Basal Receptor Activation by Locally Produced Glucagon-Like Peptide-1 Contributes to Maintaining β-Cell Function

Abstract

Glucagon-like peptide 1 (GLP-1) is a physiological stimulus of pancreatic beta-cell function. This enteroendocrine hormone is produced by intestinal L cells, and is delivered via the bloodstream to GLP-1 receptors (GLP-1Rs) on pancreatic beta-cells. In addition, there is evidence that beta-cell GLP-1Rs maintain sustained basal activity even in the absence of intestinal peptide, an observation that has raised the question whether these receptors have some degree of ligand-independent function. Here, we provide an alternative explanation for basal receptor activity based on our finding that biologically relevant amounts of fully processed GLP-1 are locally generated by insulinoma cell lines, as well as by alpha-cells of isolated rat islets in primary culture. Presence of GLP-1 was established by immunocytochemistry, as well as by selective ELISAs and bioassays of cell supernatants. A GLP-1R antagonist significantly reduced insulin secretion/production in beta-TC-6 insulinoma cells and isolated rat islets, suggesting a functionally important loop between locally produced GLP-1 and its cognate receptor. Treatment with this antagonist also inhibited the growth of beta-TC-6 cells. These observations provide novel insight into the function of insulin-producing cell lines and native beta-cells during in vitro culture, and they support the idea that locally produced GLP-1 may play a role in intra-islet regulation.

A local glucagon-like peptide 1 (GLP-1) system in human pancreatic islets

“…GLP-1-like immunoreactivity was detected by a polyclonal and a monoclonal anti-GLP-1 antibody reacting, respectively, with the mid to C terminal region of GLP-1 [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19] and the amidated C-terminus of the peptide (see the Methods). These antibodies generated similar staining patterns with NCI-H716 cells and human islets (ESM Figs 2 and 3).…”

“…The corresponding spectra of the GLP-1(7-37) and GLP-1 (7-36) tryptic sequences were matched with the putative digested sequences of the peptides using the sequence editor implemented on BIOTOOLS. GLP-1(7-37) and GLP-1(7-36) peptides were identified by the species at m/z 1105 (aa [21][22][23][24][25][26][27][28] and at m/z 2098 (aa [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20].…”

“…Primary antibodies were: guinea pig polyclonal anti-swine insulin (A0564; DAKO, Carpinteria, CA, USA); mouse monoclonal anti-human/ mouse glucagon (MAB1249, clone 181402; R&D Systems, Abingdon, UK); rat monoclonal anti-somatostatin (ab30788; Abcam, Cambridge, UK); rabbit polyclonal anti-GLP-1 (ab22625; Abcam), which reacts with the mid to C terminal region of GLP-1 [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19], enabling immunoreactivity with N terminal truncated and C terminally extended forms of GLP-1; mouse monoclonal anti-GLP-1 (HYB 147-06; BioPorto Diagnostic, Gentofte, Denmark), which is specific for the amidated C-terminus of the peptide [20]; and rabbit polyclonal antibody to PC1/3 and PC2 (Chemicon, Rosemont, IL, USA). Secondary antibodies were: Alexa Fluor 488: goat anti-guinea pig IgG, goat anti-rabbit IgG, and goat anti-mouse IgG; and Alexa Fluor 594: goat anti-mouse IgG and goat antirabbit IgG (all from Invitrogen, Molecular Probes, Leiden, the Netherlands).…”

“…In another study [12], monoclonal antibodies to GLP-1 were produced for immunocytochemistry and radioimmunoassay experiments, and immunoreactive GLP-1 was detected in rat alpha cells, with increased release upon glucose stimulation [12]. Accordingly, GLP-1 amide was found in the pancreas of fetal and neonatal rats [13], and fully processed GLP-1 was produced by insulinoma cell lines and primary rat alpha cells [14]. Furthermore, when partial beta cell loss was induced in neonatal rats by streptozotocin, islet cell regeneration was accompanied by alpha cell hyperplasia, with an altered phenotype of increased GLP-1 synthesis [15].…”

DsAAV8-mediated expression of glucagon-like peptide-1 in pancreatic beta-cells ameliorates streptozotocin-induced diabetes

“…This is consistent with previous reports indicating that rodent alpha-cells may be capable of endogenously producing small amounts of mature GLP-1. 42,43 The expression of GLP-1 within beta-cells of normoglycemic animals did not elicit significant alterations in glucose homeostasis, as indicated by the transient and small reduction in fasting blood glucose levels within treated animals and lack of difference in glucose tolerance following intraperitoneal glucose administration. In STZ-treated animals, despite protection against hyperglycemia, expression of GLP-1 in the betacells did not result in significant alteration in plasma GLP-1 levels when compared with control animals, suggesting that a local environment of increased GLP-1 contributed to the protection against STZ-induced diabetes.…”