Biology subjects

Larraufie, P.

Publications and source records attributed to Larraufie, P..

5 recordsLinked to original sources

bioRxiv2026

Non-canonical signaling mechanisms of short-chain fatty acid receptors in glucagon-like peptide-1 (GLP-1) releasing enteroendocrine cells

ObjectivesFree fatty acid receptors 2 and 3 (FFA2 and FFA3) are activated by nutrient-derived metabolites such as short-chain fatty acids (SCFAs) and ketone bodies, produced by the gut microbiota and host, respectively. This study aimed to investigate the intracellular signaling pathways recruited in glucagon-like peptide-1 (GLP-1) releasing enteroendocrine cells following activation of FFA2 and FFA3 to resolve the impact of nutrient status on enteroendocrine cell function. MethodsExperiments were performed using primary mouse colonic cultures and the mouse enteroendocrine cell line, GLUTag cells. Expression analysis by bulk RNA sequencing was used to determine expression of FFA2 and FFA3 in GLP-1 releasing cells. Measurement of GLP-1 secretion by sandwich ELISA was used to assess enteroendocrine cell function. Live-cell measurements of intracellular calcium and cAMP levels were performed to assess canonical second messenger signaling pathways. ResultsA SCFA mixture stimulated GLP-1 secretion from both primary mouse colonic cultures and GLUTag cells. In GLUTag cells, the FFA2 ligand 4-CMTB inhibited GLP-1 release independent of Gaq- and Gai-signaling as neither YM-254890 (Gaq inhibitor) nor pertussis toxin (Gai- uncoupler) altered its effect. However, 4-CMTB did elevate cAMP levels, suggesting an indirect mechanism for the increase in cAMP production. Stimulation of FFA2 with the Gai-biased ligand AZ1729 or the ketone body acetoacetate inhibited GLP-1 release and cAMP accumulation. AZ1729 was insensitive to pertussis toxin and OZITX, supporting atypical FFA2 signaling. Stimulation of FFA3 with AR420626 or the ketone body {beta}-hydroxybutyrate increased GLP-1 secretion from GLUTag cells, an effect that was not mediated by cAMP production. AR420626, but not {beta}-hydroxybutyrate increased intracellular calcium levels. ConclusionsOverall, activation of FFA2 inhibited secretory function in GLP-1-releasing enteroendocrine cells, whereas activation of FFA3 stimulated GLP-1 secretion via distinct intracellular signaling mechanisms. O_FIG O_LINKSMALLFIG WIDTH=184 HEIGHT=200 SRC="FIGDIR/small/708924v1_ufig1.gif" ALT="Figure 1"> View larger version (37K): org.highwire.dtl.DTLVardef@175dbaforg.highwire.dtl.DTLVardef@a9cc3eorg.highwire.dtl.DTLVardef@1a026e5org.highwire.dtl.DTLVardef@15997af_HPS_FORMAT_FIGEXP M_FIG Graphical Abstract C_FIG HighlightsO_LIExposure to physiological levels of SCFAs stimulates GLP-1 secretion from colonic EECs C_LIO_LIFFA2 and FFA3 regulate GLP-1 release via non-canonical signaling pathways C_LIO_LIKetone bodies activate SCFA receptors to differentially modulate GLP-1 levels C_LIO_LILigand bias enables nutrient-dependent tuning of EEC gut hormone secretion C_LI

cell biology↗

bioRxiv2024

Development of rat organoids to study intestinal adaptations after Roux-en-Y Gastric Bypass

Organoids from intestinal regions have proven to be useful tools to study intestinal epithelial responses to different conditions. Roux-en-Y gastric bypass (RYGB) has been associated with important intestinal adaptations, but the mechanisms underlying these changes are still poorly understood. Organoids could therefore be used to better decipher the intestinal adaptations associated with this surgery. Rat is a common model to assess RYGB responses in vivo, but surprisingly, very few studies managed to develop organoids from rat small intestine. The primary objective of this study was to establish a protocol for cultivating organoids derived from the small intestine of healthy rats. The second objective of this study focuses on the development of organoids from the small intestine of rats subjected to RYGB to evaluate whether phenotypic or gene expression differences emerge. We successfully devised a functional protocol for developing organoids from fresh or frozen rat small intestine tissues. The obtained organoids exhibit significant variability, making interpretation challenging. Variability is observed in size, shape, and the number of organoids developed from the same sample, but also gene expression, depending on samples prepared on different days or from fresh or frozen tissues. This protocol was then applied to the small intestine of RYGB or sham-operated rats. However, we did not detect any major difference in size between intestinal organoids derived from Sham rats and those from RYGB rats. The expression of several genes (peptide transporters, amino acid transporters, genes specific to certain types of intestinal cells, etc.) was also assessed, and inter-experiment variability was higher than any effect due to the operation on the rat the intestinal tissue was originating. In conclusion, this study has established a functional protocol to grow small intestine organoids in rats. Initial results suggest that in our experimental conditions, organoids obtained from rats subjected to RYGB do not differ from those obtained from Sham rats. However, increasing the sample size and improving reproducibility between experiments will be essential to confirm these findings.

physiology↗

bioRxiv2019

Secretin release after Roux-en-Y Gastric Bypass reveals a population of glucose-sensitive S-cells in distal small intestine

ObjectiveGastrointestinal hormones contribute to the beneficial effects of Roux-en-Y gastric bypass surgery (RYGB) on glycemic control. Secretin is secreted from duodenal S-cells in response to low luminal pH, but it is unknown whether its secretion is altered after RYGB and if secretin contributes to the post-operative improvement in glycemic control. We hypothesized that secretin secretion increases after RYGB as a result of the diversion of nutrients to more distal parts of the small intestine, and thereby affects islet hormone release.\n\nMethodsA specific secretin radioimmunoassay was developed, evaluated biochemically, and used to quantify plasma concentrations of secretin in 13 obese individuals before, 1 week after and 3 months after RYGB. Distribution of secretin and its receptor was assessed by RNA-sequencing, mass-spectrometry and in situ hybridization in human and rat tissues. Isolated, perfused rat intestine and pancreas were used to explore the molecular mechanism underlying glucose-induced secretin secretion and to study direct effects of secretin on glucagon, insulin and somatostatin secretion. Secretin was administered alone or in combination with GLP-1 to non-sedated rats to evaluate effects on glucose regulation.\n\nResultsPlasma postprandial secretin was more than doubled in humans after RYGB (P<0.001). The distal small intestine harbored secretin expressing cells in both rats and humans. Glucose increased secretion of secretin in a sodium-glucose co-transporter dependent manner when administered to the distal part but not into the proximal part of the rat small intestine. Secretin stimulated somatostatin secretion (fold change: 1.59, P<0.05) from the perfused rat pancreas but affected neither insulin (P=0.2) nor glucagon (P=0.97) secretion. When administered to rats in vivo, insulin secretion was attenuated and glucagon secretion increased (P=0.04), while blood glucose peak time was delayed (from 15 min to 45 min) and gastric emptying time prolonged (P=0.004).\n\nConclusionGlucose-sensing secretin cells located in the distal part of the small intestine may contribute to increased plasma concentrations observed after RYGB. The metabolic role of the distal S-cells warrants further studies.

physiology↗

bioRxiv2018

Comparison of human and murine enteroendocrine cells by transcriptomic and peptidomic profiling

Enteroendocrine cells (EECs) produce hormones that regulate food absorption, insulin secretion and appetite. Both EECs and their peptide products are foci of drug discovery programmes for diabetes and obesity. We compared the human and mouse EEC transcriptome and peptidome to validate mouse as a model of the human enteroendocrine axis. We present the first RNA sequencing analysis of human EECs, and demonstrate strong correlation with mouse, although with outliers including some low abundance G-protein coupled receptors. Liquid chromatography mass spectrometry (LC-MS) identified peptide hormone gradients along the human and mouse gut that should enhance progress in gut physiology and therapeutics.

cell biology↗

bioRxiv2018

Human POMC processing in vitro and in vivo revealed by quantitative peptidomics

Human obesity can result from the aberrant production or processing of proopiomelanocortin (POMC) in hypothalamic neurons, but it is unclear which human POMC-derived peptides are most relevant to body weight regulation. To address this question, we analysed both hypothalamic neurons derived from human pluripotent stem cells (hPSCs) and primary human hypothalamic tissue using quantitative liquid chromatography tandem mass spectroscopy (LC-MS/MS). In both in vitro- and in vivo-derived samples, we found that POMC was processed into {beta}-melanocyte stimulating hormone ({beta}-MSH), whose existence in the human brain has been controversial. {beta}-MSH and desacetyl -MSH (d--MSH) were produced at roughly equimolar concentrations and in vast excess to acetylated -MSH (5-to 200-fold), suggesting that the importance of both d--MSH and {beta}-MSH to human obesity has been underestimated. Since body weight is sensitive to changes in MSH concentration, we asked whether hPSC-derived hypothalamic neurons could provide mechanistic insights into the processing and secretion of MSH peptides. We found that cultured human hypothalamic neurons appropriately trafficked POMC and its derivatives, and robustly (P<0.0001) secreted them when depolarised. Furthermore, the adipocyte-derived hormone leptin significantly (P<0.01) promoted their production of both d--MSH and {beta}-MSH. These results establish hPSC-derived hypothalamic neurons as a model system for studying human-specific aspects of POMC processing that might be therapeutically harnessed to treat obesity.

neuroscience↗