Defective Resensitization in Human Airway Smooth Muscle Cells Evokes β-Adrenergic Receptor Dysfunction in Severe Asthma

Abstract

β2-adrenergic receptor (β2AR) agonists (β2-agonist) are the most commonly used therapy for acute relief in asthma, but chronic use of these bronchodilators paradoxically exacerbates airway hyper-responsiveness. Activation of βARs by β-agonist leads to desensitization (inactivation) by phosphorylation through G-protein coupled receptor kinases (GRKs) which mediate β-arrestin binding and βAR internalization. Resensitization occurs by dephosphorylation of the endosomal βARs which recycle back to the plasma membrane as agonist-ready receptors. To determine whether the loss in β-agonist response in asthma is due to altered βAR desensitization and/or resensitization, we used primary human airway smooth muscle cells (HASMCs) isolated from the lungs of non-asthmatic and fatal-asthmatic subjects. Asthmatic HASMCs have diminished adenylyl cyclase activity and cAMP response to β-agonist as compared to non-asthmatic HASMCs. Confocal microscopy showed significant accumulation of phosphorylated β2ARs in asthmatic HASMCs. Systematic analysis of desensitization components including GRKs and β-arrestin showed no appreciable differences between asthmatic and non-asthmatic HASMCs. However, asthmatic HASMC showed significant increase in PI3Kγ activity and was associated with reduction in PP2A activity. Since reduction in PP2A activity could alter receptor resensitization, endosomal fractions were isolated to assess the agonist ready β2ARs as a measure of resensitization. Despite significant accumulation of β2ARs in the endosomes of asthmatic HASMCs, endosomal β2ARs cannot robustly activate adenylyl cyclase. Furthermore, endosomes from asthmatic HASMCs are associated with significant increase in PI3Kγ and reduced PP2A activity that inhibits β2AR resensitization. Our study shows that resensitization, a process considered to be a homeostasis maintaining passive process is inhibited in asthmatic HASMCs contributing to β2AR dysfunction which may underlie asthma pathophysiology and loss in asthma control.

Inhibition of PI3K promotes dilation of human small airways in a rho kinase‐dependent manner

“…Phosphorylation of MYPT1 inactivates this key signalling event of calcium sensitization. We also demonstrated that endosomes from asthma‐derived HASM cells, as compared with non‐asthma HASM cells, manifest increased PI3Kγ activation and a reduction in protein phosphatase 2A activity that inhibits β 2 ‐adrenoceptor resensitization (Gupta et al , ). Collectively, these findings have implications providing a plausible hypothesis as to why β 2 agonists are less effective in Th2 airway inflammation (Duan et al , ; Aarthia et al , ; Townley, ).…”

“…The inflammatory milieu in asthma engenders AHR to contractile agonists and attenuates β 2 agonist‐induced bronchodilation (Cooper et al , ; Albano et al , ; Gupta et al , ). Previously, we showed that exposure to IL‐13 renders airways hyposensitive to β 2 ‐adrenoceptor agonist‐mediated airway dilation but not TAS2 receptor‐mediated airway dilation (Robinett et al , ).…”

Targeting PP2A and proteasome activity ameliorates features of allergic airway disease in mice

“…PP2A is the most abundant serine/threonine phosphatase in mammals, and numerous studies show that its activity is reduced in asthma . PP2A activity was impaired in airway smooth muscle cells of asthmatics compared to nonasthmatics, in peripheral blood mononuclear cells from severe asthma patients and in animal models of steroid‐resistant AHR .…”

β2-Adrenoceptor Function in Asthma

“…Increased activity of PI3K leading to the prevention of β2-receptor resensitization was found to be the main mechanism of β2-receptor dysfunction in these patients (Gupta et al, 2015).…”