The class of medications being recommended, known formally as GLP-1 agonists, can suppress a person’s appetite by mimicking a hormone that signals to the brain when a person is full. Food first, fizzy drinks after: Carbonated drinks can cause individuals on this medication to feel full fast. Although Medicare can now make more than 3 million patients eligible for Wegovy, this doesn’t address the gap in coverage of Ozempic and Mounjaro for diabetic patients between private and public insurance. More severe side effects are uncommon but possible. Glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are regarded as ‘incretins’ working closely to regulate glucose homeostasis. In this article we provide an overview of the pathophysiology and management of NASH, with a special focus on the pharmacological effects and possible mechanisms of GLP-1 mimetics in treating NAFLD/NASH, including dual and triple agonists at GLP-1R, glucose-dependent insulinotropic polypeptide receptor or glucagon receptor. Another class of medications, called dual GIP/GLP-1 receptor agonists, works similarly to GLP-1 receptor agonists.

We show that when the two receptors were co-expressed in HEK 293T cells with comparable receptor ratio to pancreatic cancer cells, GIP predominately induced cAMP accumulation while GLP-1 was biased towards β-arrestin 2 recruitment. AMP detection was using a Lance cAMP kit (PerkinElmer Life and Analytical Sciences), performed as previously described6. All values were converted to cAMP concentration using cAMP standard curve performed parallel and data were subsequently normalised to the response of 100 μM forskolin in each cell line, and then normalised to the WT for each agonist. Cells were stimulated with increasing concentrations of ligand, 100 μM forskolin or vehicle, and incubated for 30 min at 37 °C in 5% CO2. 1 h at 37 °C in 5% CO2. 1.5 mL) was collected and diluted to 17 mL with membrane preparation buffer followed by centrifugation at 100,000×g for 30 min at 4 °C. Cells were then incubated with coelenterazine H (5 μM) in assay buffer (1× HBSS, 10 mM HEPES, ColonBroom supplement 0.1% (w/v) BSA) for 1 h at room temperature. Plates were incubated at room temperature for 2 h before measurement of the fluorescence using an EnVision Multimode Plate (Reader PerkinElmer Life and Analytical Sciences). Cells were seeded at a density of 30,000 cells/well into 96-well culture plates and incubated overnight in DMEM containing 5% FBS at 37 °C in 5% CO2.

FlpInCHO wildtype and mutant human GLP-1R cells were seeded at a density of 25 × 104 cells/well into 24-well culture plates and [empty] incubated overnight at 37 °C in 5% CO2, washed three times in 1× PBS and fixed with 3.7% paraformaldehyde (PFA) at 4 °C for 15 min. CHOFlpIn WT GLP-1R or CHOFlpIn mutant GLP-1R cells were seeded at a density of 30,000 cells per well into a 96-well plate and incubated overnight at 37 °C in 5% CO2. Grains from GLP-1R mRNA ISH signal were rarely observed to colocalize at a higher density than background on the glucagon- or somatostatin-producing α and δ cells (Figures 2A-2C and 2G-2I, respectively), even after prolonged exposure. Membranes were increasing concentrations of peptides and RoxEx4 for 30 min prior to measurement of the BRET signal between Nluc-hGLP-1R and Rox-Ex4. The concentration-response curves were then plotted using the total area under the curve during the time of measurement post ligand addition. AMP accumulation concentration-response curves were also analysed using an operational model of agonism modified to directly estimate the ratio of τ/KA using Eq. Pharmacological data were analysed using Prism 8 (GraphPad).

Concentration-response binding and signalling data were analysed using the one-site binding inhibition and the three-parameter logistic equations in Graphpad prism, respectively. For rate analysis of G protein BRET assays, data were fitted to a one-phase association curve in Graphpad Prism. Baseline BRET measurements were taken for 2 min before the addition of the vehicle or ligand. Normalised AUC for the indicated ligand concentrations was plotted as a concentration-response curve and fitted with a three-parameter logistic curve. Luminance signals were measured using a CLariostar (BMG LabTech) at every 30 s intervals before, and every 15 s intervals after ligand addition (25 °C). This was assessed using a PHERAstar (BMG LabTech) at 10 s intervals (25 °C). Peroxidase activity was then measured using SigmaFast OPD tablets (Sigma) according to the manufacturer’s instructions, and fluorescence was detected at an emission wavelength of 492 nm. Cells were then solubilised in 0.1 M NaOH, and radioactivity was determined by gamma counting.