Inhibition of human airway smooth muscle cell proliferation by β2-adrenergic receptors and cAMP is PKA independent: Evidence for EPAC involvement

Karen M. Kassel, Todd A Wyatt, Reynold A. Panettieri, Myron Lee Toews

Research output: Contribution to journalArticle

66 Citations (Scopus)

Abstract

Mechanisms by which β-adrenergic receptor (βAR) agonists inhibit proliferation of human airway smooth muscle (HASM) cells were investigated because of their potential relevance to smooth muscle hyperplasia in asthma. We hypothesized that βAR agonists would inhibit mitogenesis in HASM cells via the β2AR, an increase in cAMP, and PKA activation. HASM cells were treated for 24 h with various agents and then analyzed for [ 3H]thymidine incorporation as a measure of cell proliferation. EGF stimulated proliferation by ∼10-fold. The nonselective βAR agonist isoproterenol and the β2AR-selective agonists albuterol and salmeterol inhibited EGF-stimulated proliferation by more than 50%, with half-maximal effects at 4.8 nM, 110 nM, and 6.7 nM, respectively. A β2AR-selective antagonist inhibited the isoproterenol effect with 100-fold greater potency than a β1AR-selective antagonist, confirming β2AR involvement in the inhibition of proliferation. The cAMP-elevating agents PGE2 and forskolin decreased EGF-induced proliferation, suggesting cAMP as the mediator. β2AR agonists and forskolin also inhibited proliferation stimulated by lysophosphatidic acid (LPA) as well as the synergistic proliferation stimulated by LPA+EGF. Importantly, PKA-selective cAMP analogs did not inhibit proliferation at concentrations that maximally activated PKA (10-100 μM), whereas a cAMP analog selective for the exchange protein directly activated by cAMP (EPAC), 8-(4-chlorophenylthio)-2′-O-methyl-cAMP, maximally inhibited proliferation at a concentration that did not activate PKA (10 μM). These data show that β2AR agonists and other cAMP-elevating agents decrease proliferation in HASM cells via a PKA-independent mechanism, and they provide pharmacological evidence for involvement of EPAC or an EPAC-like cAMP effector protein instead.

Original languageEnglish (US)
Pages (from-to)L131-L138
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
Volume294
Issue number1
DOIs
StatePublished - Jan 1 2008

Fingerprint

Epidermal Growth Factor
Adrenergic Agonists
Adrenergic Receptors
Smooth Muscle Myocytes
Cell Proliferation
Colforsin
Isoproterenol
Proteins
Albuterol
Dinoprostone
Thymidine
Hyperplasia
Smooth Muscle
Asthma
Pharmacology
lysophosphatidic acid

Keywords

  • Asthma
  • Epidermal growth factor
  • Exchange protein directly activated by cAMP
  • Lysophosphatidic acid

ASJC Scopus subject areas

  • Physiology
  • Pulmonary and Respiratory Medicine
  • Physiology (medical)
  • Cell Biology

Cite this

@article{799e57bb75a44a4092b965dbc4a4a717,
title = "Inhibition of human airway smooth muscle cell proliferation by β2-adrenergic receptors and cAMP is PKA independent: Evidence for EPAC involvement",
abstract = "Mechanisms by which β-adrenergic receptor (βAR) agonists inhibit proliferation of human airway smooth muscle (HASM) cells were investigated because of their potential relevance to smooth muscle hyperplasia in asthma. We hypothesized that βAR agonists would inhibit mitogenesis in HASM cells via the β2AR, an increase in cAMP, and PKA activation. HASM cells were treated for 24 h with various agents and then analyzed for [ 3H]thymidine incorporation as a measure of cell proliferation. EGF stimulated proliferation by ∼10-fold. The nonselective βAR agonist isoproterenol and the β2AR-selective agonists albuterol and salmeterol inhibited EGF-stimulated proliferation by more than 50{\%}, with half-maximal effects at 4.8 nM, 110 nM, and 6.7 nM, respectively. A β2AR-selective antagonist inhibited the isoproterenol effect with 100-fold greater potency than a β1AR-selective antagonist, confirming β2AR involvement in the inhibition of proliferation. The cAMP-elevating agents PGE2 and forskolin decreased EGF-induced proliferation, suggesting cAMP as the mediator. β2AR agonists and forskolin also inhibited proliferation stimulated by lysophosphatidic acid (LPA) as well as the synergistic proliferation stimulated by LPA+EGF. Importantly, PKA-selective cAMP analogs did not inhibit proliferation at concentrations that maximally activated PKA (10-100 μM), whereas a cAMP analog selective for the exchange protein directly activated by cAMP (EPAC), 8-(4-chlorophenylthio)-2′-O-methyl-cAMP, maximally inhibited proliferation at a concentration that did not activate PKA (10 μM). These data show that β2AR agonists and other cAMP-elevating agents decrease proliferation in HASM cells via a PKA-independent mechanism, and they provide pharmacological evidence for involvement of EPAC or an EPAC-like cAMP effector protein instead.",
keywords = "Asthma, Epidermal growth factor, Exchange protein directly activated by cAMP, Lysophosphatidic acid",
author = "Kassel, {Karen M.} and Wyatt, {Todd A} and Panettieri, {Reynold A.} and Toews, {Myron Lee}",
year = "2008",
month = "1",
day = "1",
doi = "10.1152/ajplung.00381.2007",
language = "English (US)",
volume = "294",
pages = "L131--L138",
journal = "American Journal of Physiology - Renal Physiology",
issn = "0363-6127",
publisher = "American Physiological Society",
number = "1",

}

TY - JOUR

T1 - Inhibition of human airway smooth muscle cell proliferation by β2-adrenergic receptors and cAMP is PKA independent

T2 - Evidence for EPAC involvement

AU - Kassel, Karen M.

AU - Wyatt, Todd A

AU - Panettieri, Reynold A.

AU - Toews, Myron Lee

PY - 2008/1/1

Y1 - 2008/1/1

N2 - Mechanisms by which β-adrenergic receptor (βAR) agonists inhibit proliferation of human airway smooth muscle (HASM) cells were investigated because of their potential relevance to smooth muscle hyperplasia in asthma. We hypothesized that βAR agonists would inhibit mitogenesis in HASM cells via the β2AR, an increase in cAMP, and PKA activation. HASM cells were treated for 24 h with various agents and then analyzed for [ 3H]thymidine incorporation as a measure of cell proliferation. EGF stimulated proliferation by ∼10-fold. The nonselective βAR agonist isoproterenol and the β2AR-selective agonists albuterol and salmeterol inhibited EGF-stimulated proliferation by more than 50%, with half-maximal effects at 4.8 nM, 110 nM, and 6.7 nM, respectively. A β2AR-selective antagonist inhibited the isoproterenol effect with 100-fold greater potency than a β1AR-selective antagonist, confirming β2AR involvement in the inhibition of proliferation. The cAMP-elevating agents PGE2 and forskolin decreased EGF-induced proliferation, suggesting cAMP as the mediator. β2AR agonists and forskolin also inhibited proliferation stimulated by lysophosphatidic acid (LPA) as well as the synergistic proliferation stimulated by LPA+EGF. Importantly, PKA-selective cAMP analogs did not inhibit proliferation at concentrations that maximally activated PKA (10-100 μM), whereas a cAMP analog selective for the exchange protein directly activated by cAMP (EPAC), 8-(4-chlorophenylthio)-2′-O-methyl-cAMP, maximally inhibited proliferation at a concentration that did not activate PKA (10 μM). These data show that β2AR agonists and other cAMP-elevating agents decrease proliferation in HASM cells via a PKA-independent mechanism, and they provide pharmacological evidence for involvement of EPAC or an EPAC-like cAMP effector protein instead.

AB - Mechanisms by which β-adrenergic receptor (βAR) agonists inhibit proliferation of human airway smooth muscle (HASM) cells were investigated because of their potential relevance to smooth muscle hyperplasia in asthma. We hypothesized that βAR agonists would inhibit mitogenesis in HASM cells via the β2AR, an increase in cAMP, and PKA activation. HASM cells were treated for 24 h with various agents and then analyzed for [ 3H]thymidine incorporation as a measure of cell proliferation. EGF stimulated proliferation by ∼10-fold. The nonselective βAR agonist isoproterenol and the β2AR-selective agonists albuterol and salmeterol inhibited EGF-stimulated proliferation by more than 50%, with half-maximal effects at 4.8 nM, 110 nM, and 6.7 nM, respectively. A β2AR-selective antagonist inhibited the isoproterenol effect with 100-fold greater potency than a β1AR-selective antagonist, confirming β2AR involvement in the inhibition of proliferation. The cAMP-elevating agents PGE2 and forskolin decreased EGF-induced proliferation, suggesting cAMP as the mediator. β2AR agonists and forskolin also inhibited proliferation stimulated by lysophosphatidic acid (LPA) as well as the synergistic proliferation stimulated by LPA+EGF. Importantly, PKA-selective cAMP analogs did not inhibit proliferation at concentrations that maximally activated PKA (10-100 μM), whereas a cAMP analog selective for the exchange protein directly activated by cAMP (EPAC), 8-(4-chlorophenylthio)-2′-O-methyl-cAMP, maximally inhibited proliferation at a concentration that did not activate PKA (10 μM). These data show that β2AR agonists and other cAMP-elevating agents decrease proliferation in HASM cells via a PKA-independent mechanism, and they provide pharmacological evidence for involvement of EPAC or an EPAC-like cAMP effector protein instead.

KW - Asthma

KW - Epidermal growth factor

KW - Exchange protein directly activated by cAMP

KW - Lysophosphatidic acid

UR - http://www.scopus.com/inward/record.url?scp=40749147710&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=40749147710&partnerID=8YFLogxK

U2 - 10.1152/ajplung.00381.2007

DO - 10.1152/ajplung.00381.2007

M3 - Article

C2 - 17993585

AN - SCOPUS:40749147710

VL - 294

SP - L131-L138

JO - American Journal of Physiology - Renal Physiology

JF - American Journal of Physiology - Renal Physiology

SN - 0363-6127

IS - 1

ER -