Dataset Information

Control of human pancreatic beta cell kinome by glucagon-like peptide-1 receptor biased agonism.

ABSTRACT:

Aim

To determine the kinase activity profiles of human pancreatic beta cells downstream of glucagon-like peptide-1 receptor (GLP-1R) balanced versus biased agonist stimulations.

Materials and methods

This study analysed the kinomic profiles of human EndoC-βh1 cells following vehicle and GLP-1R stimulation with the pharmacological agonist exendin-4, as well as exendin-4-based biased derivatives exendin-phe1 and exendin-asp3 for acute (10-minute) versus sustained (120-minute) responses, using PamChip protein tyrosine kinase and serine/threonine kinase assays. The raw data were filtered and normalized using BioNavigator. The kinase analyses were conducted with R, mainly including kinase-substrate mapping and Kyoto Encyclopedia of Genes and Genomes pathway analysis.

Results

The present analysis reveals that kinomic responses are distinct for acute versus sustained GLP-1R agonist exposure, with individual responses associated with agonists presenting specific bias profiles. According to pathway analysis, several kinases, including JNKs, PKCs, INSR and LKB1, are important GLP-1R signalling mediators, constituting potential targets for further research on biased GLP-1R downstream signalling.

Conclusion

The results from this study suggest that differentially biased exendin-phe1 and exendin-asp3 can modulate distinct kinase interaction networks. Further understanding of these mechanisms will have important implications for the selection of appropriate anti-type 2 diabetes therapies with optimized downstream kinomic profiles.

SUBMITTER: Xiao J

PROVIDER: S-EPMC10947446 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature

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Publications

Control of human pancreatic beta cell kinome by glucagon-like peptide-1 receptor biased agonism.

Xiao Jiannan J El Eid Liliane L Buenaventura Teresa T Boutry Raphaël R Bonnefond Amélie A Jones Ben B Rutter Guy A GA Froguel Philippe P Tomas Alejandra A

Diabetes, obesity & metabolism 20230425 8

<h4>Aim</h4>To determine the kinase activity profiles of human pancreatic beta cells downstream of glucagon-like peptide-1 receptor (GLP-1R) balanced versus biased agonist stimulations.<h4>Materials and methods</h4>This study analysed the kinomic profiles of human EndoC-βh1 cells following vehicle and GLP-1R stimulation with the pharmacological agonist exendin-4, as well as exendin-4-based biased derivatives exendin-phe1 and exendin-asp3 for acute (10-minute) versus sustained (120-minute) respon ...[more]

PMID: 37039251

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Project description:ObjectiveTreatment with glucagon receptor antagonists (GRAs) reduces blood glucose but causes dyslipidemia and accumulation of fat in the liver. We investigated the acute and chronic effects of glucagon on lipid metabolism in mice.MethodsChronic effects of glucagon receptor signaling on lipid metabolism were studied using oral lipid tolerance tests (OLTTs) in overnight fasted glucagon receptor knockout (Gcgr-/-) mice, and in C57Bl/6JRj mice treated with a glucagon receptor antibody (GCGR Ab) or a long-acting glucagon analogue (GCGA) for eight weeks. Following treatment, liver tissue was harvested for RNA-sequencing and triglyceride measurements. Acute effects were studied in C57Bl/6JRj mice treated with a GRA or GCGA 1 h or immediately before OLTTs, respectively. Direct effects of glucagon on hepatic lipolysis were studied using isolated perfused mouse liver preparations. To investigate potential effects of GCGA and GRA on gastric emptying, paracetamol was, in separate experiments, administered immediately before OLTTs.ResultsPlasma triglyceride concentrations increased 2-fold in Gcgr-/- mice compared to their wild-type littermates during the OLTT (P = 0.001). Chronic treatment with GCGR Ab increased, whereas GCGA treatment decreased, plasma triglyceride concentrations during OLTTs (P < 0.05). Genes involved in lipid metabolism were upregulated upon GCGR Ab treatment while GCGA treatment had opposite effects. Acute GRA and GCGA treatment, respectively, increased (P = 0.02) and decreased (P = 0.003) plasma triglyceride concentrations during OLTTs. Glucagon stimulated hepatic lipolysis, evident by an increase in free fatty acid concentrations in the effluent from perfused mouse livers. In line with this, GCGR Ab treatment increased, while GCGA treatment decreased, liver triglyceride concentrations. The effects of glucagon appeared independent of changes in gastric emptying of paracetamol.ConclusionsGlucagon receptor signaling regulates triglyceride metabolism, both chronically and acutely, in mice. These data expand glucagon´s biological role and implicate that intact glucagon signaling is important for lipid metabolism. Glucagon agonism may have beneficial effects on hepatic and peripheral triglyceride metabolism.

Project description:BackgroundInsulin secretion is enhanced upon the binding of Glucagon-like peptide-1 (GLP-1) to its receptor (GLP1R) in pancreatic β cells. Although a reduced expression of GLP1R in pancreatic islets from type 2 diabetic patients and hyperglycaemic rats has been established, it is still unknown if this is caused by differential DNA methylation of GLP1R in pancreatic islets of type 2 diabetic patients.MethodsIn this study, DNA methylation levels of 12 CpG sites close to the transcription start site of GLP1R were analysed in pancreatic islets from 55 non-diabetic and 10 type 2 diabetic human donors as well as in β and α cells isolated from human pancreatic islets. DNA methylation of GLP1R was related to GLP1R expression, HbA1c levels and BMI. Moreover, mRNA expression of MECP2, DNMT1, DNMT3A and DNMT3B was analysed in pancreatic islets of the non-diabetic and type 2 diabetic donors.ResultsOne CpG unit, at position +199 and +205 bp from the transcription start site, showed a small increase in DNA methylation in islets from donors with type 2 diabetes compared to non-diabetic donors (0.53%, p=0.02). Furthermore, DNA methylation levels of one CpG site located 376 bp upstream of the transcription start site of GLP1R correlated negatively with GLP1R expression (rho=-0.34, p=0.008) but positively with BMI and HbA1c (rho=0.30, p=0.02 and rho=0.30, p=0.03, respectively). This specific CpG site is located in an area with known SP1 and SP3 transcription factor binding sites. Moreover, when we compared the DNA methylation of the GLP1R promoter in isolated human β and α cells, we found that it was higher in α- compared with β-cells (p=0.009). Finally, there was a trend towards decreased DNMT3A expression (p=0.056) in type 2 diabetic compared with non-diabetic islets.ConclusionsTogether, our study shows that while BMI and HbA1c are positively associated with DNA methylation levels of GLP1R, its expression is negatively associated with DNA methylation of GLP1R in human pancreatic islets.

Dataset Information

Control of human pancreatic beta cell kinome by glucagon-like peptide-1 receptor biased agonism.

Aim

Materials and methods

Results

Conclusion

Publications

Control of human pancreatic beta cell kinome by glucagon-like peptide-1 receptor biased agonism.

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