Prolonged heterologous β2-adrenoceptor desensitization promotes proasthmatic airway smooth muscle function via PKA/ERK1/2-mediated phosphodiesterase-4 induction

2008 ◽  
Vol 294 (6) ◽  
pp. L1055-L1067 ◽  
Author(s):  
Aihua Hu ◽  
Gustavo Nino ◽  
Judith S. Grunstein ◽  
Sumbul Fatma ◽  
Michael M. Grunstein
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Intracellular Signaling ◽  
Prolonged Exposure ◽  
Pde4 Inhibitor ◽  
Prostaglandin E ◽  
Protein Signaling ◽  
Camp Phosphodiesterase ◽  
Phosphodiesterase 4 ◽  
Β2 Adrenergic Receptor

β2-Adrenergic receptor (β2AR) agonists acutely relieve bronchoconstriction via cAMP-mediated relaxation of airway smooth muscle (ASM). Airway constrictor responsiveness may be significantly heightened, however, following protracted exposure to these agents, presumably reflecting the effects of β2AR desensitization in ASM accompanying prolonged cAMP signaling. Because cAMP phosphodiesterase (PDE) activity can significantly modulate ASM contractility, we investigated the mechanism regulating PDE expression and its potential role in mediating changes in agonist-induced constrictor and relaxation responsiveness in ASM following its heterologous β2AR desensitization by prolonged exposure to cAMP-elevating agents. Isolated rabbit ASM tissues and cultured human ASM cells treated for 24 h with the receptor- or nonreceptor-coupled cAMP-stimulating agent, prostaglandin E2 (PGE2) or forskolin, respectively, exhibited constrictor hyperresponsiveness to acetylcholine and impaired β2AR-mediated relaxation and cAMP accumulation. These proasthmatic-like changes in ASM function were associated with upregulated PDE4 activity, reflective of increased transcription of the PDE4D5 isoform, and were prevented by pretreatment of the ASM with a PDE4 inhibitor. Extended studies using gene silencing and pharmacological approaches to inhibit specific intracellular signaling molecules demonstrated that the mechanism underlying PGE2-induced transcriptional upregulation of PDE4D5 involves PKA-dependent activation of Gi protein signaling via the βγ-subunits, the latter eliciting downstream activation of ERK1/2 and its consequent induction of PDE4D5 transcription. Collectively, these findings identify that β2AR desensitization in ASM following prolonged exposure to cAMP-elevating agents is associated with proasthmatic-like changes in ASM responsiveness that are mediated by upregulated PDE4 expression induced by activated cross talk between the PKA and ERK1/2 signaling pathways.

scholarly journals Mechanism regulating proasthmatic effects of prolonged homologous β2-adrenergic receptor desensitization in airway smooth muscle

2009 ◽  
Vol 297 (4) ◽  
pp. L746-L757 ◽  
Author(s):  
Gustavo Nino ◽  
Aihua Hu ◽  
Judith S. Grunstein ◽  
Michael M. Grunstein
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Adrenergic Receptor ◽  
Pde4 Inhibitor ◽  
Protein Signaling ◽  
Altered Expression ◽  
Asthma Morbidity ◽  
Increased Risk ◽  
Enhanced Expression ◽  
Long Acting

Use of long-acting β2-adrenergic receptor (β2AR) agonists to treat asthma incurs an increased risk of asthma morbidity with impaired bronchodilation and heightened bronchoconstriction, reflecting the adverse effects of prolonged homologous β2AR desensitization on airway smooth muscle (ASM) function. Since phosphodiesterase 4 (PDE4) regulates ASM relaxation and contractility, we examined whether the changes in ASM function induced by prolonged homologous β2AR desensitization are attributed to altered expression and action of PDE4. Cultured human ASM cells and isolated rabbit ASM tissues exposed for 24 h to the long-acting β2AR agonist salmeterol exhibited impaired acute β2AR-mediated cAMP accumulation and relaxation, respectively, together with ASM constrictor hyperresponsiveness. These proasthmatic-like changes in ASM function were associated with upregulated PDE4 activity due to enhanced expression of the PDE4D5 isoform and were prevented by pretreating the ASM preparations with the PDE4 inhibitor rolipram or with inhibitors of either PKA or ERK1/2 signaling. Extended studies using gene silencing and pharmacological approaches demonstrated that: 1) the mechanism underlying upregulated PDE4D5 expression following prolonged β2AR agonist exposure involves PKA-dependent activation of Gi protein signaling via its βγ-subunits, which elicits downstream activation of ERK1/2 and its induction of PDE4D5 transcription; and 2) the induction of PDE4 activity and consequent changes in ASM responsiveness are prevented by pretreating the β2AR agonist-exposed ASM preparations with inhibitors of Gi-βγ signaling. Collectively, these findings identify that the proasthmatic changes in ASM function resulting from prolonged homologous β2AR desensitization are attributed to upregulated PDE4 expression induced by Gi-βγ-mediated cross-talk between the PKA and ERK1/2 signaling pathways.

Qualitative immunoblot analysis of PKC isoforms expressed in airway smooth muscle

1997 ◽  
Vol 272 (4) ◽  
pp. L603-L607 ◽  
Author(s):  
H. Togashi ◽  
C. A. Hirshman ◽  
C. W. Emala
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Airway Smooth Muscle ◽  
Intracellular Signaling ◽  
Immunoblot Analysis ◽  
Calcium Dependent ◽  
Kinase C ◽  
Pkc Isoforms ◽  
Dependent Activity ◽  
Trachealis Muscle

Protein kinase C (PKC) was originally identified as a single serine/ threonine protein kinase with calcium- and phospholipid-dependent activity, but more recently PKC has been found to consist of a family of multiple isoenzymes with different biochemical characteristics, substrates, and cofactor requirements. PKC is particularly important in regulating airway smooth muscle (ASM) tone. Although a previous investigation has demonstrated PKC-beta, -delta, -epsilon, -theta and -zeta in canine trachealis muscle, additional PKC isoforms have not been characterized in ASM. Therefore, immunoblot analysis using nine isotype-specific antibodies was used to further characterize the expression of PKC isoforms in porcine ASM. In addition to the previously described beta-, delta-, epsilon-, and zeta-isoforms in ASM, the calcium-dependent alpha-isoform, and the calcium- and diacylglycerol-independent isoforms iota/lambda and mu were identified. This study demonstrates multiple PKC isoforms in porcine ASM that can participate in intracellular signaling pathways in this tissue.

scholarly journals Polymorphism of the β2-Adrenergic Receptor Gene and Desensitization in Human Airway Smooth Muscle

2000 ◽  
Vol 162 (6) ◽  
pp. 2117-2124 ◽  
Author(s):  
PAUL E. MOORE ◽  
JOHANNE D. LAPORTE ◽  
JOSEPH H. ABRAHAM ◽  
IGOR N. SCHWARTZMAN ◽  
CHANDRI N. YANDAVA ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Adrenergic Receptor ◽  
Receptor Gene ◽  
Human Airway Smooth Muscle ◽  
Human Airway ◽  
Β2 Adrenergic Receptor ◽  
Adrenergic Receptor Gene

Testosterone augments β2 adrenergic receptor genomic transcription increasing salbutamol relaxation in airway smooth muscle

2020 ◽  
Vol 510 ◽  
pp. 110801 ◽  
Author(s):  
Abril Carbajal-García ◽  
Jorge Reyes-García ◽  
María F. Casas-Hernández ◽  
Edgar Flores-Soto ◽  
Verónica Díaz-Hernández ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Adrenergic Receptor ◽  
Β2 Adrenergic Receptor

scholarly journals Targeted transgenesis identifies Gαs as the bottleneck in β2-adrenergic receptor cell signaling and physiological function in airway smooth muscle

2014 ◽  
Vol 307 (10) ◽  
pp. L775-L780 ◽  
Author(s):  
Wayne C. H. Wang ◽  
Susan H. Pauer ◽  
Dan'elle C. Smith ◽  
Madison A. Dixon ◽  
David J. Disimile ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Transgenic Mice ◽  
G Protein ◽  
Airway Smooth Muscle ◽  
Adrenergic Receptor ◽  
Coupling Efficiency ◽  
The Body ◽  
Limiting Factor ◽  
Camp Production ◽  
Β2 Adrenergic Receptor

G protein-coupled receptors are the most pervasive signaling superfamily in the body and act as receptors to endogenous agonists and drugs. For β-agonist-mediated bronchodilation, the receptor-G protein-effector network consists of the β2-adrenergic receptor (β2AR), Gs, and adenylyl cyclase, expressed on airway smooth muscle (ASM). Using ASM-targeted transgenesis, we previously explored which of these three early signaling elements represents a limiting factor, or bottleneck, in transmission of the signal from agonist binding to ASM relaxation. Here we overexpressed Gαs in transgenic mice and found that agonist-promoted relaxation of airways was enhanced in direct proportion to the level of Gαs expression. Contraction of ASM from acetylcholine was not affected in Gαs transgenic mice, nor was relaxation by bitter taste receptors. Furthermore, agonist-promoted (but not basal) cAMP production in ASM cells from Gαs-transgenic mice was enhanced compared with ASM from nontransgenic littermates. Agonist-promoted inhibition of platelet-derived growth factor-stimulated ASM proliferation was also enhanced in Gαs mouse ASM. The enhanced maximal β-agonist response was of similar magnitude for relaxation, cAMP production, and growth inhibition. Taken together, it appears that a limiting factor in β-agonist responsiveness in ASM is the expression level of Gαs. Gene therapy or pharmacological means of increasing Gαs (or its coupling efficiency to β2AR) thus represent an interface for development of novel therapeutic agents for improvement of β-agonist therapy.

β2-Agonists upregulate PDE4 mRNA but not protein or activity in human airway smooth muscle cells from asthmatic and nonasthmatic volunteers

2012 ◽  
Vol 302 (3) ◽  
pp. L334-L342 ◽  
Author(s):  
Kyoko Niimi ◽  
Qi Ge ◽  
Lyn M. Moir ◽  
Alaina J. Ammit ◽  
Thomas Trian ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Splice Variant ◽  
Adrenergic Receptor ◽  
Transforming Growth Factor ◽  
Airway Smooth Muscle Cells ◽  
Camp Phosphodiesterase ◽  
Long Acting ◽  
First Time ◽  
Β2 Agonists

β2-Adrenergic receptor (β2AR) agonists induce airway relaxation via cAMP. Phosphodiesterase (PDE)s degrade and regulate cAMP, and in airway smooth muscle (ASM) cells PDE4D degrades cAMP. Long-acting β2-agonists are now contraindicated as monotherapy for asthma, and increased PDE4D has been speculated to contribute to this phenomenon. In this study we investigated the expression of PDE4D in asthmatic and nonasthmatic ASM cells and its regulation by formoterol and budesonide. Primary ASM cells from people with or without asthma were stimulated with transforming growth factor (TGF)-β1, formoterol, and/or budesonide. PDE4D mRNA was assessed by real-time PCR, or PCR to assess splice variant production. PDE4D protein was assessed by Western blotting, and we investigated the effect of formoterol on cAMP production and PDE activity. Interleukin (IL)-6 was assessed using ELISA. PDE4D mRNA was dose dependently upregulated by formoterol, with a single splice variant, PDE4D5, present. Formoterol did not induce PDE4D protein at time points between 3 to 72 h, whereas it did induce and increase IL-6 secretion. We pretreated cells with actinomycin D and a proteasome inhibitor, MG132, and found no evidence of alterations in mRNA, protein expression, or degradation of PDE4D. Finally PDE activity was not altered by formoterol. This study shows, for the first time, that PDE4D5 is predominantly expressed in human ASM cells from people with and without asthma and that formoterol does not upregulate PDE4D protein production. This leads us to speculate that continual therapy with β2AR agonists is unlikely to cause PDE4-mediated tachyphylaxis.

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