Gut Peptides Research Articles

Increased gut permeability in cancer cachexia: mechanisms and clinical relevance.

Intestinal disorders often occur in cancer patients, in association with body weight loss, and this alteration is commonly attributed to the chemotherapy. Here, using a mouse model of cancer cachexia induced by ectopic transplantation of C26 cancer cells, we discovered a profound alteration in the gut functions (gut permeability, epithelial turnover, gut immunity, microbial dysbiosis) independently of any chemotherapy. These alterations occurred independently of anorexia and were driven by interleukin 6. Gut dysfunction was found to be resistant to treatments with an anti-inflammatory bacterium (Faecalibacterium prausnitzii) or with gut peptides involved in intestinal cell renewal (teduglutide, a glucagon-like peptide 2 analogue). The translational value of our findings was evaluated in 152 colorectal and lung cancer patients with or without cachexia. The serum level of the lipopolysaccharide-binding protein, often presented as a reflection of the bacterial antigen load, was not only increased in cachectic mice and cancer patients, but also strongly correlated with the serum IL-6 level and predictive of death and cachexia occurrence in these patients. Altogether, our data highlight profound alterations of the intestinal homeostasis in cancer cachexia occurring independently of any chemotherapy and food intake reduction, with potential relevance in humans. In addition, we point out the lipopolysaccharide-binding protein as a new biomarker of cancer cachexia related to gut dysbiosis.

Non-endocannabinoid N-acylethanolamines and 2-monoacylglycerols in the intestine.

This review focuses on recent findings of the physiological and pharmacological role of non-endocannabinoid N-acylethanolamines (NAEs) and 2-monoacylglycerols (2-MAGs) in the intestine and their involvement in the gut-brain signalling. Dietary fat suppresses food intake, and much research concerns the known gut peptides, for example, glucagon-like peptide-1 (GLP-1) and cholecystokinin (CCK). NAEs and 2-MAGs represent another class of local gut signals most probably involved in the regulation of food intake. We discuss the putative biosynthetic pathways and targets of NAEs in the intestine as well as their anorectic role and changes in intestinal levels depending on the dietary status. NAEs can activate the transcription factor PPARα, but studies to evaluate the role of endogenous NAEs are generally lacking. Finally, we review the role of diet-derived 2-MAGs in the secretion of anorectic gut peptides via activation of GPR119. Both PPARα and GPR119 have potential as pharmacological targets for the treatment of obesity and the former for treatment of intestinal inflammation. LINKED ARTICLES: This article is part of a themed section on 8th European Workshop on Cannabinoid Research. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.10/issuetoc.

Sexually Dimorphic Ano2 Expression in Nodose Neurons Determines CCK‐mediated Satiation and Obesity in Heterozygote Male Mice

The gut peptide cholecystokinin (CCK) contributes to neurally mediated post prandial satiety. Our previous results indicate that high fat diet downregulates the CCK‐sensitive anoctamins (Ano1 and Ano2) that function as calcium‐activated chloride channels (CaCCs) in intestinal vagal afferents. Using Cre‐loxP system, we generated heterozygote (HET) Nav1.8CreAno2fl/WT mice. The endogenous Scn10a promoter drives Cre expression (Nav1.8Cre) selectively in sensory neurons of the nodose, dorsal root and trigeminal ganglia where loxP sites flanked Ano2 exon 12. The male HET mice exhibited an obese phenotype and decreased sensitivity to CCK. To assess the Ano2 expression in HET vs. control (Nav1.8Cre+/−) mice, we first did qRT‐PCR on whole nodose ganglia obtained under anesthesia at 50 weeks of age and found no significant difference in Ano2 mRNA expression (n=4, p>0.05). Ano2 gene may be expressed in non‐neuronal cells of nodose ganglia while Cre expression is selective to neurons. We measured Ano2 mRNAs in nodose neurons cultured for 16–18 hours using single cell qRT‐PCR. Cre positive neurons were 57% (12 of 21) in HET and 74% (29 of 39) in control, indicating a lower efficiency of Cre expression in HET. Correspondingly, Ano2 positive neurons were 48% (10 of 21) and 62% (24 of 39), respectively, indicating the Ano2 allelic deletion in HET. The combined Cre and Ano2 positivity in HET was 29% (6 of 21) vs. 54% (21 of 39) in control. Most importantly the Ano2 mRNA levels in HET neurons (n=10) were 20% of the levels measured in control neurons (n=24, p<0.05) (0.20±0.16 vs. 1.00±0.57). The Ano2 expression in individual nodose neurons of female mice did not show a reduction or even higher in HET (n=27) vs. control (n=28) (2.58±1.48 vs. 1.42±1.27, p>0.05). The female HET mice remained CCK sensitive indicating a sexual dimorphism. We conclude that differences in expression of Ano2 mRNA support our findings of CCK insensitivity and selective obesity in male but not female HET mice (HL14388).Support or Funding InformationNIH HL14388This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Duodenal infusion of fatty acids differentially affects plasma glucagon-like peptide-1 and ghrelin concentrations in sheep.

The aim of this study was to investigate how intraduodenal infusions of fatty acids (FA) affect appetite-related gut peptides such as glucagon-like peptide-1 (GLP-1) and ghrelin in sheep. We hypothesized that these peptides can be highly reactive to unsaturated long-chain FA, because they are well known to decrease dry matter intake (DMI). Four ewes were fitted with a duodenal cannula and a jugular vein catheter for a 6-h duodenal infusion of the 9 FA (C8:0, C10:0, C12:0, C14:0, C16:0, C18:0, C18:1, C18:2, and C18:3) and water (control). The concentration of each FA was 1.6 g per metabolic body weight (BW), approximately corresponding to the amount of supplemented fat in a standard dairy cow diet. Each infusion was separated by at least 2 d. During the infusion period, blood samples were collected periodically to determine changes in plasma GLP-1, ghrelin, and metabolite concentrations. Duodenal infusions of C18:1, C18:2, and C18:3 led to higher plasma GLP-1 (P < 0.05) and lower glucose (P < 0.05) than control. Plasma ghrelin concentrations were greater in C18:1 and C18:3 infusions than control (P < 0.05). Plasma ketone bodies were higher in C8:0 and C10:0 infusions (P < 0.05), but plasma triglyceride concentrations were lower in C8:0, C10:0, C12:0, and C16:0 infusions (P < 0.05) than control. Fatty acid infusions except for C18:3 led to higher plasma NEFA concentrations than control (P < 0.05). These results confirmed that the hypophagic effect of dietary unsaturated long-chain FA is mediated by GLP-1 (an anorexigenic effect) secretion. However, we also observed higher plasma ghrelin (an orexigenic effect) partially by unsaturated long-chain FA. Thus, the gut peptide secretions when ruminant animals ingest FA supplements would complexly affect satiety and further studies are needed to determine their each impact on DMI.

Obesity Impairs Oligopeptide/Amino Acid-Induced Ghrelin Release and Smooth Muscle Contractions in the Human Proximal Stomach.

The satiation properties of proteins involve effects on gut peptide release and gastrointestinal motility which may be altered during obesity. This study compares the in vitro response and role of amino acid (AA) taste receptors (TASR) in the effect of AAs and a casein hydrolysate on ghrelin release and smooth muscle (SM) contractions in the proximal gut of lean and obese patients. Basal ghrelin release, measured from mucosal segments, is maximal in the fundus and decreased distally. Obesity selectively impaires the stimulatory effect of a casein hydrolyaste on ghrelin release in the fundus but does not affect its inhibitory effect in the small intestine (SI). The SM contractions induced by a casein hydrolysate and AAs are stronger in strips from the SI than from the fundus but are reduced in the stomach of obese patients. The region-dependent expression of AA-TASRs in the mucosa and SM layer is affected by obesity. Most of the AA-induced responses are reduced by the umami antagonist, lactisole. l-Met-induced responses involve bitter taste receptors. Region-specific targeting of AA taste receptors on both enteroendocrine and SM cells with specific AA-enriched diets might be a useful strategy to combat obesity as well as hypomotility disorders.

Biochemical properties and biological actions of obestatin and its relevence in type 2 diabetes

Obestatin was initially discovered in rat stomach extract, and although it is principally produced in the gastric mucosa, it can be found throughout the gastrointestinal tract. This 23-amino acid C-terminally amidated peptide is derived from preproghrelin and has been ascribed a wide range of metabolic effects relevant to type 2 diabetes. Obestatin reportedly inhibits gastrointestinal motility, reduces food intake and lowers body weight and improves lipid metabolism. Furthermore, it appears to exert actions on the pancreatic β-cell, most notably increasing β-cell mass and upregulating genes associated with insulin production and β-cell regeneration, with relevance to type 2 diabetes. It is becoming evident that obestatin also exerts pleiotropic effects on the cardiovascular system, possibly modulating blood pressure, endothelial function and triggering cardioprotective mechanisms, which may be important in determining cardiovascular outcomes in type 2 diabetes. Furthermore, it seems that like other gut peptides obestatin has neuroprotective properties. This review examines the biochemical properties of the obestatin peptide (its structure, sequence, stability and distribution) and the candidate receptors through which it may act. It provides a balanced examination of the reported pancreatic and extrapancreatic actions of obestatin and evaluates its potential relevance with respect to diabetes therapy, together with discussion of direct evidence linking alterations in obestatin signalling with obesity/diabetes and other diseases.

Incretin hormones: Their role in health and disease.

Incretin hormones are gut peptides that are secreted after nutrient intake and stimulate insulin secretion together with hyperglycaemia. GIP (glucose-dependent insulinotropic polypeptide) und GLP-1 (glucagon-like peptide-1) are the known incretin hormones from the upper (GIP, K cells) and lower (GLP-1, L cells) gut. Together, they are responsible for the incretin effect: a two- to three-fold higher insulin secretory response to oral as compared to intravenous glucose administration. In subjects with type 2 diabetes, this incretin effect is diminished or no longer present. This is the consequence of a substantially reduced effectiveness of GIP on the diabetic endocrine pancreas, and of the negligible physiological role of GLP-1 in mediating the incretin effect even in healthy subjects. However, the insulinotropic and glucagonostatic effects of GLP-1 are preserved in subjects with type 2 diabetes to the degree that pharmacological stimulation of GLP-1 receptors significantly reduces plasma glucose and improves glycaemic control. Thus, it has become a parent compound of incretin-based glucose-lowering medications (GLP-1 receptor agonists and inhibitors of dipeptidyl peptidase-4 or DPP-4). GLP-1, in addition, has multiple effects on various organ systems. Most relevant are a reduction in appetite and food intake, leading to weight loss in the long term. Since GLP-1 secretion from the gut seems to be impaired in obese subjects, this may even indicate a role in the pathophysiology of obesity. Along these lines, an increased secretion of GLP-1 induced by delivering nutrients to lower parts of the small intestines (rich in L cells) may be one factor (among others like peptide YY) explaining weight loss and improvements in glycaemic control after bariatric surgery (e.g., Roux-en-Y gastric bypass). GIP and GLP-1, originally characterized as incretin hormones, have additional effects in adipose cells, bone, and the cardiovascular system. Especially, the latter have received attention based on recent findings that GLP-1 receptor agonists such as liraglutide reduce cardiovascular events and prolong life in high-risk patients with type 2 diabetes. Thus, incretin hormones have an important role physiologically, namely they are involved in the pathophysiology of obesity and type 2 diabetes, and they have therapeutic potential that can be traced to well-characterized physiological effects.

Sugar sensor genes in the murine gastrointestinal tract display a cephalocaudal axis of expression and a diurnal rhythm.

Distributed along the length of the gastrointestinal (GI) tract are nutrient sensing cells that release numerous signaling peptides influencing GI function, nutrient homeostasis and energy balance. Recent studies have shown a diurnal rhythm in GI nutrient sensing, but the mechanisms responsible for rhythmicity are poorly understood. In this report we studied murine GI sugar sensor gene and protein expression levels in the morning (7 AM) and evening (7 PM). Sweet taste receptor ( tas1r2/tas1r3/gnat3/gnat1) sugar transporter ( slc5a1, slc2a2, slc2a5) and putative sugar sensor ( slc5a4a and slc5a4b) gene expression levels were highest in tongue and proximal and distal small intestine, respectively. Clock gene ( cry2/arntl) activity was detected in all regions studied. Slc5a4a and slc5a4b gene expression showed clear diurnal rhythmicity in the small intestine and stomach, respectively, although no rhythmicity was detected in SGLT3 protein expression. Tas1r2, tas1r3, gnat1, and gcg displayed a limited rhythm in gene expression in proximal small intestine. Microarray analysis revealed a diurnal rhythm in gut peptide gene expression in tongue (7 AM vs. 7 PM) and in silico promoter analysis indicated intestinal sugar sensors and transporters possessed the canonical E box elements necessary for clock gene control over gene transcription. In this report we present evidence of a diurnal rhythm in genes that are responsible for intestinal nutrient sensing that is most likely controlled by clock gene activity. Disturbances in clock gene/nutrient sensing interactions may be important in the development of diet-related diseases, such as obesity and diabetes.

Genome Wide Mapping of Peptidases in Rhodnius prolixus: Identification of Protease Gene Duplications, Horizontally Transferred Proteases and Analysis of Peptidase A1 Structures, with Considerations on Their Role in the Evolution of Hematophagy in Triatominae.

Triatominae is a subfamily of the order Hemiptera whose species are able to feed in the vertebrate blood (i.e., hematophagy). This feeding behavior presents a great physiological challenge to insects, especially in Hemipteran species with a digestion performed by lysosomal-like cathepsins instead of the more common trypsin-like enzymes. With the aim of having a deeper understanding of protease involvement in the evolutionary adaptation for hematophagy in Hemipterans, we screened peptidases in the Rhodnius prolixus genome and characterized them using common blast (NCBI) and conserved domain analyses (HMMER/blast manager software, FAT, plus PFAM database). We compared the results with available sequences from other hemipteran species and with 18 arthropod genomes present in the MEROPS database. Rhodnius prolixus contains at least 433 protease coding genes, belonging to 71 protease families. Seven peptidase families in R. prolixus presented higher gene numbers when compared to other arthropod genomes. Further analysis indicated that a gene expansion of the protease family A1 (Eukaryotic aspartyl protease, PF00026) might have played a major role in the adaptation to hematophagy since most of these peptidase genes seem to be recently acquired, are expressed in the gut and present putative secretory pathway signal peptides. Besides that, most R. prolixus A1 peptidases showed high frequencies of basic residues at the protein surface, a typical structural signature of Cathepsin D-like proteins. Other peptidase families expanded in R. prolixus (i.e., C2 and M17) also presented significant differences between hematophagous (higher number of peptidases) and non-hematophagous species. This study also provides evidence for gene acquisition from microorganisms in some peptidase families in R. prolixus: (1) family M74 (murein endopeptidase), (2) family S29 (Hepatitis C virus NS3 protease), and (3) family S24 (repressor LexA). This study revealed new targets for studying the adaptation of these insects for digestion of blood meals and their competence as vectors of Chagas disease.

Disturbance of gut satiety peptide in purging disorder.

Little is known about biological factors that contribute to purging after normal amounts of food-the central feature of purging disorder (PD). This study comes from a series of nested studies examining ingestive behaviors in bulimic syndromes and specifically evaluated the satiety peptide YY (PYY) and the hunger peptide ghrelin in women with PD (n = 25), bulimia nervosa-purging (BNp) (n = 26), and controls (n = 26). Based on distinct subjective responses to a fixed meal in PD (Keel, Wolfe, Liddle, DeYoung, & Jimerson, ), we tested whether postprandial PYY response was significantly greater and ghrelin levels significantly lower in women with PD compared to controls and women with BNp. Participants completed structured clinical interviews, self-report questionnaires, and laboratory assessments of gut peptide and subjective responses to a fixed meal. Women with PD demonstrated a significantly greater postprandial PYY response compared to women with BNp and controls, who did not differ significantly. PD women also endorsed significantly greater gastrointestinal distress, and PYY predicted gastrointestinal intestinal distress. Ghrelin levels were significantly greater in PD and BNp compared to controls, but did not differ significantly between eating disorders. Women with BNp endorsed significantly greater postprandial hunger, and ghrelin predicted hunger. PD is associated with a unique disturbance in PYY response. Findings contribute to growing evidence of physiological distinctions between PD and BNp. Future research should examine whether these distinctions account for differences in clinical presentation as this could inform the development of specific interventions for patients with PD.

Plasma ghrelin suppression as an early predictor for postoperative complications after pancreatoduodenectomy

Background/ObjectivesThe gut peptide hormone ghrelin induces appetite and exhibits an anti-inflammatory effect. Serial perioperative changes in ghrelin have been examined in several surgical procedures, but few in pancreatectomy. The present study analyzed perioperative changes in plasma ghrelin levels after pancreaduodenectomy (PD). MethodsThe study included 24 patients undergoing PD between May 2015 and January 2016 at Osaka University Hospital. Plasma ghrelin and interleukin-6 (IL-6) levels, as well as white blood cells (WBCs) and C-reactive protein (CRP), were measured preoperatively and on postoperative day (POD) 1, 3, 7, and 14 by enzyme-linked immunosorbent assay. The relationship between the individual ghrelin ratio relative to preoperative value (IGR) and the development of grade IIIa-V Clavien-Dindo (CD) complications was examined. ResultsTwelve patients (50%) developed grade IIIa CD complications (n = 6 [25%] pancreatic fistula, n = 7 [29%] intraabdominal abscess, n = 3 [13%] post-pancreatectomy hemorrhage, n = 5 [21%] wound infection, and n = 1 [4%] lymphorrhea). The IGR on POD 1 was significantly lower (p = 0.014) in patients who developed the complications compared to those who did not, but no significant differences were found in terms of WBC, CRP, or IL-6 on POD 1. When the IGR cut-off was set to 82% by receiver operative curve analysis, the sensitivity was 83%, specificity 75% and area under the curve 0.80. The lower IGR group (≤82%) had more postoperative complications and longer hospital stay. ConclusionsThe IGR on POD 1 after PD is a useful marker for predicting postoperative complications.

Pharmacological inhibition of diacylglycerol acyltransferase-1 and insights into postprandial gut peptide secretion.

AIMTo examine the role that enzyme Acyl-CoA:diacylglycerol acyltransferase-1 (DGAT1) plays in postprandial gut peptide secretion and signaling.METHODSThe standard experimental paradigm utilized to evaluate the incretin response was a lipid challenge. Following a lipid challenge, plasma was collected via cardiac puncture at each time point from a cohort of 5-8 mice per group from baseline at time zero to 10 h. Incretin hormones [glucagon like peptide-1 (GLP-1), peptide tyrosine-tyrosine (PYY) and glucose dependent insulinotropic polypeptide (GIP)] were then quantitated. The impact of pharmacological inhibition of DGAT1 on the incretin effect was evaluated in WT mice. Additionally, a comparison of loss of DGAT1 function either by genetic ablation or pharmacological inhibition. To further elucidate the pathways and mechanisms involved in the incretin response to DGAT1 inhibition, other interventions [inhibitors of dipeptidyl peptidase-IV (sitagliptin), pancreatic lipase (Orlistat), GPR119 knockout mice] were evaluated.RESULTSDGAT1 deficient mice and wildtype C57/BL6J mice were lipid challenged and levels of both active and total GLP-1 in the plasma were increased. This response was further augmented with DGAT1 inhibitor PF-04620110 treated wildtype mice. Furthermore, PF-04620110 was able to dose responsively increase GLP-1 and PYY, but blunt GIP at all doses of PF-04620110 during lipid challenge. Combination treatment of PF-04620110 and Sitagliptin in wildtype mice during a lipid challenge synergistically enhanced postprandial levels of active GLP-1. In contrast, in a combination study with Orlistat, the ability of PF-04620110 to elicit an enhanced incretin response was abrogated. To further explore this observation, GPR119 knockout mice were evaluated. In response to a lipid challenge, GPR119 knockout mice exhibited no increase in active or total GLP-1 and PYY. However, PF-04620110 was able to increase total GLP-1 and PYY in GPR119 knockout mice as compared to vehicle treated wildtype mice.CONCLUSIONCollectively, these data provide some insight into the mechanism by which inhibition of DGAT1 enhances intestinal hormone release.

Anxiety, Depression, and the Microbiome: A Role for Gut Peptides.

The complex bidirectional communication between the gut and the brain is finely orchestrated by different systems, including the endocrine, immune, autonomic, and enteric nervous systems. Moreover, increasing evidence supports the role of the microbiome and microbiota-derived molecules in regulating such interactions; however, the mechanisms underpinning such effects are only beginning to be resolved. Microbiota-gut peptide interactions are poised to be of great significance in the regulation of gut-brain signaling. Given the emerging role of the gut-brain axis in a variety of brain disorders, such as anxiety and depression, it is important to understand the contribution of bidirectional interactions between peptide hormones released from the gut and intestinal bacteria in the context of this axis. Indeed, the gastrointestinal tract is the largest endocrine organ in mammals, secreting dozens of different signaling molecules, including peptides. Gut peptides in the systemic circulation can bind cognate receptors on immune cells and vagus nerve terminals thereby enabling indirect gut-brain communication. Gut peptide concentrations are not only modulated by enteric microbiota signals, but also vary according to the composition of the intestinal microbiota. In this review, we will discuss the gut microbiota as a regulator of anxiety and depression, and explore the role of gut-derived peptides as signaling molecules in microbiome-gut-brain communication. Here, we summarize the potential interactions of the microbiota with gut hormones and endocrine peptides, including neuropeptide Y, peptide YY, pancreatic polypeptide, cholecystokinin, glucagon-like peptide, corticotropin-releasing factor, oxytocin, and ghrelin in microbiome-to-brain signaling. Together, gut peptides are important regulators of microbiota-gut-brain signaling in health and stress-related psychiatric illnesses.

RD Lawrence Lecture 2017 Incretins: the intelligent hormones in diabetes.

The incretin hormones glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) have attracted considerable scientific and clinical interest due largely to their insulin-releasing and glucose-lowering properties. Indeed, GLP-1-based therapies are now key treatment options for many people with diabetes worldwide. In contrast, GIP-based agents have yet to reach the clinic based primarily on the impaired insulinotropic action of GIP observed in people with diabetes. Nevertheless, GIP is a key physiological regulator of insulin secretion and stable forms of GIP show much promise in rodent models to alleviate diabetes-obesity. Recent studies suggest that GIP may have an important role to play in a combination therapeutic approach or bioengineered with other gut peptides. Moreover, recent experimental studies indicate that incretins also exert pleiotropic effects in regions of the brain associated with learning and memory, thereby supporting preclinical data demonstrating that incretin-based drugs improve cognitive function. This review article, based on the RD Lawrence Lecture presented at Diabetes UK Annual Professional Conference (2017), provides a brief overview of incretins with a major focus on GIP, the development of designer GIP analogues, and how these molecules can improve cognition. Thus, incretins can be considered as 'the intelligent hormones' and may hold the key to successfully treating the alarming rise in neurodegenerative disorders.

The Role of GLP-1 in the Metabolic Success of Bariatric Surgery.

Two of the most popular bariatric procedures, vertical sleeve gastrectomy (VSG) and Roux-en-Y gastric bypass (RYGB), are commonly considered metabolic surgeries because they are thought to affect metabolism in a weight loss-independent manner. In support of this classification, improvements in glucose homeostasis, insulin sensitivity, and even discontinuation of type 2 diabetes mellitus (T2DM) medication can occur before substantial postoperative weight loss. The mechanisms that underlie this effect are unknown. However, one of the common findings after VSG and RYGB in both animal models and humans is the sharp postprandial rise in several gut peptides, including the incretin and satiety peptide glucagonlike peptide-1 (GLP-1). The increase in endogenous GLP-1 signaling has been considered a primary pathway leading to postsurgical weight loss and improvements in glucose metabolism. However, the degree to which GLP-1 and other gut peptides are responsible for the metabolic successes after bariatric surgery is continually debated. In this review we discuss the mechanisms underlying the increase in GLP-1 and its potential role in the metabolic improvements after bariatric surgery, including remission of T2DM. Understanding the role of changes in gut peptides, or lack thereof, will be crucial in understanding the critical factors necessary for the metabolic success of bariatric surgery.

Alterations in circadian and meal-induced gut peptide levels in lean and obese rats.

Alterations in gut hormone signaling are a likely contributing factor to the metabolic disturbances associated with overweight/obesity as they coordinate the timing of feeding behavior, absorption, and utilization of nutrients. These hormones are released in response to food intake, or follow a circadian or anticipatory pattern of secretion that is independent of nutrient stimulation. The aim of this study was to identify the degree to which high-fat diet-induced obesity would alter the daily rhythm of gut peptide plasma levels (glucagon-like peptide-1 [GLP-1], peptide YY [PYY], insulin or amylin [AMY]) or meal-induced levels in the middle of the light or dark cycle. Male Sprague-Dawley rats were fed a high-fat diet (OBESE) or chow (LEAN), implanted with jugular catheters, and blood samples were taken every 2 h throughout the light/dark cycle while freely feeding or after an Ensure liquid meal. We found that even when OBESE and LEAN animals ate the same kcals and have a similar pattern of food intake, there is a difference in both the levels and rhythm of plasma gut peptides. GLP-1 and PYY are higher during the light cycle in LEAN animals and AMY is higher in the OBESE group throughout the light/dark cycle. There was also a differential response of plasma gut signals after the Ensure meal, even though the composition and amount of intake of the meal were the same in both groups. These changes occur prior to the high-fat diet induced loss of glycemic control and may be a target for early intervention. Impact statement The aim of this study was to test if obesity would alter the daily rhythm of gut peptides or meal-induced levels in the middle of the light or dark cycle. We found that even when animals are eating the same amount (in kcal) of food that the obese animals have altered daily rhythms and meal-induced gut peptide levels. In particular, we are the first to show that obesity induces increases in peptide YY levels during the light cycle and amylin remains high throughout the light and dark cycle in obese animals. These changes occurred prior to a loss of glycemic control. Thus, the rhythm of gut peptides could be used as an early indicator of later and more serious metabolic disturbances and may be a target for early intervention.

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.

Chylomicrons stimulate incretin secretion in mouse and human cells

Aims/hypothesisLipids are a potent stimulus for the secretion of glucagon-like peptide (GLP)-1 and glucose-dependent insulinotropic peptide (GIP). Traditionally, this effect was thought to involve the sensing of lipid digestion products by free fatty acid receptor 1 (FFA1) and G-protein coupled receptor 119 (GPR119) on the apical surface of enteroendocrine cells. However, recent evidence suggests that lipids may in fact be sensed basolaterally, and that fatty acid absorption and chylomicron synthesis may be a prerequisite for their stimulatory effect on gut peptide release. Therefore, we investigated the effect of chylomicrons on GLP-1 and GIP secretion in vitro.MethodsThe effect of chylomicrons on incretin secretion was investigated using GLUTag cells and duodenal cultures of both murine and human origin. The role of lipoprotein lipase (LPL) and FFA1 in GLUTag cells was assessed by pharmacological inhibition and small (short) interfering RNA (siRNA)-mediated knockdown. The effect of chylomicrons on intracellular calcium concentration ([Ca2+]i) was determined by imaging GLUTag cells loaded with Fura-2. In the primary setting, the contributions of FFA1 and GPR119 were investigated using L cell-specific Gpr119 knockout cultures treated with the FFA1 antagonist GW1100.ResultsChylomicrons stimulated GLP-1 release from GLUTag cells, and both GLP-1 and GIP secretion from human and murine duodenal cultures. Chylomicron-triggered GLP-1 secretion from GLUTag cells was largely abolished following lipase inhibition with orlistat or siRNA-mediated knockdown of Lpl. In GLUTag cells, both GW1100 and siRNA-mediated Ffar1 knockdown reduced GLP-1 secretion in response to chylomicrons, and, consistent with FFA1 Gq-coupling, chylomicrons triggered an increase in [Ca2+]i. However, LPL and FFA1 inhibition had no significant effect on chylomicron-mediated incretin secretion in murine cultures. Furthermore, the loss of GPR119 had no impact on GLP-1 secretion in response to chylomicrons, even in the presence of GW1100.Conclusions/interpretationChylomicrons stimulate incretin hormone secretion from GLUTag cells as well as from human and murine duodenal cultures. In GLUTag cells, the molecular pathway was found to involve LPL-mediated lipolysis, leading to the release of lipid species that activated FFA1 and elevated intracellular calcium.

The glucagon-like peptide 1 receptor agonist Exendin-4 decreases relapse-like drinking in socially housed mice

Glucagon-like peptide-1 (GLP-1) is a gut peptide that regulates food intake and glucose metabolism. GLP-1 is also produced and released in the brain, and GLP-1 receptors are expressed in brain regions important for alcohol and drug reward, and for the development of addiction. GLP-1 receptor agonists can decrease alcohol intake acutely in rodents. However, alcohol use disorder is a chronic condition that requires treatments to be effective in promoting abstinence from excessive alcohol consumption over time. Here, we assessed the effect of daily treatment with the GLP-1 receptor agonist Exendin-4 in an assay of relapse-like drinking in socially housed mice. Male C57BL/6NTac mice were allowed continuous access to alcohol without tastant in the home cage for 37days. Then, alcohol bottles were removed and Exendin-4 (1.5μg/kg/day) or saline was administered subcutaneously for 8days during alcohol deprivation. Treatment continued for 8 additional days after reintroducing access to alcohol. A high-precision automated fluid consumption system was used to monitor intake of alcohol and water, drinking kinetics, and locomotor activity. Exendin-4 prevented the deprivation-induced increase in alcohol intake observed in control mice, without significantly affecting total fluid intake, body weight, or locomotor activity. The reduced alcohol intake was caused by a protracted latency to the first drink of alcohol and a reduced number of drinking bouts, while bout size and duration were not affected. The effect was maintained undiminished throughout the treatment period. These findings support the possible use of GLP-1 receptor agonists in the treatment of alcohol use disorder.

Structure of protein emulsion in food impacts intestinal microbiota, caecal luminal content composition and distal intestine characteristics in rats.

Few studies have evaluated in vivo the impact of food structure on digestion, absorption of nutrients and on microbiota composition and metabolism. In this study we evaluated in rat the impact of two structures of protein emulsion in food on gut microbiota, luminal content composition, and intestinal characteristics. Rats received for 3 weeks two diets of identical composition but based on lipid-protein matrices of liquid fine (LFE) or gelled coarse (GCE) emulsion. LFE diet led to higher abundance, when compared to the GCE, of Lactobacillaceae (Lactobacillus reuteri) in the ileum, higher β-diversity of the caecum mucus-associated bacteria. In contrast, the LFE diet led to a decrease in Akkermansia municiphila in the caecum. This coincided with heavier caecum content and higher amount of isovalerate in the LFE group. LFE diet induced an increased expression of (i) amino acid transporters in the ileum (ii) glucagon in the caecum, together with an elevated level of GLP-1 in portal plasma. However, these intestinal effects were not associated with modification of food intake or body weight gain. Overall, the structure of protein emulsion in food affects the expression of amino acid transporters and gut peptides concomitantly with modification of the gut microbiota composition and activity. Our data suggest that these effects of the emulsion structure are the result of a modification of protein digestion properties.