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Epinephrine Induces Human Airway Smooth Muscle Contraction Through the Alpha-1 Adrenergic Receptor After Beta-2 Adrenergic Receptor Desensitization
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A1218 - Epinephrine Induces Human Airway Smooth Muscle Contraction Through the Alpha-1 Adrenergic Receptor After Beta-2 Adrenergic Receptor Desensitization
Author Block: B. Deeney1, G. Cao1, B. E. Himes2, S. Orfanos1, R. A. Panettieri1; 1Institute for Translational Medicine and Science, Rutgers University, New Brunswick, NJ, United States, 2Department of Biostatistics and Epidemiology, University of Pennsylvania, Philadelphia, PA, United States.
Rationale: Human airway smooth muscle (HASM) cells express β2 adrenergic receptors (β2AR) that promote airway smooth muscle relaxation, and α1 adrenergic receptors (α1AR), that may evoke smooth muscle contraction. Little research has been done to determine if the α1AR plays a role in evoking bronchoconstriction. Epinephrine (epi), an adrenergic receptor agonist, binds to α and β adrenergic receptors. We posit that α1AR agonists induce bronchoconstriction after desensitization of the β2AR in HASM cells.
Methods: RNA sequencing was performed on asthmatic and non-asthmatic HASM cells. Human tracheal rings were immunostained for α1AR. HASM cells were treated overnight with β2 agonist to desensitize the β2AR and were then stimulated with epinephrine either with or without pretreatment with Doxazonin mesylate, an α1AR antagonist. Lysates were then collected for immunoblot. Blots were probed for phosphorylated myosin light chain (MLC) as the surrogate for bronchoconstriction.
Results: RNAseq and immunostaining of tissue from human trachea show high levels of α1AR mRNA and protein, respectively. Epinephrine increases MLC phosphorylation (2.16 ± 1.27 fold change non-β2AR desensitized vs β2AR desensitized, p=0.0009) and calcium release (AUC 365,799 ± 25,777 Control vs 474,587 ± 74,065 β2AR desensitized) of HASM cells after β2AR desensitization. Epinephrine also evokes a decrease in airway luminal area after β2AR desensitization in hPCLS. Doxazosin mesylate abrogates both MLC phosphorylation (0.57 ± 0.08 fold β2AR desensitized+Doxazosin+Epi vs β2AR desensitized+Epi, p=0.03) as well as calcium levels (AUC 474,587 ± 74,065 β2AR desensitized vs 183,564 ± 43,361 non-β2AR desensitized, p=0.007) induced by epinephrine in β2AR desensitized HASM cells.
Conclusions: These findings suggest that epinephrine evokes airway smooth muscle cell shortening after β2AR desensitization in an α1AR dependent manner.
Author Block: B. Deeney1, G. Cao1, B. E. Himes2, S. Orfanos1, R. A. Panettieri1; 1Institute for Translational Medicine and Science, Rutgers University, New Brunswick, NJ, United States, 2Department of Biostatistics and Epidemiology, University of Pennsylvania, Philadelphia, PA, United States.
Rationale: Human airway smooth muscle (HASM) cells express β2 adrenergic receptors (β2AR) that promote airway smooth muscle relaxation, and α1 adrenergic receptors (α1AR), that may evoke smooth muscle contraction. Little research has been done to determine if the α1AR plays a role in evoking bronchoconstriction. Epinephrine (epi), an adrenergic receptor agonist, binds to α and β adrenergic receptors. We posit that α1AR agonists induce bronchoconstriction after desensitization of the β2AR in HASM cells.
Methods: RNA sequencing was performed on asthmatic and non-asthmatic HASM cells. Human tracheal rings were immunostained for α1AR. HASM cells were treated overnight with β2 agonist to desensitize the β2AR and were then stimulated with epinephrine either with or without pretreatment with Doxazonin mesylate, an α1AR antagonist. Lysates were then collected for immunoblot. Blots were probed for phosphorylated myosin light chain (MLC) as the surrogate for bronchoconstriction.
Results: RNAseq and immunostaining of tissue from human trachea show high levels of α1AR mRNA and protein, respectively. Epinephrine increases MLC phosphorylation (2.16 ± 1.27 fold change non-β2AR desensitized vs β2AR desensitized, p=0.0009) and calcium release (AUC 365,799 ± 25,777 Control vs 474,587 ± 74,065 β2AR desensitized) of HASM cells after β2AR desensitization. Epinephrine also evokes a decrease in airway luminal area after β2AR desensitization in hPCLS. Doxazosin mesylate abrogates both MLC phosphorylation (0.57 ± 0.08 fold β2AR desensitized+Doxazosin+Epi vs β2AR desensitized+Epi, p=0.03) as well as calcium levels (AUC 474,587 ± 74,065 β2AR desensitized vs 183,564 ± 43,361 non-β2AR desensitized, p=0.007) induced by epinephrine in β2AR desensitized HASM cells.
Conclusions: These findings suggest that epinephrine evokes airway smooth muscle cell shortening after β2AR desensitization in an α1AR dependent manner.
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