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¹ Department of Endocrinology, Barts and the Royal London School of Medicine and Dentistry, West Smithfield, London EC1A 7BE, UK
² Veterinary Basic Sciences, Royal Veterinary College, Royal College Street, London NW1 0TU, UK

(Requests for offprints should be addressed to K K Sidhu, Centre for Molecular Endocrinology, William Harvey Research Institute, Queen Mary University of London, St Bartholomew’s Hospital, West Smithfield, London EC1A 7BE, UK; Email: k.k.sidhu{at}qmul.ac.uk)

The insulinotrophic effects of glucagon-like peptide 1 (GLP-1) are mediated by its seven-transmembrane receptor (GLP-1R) in pancreatic ß-cells. We have transiently transfected the GLP-1R and a proopiomelanocortin (POMC) promoter-driven human preproinsulin gene vector (pIRES) into the AtT-20 pituitary corticotrophic cell line, to investigate the possibility of creating a regulated, insulin-expressing cell line. Receptor expression was confirmed by RT-PCR and functionality was demonstrated by measuring changes in cAMP levels in response to GLP-1. Rapid (5 min) stimulation of cAMP production was observed with 100 nM GLP-1, 24 h after transfection of 2 µg GLP-1R DNA. AtT-20 cells co-transfected with GLP-1R and human glycoprotein hormone -subunit or rat POMC promoters revealed GLP-1-stimulated cAMP activation of transcription. Co-transfection of the pIRES vector with the GLP-1R resulted in GLP-1-stimulated activation of POMC promoter-driven preproinsulin gene transcription but insulin secretion was not detected. However, using an adenoviral expression system to infect AtT-20 cells with GLP-1R and the preproinsulin gene (including 120 bp of its own promoter) resulted in a 6.4 ± 0.6-fold increase in cAMP and a 4.9 ± 0.8-fold increase in insulin secretion in response to 100 nM GLP-1. These results demonstrate, for the first time, functional GLP-1R-mediated preproinsulin gene transcription and secretion in a transplantable cell line.

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