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Plasma adenosine concentration increases during exercise: A possible contributing factor in exercise-induced bronchoconstriction in asthma

      Abstract

      We investigated whether the level of plasma adenosine (ADO) changed during exercise and whether this could be related to exercise-induced bronchoconstriction. Baseline levels of ADO did not differ, but exercise resulted in higher ADO in patients with asthma than in healthy subjects (86 ± 35 vs 59 ± 16 nmol/L; P < .001). In patients with asthma, the increase in ADO was related to decreases in FEV1 (r 2 = 0.475; P < .05) and SaO2 (r 2 = 0.693; P < .05). These data suggest that adenosine might be involved in the development of exercise-induced bronchoconstriction. (J Allergy Clin Immunol 2002;109:446-8.)

      Keywords

      Abbreviations:

      ADO: (Plasma adenosine), EIB: (Exercise-induced bronchoconstriction), HXN: (Plasma hypoxanthine), INO: (Plasma inosine), SaO2: (Hemoglobin oxygen saturation)
      Exercise induces bronchoconstriction in many patients with asthma. The mechanism of exercise-induced bronchoconstriction (EIB) is not completely understood, but available data indicate that cooling and dehydration of the airways might stimulate pulmonary inflammatory cells to release bronchoconstrictor mediators.
      • Anderson SD
      • Holzer K.
      Exercise-induced asthma: is it the right diagnosis in elite athletes?.
      Adenosine produces a dose-related airway obstruction when it is inhaled by patients with asthma but not when it is inhaled by healthy volunteers.
      • Cusley MJ
      • Tattersfield AE
      • Holgate ST.
      Inhaled adenosine and guanosine on airway resistance in normal and asthmatic subjects.
      It is likely that binding of adenosine to A3 and/or A2b adenoreceptor subtypes on immunologically activated lung inflammatory cells induces the release of bronchoconstrictor mediators.
      • Olah ME
      • Stiles GL.
      Adenosine receptor subtypes: characterization and therapeutic regulation.
      • Feoktistov I
      • Polosa R
      • Holgate ST
      • Biaggioni I.
      Adenosine A2B receptors: a novel therapeutic target in asthma?.
      Allergen-induced bronchospasm is known to be associated with an elevation in levels of plasma adenosine (ADO) in patients with asthma.
      • Mann JS
      • Holgate ST
      • Renewich AG
      • Cushley MJ.
      Airway effects of purine nucleosides and nucleotides and release with bronchial provocation in asthma.
      Furthermore, such an increase in ADO can enhance the release of bronchoconstrictor mediators during the allergen-induced immediate airway response.
      • Huszár É
      • Horváth I
      • Barát E
      • Herjavecz I
      • Böszörményi-Nagy GY
      • Kollai M.
      Elevated circulating adenosine level potentiates antigen-induced immediate bronchospasm and bronchoconstrictor mediator release in sensitized guinea pigs.
      Adenosine itself is produced by and released from exercising tissues; thus it might play a homeostatic role through its effect as a potent vasodilator.
      • Fuller RW
      • Maxwell DL
      • Conradson T-BG
      • Dixon CMS
      • Barnes PJ.
      Circulatory and respiratory effects of infused adenosine in conscious man.
      The aims of the present study were to examine whether exercise is associated with increases in ADO in patients with asthma and in healthy control subjects and to determine whether changes in ADO are related to the degree of EIB observed in patients with asthma. We measured ADO and FEV1 values in subjects at rest and after they had undergone treadmill exercise testing. To estimate the total amount of adenosine released, the plasma concentrations of the metabolites of adenosine, including inosine and hypoxanthine, were also determined. To monitor oxygen availability, hemoglobin oxygen saturation (SaO2) and plasma lactate levels were also measured in these subjects.

      Methods

      Subjects

      Nine nonatopic healthy volunteers (age, 18 to 42 years; 4 male and 5 female) and 9 patients with stable mild asthma (age, 20 to 32 years; 6 male and 3 female) participated in the study. All patients met the American Thoracic Society diagnostic criteria for bronchial asthma, and all had histories of EIB. All patients were atopic, as demonstrated by at least 2 positive results to skin tests with the following common aeroallergens: house dust mite, cat dander, dog dander, grass pollen, and Aspergillus fumigatus (Soluprick; ALK Abello, Copenhagen, Denmark). Treatment was limited to inhalation of short-acting β2 agonists and corticosteroids. β2 agonists were withdrawn at least 12 hours and steroids (4 of 9 patients) 24 hours before the study.
      The protocol was approved by the local ethics committee, and written informed consent was obtained from each subject before the study. The subjects, all of whom were nonsmokers, were instructed to avoid taking caffeinated drinks and foods and refrain from exercise for at least 12 hours before the study. No patients were studied within 6 weeks of an upper respiratory tract infection or an exacerbation of asthma.

      Study design and protocol

      All subjects completed a standardized treadmill exercise test at room temperature; the tests were administered at the same time of day, and nose clips were worn. The speed, gradient, time, and heart rate were displayed continuously during running. The exercise intensity was set to raise the subjects’ heart rate to 90% of the age-adjusted maximum target level. Each subject exercised for 6 minutes under these target conditions. There were no significant differences in fitness levels between the 2 groups.
      SaO2 was determined by means of a pulse oxymeter before and at the termination of exercise. A venous blood sample was taken and FEV1 was measured as soon as technically possible after the determination of post-exercise SaO2 (within 6 minutes). Healthy volunteers and asthmatic subjects did not differ with respect to exercise measurement time.

      Lung function test

      FEV1 was measured by means of an electronic spirometer (Medicor MS-11; Budapest, Hungary). The best of 3 consecutive maneuvers was accepted for evaluation. Each pre-exercise FEV1 value was expressed as a percentage of the predicted normal value for the subject's height and age, according to European Community for Coal and Steel reference values for FEV1.

      Determination of ADO, plasma inosine, plasma hypoxanthine, and plasma lactate concentrations

      Analysis of plasma samples for adenosine and its metabolites were carried out through use of an isocratic reverse-phase HPLC method, as described previously.
      • Huszar É
      • Barát E
      • Kollai M.
      Isocratic high-performance liquid chromatographic determination of plasma adenosine.
      Plasma lactate concentrations were determined by enzymatic analysis (Boehringer Mannheim, Mannheim, Germany). UV absorption was measured at 340 nm.
      Each result is expressed as the mean ± 1 SD. Two-by-two repeated-measures ANOVA ([SUBJECT : control/patient] × [EXERCISE : before/after]) followed by Tukey-Kramer multiple-comparison testing was performed to compare baseline and postexercise data from healthy and asthmatic subjects within and between groups. Multiple regression analysis was performed to determine the intercorrelations between changes in ADO and FEV1 or SaO2. All statistical analyses were performed through use of an NCSS 2000 computer software package (Statistical Solutions Ltd, Cork, Ireland).

      Results

      Physiologic variables before exercise

      Resting FEV1 was significantly lower in patients with asthma than in healthy volunteers (Fig 1).
      Figure thumbnail gr1
      Fig. 1Individual and mean (± 1 SD) values of FEV1 and concentrations of ADO in healthy volunteers (Control) and patients with asthma (Patient) before (B) and after (PE) exercise. Differences in FEV1% and/or ADO within groups: *P < .05; ***P < .001. Differences in FEV1% and/or ADO between groups: ΔP < .05; ΔΔΔP < .001.
      There were no differences in pre-exercise values for ADO concentration (Fig 1), plasma inosine and plasma hypoxanthine (INO + HXN) concentration, plasma lactate concentration, or SaO2 between patients with asthma and healthy volunteers.

      Effects of exercise

      After exercise, FEV1 did not change in control subjects; however, there was a decrease in patients with asthma (83% ± 9% vs 64% ± 18%; Fig 1). ADO and plasma lactate levels were increased after exercise in both healthy subjects (P < .05 and P < .001 for ADO and lactate, respectively) and patients with asthma (P < .001 for both ADO and lactate). INO + HXN was elevated in the patients with asthma (P < .001) but not in the healthy controls. These elevated levels of postexercise plasma concentrations were significantly higher in patients with asthma than in healthy controls (ADO, 86 ± 35 nmol/L vs 59 ± 16 nmol/L [Fig 1]; INO + HXN, 8.5 ± 2.9 vs 4.4 ± 2.4 μmol/L [P < .01]; lactate, 9.6 ± 3.9 vs 5.8 ± 2.0 mmol/L [P < .01]). There was a decrease in SaO2 in patients with asthma (P < .001) after exercise. This was reflected by lower postexercise SaO2 in patients with asthma than in control subjects (91.1% ± 4.5% vs 95.3% ± 1.4% [P < .01]).

      Anova main effects

      Because there were only 2 levels of the repeated-measures variable, there were no concerns regarding the nonsphericity assumption. Significant main effects of the SUBJECT factor were observed in all variables except ADO, and mean effects of the EXERCISE factor were significant in all variables (with power values [1 - β] between 0.61 and 0.99). The interactions of these 2 factors (SUBJECT /EXERCISE ) were significant as follows:
      • for FEV1: F(1,16) = 32.47; P < .00003; 1 - β = 0.99
      • for ADO: F(1,16) = 6.28; P < .023; 1 - β = 0.65
      • for INO + HXN: F(1,16) = 12.10; P < .003; 1 - β = 0.90
      • for SaO2: F(1,16) = 7.14; P < .0167; 1 - β = 0.71
      • for lactate: F(1,16) = 6.91; P < .018; 1 - β = 0.69.

      Relations between changes in SaO2, ADO, and FEV 1

      In healthy subjects, there were no correlations between exercise-induced changes in SaO2, ADO, and FEV1. In patients with asthma, however, the increase in ADO was proportional to the percent fall in SaO2(Fig 2, A ).
      Figure thumbnail gr2
      Fig. 2A, Relationship between increase in ADO and percent fall in SaO2 in patients with asthma (r 2 = 0.693; P < .05). B, Relationship between percent fall in FEV1 and increase in ADO in patients with asthma (r 2 = 0.475; P < .05).
      Similarly, the percent fall in FEV1 was proportional to the increase in ADO (Fig 2, B ).

      Discussion

      Our results indicate that exercise-induced increases in ADO and INO + HXN are significantly higher in patients with asthma than in healthy subjects. In patients with asthma, the increase in ADO was related to the decrease in SaO2; however, the degree of airway obstruction was related to the increase in ADO.
      The larger increase in ADO and its metabolites in response to exercise in asthmatic subjects was probably the result of enhanced adenosine production in these subjects in comparison with healthy volunteers. It seems likely that reduced oxygen availability might contribute to the facilitated adenosine production, inasmuch as both the fall in SaO2 and the increase in lactate accumulation were greater in patients with asthma than in healthy volunteers. Moreover, the fall in SaO2 was also related to the increase in ADO. Elevation in ADO, together with all of the other changes associated with exercise, did not cause airway obstruction in healthy controls; in contrast, FEV1 decreased both significantly and in proportion to the increase in ADO in patients with asthma.
      Although association between FEV1 and ADO does not necessarily mean a causative relationship, this finding supports the hypothesis that adenosine might participate in the development of EIB. It can be hypothesized that the increased adenosine concentration enhances the effect of physical triggers activated by exercise and that the summated effect of adenosine and physical stimulus on primed inflammatory cells induces the release of bronchoconstrictor mediators and leads to the development of airway obstruction.
      • Van Schoor J
      • Joos GF
      • Kips JC
      • Drajesk JF
      • Carpenter PJ
      • Pauwels PA.
      The effect of ABT-761, a novel 5-lipoxygenase inhibitor, on exercise- and adenosine-induced bronchoconstriction in asthmatic subjects.
      • Finnerty JP
      • Polosa R
      • Holgate ST.
      Repeated exposure of asthmatic airways to inhaled adenosine 5′-monophosphate attenuates bronchoconstriction provoked by exercise.
      This proposed mechanism is supported by our previous data obtained from guinea pigs, in which we found that ovalbumin challenge produced a greater bronchoconstrictor mediator release and airway narrowing after adenosine administration.
      • Huszár É
      • Horváth I
      • Barát E
      • Herjavecz I
      • Böszörményi-Nagy GY
      • Kollai M.
      Elevated circulating adenosine level potentiates antigen-induced immediate bronchospasm and bronchoconstrictor mediator release in sensitized guinea pigs.
      In conclusion, EIB is closely related to the increase in ADO concentration, indicating a possible role for adenosine in the development of EIB.

      Acknowledgements

      We thank N. Szilagyi, PhD, for statistical analysis; L. E. Donnelly, PhD, for grammatical correction; Mrs R. Nagy and Mrs M. Czok for lung function measurements; and Mrs M. Leisz,, Mrs M. Hernádi, and Mrs M. Mikoss for the collection and processing of blood samples.

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