The mechanisms of chronic nadolol treatment on the regulation of bronchial tone in a murine model of asthma
Statement of the problem: Using a murine model of asthma, chronic beta blocker treatment was found to improve airway hyperresponsiveness (AHR). This finding completed a paradigm shift in the treatment of asthma using β adrenoceptor (AR) ligands, which was an analogy of the paradigm shift that occurred in congestive heart failure. Therefore, we hypothesized that chronic beta blocker treatment altered the cellular signaling modulating airway smooth muscle tone. Procedure or methods: Radioligand binding assays were used to measure β AR number and further identify the subtype of β AR regulated by chronic beta blocker treatment. To evaluate the effect of chronic nadolol treatment on Gs signaling pathways, cAMP accumulation in the lung homogenates was measured by direct enzyme immunoassay; the cAMP-PKA dependent signaling was evaluated by isoproterenol induced tracheal relaxation; Gs, GRK2, GRK3, PDE4D and β arrestin 2, important proteins regulating the Gs signaling and playing important roles in lung physiology were measured by immunoblotting. To evaluate the effect of chronic nadolol treatment on Gi and Gq signaling pathways, Gαi3, Gαi2, Gq, GRK5, PLCβ1, PKCα were also measured by immunoblotting. To evaluate the effect of chronic nadolol treatment on arachidonic acid metabolism, cysteinyl leukotrines were measured in bronchoalveolar lavage fluid (BALF), while cPLA2, COX2 and 5-LO were also measured by immunoblotting. In addition, β2 AR knockout mice were treated with the same protocol as the wildtype mice to confirm that the effects of chronic nadolol treatment on PDE4D, Gαi3 and PLCβ1 were β2 AR dependent.
Results: Chronic beta blocker treatment increased β2 AR density and decreased PDE4D expression. Although chronic beta blocker treatment showed no effect on cAMP-PKA dependent airway smooth muscle relaxation, the regulation of Gi expression and Gi signaling in airway smooth muscle were found to be altered with chronic nadolol treatment. Also, chronic nadolol treatment decreased PLCβ1 expression and PKCα, 5- LO translocation. While a reduction in AHR, mucous metaplasia, and inflammatory cell counts in bronchoalveolar lavage fluid (BALF) was observed in wildtype mice, chronic nadolol treatment had no effect on PDE4D, Gi3 and PLCβ1 expression in β2 AR null mice. Conclusion: Chronic nadolol treatment decreased airway tone in a murine model of asthma by decreasing cAMP breakdown and Gi, Gq signaling. These effects are possibly dependent on its chronic blockade of β2 AR.