with the inhalation day, which indicated no increased histamine release during the late obstructive reaction. In Table II the mean NT-methylhistamine excretion before allergen inhalation and the increase in NTmethylhistamine excretion after allergen inhalation are listed for the whole group of patients with an early bronchial-obstructive reaction (n = 28), for the patients with a moderate reaction (n = 14), and for the patients with a severe reaction (n = 14). The normal range for urinary N7-methylhistamine excretion is 40 to 160 pmol/mol of creatinine.27 Each patient demonstrated prechallenge values within this normal range. Between the severe and the moderate reactors, no significant difference in the increase of Nr-methylhistamine excretion after provocation could be observed (coefficient of correlation between the fall in FEV, and the increase in NT-methylhistamine excretion: 0.21). However, the difference in histamine reactivity (threshold values) between the severe and the moderate reactors was significant (nonparametric test of Wilcoxon, p < 0.05). DISCUSSION Recently, plasma histamine measurements, which were hampered by a poor quality control,28 have been technically improved. r4, l5 However, histamine in the blood stream is rapidly metabolized, resulting in a half-life time of approximately 1 min,r7 and plasma histamine determinations can easily have interference by artifacts as a result of “breakage ” of basophilic

6

@

10

12

Time

(hours)

14

16

16

-

NT-methylhistamine excretion inhalation day. Thearrows

for patient 11 indicate the start

granulo~cytes. 15, 2g The extent to which plasma histamine represents basophil disruption during sampling and plasma separation procedures remains difficult to assess. In this view it is notable that measurements of histamine metabolites in urine, which can provide a reliable parameter for the histamine liberation and/or production rate, 27*3o have attracted little attention. Intrabronchially administered histamine, previously mentioned, is excreted, to a substantial degree, as NTmethylhistamine in urine.20 Therefore, in the assumption that the metabolic fate of histamine released in response to antigen challenge is the same as that of histamine administered to the bronchial lumen, selective and sensitive determination of N7-methylhistamine in urine may be considered as a parameter for lung mast cell degranulation. Our observation that, during allergen-induced early bronchial-obstructive reactions, urinary NT-methylhistamine levels are enhanced in comparison with prechallenge values is in accordance with the study of Ltiwhagen et a121 The absence of a significant correlation between the fall in FEV, and the increase in urinary N7-methylhistamine indicates that the amount of histamine released by an allergen provocation is not the sole determinant of the severity of bronchial obstruction. This is well explained by the phenomenon that the group of patients with a moderate reaction (decrease in FEV1 20% to 35%) had a significantly lower histamine reactivity than the group of patients with a more severe reaction (decrease in FEV, 235%). This observation indicates that, in addition to mast cellbound IgE, the degree of bronchial hyperreactivity is an important factor in the obstructive reaction after

244

Keyzer

et al.

J. ALLERGY

F rloO

5

t,

Early I

reaction

n,19

Late

CLIN. IMMUNOL. SEPTEMBER 1984

reaction

n=ll /

-40. control

solution

allergen

FIG. 2. Change in hourly urinary NT-methylhistamine before (at 8 A.M.) and after inhalation of the control early and late reactions, the urines after inhalation decrease in FEV, was registered. Individual values

allergen inhalation as it was previously demonstrated in other studiesU31P3j

control

solution

allergen

excretion between the urines collected solution and the allergen extracts. For both were those collected as soon as maximal and meal1 ? 1 SD are depicted.

nates from lung mast cell degranulation, it cannot be completely excluded that some allergen may reach the

be a prerequisite for the occurrence of the late asthmatic response.** 6 It can be concluded that the determination of NTmethylhistamine in fractionated urines can be of importance to assess the role of histamine liberation in asthmatic reactions. Although the interindividual increase in NT-methylhistamine excretion after allergen

circulation after inhalation. If this is correct, IgEmediated basophil degranulation may also be, at least partly, the cause of increased urinary NT-methylhis-

rameter may be used for the objective evaluation of anti-allergic drugs in a statistically accountable group

tamine,

of patients.

Although

the increased production

of histamine

metabolites after the early phase of an allergeninduced bronchial obstruction most probably origi-

In contrast to the early obstructive reaction, the late reaction is not attended with increased urinary NTmethylhistamine levels when these levels are compared with the control day (Fig. 2). This observation does not support a role of renewed mast cell degranu-

lation during the late bronchial-obstructive reaction and is in concert with two other studies.ga l2 Nagy et aL8 demonstrated enhanced circulating neutrophil chemotactic activity and hence suggested repeated, spontaneous degranulation of lung mast cells during the late response. However, the origin of the highmolecular-weight neutrophil chemotactic factor is not well documented.36 Nevertheless some degree of mast cell degranulation during the early reaction appears to

inhalation

varies considerably,

this biochemic

pa-

This investigation was performed in collaboration with the Department of Pulmonology (Head Professor Dr. H. J. Sluiter), Clinic for Internal Medicine, University Hospital, Groningen, The Netherlands. We thank Mrs. M. L. J. de Bruin and Mr. H. Breukelman for their excellent technical assistance. REFERENCES

1. van Lookeren CampagneJG, Knol K, de Vries K: House dust provocationin children. Stand J Respir Dis 50:76, 1969 2. Lichtenstein ML, Austen KF: Asthma, physiology, immunopharmacology,and treatment. New York, San Francisco, London, 1977,AcademicPress 3. AustenKF, OrangeRP: Bronchialasthma:the possiblerole of

VOLUME NUMBER

4.

5. 6.

7.

8.

9.

10.

11.

12.

13.

14.

15.

16.

17.

18.

19. 20.

74 3, PART 1

chemical mediators of immediate hypersensitivity in the patbogenesis of subacute chronic disease. Am Rev Respir Dis 112:423, 1975 Booy-Noord H, Orie NGM, de Vries K: Immediate and late bronchial-obstructive reactions to inhalation of house dust and protective effects of disodium cromoglycate and prednisolone. J ALLERGY CLIN IMMUNOL 48:344, 1971 Warner JO: Significance of late reactions after bronchial challenge with house dust mite. Arch Dis Child 51:905, 1976 Booy-Noord H, de Vries K, Sluiter HJ, Orie NGM: Late bronchial obstructive-reaction to experimental inhalation of house dust extract. Clin Allergy 2:43, 1972 Pepys J, Hutchcroft BJ: Bronchial provocation tests in etiologic diagnosis and analysis of asthma. Am Rev Respir Dis 112:829, 1975 Nagy L, Tak H, Lee MB, Kay AB: Neutmphil chemotactic activity in antigen-induced late asthmatic reactions. N Engl J Med 306497, 1982 Allen DH, Mathison DA, Wagner PD, Arroyave CM, Tan EM: Mediator release during allergen-induced bronchoconstriction in asthmatic subjects. Chest 75235, 1979 Fairfax AJ, Hanson JM, Morley J: The late reaction following bronchial provocation with house dust mite allergen. Dependence on arachidonic acid metabolism. Clin Exp Immunol 52:393, 1983 Bhat KN, Arroyave CM, Mamey SR, Stevenson DD, Tan EM Plasma histamine changes during provoked bronchospasm in asthmatic patients. J ALLERGY CLIN IMMUNOL 58:647, 1976 Kauffman HF. van der Heide S, de Monchy JGR, de Vries K: Plasma histamine concentrations and complement activation during house dust mite-provoked bronchial-obstructive reactions. Clin Allergy 13:219, 1983 Martin GL, Atkins PC, Dunsky EH, Zweiman B: Effects of theophylline, terbutaline, and prednisolone on antigen-induced bronchospasm and mediator release. J ALLERGY CLIN IMMUNOL 66204, 1980 Brown MJ, Ind PW, Causon R, Lee TH: A novel doubleisotope technique for the enzymatic assay of plasma histamine: application to estimation of mast cell activation by antigen challenge in asthmatics. J ALLERGY CLIN IMMUNOL 69:20, 1982 Ind PW, Barnes PJ, Brown MJ, Causon R, Dollery CT: Measurement of plasma histamine in asthma. Clin Allergy 13:61, 1983 Lee TH, Nagy L, Brown MJ, O’Driscoll BR, Causon R, Kay AB: Mechanism of antigen-induced late asthmatic reactions. Agents Actions 13(suppl):273, 1983 Ind PW, Brown MJ, Lhoste FJM, Macquin IM, Dollery CT: Concentration effect relationships of infused histamine in normal volunteers. Agents Actions 1212, 1982 Lilja B, Lindell SE, Salden F: Formation and destruction of i4C-histamine in human lung tissue in vitro. J ALLERGY 31:492, 1960 Lilja B, Lindell SE: Metabolism of ‘%histamine in heartlung-liver preparation of cats. Br J Pharmacol 16203, 1961 Granerus G, Bergmark J, Lijwhagen 0, Thiringer G: Metabo-

Urinary

21.

22.

23.

24.

25.

26.

27.

28. 29. 30.

31. 32.

33.

34. 35.

36.

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during

bronchial

reactions

245

lism ‘of i4C-histamine given intra bronchially to asthmatic patients. Allergy 35:31, 1980 Liiwhagen 0, Granerus G, Wetterquist H: Studies on histamine metabolism in allergen-induced asthma. Allergy 35:521, 1980 Keyzer JJ, Wolthers BG, Muskiet FAJ, Kauffman HF, Green A: &termination of N7-methylhistamine in plasma and urine by isotope dilution mass fragmentography. Clin Chim Acta 113:165, 1981 Keyzer JJ, Wolthers BG, Breukelman H, van der Slik W, de Vries K: Determination of NT-methylhistamine in urine by gas chromatography using nitrogen-phosphorus detection. J Chronatogr 275:261, 1983 Meurs H, Koeter GH, de Vries K, Kauffman HF: The betaadrenergic system and allergic bronchial asthma: changes in lymphocyte beta-adrenergic receptor number and adenylate cyclase activity after an allergen-induced asthmatic attack. J ALLE.RGY CLIN IMMUNOL 70:272, 1982 Keyzer JJ, Breukelman H, Wolthers BG, van der Heuvel M, Kromme N, Berg WChr: Urinary excretion of histamine and some of its metabolites in man: influence of the diet. Agents Actions (in press) Chas;ron AL, Grady HT, Stanley MA: Determination of creatinine by means of automatic chemical analysis. Tech Bull Regist Med Techn 30:207, 1960 Keyzer JJ, de Monchy JGR, van Doormaal JJ, van Voorst Vader PC: Improved diagnosis of mastocytosis by measurement of urinary histamine metabolites. N Engl J Med 309: 1603, 1983 Gleich GJ, Hull WM: Measurement of histamine: a quality control study. J ALLERGY CLIN IMMUNOL 66295, 1980 Findlay SR, Lichtenstein LM: Basophil “releasability” in patienh with asthma. Am Rev Respir Dis 12253, 1980 Keyzer JJ, Udding H, de Vries K: Measurement of NT-methmethylhistamine concentrations in urine as a parameter for histanine release by radiographic contrast media. Diagn Imaging 53:67, 1984 Gokemeyer JDM: Hyperreactiviteit van de luchtwegen. Groningen, The Netherlands, 1976 (thesis) Neijens HG, Degenhart HG, Raatgeep HC, Kerrebijn KI? Study on the significance of bronchial hyperreactivity in the bronchus obstruction after inhalation of cat dander allergen. J ALLERGY CLIN IMMUNOL 64507, 1979 Cockcroft DW, Ruffin RE, Fiih PA, Camier A, Jupiner EF, Dolovick J, Hargreave FE: Determinants of allergen-induced asthma: dose of allergen, circulating IgE antibody concentration, and bronchial responsiveness to inhaled histamine. Am Rev Respir Dis 120: 1053, 1979 Boushey HA, Holtzmann UT, Sheller JR, Nadel JA: Bronchial hyperreactivity. Am Rev Respir Dis 121:389, 1980 Cockcroft DW, Killian DN, Mellon JJA, Hargreave FE: Bronchial reactivity to inhaled histamine: a method and clinical survey. Clin Allergy 7:235, 1977 Lee ‘TH, Nagakura T, Papageorgiou N, Iikura Y, Kay AB: Excercise-induced late asthmatic reactions with neutrophil chemotactic activity. N Engl J Med 3081502, 1983