The endogenous preproglucagon system is not essential for gut growth homeostasis in mice

Objective: The prevalence of obesity and related co-morbidities is reaching pandemic proportions. Today, the most effective obesity treatments are glucagon-like peptide 1 (GLP-1) analogs and bariatric surgery. Interestingly, both intervention paradigms have been associated with adaptive growth responses in the gut; however, intestinotrophic mechanisms associated with or secondary to medical or surgical obesity therapies are poorly understood. Therefore, the objective of this study was to assess the local basal endogenous and pharmacological intestinotrophic effects of glucagon-like peptides and bariatric surgery in mice. Methods: We used in situ hybridization to provide a detailed and comparative anatomical map of the local distribution of GLP-1 receptor (Glp1r), GLP-2 receptor (Glp2r), and preproglucagon (Gcg) mRNA expression throughout the mouse gastrointestinal tract. Gut development in GLP-1R-, GLP-2R-, or GCG-deficient mice was compared to their corresponding wild-type controls, and intestinotrophic effects of GLP-1 and GLP-2 analogs were assessed in wild-type mice. Lastly, gut volume was determined in a mouse model of vertical sleeve gastrectomy (VSG). Results: Comparison of Glp1r, Glp2r, and Gcg mRNA expression indicated a widespread, but distinct, distribution of these three transcripts throughout all compartments of the mouse gastrointestinal tract. While mice null for Glp1r or Gcg showed normal intestinal morphology, Glp2r−/− mice exhibited a slight reduction in small intestinal mucosa volume. Pharmacological treatment with GLP-1 and GLP-2 analogs significantly increased gut volume. In contrast, VSG surgery had no effect on intestinal morphology. Conclusion: The present study indicates that the endogenous preproglucagon system, exemplified by the entire GCG gene and the receptors for GLP-1 and GLP-2, does not play a major role in normal gut development in the mouse. Furthermore, elevation in local intestinal and circulating levels of GLP-1 and GLP-2 achieved after VSG has limited impact on intestinal morphometry. Hence, although exogenous treatment with GLP-1 and GLP-2 analogs enhances gut growth, the contributions of endogenously-secreted GLP-1 and GLP-2 to gut growth may be more modest and highly context-dependent

Functional Consequences of Glucagon-like Peptide-1 Receptor Cross-talk and Trafficking

The signaling capacity of seven-transmembrane/G-protein-coupled receptors (7TM/GPCRs) can be regulated through ligand-mediated receptor trafficking. Classically, the recycling of internalized receptors is associated with resensitization, whereas receptor degradation terminates signaling. We have shown previously that the incretin glucagon-like peptide-1 receptor (GLP-1R) internalizes fast and is primarily resensitized through recycling back to the cell surface. GLP-1R is expressed in pancreatic islets together with the closely related glucose-dependent insulinotropic polypeptide (GIPR) and glucagon (GCGR) receptors. The interaction and cross-talk between coexpressed receptors is a wide phenomenon of the 7TM/GPCR superfamily. Numerous reports show functional consequences for signaling and trafficking of the involved receptors. On the basis of the high structural similarity and tissue coexpression, we here investigated the potential cross-talk between GLP-1R and GIPR or GCGR in both trafficking and signaling pathways. Using a real-time time-resolved FRET-based internalization assay, we show that GLP-1R, GIPR, and GCGR internalize with differential properties. Remarkably, upon coexpression of the internalizing GLP-1R and the non-internalizing GIPR, GLP-1-mediated GLP-1R internalization was impaired in a GIPR concentration-dependent manner. As a functional consequence of such impaired internalization capability, GLP-1-mediated GLP-1R signaling was abrogated. A similar compromised signaling was found when GLP-1R internalization was abrogated by a dominant-negative version of dynamin (dynamin-1 K44E), which provides a mechanistic link between GLP-1R trafficking and signaling. This study highlights the importance of receptor internalization for full functionality of GLP-1R. Moreover, cross-talk between the two incretin receptors GLP-1R and GIPR is shown to alter receptor trafficking with functional consequences for GLP-1R signaling

Long-Term Treatment with Liraglutide, a Glucagon-Like Peptide-1 (GLP-1) Receptor Agonist, Has No Effect on β-Amyloid Plaque Load in Two Transgenic APP/PS1 Mouse Models of Alzheimer's Disease.

One of the major histopathological hallmarks of Alzheimer's disease (AD) is cerebral deposits of extracellular β-amyloid peptides. Preclinical studies have pointed to glucagon-like peptide 1 (GLP-1) receptors as a potential novel target in the treatment of AD. GLP-1 receptor agonists, including exendin-4 and liraglutide, have been shown to promote plaque-lowering and mnemonic effects of in a number of experimental models of AD. Transgenic mouse models carrying genetic mutations of amyloid protein precursor (APP) and presenilin-1 (PS1) are commonly used to assess the pharmacodynamics of potential amyloidosis-lowering and pro-cognitive compounds. In this study, effects of long-term liraglutide treatment were therefore determined in two double APP/PS1 transgenic mouse models of Alzheimer's disease carrying different clinical APP/PS1 mutations, i.e. the 'London' (hAPPLon/PS1A246E) and 'Swedish' mutation variant (hAPPSwe/PS1ΔE9) of APP, with co-expression of distinct PS1 variants. Liraglutide was administered in 5 month-old hAPPLon/PS1A246E mice for 3 months (100 or 500 ng/kg/day, s.c.), or 7 month-old hAPPSwe/PS1ΔE9 mice for 5 months (500 ng/kg/day, s.c.). In both models, regional plaque load was quantified throughout the brain using stereological methods. Vehicle-dosed hAPPSwe/PS1ΔE9 mice exhibited considerably higher cerebral plaque load than hAPPLon/PS1A246E control mice. Compared to vehicle-dosed transgenic controls, liraglutide treatment had no effect on the plaque levels in hAPPLon/PS1A246E and hAPPSwe/PS1ΔE9 mice. In conclusion, long-term liraglutide treatment exhibited no effect on cerebral plaque load in two transgenic mouse models of low- and high-grade amyloidosis, which suggests differential sensitivity to long-term liraglutide treatment in various transgenic mouse models mimicking distinct pathological hallmarks of AD

Stereological analysis of brain regional volume and plaque volume in transgenic hAPP_Lon/PS1_A246E and hAPP_Swe/PS1_ΔE9 mice.

<p>Stereological assessments were only performed in brain structures showing presence of β-amyloid plaques. Liraglutide treatment had no effect on brain regional volume and plaque load in hAPP_Lon/PS1_A246E (A, B) and hAPP_Swe/PS1_ΔE9 (C, D) mice.</p

Representative photomicrographs depicting β-amyloid plaque levels in eight month-old transgenic hAPP_Lon/PS1_A246E mice.

<p>(A-C) Plaque depositions in vehicle-dosed hAPP_Lon/PS1_A246E mice, indicated at three individual coronal levels. (D) vehicle-dosed hAPP_Lon/PS1_A246E mouse (E) liraglutide-treated hAPP_Lon/PS1_A246E mouse (100 μg/kg/day); (F) liraglutide-treated hAPP_Lon/PS1_A246E mouse (500 μg/kg/day).</p

Representative photomicrographs depicting β-amyloid plaque levels in 12 month-old transgenic hAPP_Swe/PS1_ΔE9 mice.

<p>(A-C) Plaque depositions in vehicle-dosed hAPP_Swe/PS1_ΔE9 mice, indicated at three individual coronal levels. (D) vehicle-dosed wild-type mouse, (E) vehicle-dosed hAPP_Swe/PS1_ΔE9 mouse, (F) liraglutide-treated hAPP_Swe/PS1_ΔE9 mouse (500 μg/kg/day).</p

Total brain volume and plaque levels in two transgenic hAPP/PS1 mouse models of Alzheimer’s disease, as determined by stereological means.

<p>Total brain volume and plaque levels in two transgenic hAPP/PS1 mouse models of Alzheimer’s disease, as determined by stereological means.</p