Asthma and lower airway disease
Steroids completely reverse albuterol-induced β2-adrenergic receptor tolerance in human small airways

https://doi.org/10.1016/j.jaci.2008.07.040Get rights and content

Background

Evidence suggests that chronic stimulation of β2-adrenergic receptors (β2-ARs) induces receptor tolerance that limits the efficacy of β-agonists in the treatment of asthma. The precise mechanisms that induce β2-AR tolerance remain unclear.

Objective

We sought to determine whether steroids modulate albuterol-induced β2-AR tolerance in human small airways.

Methods

β2-AR responsiveness to isoproterenol was characterized in human precision-cut lung slices (PCLSs) precontracted to carbachol after pretreatment with albuterol.

Results

Incubation of PCLSs with albuterol for 3, 6, or 12 hours attenuated subsequent isoproterenol-induced relaxation in a dose- and time-dependent manner. A 40% decrease (P < .0001) in maximum relaxation and a 45% decrease (P = .0011) in airway sensitivity from control values occurred after the maximum time and concentration of albuterol incubation. Desensitization was not evident when airways were relaxed to forskolin. Dexamethasone pretreatment of PCLSs (1 hour) prevented albuterol-induced β2-AR desensitization by increasing the maximum drug effect (P = .0023) and decreasing the log half-maximum effective concentration values (P < .0001) from that of albuterol alone. Albuterol (12-hour incubation) decreased the β2-AR cell-surface number (P = .013), which was not significantly reversed by 1 hour of preincubation with dexamethasone.

Conclusion

These data suggest that β2-AR desensitization occurs with prolonged treatment of human small airways with albuterol through mechanisms upstream of protein kinase A and that steroids prevent or reverse this desensitization. Clarifying the precise molecular mechanisms by which β2-AR tolerance occurs might offer new therapeutic approaches to improve the efficacy of bronchodilators in asthma and chronic obstructive pulmonary disease.

Section snippets

Reagents

Reagents used were carbachol, isoproterenol, low-melting-point agarose (IX-A), dexamethasone, and Ham's F-12 medium (supplemented with 2 mmol/L glutamine, 100 U/mL penicillin, 100 μg/mL streptomycin, 2.5 μg/mL Fungizone, 50 μg/mL gentamycin, and 1 mol/L HEPES [pH 7.6]). All reagents were obtained from Sigma (St Louis, Mo), unless otherwise stated.

PCLS preparation and airway function

Healthy whole lungs were received from the National Disease Research Interchange. The smallest lobe was cut free, exposing its main bronchiole, and

Carbachol induces airway luminal diameter narrowing in a dose-dependent manner

Slices were incubated with cumulative doses of carbachol and luminal narrowing was determined as shown in Fig 1, A, to determine whether agonists induce small airway narrowing in PCLSs. Carbachol abrogated airway luminal diameter with a log EC50 value of −0.39 ± 0.06 μmol/L. Additionally, the Emax (86.0% ± 3.1%) of this effect was observed at a concentration of 30 μmol/L. Carbachol dose responses inducing luminal diameter narrowing were performed at 24, 48, 72, and 96 hours ex vivo to assess

Discussion

This study is the first to demonstrate a model of β2-AR tolerance in human small airways. The short-acting bronchodilator albuterol induced a concentration- and time-dependent decrease in β2-AR activity at 3, 6, and 12 hours and at concentrations of 0.01, 0.1, and 1.0 μmol/L, with no difference in relaxation to forskolin shown after incubation with albuterol versus control values. This study has also shown that a 1-hour incubation with dexamethasone prevented the β2-AR desensitization from

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Supported by HL080676, HL064063, HL081824, and ES013508.

Disclosure of potential conflict of interest: The authors have declared that they have no conflict of interest.

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