Expression of functional beta 2-adrenergic receptors in a rat airway epithelial cell line (SPOC1) and cell density-dependent induction by glucocorticoids.
The beta 2-adrenergic receptor (beta 2AR) signal transduction system regulates many key functions of airway epithelium. In this study, we have pharmacologically characterized the beta 2AR and determined the impact of glucocorticoids on beta 2AR gene transcription in SPOC1 cells, a continuous cell line derived from the tracheal epithelium of rats. [125I]Cyanoiodopindolol assays demonstrated that binding to SPOC1 cell membranes was saturable (Bmax = 62.6 +/- 6 fmol/mg protein) and of high affinity (Kd = 6.3 +/- 0.8 pM). From competition experiments, the rank order of potency of agonists (isoproterenol > epinephrine >> norepinephrine) and the high affinity (Ki = 0.37 +/- 0.05 nM) of the beta 2-selective antagonist ICI 118,551 suggested the predominance of the beta 2AR subtype. Two isoforms of the alpha subunit of Gs (45 and 52 kDa) were identified by Western blot analysis. Isoproterenol-stimulated cyclic AMP levels increased in a dose-dependent manner, confirming that SPOC1 cell beta 2ARs are functionally coupled to adenylyl cyclase. The effect of glucocorticoids on beta 2AR expression was assessed in radioligand and transient transfection assays. Dexamethasone treatment of SPOC1 cells increased both beta 2AR protein and beta 2AR-luciferase fusion gene expression 1.6- to 3.1-fold, with the greatest increase demonstrated in cells cultured at low density compared to cells grown at high density.
Cao
W
W
Department of Physiology and Biophysics, University of Arkansas for Medical Sciences, Little Rock, USA.
McGraw
D W
DW
Lee
T T
TT
Dicker-Brown
A
A
Hiller
F C
FC
Cornett
L E
LE
Jones
S M
SM
D058666:Adrenergic beta-2 Receptor Agonists; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002452:Cell Count; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D003907:Dexamethasone / Q000494:pharmacology*; D004837:Epinephrine / Q000494:pharmacology; D004848:Epithelium / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D005938:Glucocorticoids / Q000494:pharmacology*; D007545:Isoproterenol / Q000494:pharmacology; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D009638:Norepinephrine / Q000494:pharmacology; D011433:Propranolol / Q000494:pharmacology; D051381:Rats; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D014132:Trachea / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection
D058666:Adrenergic beta-2 Receptor Agonists; D000318:Adrenergic beta-Agonists; D005938:Glucocorticoids; D018343:Receptors, Adrenergic, beta-2; D003907:Dexamethasone; D011433:Propranolol; D000242:Cyclic AMP; D008156:Luciferases; D007545:Isoproterenol; D009638:Norepinephrine; D004837:Epinephrine