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. 2017 Dec 5;12(12):e0189060.
doi: 10.1371/journal.pone.0189060. eCollection 2017.

The acute glucose lowering effect of specific GPR120 activation in mice is mainly driven by glucagon-like peptide 1

Linda Sundström  1 , Susanna Myhre  2 , Monika Sundqvist  2 , Andrea Ahnmark  2 , William McCoull  3 , Piotr Raubo  3 , Sam D Groombridge  3 , Magnus Polla  2 , Ann-Christin Nyström  2 , Lisbeth Kristensson  1 , Mats Någård  2 , Maria Sörhede Winzell  2
Affiliations

The acute glucose lowering effect of specific GPR120 activation in mice is mainly driven by glucagon-like peptide 1

Linda Sundström et al. PLoS One. .

Abstract

The mechanism behind the glucose lowering effect occurring after specific activation of GPR120 is not completely understood. In this study, a potent and selective GPR120 agonist was developed and its pharmacological properties were compared with the previously described GPR120 agonist Metabolex-36. Effects of both compounds on signaling pathways and GLP-1 secretion were investigated in vitro. The acute glucose lowering effect was studied in lean wild-type and GPR120 null mice following oral or intravenous glucose tolerance tests. In vitro, in GPR120 overexpressing cells, both agonists signaled through Gαq, Gαs and the β-arrestin pathway. However, in mouse islets the signaling pathway was different since the agonists reduced cAMP production. The GPR120 agonists stimulated GLP-1 secretion both in vitro in STC-1 cells and in vivo following oral administration. In vivo GPR120 activation induced significant glucose lowering and increased insulin secretion after intravenous glucose administration in lean mice, while the agonists had no effect in GPR120 null mice. Exendin 9-39, a GLP-1 receptor antagonist, abolished the GPR120 induced effects on glucose and insulin following an intravenous glucose challenge. In conclusion, GLP-1 secretion is an important mechanism behind the acute glucose lowering effect following specific GPR120 activation.

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Conflict of interest statement

Competing Interests: The authors of the paper are AstraZeneca employees and shareholders. The presented data and chemical compounds are not patented and are not products in development or marketed products. The presented compounds are tools to be used for exploring and evaluation the biology of GPR120 signaling. We fully adhere to all PLOS ONE policies on sharing data and materials.

Figures

Fig 1
Fig 1. Structure of AZ13581837 and Metabolex-36 and specificity of the compounds for human and mouse GPR120 and human or mouse GPR40.
A) Chemical structure of AZ13581837 and B) Metabolex-36.C) Effect of AZ13581837 (squares) and Metabolex-36 (circles) on DMR response in CHO-hGPR120 (filled symbols) and CHO (open symbols). D) Activity in CHO-GPR40 cells for AZ13581837 (filled squares), Metabolex-36 (filled circles) and GW9508 (filled triangles). Activity in CHO-hGPR120 cells is shown as reference for AZ13581837 (open squares), Metabolex-36 (open circles) and GW9508 (open triangles). E) Cross species selectivity evaluated in CHO-mGPR120 cells using a DMR assay. Activity of AZ13581837 (squares) and Metabolex-36 (circles) on mouse GPR120 (filled symbols) compared to human GPR120 (open symbols). F) Cross species selectivity for GPR40 evaluated using a calcium mobilization assay. Effect of AZ13581837 (filled squares) and Metabolex-36 (filled circles) on mouse GPR40 with GW9508 (filled triangles) as reference. Activity in CHO-hGPR120 cells is shown as comparison for AZ13581837 (open squares) and Metabolex-36 (open circles). Data are shown as mean ± SEM run in duplicates or more and representative for two or more independent experiments.
Fig 2
Fig 2. AZ13581837 and Metabolex-36 induce GPR120 dependent calcium mobilization, DMR response and cAMP production in GPR120 overexpressing cells.
A) Effect of AZ13581837 (squares) and Metabolex-36 (circles) on calcium mobilization in CHO-hGPR120 (filled symbols) and CHO cells (open symbols). B) Calcium mobilization induced by AZ13581837 (squares) and Metabolex-36 (circles) in untreated (open symbols) and QIC treated (filled symbols) CHO-hGPR120 cells. C) DMR response from stimulation with AZ13581837 (squares) and Metabolex-36 (circles) in untreated (open symbols) and QIC treated (filled symbols) CHO-hGPR120 cells. D) DMR response from AZ13581837 (squares) and Metabolex-36 (circles) in untreated (open symbols) and CTX treated (filled symbols) CHO-hGPR120 cells. E) DMR response from stimulation with AZ13581837 (squares) and Metabolex-36 (circles) in untreated (open symbols) and PTX treated (filled symbols) CHO-hGPR120 cells. F) Effect of AZ13581837 (squares) and Metabolex-36 (circles) on cAMP production in CHO-hGPR120 (filled symbols) and CHO cells (open symbols). G) Recruitment of β-arrestin in U2OS-hGPR120 cells induced by AZ13581837 (filled squares) and Metabolex-36 (filled circles). Data are mean ± SEM of experiments run in duplicates or more and representative for two or more independent experiments.
Fig 3
Fig 3. AZ13581837 and Metabolex-36 reduced cAMP production in mouse islets and induced GLP-1 secretion from STC-1 cells.
Effect of 10 μM AZ13581837, 10 μM Metabolex-36, 50 nM Exendin-4 or vehicle control on cAMP production in dispersed islets from wild type (A) and GP120 null mice (B). Data represent mean ± SEM from three independent experiments where islet were isolated from two or four mice of each genotype. cAMP was measured in at least triplicates for both wild type and GPR120 null islet in each experiment. STC-1 cells were stimulated with Metabolex-36, AZ13581837 or vehicle control (0.1% DMSO) for 2 hours and secreted active GLP-1 was measured by ELISA (C). Three independent GLP-1 secretion experiments were run where n = 3 of each control and compound treatment. *p<0.05, **p<0.01, ***p<0.001 and ****p<0.0001 versus vehicle control (two sample, two sided t-test).
Fig 4
Fig 4. Effect of Metabolex-36 and AZ13581837 on oral glucose tolerance in mice.
Effect of Metabolex-36 (A) and AZ13581837 (C) on glucose response after an oral glucose challenge (2g/kg) in male mice and the corresponding unbound circulating concentrations of Metabolex-36 (B) and AZ13581837 (D) during the experiment. AZ13581837 and Metabolex-36 were given in different doses as indicated in the figures with n = 10 mice group and compared to vehicle treated mice (n = 12 mice per group). The EC50 value for each GPR120 agonist assessed on mouse GPR120 using a DMR assay is indicated in figures. Blood glucose levels following oral glucose administration in GPR120 null mice (E) and wild type mice (F) were determined for vehicle (open squares) and Metabolex-36 (filled squares).
Fig 5
Fig 5. Metabolex-36 and AZ13581837 increased insulin secretion in IVGTT in lean mice.
Insulin (A) and blood glucose (C) levels following an intravenous glucose challenge after oral administration of Metabolex-36 and AZ13581837 in lean female mice and corresponding AIR (B) and glucose elimination (D). Data represent six (Metabolex-36, n = 33, vehicle n = 34) and two (AZ13581837, n = 14) independent experiments and data are presented as mean ± SEM. Plasma levels of total GLP-1 (E) at time point was determined in separate experiments with n = 10 mice per group. ***p<0.001 and **p<0.01versus vehicle control.
Fig 6
Fig 6. AZ13581837 had no effect on insulin secretion in GPR120 null mice.
Insulin (A) and blood glucose (C) levels following an intravenous glucose challenge after administration of AZ13581837 in female, lean wt or GPR120 null mice and corresponding AIR (B) and glucose elimination (D). The results are from one experiment with n = 4 per group. Data are presented as mean ± SEM.**p<0.01, *p<0.05.
Fig 7
Fig 7. Exendin 9–39 blocked the AZ13581837 induced potentiation of insulin secretion in lean mice.
Insulin levels following intravenous glucose challenge (A) and corresponding blood glucose (C), after administration of AZ13581837, exendin 9–39 or a co-administration of both, with corresponding calculations of AIR (B) and glucose elimination (D). The IVGTT data are from two independent experiments with n = 10 mice per group. Data are presented as mean ± SEM.***p<0.001 and *p<0.05 versus vehicle control.
Fig 8
Fig 8. Exendin 9–39 blocked the Metabolex-36 induced potentiation of insulin secretion in lean mice.
Insulin levels following intravenous glucose challenge (A) and corresponding blood glucose (C), after administration of Metabolex-36, exendin 9–39 or a co-administration of both, with corresponding calculations of AIR (B) and glucose elimination (D). The IVGTT data are from two independent experiments with 6–7 mice per group. Data are presented as mean ± SEM.**p<0.01 and *p<0.05 versus vehicle control.
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