Beta2-adrenoceptor signaling is required for the development of an asthma phenotype in a murine model

Abstract

Chronic regular use of beta(2)-adrenoceptor (beta(2)-AR) agonists in asthma is associated with a loss of disease control and increased risk of death. Conversely, we have found that administration of beta(2)-AR inverse agonists results in attenuation of the asthma phenotype in an allergen-driven murine model. Besides antagonizing agonist-induced signaling and reducing signaling by empty receptors, beta-AR inverse agonists can also activate signaling by novel pathways. To determine the mechanism of the beta-AR inverse agonists, we compared the asthma phenotype in beta(2)-AR-null and wild-type mice. Antigen challenge of beta(2)-AR-null mice produced results similar to what was observed with chronic beta(2)-AR inverse agonist treatment, namely, reductions in mucous metaplasia, airway hyperresponsiveness (AHR), and inflammatory cells in the lungs. These results indicate that the effects of beta(2)-AR inverse agonists are caused by inhibition of beta(2)-AR signaling rather than by the induction of novel signaling pathways. Chronic administration of alprenolol, a beta-blocker without inverse agonist properties, did not attenuate the asthma phenotype, suggesting that it is signaling by empty receptors, rather than agonist-induced beta(2)-AR signaling, that supports the asthma phenotype. In conclusion, our results demonstrate that, in a murine model of asthma, beta(2)-AR signaling is required for the full development of three cardinal features of asthma: mucous metaplasia, AHR, and the presence of inflammatory cells in the lungs.

Fig. 1.

Effect of β₂-AR gene disruption and chronic administration of the inverse agonist nadolol on mucin content in the airway epithelium. (A) Mucin content in the airway epithelia of FVB/N β₂-AR^−/− and FVB/N mice was measured with PAFS from saline-challenged mice (control), antigen-challenged mice (S/C) administered with either vehicle, or antigen-challenged mice administered nadolol for 28 days (Nad 28 d). (Scale bar, 20 μm.) (B) Morphometric quantification of the mucin volume density was assessed from the various treatment groups. Values represent the mean ± SEM of data from 6–8 mice in each group. #, P < 0.05 vs. control FVB/N β₂-AR^−/− and FVB/N mice; @, P < 0.05 vs. SC vehicle-treated FVB/N β₂-AR^−/− mice; *, P < 0.05 vs. SC vehicle-treated FVB/N mice.

Fig. 2.

Effect of β₂-AR gene disruption and chronic administration of the inverse agonist nadolol on AHR. FVB/N β₂-AR^−/− and FVB/N mice were saline-challenged (control) or antigen-challenged (S/C) and administered either vehicle or nadolol for 28 days before receiving methacholine. (A–D) Values for R_aw were recorded by using a computer-controlled ventilator apparatus comparing antigen-challenged FVB/N mice (filled circles) with saline-challenged FVB/N mice (A, open circles), antigen-challenged FVB/N mice treated with nadolol (B, open squares), antigen-challenged FVB/N β₂-AR^−/− mice (C, open diamonds), or antigen-challenged FVB/N β₂-AR^−/− mice treated with nadolol (D, open triangles). (E) Values for peak R_aw were determined for each mouse by choosing the highest R_aw value produced by any of the methacholine doses (most often the next to last dose, 120 mg/mL) from the individual methacholine dose–response curves. Values represent the mean ± SEM of data from 8 mice in each group. #, P < 0.05 vs. control FVB/N β₂-AR^−/− and FVB/N mice; @, P < 0.05 vs. SC vehicle-treated FVB/N β₂-AR^−/− mice; *, P < 0.05 vs. SC vehicle-treated FVB/N mice.

Fig. 3.

Effect of β₂-AR gene disruption and chronic administration of the inverse agonist nadolol on cell count and eosinophils. Total cell count (A) and eosinophils in BALF (B) from saline-challenged (control) mice, and antigen-challenged (S/C) mice administered with either vehicle or nadolol for 28 days (Nad 28 d). BALF was collected 24 h after the last challenge. Values represent the mean ± SEM of data from 8–12 mice in each group. #, P < 0.05 vs. control FVB/N β₂-AR^−/− and FVB/N mice; @, P < 0.05 vs. SC vehicle-treated FVB/N β₂-AR^−/− mice; *, P < 0.05 vs. SC vehicle-treated FVB/N mice.

Fig. 4.

Effect of chronic administration of nadolol and alprenolol on mucin content in the airway epithelium and BALF cell counts. (A) Mucin content in the airway epithelium of BALB/c mice was measured by using PAFS from antigen-challenged (S/C) mice administered the inverse agonist nadolol (Nad), alprenolol (a β-blocker with weak agonist properties) (Alp), or nadolol and alprenolol for 28 days. (Scale bar, 20 μm.) (B) Morphometric quantification of the mucin volume density (Right) and cell counts (Left) were assessed from the various treatment groups. Values represent the mean ± SEM of data from 5–7 mice in each group. #, P < 0.05 vs. control mice; *, P < 0.05 vs. SC vehicle-treated mice.