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A7175 - In Vitro Effects of Glucagon-Like Peptide 1 on Human Bronchial Epithelial and Smooth Muscle Cells
Author Block: D. Nguyen1, G. R. Reddy2, A. L. Linderholm1, Y. K. Xiang2, N. J. Kenyon1; 1Pulmonary, Critical Care, and Sleep Medicine, University of California Davis, Sacramento, CA, United States, 2Pharmacology, University of California Davis, Davis, CA, United States.
Rationale:
Glucagon-like peptide 1 (GLP-1) is an anti-inflammatory incretin used to treat diabetes and obesity that may have beneficial effects within the lung, particularly for obesity-related asthma. It has been found to stimulate bronchodilation and surfactant production via its ubiquitously expressed receptor. Studies indicate that this may occur through activation of the secondary messengers, cyclic adenosine monophosphate (cAMP) and guanosine monophosphate (cGMP). Using fluorescence resonance energy transfer (FRET), human bronchial epithelial (HBE1) and airway smooth muscle (HASM) cells were evaluated in vitro for real-time activation of cAMP and cGMP. We hypothesized that GLP-1 would directly activate these secondary messengers, similar to the beta-adrenoreceptor agonist, isoproterenol.
Methods:
HBE1 and HASM were cultured on glass coverslips and infected with an adenovirus containing a cAMP or cGMP biosensor for 36 hours. The coverslips were then maintained in calcium-free phosphate-buffered saline for FRET recording. Cells were treated with isoproterenol (1 μM), GLP-1 (1 μM), or a combination of the two with and without pre-treatment with the GLP-1 receptor antagonist exendin 9-39 (10 ng/mL). Images were acquired using a Leica DMI3000 B inverted fluorescence microscope (Leica Biosystems, Buffalo Grove, IL) with a 40×/1.3 numerical aperture oil-immersion objective lens and a charge-coupled device camera controlled by Metafluor software (Molecular Devices, Sunnyvale, CA). Images were subjected to background subtraction, and were acquired every 20 seconds with an exposure time of 200 milliseconds. Donor/acceptor FRET ratios were calculated and normalized to the ratio values of baseline prior to medication administration. Activation of cAMP or cGMP decreased CFP-YFP FRET efficiency.
Results:
Isoproterenol administration caused significant cAMP and cGMP activation, as seen by FRET ratio changes, in HBE1 and HASM cells. GLP-1 alone did not alter the FRET ratios of the secondary messengers in either cell line. However, co-administration with isoproterenol amplified cAMP and cGMP stimulation in both cell lines. This enhancement was then attenuated in the presence of GLP-1 receptor antagonist, exendin 9-39.
Conclusions:
Glucagon-like peptide 1 augmented isoproterenol activation of cAMP and cGMP in both human bronchial and airway smooth muscle cells. We believe this synergy can be beneficial and may provide a rationale for GLP-1 adjunctive therapy for patients with obesity-related asthma. We will further investigate primary human bronchial and airway smooth muscle cells to elucidate the underlying interactive mechanisms of GLP-1 and beta agonists.