Studies on GLP-1 produced within the pancreas and its significance for the function and regulation of islets of Langerhans

Question: Targeting incretin receptors is a promising approach in current therapeutic regimes for the treatment of type 2 diabetes mellitus and obesity. Interestingly, pancreatic islet cells themselves are capable to produce glucagon-like peptide 1 (GLP-1) [1, 2]. As function and regulation of this local incretin synthesis are not clear, the aim of this study is to elucidate the role of intra-islet GLP-1 production on paracrine signaling. Therefore, the effects of GLP-1 and exendin (9-39) amide, a truncated peptide competitively inhibiting GLP-1 receptors, were investigated in mouse islets cultured under physiological conditions and in a milieu mimicking a hyperglycemic environment.

Methods: Islets or beta cells were isolated from C57BL/6N mice. Experiments were performed after 7 days of cultivation under control conditions or in a toxic model. To mimic a diabetic environment, islets were treated either with a high concentration of glucose (33 mM) or glucose and palmitate (500 µM, BSA-bound). Glucose-stimulated insulin release and GLP-1 secretion were measured by radioimmunoassay and ELISA. Gene expression levels were detected by RT-qPCR. Apoptosis was measured by TUNEL-labeling followed by Hoechst staining and calcium oscillations in islets were monitored using the fluorescent dye FURA-2-AM.

Results: Acute inhibition of GLP-1 receptors in islets with exendin 9-39 (Ex 9-39,100 nM) decreases insulin secretion (15 mM glucose, G15) after cultivation in control and glucotoxic conditions, revealing effects of intra-islet GLP-1 already under non-pathological circumstances (G15: 4.7±0.5 vs. G15+Ex 9-39: 2.6±0.3 (ng/(islet*h)), n=3, p < 0.01). Accordingly, acute suppression of GLP 1 receptors reduces the calcium concentration (10 mM glucose, G10) in islets (AUC G10: 2.2±0.9 vs. AUC G10+Ex 9-39: 1.6±0.5, p < 0.001, n=19). External addition of GLP-1 (50 nM) induces a stronger increase in glucose-mediated insulin release (10 mM glucose, G10) after glucotoxic pretreatment compared to islets cultured under control conditions (G10+GLP-1: 5±2 vs. control: 1.5±0.8 (ng/(islet*h)), n=5, p < 0.001), suggesting an altered sensitivity of the GLP-1 signaling cascade. GLP-1 secretion is already present in the physiological model, and does not raise after preincubation of the islets in medium supplemented with 33 mM glucose (6±1 vs. 6±1 (pM), n=5). Consistently, gene expression of PC1/3 is not upregulated by glucotoxicity (n=5). These data show, that GLP-1 production in islets is not restricted to a pathological environment. In line with this, treatment of islet cells with exendin 9-39 (24 h) elevates the number of apoptotic cells (n=5, p < 0.001), suggesting that locally released GLP-1 is physiologically required for maintaining beta cell mass.

Conclusion: Intra-islet GLP-1 secretion is essential to ensure proper islet function and to maintain beta cell homeostasis even under non-toxic conditions. Our studies show that islets secrete GLP-1 permanently and not only when they are challenged by a diabetic environment, pointing to a more relevant role of local GLP-1 production than previously assumed. As metabolic stress increases the sensitivity of islets to GLP-1, further investigations are required on how the GLP-1 signaling cascade changes during progression of diabetes.