Common food additive carrageenan inhibits proglucagon expression and GLP-1 secretion by human enteroendocrine L-cells.

Proglucagon mRNA expression and GLP-1 secretion by cultured human L-cells (NCI-H716) were inhibited following exposure to λ-carrageenan, a commonly used additive in processed foods. Carrageenan is composed of sulfated or unsulfated galactose residues linked in alternating alpha-1,3 and beta-1,4 bonds and resembles the endogenous sulfated glycosaminoglycans. However, carrageenan has unusual alpha-1,3-galactosidic bonds, which are not innate to human cells and are implicated in immune responses. Exposure to carrageenan predictably causes inflammation, and carrageenan impairs glucose tolerance and contributes to insulin resistance. When cultured human L-cells were deprived overnight of glucose and serum and then exposed to high glucose, 10% FBS, and λ-carrageenan (1 µg/ml) for 10 minutes, 1 h, and 24 h, mRNA expression of proglucagon and secretion of GLP-1 were significantly reduced, compared to control cells not exposed to carrageenan. mRNA expression of proglucagon by mouse L-cells (STC-1) was also significantly reduced and supports the findings in the human cells. Exposure of co-cultured human intestinal epithelial cells (LS174T) to the spent media of the carrageenan-treated L-cells led to a decline in mRNA expression of GLUT-2 at 24 h. These findings suggest that ingestion of carrageenan-containing processed foods may impair the production of GLP-1, counteract the effect of GLP-1 receptor agonists and induce secondary effects on intestinal epithelial cells.

Proglucagon signalling in the rat Dorsomedial Hypothalamus - Physiology and high-fat diet-mediated alterations.

A relatively new pharmacological target in obesity treatment has been the preproglucagon (PPG) signalling, predominantly with glucagon-like peptide (GLP) 1 receptor agonists. As far as the PPG role within the digestive system is well recognised, its actions in the brain remain understudied. Here, we investigated PPG signalling in the Dorsomedial Hypothalamus (DMH), a structure involved in feeding regulation and metabolism, using in situ hybridisation, electrophysiology, and immunohistochemistry. Our experiments were performed on animals fed both control, and high-fat diet (HFD), uncovering HFD-mediated alterations. First, sensitivity to exendin-4 (Exn4, a GLP1R agonist) was shown to increase under HFD, with a higher number of responsive neurons. The amplitude of the response to both Exn4 and oxyntomodulin (Oxm) was also altered, diminishing its relationship with the cells' spontaneous firing rate. Not only neuronal sensitivity, but also GLP1 presence, and therefore possibly release, was influenced by HFD. Immunofluorescent labelling of the GLP1 showed changes in its density depending on the metabolic state (fasted/fed), but this effect was eliminated by HFD feeding. Interestingly, these dietary differences were absent after a period of restricted feeding, allowing for an anticipation of the alternating metabolic states, which suggests possible prevention of such outcome.

Blockade of glucagon increases muscle mass and alters fiber type composition in mice deficient in proglucagon-derived peptides.

AIMS/INTRODUCTION: Glucagon is secreted from pancreatic α-cells and plays an important role in amino acid metabolism in liver. Various animal models deficient in glucagon action show hyper-amino acidemia and α-cell hyperplasia, indicating that glucagon contributes to feedback regulation between the liver and the α-cells. In addition, both insulin and various amino acids, including branched-chain amino acids and alanine, participate in protein synthesis in skeletal muscle. However, the effect of hyperaminoacidemia on skeletal muscle has not been investigated. In the present study, we examined the effect of blockade of glucagon action on skeletal muscle using mice deficient in proglucagon-derived peptides (GCGKO mice). MATERIALS AND METHODS: Muscles isolated from GCGKO and control mice were analyzed for their morphology, gene expression and metabolites. RESULTS: GCGKO mice showed muscle fiber hypertrophy, and a decreased ratio of type IIA and an increased ratio of type IIB fibers in the tibialis anterior. The expression levels of myosin heavy chain (Myh) 7, 2, 1 and myoglobin messenger ribonucleic acid were significantly lower in GCGKO mice than those in control mice in the tibialis anterior. GCGKO mice showed a significantly higher concentration of arginine, asparagine, serine and threonine in the quadriceps femoris muscles, and also alanine, aspartic acid, cysteine, glutamine, glycine and lysine, as well as four amino acids in gastrocnemius muscles. CONCLUSIONS: These results show that hyperaminoacidemia induced by blockade of glucagon action in mice increases skeletal muscle weight and stimulates slow-to-fast transition in type II fibers of skeletal muscle, mimicking the phenotype of a high-protein diet.

High Protein Diet Feeding Aggravates Hyperaminoacidemia in Mice Deficient in Proglucagon-Derived Peptides.

(1) Background: Protein stimulates the secretion of glucagon (GCG), which can affect glucose metabolism. This study aimed to analyze the metabolic effect of a high-protein diet (HPD) in the presence or absence of proglucagon-derived peptides, including GCG and GLP-1. (2) Methods: The response to HPD feeding for 7 days was analyzed in mice deficient in proglucagon-derived peptides (GCGKO). (3) Results: In both control and GCGKO mice, food intake and body weight decreased with HPD and intestinal expression of Pepck increased. HPD also decreased plasma FGF21 levels, regardless of the presence of proglucagon-derived peptides. In control mice, HPD increased the hepatic expression of enzymes involved in amino acid metabolism without the elevation of plasma amino acid levels, except branched-chain amino acids. On the other hand, HPD-induced changes in the hepatic gene expression were attenuated in GCGKO mice, resulting in marked hyperaminoacidemia with lower blood glucose levels; the plasma concentration of glutamine exceeded that of glucose in HPD-fed GCGKO mice. (4) Conclusions: Increased plasma amino acid levels are a common feature in animal models with blocked GCG activity, and our results underscore that GCG plays essential roles in the homeostasis of amino acid metabolism in response to altered protein intake.

Blue Whiting (Micromesistius poutassou) Protein Hydrolysates Increase GLP-1 Secretion and Proglucagon Production in STC-1 Cells Whilst Maintaining Caco-2/HT29-MTX Co-Culture Integrity.

Inducing the feeling of fullness via the regulation of satiety hormones presents an effective method for reducing excess energy intake and, in turn, preventing the development of obesity. In this study, the ability of blue whiting soluble protein hydrolysates (BWSPHs) and simulated gastrointestinal digested (SGID) BWSPHs, to modulate the secretion and/or production of satiety hormones, such as glucagon-like peptide-1 (GLP-1), cholecystokinin (CCK) and peptide YY (PYY), was assessed in murine enteroendocrine STC-1 cells. All BWSPHs (BW-SPH-A to BW-SPH-F) (1.0% w/v dw) increased active GLP-1 secretion and proglucagon production in STC-1 cells compared to the basal control (Krebs-Ringer buffer) (p < 0.05). The signaling pathway activated for GLP-1 secretion was also assessed. A significant increase in intracellular calcium levels was observed after incubation with all BWSPHs (p < 0.05) compared with the control, although none of the BWSPHs altered intracellular cyclic adenosine monophosphate (cAMP) concentrations. The secretagogue effect of the leading hydrolysate was diminished after SGID. Neither pre- nor post-SGID hydrolysates affected epithelial barrier integrity or stimulated interleukin (IL)-6 secretion in differentiated Caco-2/HT-29MTX co-cultured cells. These results suggest a role for BWSPH-derived peptides in satiety activity; however, these peptides may need to be protected by some means to avoid loss of activity during gastrointestinal transit.

Altered islet prohormone processing: a cause or consequence of diabetes?

Peptide hormones are first produced as larger precursor prohormones that require endoproteolytic cleavage to liberate the mature hormones. A structurally conserved but functionally distinct family of nine prohormone convertase enzymes (PCs) are responsible for cleavage of protein precursors, of which PC1/3 and PC2 are known to be exclusive to neuroendocrine cells and responsible for prohormone cleavage. Differential expression of PCs within tissues defines prohormone processing; whereas glucagon is the major product liberated from proglucagon via PC2 in pancreatic α-cells, proglucagon is preferentially processed by PC1/3 in intestinal L cells to produce glucagon-like peptides 1 and 2 (GLP-1, GLP-2). Beyond our understanding of processing of islet prohormones in healthy islets, there is convincing evidence that proinsulin, pro-islet amyloid polypeptide (proIAPP), and proglucagon processing is altered during prediabetes and diabetes. There is predictive value of elevated circulating proinsulin or proinsulin-to-C-peptide ratio for progression to type 2 diabetes, and elevated proinsulin or proinsulin-to-C-peptide ratio is predictive for development of type 1 diabetes in at-risk groups. After onset of diabetes, patients have elevated circulating proinsulin and proIAPP, and proinsulin may be an autoantigen in type 1 diabetes. Furthermore, preclinical studies reveal that α-cells have altered proglucagon processing during diabetes, leading to increased GLP-1 production. We conclude that despite strong associative data, current evidence is inconclusive on the potential causal role of impaired prohormone processing in diabetes and suggest that future work should focus on resolving the question of whether altered prohormone processing is a causal driver or merely a consequence of diabetes pathology.

Activation of PPG neurons following acute stressors differentially involves hindbrain serotonin in male rats.

Within the hindbrain, serotonin (5-HT) functions as a modulator of the central glucagon-like peptide-1 (GLP-1) system. This interaction between 5-HT and GLP-1 is achieved via 5-HT2C and 5-HT3 receptors and is relevant for GLP-1-mediated feeding behavior. The central GLP-1 system is activated by various stressors, activates the hypothalamic pituitary adrenocortical (HPA) axis, and contributes to stress-related behaviors. Whether 5-HT modulates GLP-1's role in the stress response in unknown. We hypothesized that the serotonergic modulation of GLP-1-producing neurons (i.e., PPG neurons) is stimuli-specific and that stressed-induced PPG activity is one of the modalities in which 5-HT plays a role. In this study, we investigated the roles of 5-HT2C and 5-HT3 receptors in mediating the activation of PPG neurons in the nucleus tractus solitarius (NTS) following exposure to three different acute stressors: lithium chloride (LiCl), noncontingent cocaine (Coc), and novel restraint stress (RES). Results showed that increased c-Fos expression in PPG neurons following LiCl and RES-but not Coc-is dependent on hindbrain 5-HT2C and 5-HT3 receptor signaling. Additionally, stressors that depend on 5-HT signaling to activate PPG neurons (i.e., LiCl and RES) increased c-Fos expression in 5-HT-expressing neurons within the caudal raphe (CR), specifically in the raphe magnus (RMg). Finally, we showed that RMg neurons innervate NTS PPG neurons and that some of these PPG neurons lie in close proximity to 5-HT axons, suggesting RMg 5-HT-expressing neurons are the source of 5-HT input responsible for engaging NTS PPG neurons. Together, these findings identify a direct RMg to NTS pathway responsible for the modulatory effect of 5-HT on the central GLP-1 system-specifically via activation of 5-HT2C and 5-HT3 receptors-in the facilitation of acute stress responses.

Central and peripheral GLP-1 systems independently suppress eating.

The anorexigenic peptide glucagon-like peptide-1 (GLP-1) is secreted from gut enteroendocrine cells and brain preproglucagon (PPG) neurons, which, respectively, define the peripheral and central GLP-1 systems. PPG neurons in the nucleus tractus solitarii (NTS) are widely assumed to link the peripheral and central GLP-1 systems in a unified gut-brain satiation circuit. However, direct evidence for this hypothesis is lacking, and the necessary circuitry remains to be demonstrated. Here we show that PPGNTS neurons encode satiation in mice, consistent with vagal signalling of gastrointestinal distension. However, PPGNTS neurons predominantly receive vagal input from oxytocin-receptor-expressing vagal neurons, rather than those expressing GLP-1 receptors. PPGNTS neurons are not necessary for eating suppression by GLP-1 receptor agonists, and concurrent PPGNTS neuron activation suppresses eating more potently than semaglutide alone. We conclude that central and peripheral GLP-1 systems suppress eating via independent gut-brain circuits, providing a rationale for pharmacological activation of PPGNTS neurons in combination with GLP-1 receptor agonists as an obesity treatment strategy.

The RNA-binding protein, HuD regulates proglucagon biosynthesis in pancreatic α cells.

Glucagon is a peptide hormone generated by pancreatic α cells. It is the counterpart of insulin and plays an essential role in the regulation of blood glucose level. Therefore, a tight regulation of glucagon levels is pivotal to maintain homeostasis of blood glucose. However, little is known about the mechanisms regulating glucagon biosynthesis. In this study, we demonstrate that the RNA-binding protein HuD regulates glucagon expression in pancreatic α cells. HuD was found in α cells from mouse pancreatic islet and mouse glucagonoma αTC1 cell line. Ribonucleoprotein immunoprecipitation analysis, followed by RT-qPCR showed the association of HuD with glucagon mRNA. Knockdown of HuD resulted in a reduction in both proglucagon expression and cellular glucagon level by decreasing its de novo synthesis. Reporter analysis using the EGFP reporter containing 3' untranslated region (3'UTR) of glucagon mRNA showed that HuD regulates proglucagon expression via its 3'UTR. In addition, the relative level of glucagon in the islets and plasma was lower in HuD knockout (KO) mice compared to age-matched control mice. Taken together, these results suggest that HuD is a novel factor regulating the biosynthesis of proglucagon in pancreatic α cells.

Glucagon receptor antagonism promotes the production of gut proglucagon-derived peptides in diabetic mice.

Glucagon is an essential regulator of glucose homeostasis, particularly in type 2 diabetes (T2D). Blocking the glucagon receptor (GCGR) in diabetic animals and humans has been shown to alleviate hyperglycemia and increase circulating glucagon-like peptide-1 (GLP-1) levels. However, the origin of the upregulated GLP-1 remains to be clarified. Here, we administered high-fat diet + streptozotocin-induced T2D mice and diabetic db/db mice with REMD 2.59, a fully competitive antagonistic human GCGR monoclonal antibody (mAb) for 12 weeks. GCGR mAb treatment decreased fasting blood glucose levels and increased plasma GLP-1 levels in the T2D mice. In addition, GCGR mAb upregulated preproglucagon gene expression and the contents of gut proglucagon-derived peptides, particularly GLP-1, in the small intestine and colon. Notably, T2D mice treated with GCGR mAb displayed a higher L-cell density in the small intestine and colon, which was associated with increased numbers of LK-cells coexpressing GLP-1 and glucose-dependent insulinotropic polypeptide and reduced L-cell apoptosis. Furthermore, GCGR mAb treatment upregulated GLP-1 production in the pancreas, which was detected at lower levels than in the intestine. Collectively, these results suggest that GCGR mAb can increase intestinal GLP-1 production and L-cell number by enhancing LK-cell expansion and inhibiting L-cell apoptosis in T2D.

In vitro modulation of glucagon-like peptide release by DPP-IV inhibitory polyphenol-polysaccharide conjugates of sprouted quinoa yoghurt.

Glucagon-like peptide-1 (GLP-1) is an important signal in the peripheral and neural systems, which contributes to the maintenance of glucose and energy homeostasis. In this study, 1H NMR validated polyphenols and polysaccharides extracted from sprouted quinoa yoghurt were used as isolates and conjugates to upregulate the stimulation of GLP-1 release in NCI-H716 cells. In addition, we explored their effect on proglucagon and prohormone convertase 3 mRNA expressions, HNF-3γ and CCK-2R gene protein expression, as well as cytosolic calcium release. Variations in concentration showed a dose-dependent GLP-1 stimulation, and were significantly optimized by germination. Proglucagon mRNA expression in NCI-H716 cells was upregulated, and was relatively highest with QYPSP1 treatments in a 2.68 fold. The results suggested that the conjugates had greater potential to stimulate GLP-1 release than their isolates. Sprouted quinoa yoghurt could therefore be a potential functional food useful to regulate glucose and energy homeostasis.

Proglucagon-Derived Peptides, Glucose-Dependent Insulinotropic Polypeptide, and Dipeptidyl Peptidase-4-Mechanisms of Action in Adipose Tissue.

Proglucagon-derived peptides (PGDPs) and related gut hormones exemplified by glucose-dependent insulinotropic polypeptide (GIP) regulate energy disposal and storage through actions on metabolically sensitive organs, including adipose tissue. The actions of glucagon, glucagon-like peptide (GLP)-1, GLP-2, GIP, and their rate-limiting enzyme dipeptidyl peptidase-4, include direct and indirect regulation of islet hormone secretion, food intake, body weight, all contributing to control of white and brown adipose tissue activity. Moreover, agents mimicking actions of these peptides are in use for the therapy of metabolic disorders with disordered energy homeostasis such as diabetes, obesity, and intestinal failure. Here we highlight current concepts and mechanisms for direct and indirect actions of these peptides on adipose tissue depots. The available data highlight the importance of indirect peptide actions for control of adipose tissue biology, consistent with the very low level of endogenous peptide receptor expression within white and brown adipose tissue depots. Finally, we discuss limitations and challenges for the interpretation of available experimental observations, coupled to identification of enduring concepts supported by more robust evidence.

Aspartame supplementation in starter accelerates small intestinal epithelial cell cycle and stimulates secretion of glucagon-like peptide-2 in pre-weaned lambs.

The objective of this study was to test the hypothesis that aspartame supplementation in starter diet accelerates small intestinal cell cycle by stimulating secretion and expression of glucagon-like peptide -2 (GLP-2) in pre-weaned lambs using animal and cell culture experiments. In vivo, twelve 14-day-old lambs were selected and allocated randomly to two groups; one was treated with plain starter diet (Con, n = 6) and the other was treated with starter supplemented with 200 mg of aspartame/kg starter (APM, n = 6). Results showed that the lambs received APM treatment for 35 d had higher (p < .05) GLP-2 concentration in the plasma and greater jejunum weight/live body weight (BW) and jejunal crypt depth. Furthermore, APM treatment significantly upregulated (p < .05) the mRNA expression of cyclin D1 in duodenum; and cyclin A2, cyclin D1, cyclin-dependent kinases 6 (CDK6) in jejunum; and cyclin A2, cyclin D1, CDK4 in ileum. Moreover, APM treatment increased (p < .05) the mRNA expression of glucagon (GCG), insulin-like growth factor 1 (IGF-1) in the jejunum and ileum and mRNA expression of GLP-2 receptor (GLP-2R) in the jejunum. In vitro, when jejunal cells were treated with GLP-2 for 2 hr, the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) OD, IGF-1 concentration, and the mRNA expression of IGF-1, cyclin D1 and CDK6 were increased (p < .05). Furthermore, IGF-1 receptor (IGF-1R) inhibitor decreased (p < .05) the mRNA expression of IGF-1, cyclin A2, cyclin D1 and CDK6 in GLP-2 treatment jejunal cells. These results suggest that aspartame supplementation in starter accelerates small intestinal cell cycle that may, in part, be related to stimulate secretion and expression of GLP-2 in pre-weaning lambs. Furthermore, GLP-2 can indirectly promote the proliferation of jejunal cells mainly through the IGF-1 pathway. These findings provide new insights into nutritional interventions that promote the development of small intestines in young ruminants.

Short communication: Expression of transcripts for proglucagon, glucose-dependent insulinotropic peptide, peptide YY, and their cognate receptors, in feline peripheral tissues.

Gastrointestinal hormone based therapies are being investigated for treating diabetes in cats; however, the tissue distribution of these hormones and their cognate receptors remain largely understudied. We determined the distribution of transcripts for the gut hormones proglucagon (Gcg), glucose-dependent insulinotropic peptide (Gip), peptide YY (Pyy), and their receptors (Glp1r, Gipr, Npy2r), in feline peripheral tissues. The Gcg, Gip and Pyy mRNA were expressed in the gut, with higher Gcg and Pyy abundance in the lower gut. Interestingly, Glp1r and Npy2r mRNA were expressed in multiple peripheral tissues including the gut, pancreas and liver, whereas, Gipr mRNA was restricted to the stomach and adipose tissues. The localized mRNA expression of Gcg and Pyy in the gut, but the extensive distribution of Glp1r and Npy2r in several peripheral tissues suggests that these hormones may have pleiotropic physiological functions in cats.

ß Cell tone is defined by proglucagon peptides through cAMP signaling.

Paracrine interactions between pancreatic islet cells have been proposed as a mechanism to regulate hormone secretion and glucose homeostasis. Here, we demonstrate the importance of proglucagon-derived peptides (PGDPs) for α to ß cell communication and control of insulin secretion. Signaling through this system occurs through both the glucagon-like peptide receptor (Glp1r) and glucagon receptor (Gcgr). Loss of PGDPs, or blockade of their receptors, decreases insulin secretion in response to both metabolic and nonmetabolic stimulation of mouse and human islets. This effect is due to reduced ß cell cAMP and affects the quantity but not dynamics of insulin release, indicating that PGDPs dictate the magnitude of insulin output in an isolated islet. In healthy mice, additional factors that stimulate cAMP can compensate for loss of PGDP signaling; however, input from α cells is essential to maintain glucose tolerance during the metabolic stress induced by high-fat feeding. These findings demonstrate an essential role for α cell regulation of ß cells, raising the possibility that abnormal paracrine signaling contributes to impaired insulin secretion in diabetes. Moreover, these findings support reconsideration of the role for α cells in postprandial glucose control.

GLP-2 receptor signaling controls circulating bile acid levels but not glucose homeostasis in Gcgr-/- mice and is dispensable for the metabolic benefits ensuing after vertical sleeve gastrectomy.

OBJECTIVE: Therapeutic interventions that improve glucose homeostasis such as attenuation of glucagon receptor (Gcgr) signaling and bariatric surgery share common metabolic features conserved in mice and humans. These include increased circulating levels of bile acids (BA) and the proglucagon-derived peptides (PGDPs), GLP-1 and GLP-2. Whether BA acting through TGR5 (Gpbar1) increases PGDP levels in these scenarios has not been examined. Furthermore, although the importance of GLP-1 action has been interrogated in Gcgr-/- mice and after bariatric surgery, whether GLP-2 contributes to the metabolic benefits of these interventions is not known. METHODS: To assess whether BA acting through Gpbar1 mediates improved glucose homeostasis in Gcgr-/- mice we generated and characterized Gcgr-/-:Gpbar1-/- mice. The contribution of GLP-2 receptor (GLP-2R) signaling to intestinal and metabolic adaptation arising following loss of the Gcgr was studied in Gcgr-/-:Glp2r-/- mice. The role of the GLP-2R in the metabolic improvements evident after bariatric surgery was studied in high fat-fed Glp2r-/- mice subjected to vertical sleeve gastrectomy (VSG). RESULTS: Circulating levels of BA were markedly elevated yet similar in Gcgr-/-:Gpbar1+/+ vs. Gcgr-/-:Gpbar1-/- mice. Loss of GLP-2R lowered levels of BA in Gcgr-/- mice. Gcgr-/-:Glp2r-/- mice also exhibited shifts in the proportion of circulating BA species. Loss of Gpbar1 did not impact body weight, intestinal mass, or glucose homeostasis in Gcgr-/- mice. In contrast, small bowel growth was attenuated in Gcgr-/-:Glp2r-/- mice. The improvement in glucose tolerance, elevated circulating levels of GLP-1, and glucose-stimulated insulin levels were not different in Gcgr-/-:Glp2r+/+ vs. Gcgr-/-:Glp2r-/- mice. Similarly, loss of the GLP-2R did not attenuate the extent of weight loss and improvement in glucose control after VSG. CONCLUSIONS: These findings reveal that GLP-2R controls BA levels and relative proportions of BA species in Gcgr-/- mice. Nevertheless, the GLP-2R is not essential for i) control of body weight or glucose homeostasis in Gcgr-/- mice or ii) metabolic improvements arising after VSG in high fat-fed mice. Furthermore, despite elevations of circulating levels of BA, Gpbar1 does not mediate elevated levels of PGDPs or major metabolic phenotypes in Gcgr-/- mice. Collectively these findings refine our understanding of the relationship between Gpbar1, elevated levels of BA, PGDPs, and the GLP-2R in amelioration of metabolic derangements arising following loss of Gcgr signaling or after vertical sleeve gastrectomy.

ß Cell GLP-1R Signaling Alters α Cell Proglucagon Processing after Vertical Sleeve Gastrectomy in Mice.

Bariatric surgery, such as vertical sleeve gastrectomy (VSG), causes high rates of type 2 diabetes remission and remarkable increases in postprandial glucagon-like peptide-1 (GLP-1) secretion. GLP-1 plays a critical role in islet function by potentiating glucose-stimulated insulin secretion; however, the mechanisms remain incompletely defined. Therefore, we applied a murine VSG model to an inducible ß cell-specific GLP-1 receptor (GLP-1R) knockout mouse model to investigate the role of the ß cell GLP-1R in islet function. Our data show that loss of ß cell GLP-1R signaling decreases α cell GLP-1 expression after VSG. Furthermore, we find a ß cell GLP-1R-dependent increase in α cell expression of the prohormone convertase required for the production of GLP-1 after VSG. Together, the findings herein reveal two concepts. First, our data support a paracrine role for α cell-derived GLP-1 in the metabolic benefits observed after VSG. Second, we have identified a role for the ß cell GLP-1R as a regulator of α cell proglucagon processing.

Gastrectomy with Roux-en-Y reconstruction as a lean model of bariatric surgery.

BACKGROUND: Altered enteroendocrine hormone responses are widely believed to underlie the beneficial effects of bariatric surgery in type 2 diabetes. While elevated postprandial glucagon-like peptide-1 (GLP-1) is considered one of the mediators, increased postprandial glucagon levels have recently been implicated. OBJECTIVES: We investigated hormonal responses in lean patients after prophylactic total gastrectomy (PTG), as a model of Roux-en-Y gastric bypass without the confounding effects of obesity or massive weight loss. SETTING: University hospital, United Kingdom. METHODS: Ten participants after PTG and 9 healthy volunteers were recruited for oral glucose tolerance tests. Plasma glucose, insulin, GLP-1, peptide YY, glucose-dependent insulinotropic-polypeptide, glucagon, oxyntomodulin, glucagon(1-61), and glicentin levels were assessed using immunoassays and/or mass spectrometry. RESULTS: PTG participants exhibited accelerated plasma glucose appearance, followed, in 3 of 10 cases, by hypoglycemia (<3 mM glucose). Plasma GLP-1, peptide YY, glucose-dependent insulinotropic-polypeptide, glicentin, and oxyntomodulin responses were elevated, and glucagon appeared to rise in PTG participants when measured with a glucagon-specific enzyme-linked immunosorbent assay. We revisited the specificity of this assay, and demonstrated significant cross-reactivity with glicentin and oxyntomodulin at concentrations observed in PTG plasma. Reassessment of glucagon with the same assay using a modified protocol, and by liquid chromatography-mass spectrometry, demonstrated suppression of glucagon secretion after oral glucose tolerance tests in both PTG and control cohorts. CONCLUSIONS: Care should be taken when assessing glucagon levels in the presence of elevated plasma levels of other proglucagon products. Substantial elevation of GLP-1 and insulin responses after PTG likely contribute to the observed hypoglycemia, and mirror similar hormone levels and complications observed in bariatric weight loss patients.

Enteroendocrine L Cells Sense LPS after Gut Barrier Injury to Enhance GLP-1 Secretion.

Glucagon-like peptide 1 (GLP-1) is a hormone released from enteroendocrine L cells. Although first described as a glucoregulatory incretin hormone, GLP-1 also suppresses inflammation and promotes mucosal integrity. Here, we demonstrate that plasma GLP-1 levels are rapidly increased by lipopolysaccharide (LPS) administration in mice via a Toll-like receptor 4 (TLR4)-dependent mechanism. Experimental manipulation of gut barrier integrity after dextran sodium sulfate treatment, or via ischemia/reperfusion experiments in mice, triggered a rapid rise in circulating GLP-1. This phenomenon was detected prior to measurable changes in inflammatory status and plasma cytokine and LPS levels. In human subjects, LPS administration also induced GLP-1 secretion. Furthermore, GLP-1 levels were rapidly increased following the induction of ischemia in the human intestine. These findings expand traditional concepts of enteroendocrine L cell biology to encompass the sensing of inflammatory stimuli and compromised mucosal integrity, linking glucagon-like peptide secretion to gut inflammation.

Detailed characterisation of STC-1 cells and the pGIP/Neo sub-clone suggests the incretin hormones are translationally regulated.

STC-1 is a heterogeneous plurihormonal cell line producing several prominent gut peptide hormones. pGIP/Neo is a genetically selected sub-clone of STC-1 with augmented levels of glucose-dependent insulinotropic peptide (GIP). Morphometric parameters, hormone concentrations, mRNA transcripts, hormone immunocytochemistry and nutrient utilisation/production of these two cell lines were compared. Proglucagon-derived peptides (Glucagon-like peptide-1 (GLP-1) and - 2(GLP-2)) were lower in sub-clone cells than progenitor cells. High Content Analysis found altered intracellular GLP-1, GIP, cholecystokinin (CCK) and peptide YY (PYY) levels and differing hormone co-localisation. The proportion pGIP/Neo cells containing GIP immunoreactivity (82%) was greater than STC-1 (65%), as were the proportion with 'GIP only', 'GLP-1+GIP' or 'GIP+PYY' immunoreactivity. Most surprisingly mRNA transcripts of the proglucagon and GIP genes were inversely correlated to the levels of their translated peptides. This strongly suggests that proglucagon and GIP are encoded on 'translationally regulated genes' - a characteristic possessed by other endocrine hormones. Metabolomic profiling revealed differences in cellular nutrient utilisation/production and that under normal culture conditions both cell lines exhibit signs of overflow metabolism. These studies provide an insight into the metabolism and properties of these valuable cells, suggesting for the first time that incretin hormone genes are translationally regulated.