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(Chest. 1999;115:352-356.)
© 1999 American College of Chest Physicians

Effect of Infused Angiotensin II on the Bronchoconstrictor Activity of Inhaled Endothelin-1 in Asthma*

George W. Chalmers, MD; Evelyn A. Millar, MD; Stuart A. Little, MD; Malcolm C. Shepherd, MD and Neil C. Thomson, MD

* From the Department of Respiratory Medicine, West Glasgow Hospitals University NHS Trust, Glasgow, Scotland, UK. Supported by grants from Chest Heart & Stroke Scotland and the National Asthma Campaign (UK).


   Abstract
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 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Study objectives: Endothelin (ET)-1 is a potent bronchoconstrictor, and asthmatics demonstrate bronchial hyperresponsiveness to ET-1 given by inhalation. Angiotensin II (Ang II) is increased in plasma in acute severe asthma, causes bronchoconstriction in asthmatics, and potentiates contractions induced by ET-1 in bovine bronchial smooth muscle in vitro, and contractions induced by methacholine both in vitro and in vivo. We wished to examine any potentiation of the bronchoconstrictor activity of inhaled ET-1 by infused Ang II at subbronchoconstrictor doses.

Design: Double-blind randomized placebo-controlled study.

Setting: Asthma research unit in university hospital.

Patients: Eight asthmatic subjects with baseline FEV1 88% predicted, bronchial hyperreactivity (geometric mean, concentration of methacholine producing 20% fall, methacholine PC20 2.5 mg/mL), and mean age 37.1 years.

Interventions: We examined the effect of subbronchoconstrictor doses of infused Ang II (1 ng/kg/min and 2 ng/kg/min) or placebo on bronchoconstrictor responses to inhaled ET-1 (dose range, 0.96 to 15.36 nmol).

Measurements: Oxygen saturation, noninvasive BP, and spirometric measurements were made throughout the study visits. Blood was sampled for plasma Ang II levels at baseline and before and after ET-1 inhalation.

Results: Ang II infusion did not produce bronchoconstriction per se at either dose prior to ET-1 challenge. Bronchial challenge with inhaled ET-1 produced dose-dependent bronchoconstriction, but there was no difference in bronchial responsiveness to ET-1 comparing infusion of placebo with Ang II at 1 ng/kg/min or 2 ng/kg/min (geometric mean, concentration of ET-1 producing 15% fall, 5.34 nmol, 4.95 nmol, and 4.96 nmol, respectively) (analysis of variance, p > 0.05). There was an increase in systolic and diastolic BP at the higher dose of Ang II compared to placebo (mean 136/86 vs 117/75 mm Hg, respectively). Plasma Ang II was elevated following infusion of both doses of Ang II compared to placebo.

Conclusions: In contrast to the potentiating effect on methacholine-induced bronchoconstriction, Ang II at subbronchoconstrictor doses does not potentiate ET-1-induced bronchoconstriction in asthma.

Key Words: angiotensin • asthma • bronchoconstriction • endothelin


   Introduction
TOP
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
The 21-amino acid peptide endothelin (ET)-1 is produced by a number of pulmonary cells, including bronchial epithelium1 and pulmonary vascular endothelium; ET-1 is also a highly potent bronchoconstrictor in human airways in vitro.2 ET-1 has a number of properties that would suggest a pathologic role in asthma, including direct bronchoconstriction, mitogenic activity, and potentiation of cholinergic nerve function,3 although the exact nature of its role in asthma is yet to be clarified. We have demonstrated that asthmatics exhibit bronchial hyperreactivity to ET-1 given by inhalation in vivo, with a bronchoconstrictor potency around 100 times that of methacholine,4 and there is evidence for increased production of ET-1 in asthmatic airways,5 with a relationship between ET-1 in BAL fluid and airflow obstruction in asthma.6 Plasma ET-1 is increased in acute severe asthma,7 and a reduction in BAL ET-1 following treatment with oral steroids and inhaled bronchodilators has been described.8 These data together suggest a potential role for ET-1 in the pathophysiology of asthma.

Angiotensin II (Ang II) is a product of the renin-angiotensin system (RAS), and like ET-1, it is a potent vasoconstrictor which also has bronchoconstrictor activity, causing contraction of isolated human bronchial rings.9 Our group has shown that the RAS is activated in acute severe asthma with increased plasma levels of Ang II,10 ,11 but not in chronic stable asthma, and that infusion of Ang II in mild asthmatics to plasma levels found in acute severe asthma causes bronchoconstriction.10 In addition, there is evidence of synergy in bronchoconstriction between Ang II and other bronchoconstrictors, including the acetylcholine analog, methacho-line, and ET-1. Ang II potentiates methacholine-induced bronchoconstriction in human airway, both in vitro and in mild asthmatics in vivo,9 and potentiates ET-1-induced bronchoconstriction in bovine bronchial rings.12 We sought to extend this last observation, and our work on the bronchoconstrictor activity of ET-1, by examining the potential interactive effect of Ang II on ET-1-induced bronchoconstriction in mild asthmatics in vivo. Ang II infusion was used to simulate the elevation of plasma Ang II observed in acute severe asthma, but subbronchoconstrictor doses of Ang II were used to ensure that bronchoconstrictor effects were not simply additive to the effects of inhaled ET-1. ET-1 was given by inhalation as a bronchial challenge test. Interactions between potential mediators in asthma may provide important information about the pathophysiology of the condition, and we felt that the in vitro bronchoconstrictor synergism between ET-1 and Ang II required investigation in vivo.


   Materials and Methods
TOP
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Patients
Eight mild asthmatics were recruited, with stable symptoms at the time of study, and no history within the preceding month of respiratory infection, antibiotics or oral corticosteroid use. Asthma was defined according to the American Thoracic Society definition,13 baseline lung function was recorded, and nonspecific bronchial hyperresponsiveness was established using a methacholine challenge test, with all the asthmatic subjects having a methacholine concentration producing 20% fall < 8 mg/mL. Long-acting ß-agonist bronchodilators were withheld for 24 h, and short-acting ß-agonist bronchodilators for 6 h prior to each study visit. The study was approved by the West Ethics Committee, West Glasgow Hospitals University NHS Trust, and each subject gave written informed consent.

Protocol
Asthmatic subjects attended for methacholine screening test, followed by three visits for ET-1 inhalation, and infusion of Ang II at either 1 ng/kg/min or 2 ng/kg/min or placebo in a randomized double-blind fashion. The interval between visits was not fixed but was generally around 1 week. Randomization and preparation of Ang II was carried out by sterile pharmacy in our institution. On attending the laboratory, spirometry was checked, with all subjects having a prechallenge FEV1 of >= 70% predicted. After a period of rest (15 min) in a recumbent position, blood was sampled for baseline measurement of plasma Ang II, and the infusion of Ang II or placebo was started, using a syringe pump (IVAC P2000; IVAC Ltd; Hampshire, UK). After a further 30 min to reach steady state, another blood sample was taken, and the bronchial challenge with ET-1 was started. Dried purified ET-1 was reconstituted using 0.9% saline solution prior to nebulization to a concentration of 0.2 mg/mL, and administered using an air-driven dosimeter calibrated to deliver 0.006 mL/breath, with a doubling dose range for ET-1 of 0.96 to 15.36 nmol. Spirometry was checked 1, 3, 5, 10, and 15 min after each dose, and the challenge test was discontinued once a 15% fall in FEV1 had been observed, or at the maximum dose in the dosing schedule, whichever came first. We have previously reported the use of ET-1 as a bronchial challenge test, and although we observed bronchoconstriction which persisted for up to 1 h, the onset of bronchoconstriction was within 5 min in each case.4 A final blood sample for plasma Ang II was taken at the conclusion of the ET-1 challenge test (defined as a fall in FEV1 of >= 15% or on reaching the maximum dose in the dosing schedule). Pulse oximetry, noninvasive BP, pulse rate, and spirometry were monitored at regular intervals throughout the study visit, with the patient in a recumbent position throughout. Albuterol, 200 µg, was given, and spirometry was repeated to ensure that the bronchoconstriction had been reversed.

Laboratory Processing and Assays
Plasma Ang II was assayed using an in-house assay which has previously been described,14 with interassay and intra-assay coefficients of variation of < 10% in each case.

Data Handling and Statistical Analysis
Statistics were performed on a desktop computer (Apple Macintosh; Apple Computer Inc; Cupertino, CA) using a statistical software package (Minitab Statistical Software; Minitab Inc; State College, PA). Demographic factors and baseline and maximum fall in lung function parameters were analyzed using parametric statistics, plasma Ang II levels by nonparametric Mann-Whitney U test and values for provoking doses of ET-1 on each visit compared by analysis of variance with significance accepted at the 95% level in each case. Provoking concentrations of bronchoconstrictor substances are expressed as geometric mean (range). The geometric mean is used because the scale of dose increase is nonlinear, and it is obtained by calculating the antilog of the mean log provoking concentration values.


   Results
TOP
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Demographics and Lung Function
The eight subjects had a mean (± SD) age of 37.1 (± 9.8) years, and spirometry showed a mean (± SD) FEV1 of 2.72 (± 0.5) L (87.9 [± 12.8]% predicted). (Table 1 provides demographic factors and asthma treatment). All subjects had bronchial hyperreactivity with geometric mean (range) concentration of methacholine producing 20% fall in FEV1 (PC20FEV1) of 1.8 (0.47 to 8.0) mg/mL. There were no differences in mean (±SD) FEV1 at baseline on each study day (placebo, 84 [±11]% predicted; Ang II 1 ng/kg/min, 84 (±15)% predicted; Ang II 2 ng/kg/min, 80 [±16]% predicted).


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  		Table 1. Demographics, Treatment, and Bronchial Responsiveness to ET-1*

 
Plasma Angiotensin II
Median (interquartile range) plasma Ang II concentrations (at baseline, prior to, and on completion of each ET-1 inhalation) were 8.1 (6.2 to 13.8), 11.25 (6.1 to 15.8) and 5.7 (4.1 to 9.0) pg/mL after placebo; 9.3 (6.8 to 10.8), 20.8 (12.2 to 25.4), and 15.3 (12.0 to 19.6) pg/mL after Ang II 1 ng/kg/min; and 8.4 (6.3 to 11.5), 28.4 (18.4 to 51.3), and 23.8 (17.1 to 53.3) pg/mL after Ang II 2 ng/kg/min, respectively (Fig 1 —presented as mean [SEM] for clarity). Plasma levels were not different to placebo at baseline for either Ang II infusion dose, and greater than placebo pre- and post-ET-1 inhalation for both Ang II doses.



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  		Figure 1. Plasma Ang II levels (mean [SEM]) at baseline, steady-state infusion (pre-ET-1 inhalation) and post-ET-1 inhalation, during infusion of placebo, Ang II, 1 ng/kg/min, and Ang II, 2 ng/kg/min.

 
Airway Responses
Ang II infusion per se did not result in bronchoconstriction at either 1 ng/kg/min (mean [±SD] FEV1 85 [±13]% predicted) or 2 ng/kg/min (mean [±SD] FEV1 80 [±15]% predicted). ET-1 inhalation produced dose-dependent bronchoconstriction in all subjects as previously demonstrated,4 with geometric mean (range) concentration of ET-1 producing 15% fall in FEV1 (PC15FEV1 ET-1) of 3.20 (0.48 to 15.36) nmol for placebo, 3.76 (1.04 to 10.86) nmol for Ang II at 1 ng/kg/min, and 3.53 (0.64 to 11.3) nmol for Ang II at 2 ng/kg/min (Fig 2 and Table 1 ), but there was no difference in bronchial responsiveness to ET-1 (analysis of variance p > 0.05) comparing placebo infusion with the two doses of Ang II.



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  		Figure 2. PC15 FEV1 during infusion of placebo, Ang II, 1 ng/kg/min, and Ang II, 2 ng/kg/min. Log scale with geometric mean values shown for placebo and Ang II 2 ng/kg/min. There were no significant differences in bronchial reactivity to ET-1 on any of the study days.

 
BP, Pulse, and Oxygen Saturation
There were no differences between mean BP readings at baseline on each of the three study visits (placebo, Ang II 1 ng/kg/min, and Ang II 2 ng/kg/min). The higher dose of infused Ang II resulted in an increase in mean (SEM) diastolic BP compared with placebo prior to the ET-1 inhalation (82 [2.7] vs 73 [2.9] mm Hg, respectively), and an increase in both systolic and diastolic mean (SEM) BP at the end of the study visit compared with placebo (systolic BP, 136 [6.6] vs 117 [4.7]; diastolic BP, 86 [2.4] vs 75 [2.1] mm Hg, respectively). Pulse rates and oxygen saturation measured by pulse oximetry did not change significantly on any of the study days (data not presented).


   Discussion
TOP
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
We have demonstrated that inhaled ET-1 produces bronchoconstriction in mild asthmatics, but did not show any potentiation of this effect by infusion of sub-bronchoconstrictor doses of Ang II.

The lack of potentiation of ET-1-induced bronchoconstriction by Ang II was contrary to our expectations, having previously demonstrated potentiation by Ang II of ET-1-induced bronchoconstriction in bovine bronchial preparations in vitro.12 Similarly, our group has demonstrated potentiation by Ang II of methacholine-induced bronchoconstriction in both human bronchial rings in vitro and in asthma in vivo,9 although in contrast, there was no evidence of potentiation by Ang II of histamine-induced bronchoconstriction, either in human bronchi in vitro or in vivo in asthma,15 suggesting that potentiation of bronchoconstriction by Ang II may vary according to the spasmogen used.

ET-1-induced bronchoconstriction in human airways is mediated mainly by the endothelin B receptor,16 but the exact mechanism of bronchoconstriction is not known in man. In contrast to animal tissues, ET-1 appears to exert bronchoconstrictor activity in human airways directly on smooth muscle,2 without the involvement of acetylcholine, leukotrienes, histamine, or platelet-activating factor,17 although there is evidence that endothelin B receptor activation may potentiate cholinergic nerve-mediated contraction in human bronchial preparations.18 Interestingly, Ang II has also been shown to potentiate neural cholinergic bronchoconstriction evoked by electrical field stimulation19 in rabbit airway smooth muscle. The finding that methacholine-induced bronchoconstriction is potentiated by Ang II in asthmatics in vivo, while histamine and ET-1-induced bronchoconstriction are not, may therefore suggest that potentiation of bronchoconstriction by Ang II is specific to cholinergic agents. In addition, while the in vitro component of the study by Millar et al9 suggests that Ang II potentiates methacholine bronchoconstriction postjunctionally, the influence of prejunctional factors in vivo is not known and may account for differences in potentiation between different bronchoconstrictors.

Endothelin and histamine receptors in the airway are coupled to specific G proteins, with signal transduction in each case involving (among other pathways) stimulation of phospholipase C with subsequent synthesis of 1,4,5-inositol triphosphate and diacylglycerol with activation of protein kinase C.20 ,21 Similarly, cholinergic muscarinic receptors in the airways are also coupled to membrane phospholipid hydrolysis to form 1,4,5-inositol triphoshate, but there are differences in the pathways involved,21 and cross talk at the second messenger level between Ang II intracellular pathways and endothelin or cholinergic second messenger pathways may account for the differences in interactions between Ang II and endothelin or methacholine in the airways.

Ang II is a weak bronchoconstrictor, and the infusion doses of Ang II were deliberately chosen to fall below the levels required to produce bronchoconstriction per se. We found no bronchoconstriction in our subjects that could be attributed to the effects of Ang II alone, but it could be argued that potentiation of ET-1-induced bronchoconstriction might occur at higher plasma levels of Ang II. Potentiation of methacholine-induced bronchoconstriction in asthma by subbronchoconstrictor doses of Ang II was observed in a previous study from our group9 in which an increase in bronchial responsiveness to methacholine was observed in six of seven patients during infusion of Ang II at 2 ng/kg/min. Comparing plasma Ang II levels with this study showed that we achieved similar elevation in mean plasma levels of Ang II prior to the ET-1 inhalation and elevated but slightly lower levels at completion of the study for both doses of Ang II. The reasons for this small difference are not clear, and while this could account for a difference in potentiation of bronchoconstriction, we observed no evidence of potentiation in those patients whose plasma Ang II levels exceeded the mean levels observed in the previous study. The median interquartile range peak plasma levels of Ang II observed in this study (28.4 [18.4 to 51.3] pg/mL during infusion of Ang II, 2 ng/kg/min) were lower than those reported in acute severe asthma (median 56 [12 to 109] pg/mL)10 and there is therefore potential for interaction in acute severe asthma which could not be demonstrated in this study, if such interaction is dependent merely on the plasma concentration of Ang II. In the previous study, showing potentiation by Ang II of ET-1-induced bronchoconstriction in bovine airways,12 the levels of Ang II used in vitro (10-7 or 3 x 10-7 M) were higher than plasma levels in our study at baseline (around 8 x 10-12 M) or at peak levels (around 3 x 10-11 M), and higher also than plasma levels in acute severe asthma (around 5 x 10-11 M),10 and it is possible that this difference in concentration of Ang II accounts for the lack of interaction with ET-1 in asthma in vivo.

In conclusion, the role of the RAS in asthma is not fully understood, and in particular, interactions between Ang II and bronchoconstrictors that may be implicated in asthma appear to be diverse, with potentiation of the effects of methacholine, but not histamine or ET-1 in asthma.


   Acknowledgements
 
We are grateful to Dr. J.J. Morton who performed the plasma Ang II assays, and Ms. Wendy Fallon for her help in blinding of the study and preparation of the infusion solutions.


   Footnotes
 
Correspondence to: George W. Chalmers, MD, Chest Heart & Stroke Scotland Research Fellow, West Glasgow Hospitals University NHS Trust, 1053 Great Western Rd, Glasgow G12 0YN, Scotland, UK; e-mail: gchalmers@btinternet.com

Abbreviations: Ang II = angiotensin II; ET-1 = endothelin-1; PC15FEV1 ET-1 = concentration of endothelin-1 producing 15% fall in FEV1; PC20FEV1 methacholine = concentration of methacholine producing 20% fall in FEV1; RAS = renin-angiotensin system

Received for publication April 30, 1998. Accepted for publication September 10, 1998.


   References
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 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Mattoli, S, Mezzetti, M, Riva, G, et al (1990) Specific binding of endothelin on human bronchial smooth muscle cells in culture and secretion of endothelin-like material from bronchial epithelial cells. Am J Respir Cell Mol Biol 3,145-151
  2. Nally, JE, McCall, R, Young, LC, et al (1994) Mechanical and biochemical responses to endothelin-1 and endothelin-3 in human bronchi. Eur J Pharmacol 288,53-60[CrossRef][ISI][Medline]
  3. Hay, DWP, Henry, PJ, Goldie, RG (1996) Is endothelin-1 a mediator in asthma? Am J Respir Crit Care Med 154,1594-1597[Abstract]
  4. Chalmers, GW, Little, SA, Patel, KR, et al (1997) Endothelin-1-induced bronchoconstriction in asthma. Am J Respir Crit Care Med 156,382-388[Abstract/Free Full Text]
  5. Springall, DR, Howarth, PH, Counihan, H, et al (1991) Endothelin immunoreactivity of airway epithelium in asthmatic patients. Lancet 337,697-701[CrossRef][ISI][Medline]
  6. Redington, AE, Springall, DR, Ghatei, MA, et al (1995) Endothelin in bronchoalveolar lavage fluid and its relation to airflow obstruction in asthma. Am J Respir Crit Care Med 151,1034-1039[Abstract]
  7. Aoki, T, Kojima, T, Ono, A, et al (1994) Circulating endothelin-1 levels in patients with bronchial asthma. Ann Allergy 73,365-369[ISI][Medline]
  8. Mattoli, S, Soloperto, M, Maurini, M, et al (1991) Levels of endothelin-1 in the bronchoalveolar lavage fluid of patients with symptomatic asthma and reversible airflow obstruction. J Allergy Clin Immunol 88,376-384[CrossRef][ISI][Medline]
  9. Millar, EA, Nally, JE, Thomson, NC (1995) Angiotensin II potentiates methacholine-induced bronchoconstriction in human airway both in vitro and in vivo. Eur Respir J 8,1838-1841[Abstract]
  10. Millar, EA, Angus, RA, Hulks, G, et al (1994) Activity of the renin-angiotensin system in acute severe asthma and the effect of angiotensin II on lung function. Thorax 49,492-495[Abstract]
  11. Ramsay, SG, Dagg, KD, McKay, IC, et al (1997) Investigations on the renin-angiotensin system in acute severe asthma. Eur Respir J 10,2766-2771[Abstract]
  12. Nally, JE, Clayton, RA, Wakelam, MJO, et al (1994) Angiotensin II enhances response to endothelin-1 in bovine smooth muscle. Pulm Pharmacol 7,409-413[CrossRef][ISI][Medline]
  13. . American Thoracic Society. (1987) Standards for the diagnosis and care of patients with chronic obstructive pulmonary disease (COPD) and asthma. Am Rev Respir Dis 136,225-234[ISI][Medline]
  14. Morton, JJ, Webb, DJ (1985) Measurement of plasma angiotensin II. Clin Sci 68,483-484[Medline]
  15. Ramsay, SG, Clayton, RA, Dagg, KD, et al (1997) Effect of angiotensin II on histamine-induced bronchoconstriction in the human airway both in vitro and in vivo. Respir Med 91,609-615[CrossRef][ISI][Medline]
  16. Hay, DWP, Luttmann, MA, Hubbard, WC, et al (1993) Endothelin receptor subtypes in human and guinea-pig pulmonary tissues. Br J Pharmacol 110,1175-1183[ISI][Medline]
  17. Hay, DWP, Hubbard, WC, Undem, BJ (1993) Endothelin-induced contraction and mediator release in human bronchus. Br J Pharmacol 110,392-398[ISI][Medline]
  18. Fernandes, LB, Henry, PJ, Rigby, PJ, et al (1996) Endothelin B (ETB) receptor-activated potentiation of cholinergic nerve-mediated contraction in human bronchus. Br J Pharmacol 118,1873-1874[ISI][Medline]
  19. Yamawaki, I, Tamaoki, J, Yamauchi, F, et al (1992) Angiotensin II potentiates neurally-mediated contraction of rabbit airway smooth muscle. Respir Physiol 89,239-247[CrossRef][ISI][Medline]
  20. Ambar, I, Sokolovsky, M (1993) Endothelin receptors stimulate both phospholipase C and phospholipase D activities in different cell lines. Eur J Pharmacol 245,31-41[CrossRef][ISI][Medline]
  21. Hill, JS (1990) Distribution, properties and functional characteristics of three classes of histamine receptor. Pharmacol Rev 42,45-83[Abstract]



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