Number Of Enterochromaffin Cells Research Articles

Morphometric features of the duodenal wall in piglets during different periods of postnatal and neonatal ontogenesis under the influence of the ‘Globigen Jump Start’ feed additive

The morphological parameters of the small intestinal mucosa, such as villi size and crypts, play a key role in the formation of the intestinal tube’s absorption surface. The number of goblet and enterochromaffin cells indicates the epithelium’s functional condition in terms of mucus secretion and production of catecholamines and hormones (serotonin, dopamine). That is why it is important to study and determine certain morphometric parameters of the duodenal wall in piglets during different periods of neonatal and postnatal development, especially during stress situations, namely weaning and transition to protein feeding. The article presents the resulting effect studies of ‘Globigen Jump Start’ feed additive on the histo-morphological parameters of the piglets’ duodenal mucosa on day 7, 14 and 28 of life. A positive effect on mitigating weaning stress was manifested by a decrease in the quantitative and linear indicators of histoarchitectonics of intestinal wall’s individual morphological components. A significant increase in goblet cells, especially in the experimental group, had a positive impact on the mucopolysaccharide synthesis. The piglets’ gastrointestinal tract physiology involves a complex interaction between the central nervous system, metasympathetic nervous system, APUD system, and endocrine system. Due to these systems, the information is transmitted according to the direct and reverse communication mechanisms in the regulation of the gastrointestinal tract function. An increase in the number of enterochromaffin cells and their nuclei diameter in the experimental group of piglets indicated an increased synthesis of catecholamines and hormones. It has an extremely positive effect on the immune and physiological status of piglets, thus emphasizing the crucial role of serotonin in neuronal metabolism and the formation of stress resistance. The increase in the thickness of the duodenal wall muscle layer, in our opinion, occured due to the active peristalsis, which was enhanced by the action of some catecholamines, which were synthesized in a slightly larger amount under the influence of nutrients making up the ‘Globigen Jump Start’ feed additive.

Microbial Indoles: Key Regulators of Organ Growth and Metabolic Function.

Gut microbes supporting body growth are known but the mechanisms are less well documented. Using the microbial tryptophan metabolite indole, known to regulate prokaryotic cell division and metabolic stress conditions, we mono-colonized germ-free (GF) mice with indole-producing wild-type Escherichia coli (E. coli) or tryptophanase-encoding tnaA knockout mutant indole-non-producing E. coli. Indole mutant E. coli mice showed multiorgan growth retardation and lower levels of glycogen, cholesterol, triglycerides, and glucose, resulting in an energy deficiency despite increased food intake. Detailed analysis revealed a malfunctioning intestine, enlarged cecum, and reduced numbers of enterochromaffin cells, correlating with a metabolic phenotype consisting of impaired gut motility, diminished digestion, and lower energy harvest. Furthermore, indole mutant mice displayed reduction in serum levels of tricarboxylic acid (TCA) cycle intermediates and lipids. In stark contrast, a massive increase in serum melatonin was observed-frequently associated with accelerated oxidative stress and mitochondrial dysfunction. This observational report discloses functional roles of microbe-derived indoles regulating multiple organ functions and extends our previous report of indole-linked regulation of adult neurogenesis. Since indoles decline by age, these results imply a correlation with age-linked organ decline and levels of indoles. Interestingly, increased levels of indole-3-acetic acid, a known indole metabolite, have been shown to correlate with younger biological age, further supporting a link between biological age and levels of microbe-derived indole metabolites. The results presented in this resource paper will be useful for the future design of food intervention studies to reduce accelerated age-linked organ decline.

Serotonin Immunoreactive Cells in Extrahepatic Bile Ducts, Major Duodenal Papilla and Gallbladder in the Domestic Pig

Abstract The main part of serotonin in the body is synthesized and released by a certain type of enteroendocrine cells in the intestinal mucosa called enterochromaffin cells. The scarce qualitative and quantitative data on enterochromaffin and serotonin-positive mast cells in porcine extrahepatic bile ducts and gallbladder, motivated us to undertake the present study. The aim of this study was to determine the localization and density of serotonin-positive cells in the wall of the extrahepatic bile ducts and gallbladder in pigs. An immunohistochemical method was used to identify enterochromaffin cells and determine their percentage relative to the total number of endocrine cells labeled with chromogranin A. Serotonin-positive mast cells were identified after tryptase staining of serial sections. The endocrine function of mast cells was demonstrated by chromogranin A immunolabeling. The highest number of enterochromaffin cells were found in the intramural part of the ductus choledochus, followed by the papilla duodeni major, extramural part of the ductus choledochus, ductus hepaticus comunis, ductus cysticus, and gallbladder. In all parts of the extrahepatic bile ducts, the highest number of mast cells was found in the muscle layer, followed by the serosal layer and the propria. The expression of serotonin in the enterochromaffin cells of the biliary glands and in the mast cells of the analyzed organs suggests a possible synthesis of serotonin, which probably regulates physiological and pathological processes.

No synergistic effect of fecal microbiota transplantation and shugan decoction in water avoidance stress-induced IBS-D rat model.

BackgroundIt has been reported that 5-hydroxytryptamine (5-HT, serotonin) metabolism is involved in the pathogenesis of irritable bowel syndrome (IBS) and that either Shugan decoction (SGD) or fecal microbiota transplantation (FMT) can alleviate the symptoms of IBS in patients and animal models. But the synergistic effect of FMT and SGD on 5-HT metabolism and IBS symptoms has not been investigated.AimThe main purpose of this study is to observe the synergistic effect of FMT with SGD on symptoms and 5-HT metabolism in IBS-D rats induced by water avoidance stress (WAS). Moreover, the possible material basis of the FMT was investigated.MethodsIn experiment I, rats were randomly divided into seven groups. Control group: routine feeding; WAS→ Control group: routine feeding with fecal microbiota liquid (FML) 1 (derived from rats in WAS group) gavage since the fourth day; WAS group: 10 days WAS with routine feeding; SGD group: 10 days WAS with SGD gavage since the fourth day on the base of routine feeding; Control→ WAS group: 10 days WAS with FML2 (derived from rats in Control group) gavage since the fourth day with routine feeding; SGD→ WAS group: 10 days WAS with FML3 (derived from rats in SGD group) gavage since the fourth day with routine feeding; SGD + (Control→ WAS) group: 10 days WAS with SGD and FML2 (derived from rats in Control group) gavage since the fourth day with routine feeding. In experiment II, rats were randomly divided into three groups. Control group: routine feeding; Control→ WAS group: 10 days WAS with FML2 gavage since the fourth day with routine feeding; FControl→ WAS group: 10 days WAS with FML2 filtrate gavage since the fourth day. The number of fecal pellets output (FPT) and the pain pressure threshold (PPT) were recorded. The histological changes in colon mucosa were observed by hematoxylin-eosin (HE) stain. The number of enterochromaffin cells (ECs), the content of 5-HT, and the expression of serotonin reuptake transporter (SERT) protein in the colon were measured by immunofluorescence or western blotting.ResultsCompared with that in the control group, the PPT and the expression of SERT in the WAS group and that in the WAS→ Control group were decreased with the increased number of ECs and the level of 5-HT in colon. But the FPT was not increased in the WAS→ Control group although that was increased in the WAS group. Compared with that in the WAS group, the FPT, the PPT, the number of ECs, the level of 5-HT, and the expression of SERT protein in colon in the SGD group, control→ WAS group, SGD→ WAS group, and SGD+(Control→ WAS) group were all recovered. The recovery of these indicators in the Control→ WAS group and that in the FControl→ WAS group was not significantly different.ConclusionNo synergistic effect of SGD with FMT on IBS symptoms induced by WAS was found. The metabolites of intestinal microbiota may be the main active substances of the FML derived from normal rats to alleviate WAS-induced IBS symptoms.

Ameliorative effect and mechanism of Si-Ni-San on chronic stress-induced diarrhea-irritable bowel syndrome in rats.

Background: Chronic stress-induced diarrhea is a common clinical condition, characterized by an abnormal bowel movement and loose stools, which lacks effective treatment in the clinic. Si-Ni-San (SNS) is a compound traditional Chinese medicine extensively used in China for stress-related diarrhea. However, the mechanism is unclear. Methods: Male Wistar rats (200 ± 20 g) were placed in a restraint cylinder and fixed horizontally for 3 h once daily for 21 consecutive days to establish a chronic restraint stress (CRS) rat model. SNS (0.6944 g/kg or 1.3888 g/kg) was given by gavage 1 h before the restraint once daily for 21 consecutive days. We examined the fecal score, dopamine β hydroxylase (DβH), and c-fos expression in locus coeruleus, norepinephrine (NE) content in ileum and plasma, expression of α1 adrenergic receptors, MLCK, MLC, and p-MLC in the colon and mesenteric arteries, contraction of isolated mesenteric arteries, The expression of subunit δ of ATP synthase (ATP5D) in intestinal tissues, ATP, ADP, and AMP content in the ileum and colon, occludin expression between ileum epithelial cells, the number of enterochromaffin cells (ECs) and mast cells (MCs) in the ileum, and 5-hydroxytryptamine (5-HT) content in the ileum and plasma. Results: After SNS treatment, the fecal score was improved. The increased expression of DβH and c-fos in locus coeruleus was inhibited. SNS suppressed the increased NE content in the ileum and plasma, down-regulated α1 adrenergic receptors in mesenteric arteries and MLCK, MLC, p-MLC in the colon and mesenteric arteries, and inhibited the contraction of mesenteric arteries. SNS also increased the ATP content in the ileum and colon, inhibited low expression of ATP5D in intestinal tissues, inhibited the decrease of ATP/ADP in the ileum and ATP/AMP in the colon, and up-regulated the occludin expression between ileum epithelial cells. In addition, SNS inhibited the increase of ECs and MCs in the ileum and the increase of 5-HT content in the ileum and plasma. Conclusion: This study demonstrated that SNS could improve CRS-induced abnormal feces in rats. This effect was related to the inhibition of CRS-induced increased expression of DβH and c-fos in the locus coeruleus, NE content in the ileum and plasma, and the contraction of isolated mesenteric arteries; inhibition of energy metabolism abnormality and decreased occludin expression; inhibition of increased ECs and MCs in the ileum, and 5-HT content in the ileum and plasma.

Subepithelial Serotonin Reduces Small Intestinal Epithelial Cell Tightness via Reduction of Occluding Expression.

The precise pathogenesis of irritable bowel syndrome (IBS) remains unresolved; however, recent studies have reported that patients with diarrhea-predominant IBS exhibit an increased small intestinal permeability and increased number of enterochromaffin cells containing high 5-hydroxytryptamine (5HT; serotonin) levels. In this study, we investigated whether 5HT has the potential to modulate small intestinal epithelial cell permeability, focusing on tight junction-associated proteins. The differentiated Caco-2 cell monolayer on porous filters (Millicell) was used. Then, 5HT was added to the lower Millicell compartment for 7 days. Intestinal epithelial cell permeability was assessed by measuring the flux of paracellular permeability markers. We further assessed the expression of occludin in the 5HT-stimulated Caco-2 monolayer. We found that 5HT did not affect the viability of Caco-2 cells at concentrations up to 100 µM during the experimental period. Administration of 5HT to the basal side of Caco-2 cells increased the flux of 3H-labeled mannitol (182 Da) but did not increase that of FITC-dextran (4000 Da). Among the tight junction proteins, the expression of occludin was specifically decreased by stimulation with 5HT at a concentration of 100 µM. In conclusion, excessive 5HT in the basal side increased the permeability of intestinal epithelial cells via reduction of occludin expression.

Dietary Supplementation throughout Life with Non-Digestible Oligosaccharides and/or n-3 Poly-Unsaturated Fatty Acids in Healthy Mice Modulates the Gut-Immune System-Brain Axis.

The composition and activity of the intestinal microbial community structures can be beneficially modulated by nutritional components such as non-digestible oligosaccharides and omega-3 poly-unsaturated fatty acids (n-3 PUFAs). These components affect immune function, brain development and behaviour. We investigated the additive effect of a dietary combination of scGOS:lcFOS and n-3 PUFAs on caecal content microbial community structures and development of the immune system, brain and behaviour from day of birth to early adulthood in healthy mice. Male BALB/cByJ mice received a control or enriched diet with a combination of scGOS:lcFOS (9:1) and 6% tuna oil (n-3 PUFAs) or individually scGOS:lcFOS (9:1) or 6% tuna oil (n-3 PUFAs). Behaviour, caecal content microbiota composition, short-chain fatty acid levels, brain monoamine levels, enterochromaffin cells and immune parameters in the mesenteric lymph nodes (MLN) and spleen were assessed. Caecal content microbial community structures displayed differences between the control and dietary groups, and between the dietary groups. Compared to control diet, the scGOS:lcFOS and combination diets increased caecal saccharolytic fermentation activity. The diets enhanced the number of enterochromaffin cells. The combination diet had no effects on the immune cells. Although the dietary effect on behaviour was limited, serotonin and serotonin metabolite levels in the amygdala were increased in the combination diet group. The combination and individual interventions affected caecal content microbial profiles, but had limited effects on behaviour and the immune system. No apparent additive effect was observed when scGOS:lcFOS and n-3 PUFAs were combined. The results suggest that scGOS:lcFOS and n-3 PUFAs together create a balance—the best of both in a healthy host.

Involvement of mucosal flora and enterochromaffin cells of the caecum and descending colon in diarrhoea-predominant irritable bowel syndrome

BackgroundAccumulating evidence supports the pivotal role of intestinal flora in irritable bowel syndrome (IBS). Serotonin synthesis by enterochromaffin (EC) cells is influenced by the gut microbiota and has been reported to have an interaction with IBS. The comparison between the microbiota of the caecal and colonic mucosa in IBS has rarely been studied. The aim of this study was to investigate the relationship between the gut microbiota, EC cells in caecum and descending colon, and diarrhoea-predominant IBS (IBS-D) symptoms.ResultsA total of 22 IBS-D patients and 22 healthy controls (HCs) were enrolled in our study. Hamilton anxiety (HAM-A) and Hamilton depression (HAM-D) grades increased significantly in IBS-D patients. In addition, the frequency of defecation in IBS-D patients was higher than that in HCs. Among the preponderant bacterial genera, the relative abundance of the Ruminococcus_torques_ group increased in IBS-D patients in caecum samples while Raoultella and Fusobacterium were less abundant. In the descending colon, the abundance of the Ruminococcus_torques_group and Dorea increased in IBS-D patients and Fusobacterium decreased. No difference was observed between the descending colon and caecum in regards to the mucosal-associated microbiota. The number of EC cells in the caecum of IBS-D patients was higher than in HCs and the expression of TPH1 was higher in IBS-D patients both in the caecum and in the descending colon both at the mRNA and protein level. Correlation analysis showed that the Ruminococcus_torques_group was positively associated with HAM-A, HAM-D, EC cell number, IBS-SSS, degree of abdominal pain, frequency of abdominal pain and frequency of defecation. The abundance of Dorea was positively associated with EC cell number, IBS-SSS, HAM-A, HAM-D and frequency of abdominal pain.ConclusionsEC cell numbers increased in IBS-D patients and the expression of TPH1 was higher than in HCs. The Ruminococcus torques group and Dorea furthermore seem like promising targets for future research into the treatment of IBS-D patients.

Acute infection with Toxoplasma gondii oocysts preferentially activates non-neuronal cells expressing serotonin in the jejunum of rats

The interaction of Toxoplasma gondii with the gastrointestinal tract of its host is highly regulated. Once ingested, the parasite crosses the epithelium without altering the permeability of the intestinal barrier. Nevertheless, many studies report alterations ranging from structural to functional damage in cells and tissues that make up the wall of the small and large intestine. Although the immune response to the parasite has been extensively studied, the role of serotonin (5-HT) in toxoplasmosis is poorly understood. Here we investigate the distribution of cells expressing 5-HT and its effects on cells and tissues of the jejunal wall of rats after 2, 3, or 7 days of T. gondii infection. Key resultsOur results show that transposition of the jejunal epithelium by T. gondii leads to ruptures in the basement membrane and activation of the immune system, as confirmed by the decrease in laminin immunostaining and the increase in the number of mast cells, respectively. Conclusions and inferencesWe showed an increase in the number of enterochromaffin cells and mast cells expressing 5-HT in the jejunal wall. We also observed that the percentage of serotonergic mast cells increased in the total population. Thus, we can suggest that oral infection by T. gondii oocysts preferentially activates non-neuronal cells expressing 5-HT. Together, these results may explain both the changes in the extracellular matrix and the morphology of the enteric ganglia.

Female serotonin transporter-knockout rat: A potential model of irritable bowel syndrome.

Irritable bowel syndrome (IBS) is a common functional gastrointestinal disease. Although visceral hypersensitivity (VH) and disturbed gastrointestinal motility are typical pathophysiological features of IBS, the pathological mechanisms underlying this disease remain unclear. Serotonin system abnormalities are considered to play an important role in the pathomechanisms of IBS. Here, we hypothesize that similar alterations, including VH and colonic motility, induced by serotonin transporter (SERT) knockout result from altered serotonin signaling. We sought to determine the molecular mechanism underlying VH and colonic dysmotility induced by SERT knockout. We found that female SERT (slc6a4)-knockout (KO; ie, slc6a4-/- ) rats exhibited lower pain pressure thresholds (PPTs) than wild-type (WT; ie, slc6a4+/+ ) rats in response to colorectal distension (CRD). Significantly increased fecal pellet output and reduced concentration of serum tryptophan were observed in the female SERT KO rats. The concentrations of 5-hydroxytryptamine (5-HT) in platelet-rich plasma (PRP) and serum in SERT KO rats were lower than those in WT rats, but the numbers of enterochromaffin cells (ECs) and the concentrations of 5-HT in colon of SERT KO rats were higher than those of WT rats. Finally, increased expression levels of 5-HT1B receptors, 5-HT2C receptors, 5-HT3A receptors, 5-HT3B receptors, 5-HT6 receptors, 5-HT7 receptors, and glycosylated dopamine transporters (DATs) were found in the female SERT KO rats. We concluded that alterations in the serotonin system induced by the knockout of slc6a4 might result in VH and accelerated gastrointestinal motility in female SERT KO rats, which can be used as an animal model of IBS.

Consumption of indigestible saccharides and administration of Bifidobacterium pseudolongum reduce mucosal serotonin in murine colonic mucosa.

SCFA increase serotonin (5-hydroxytryptamine, 5-HT) synthesis and content in the colon in vitro and ex vivo, but little is known in vivo. We tested whether dietary indigestible saccharides, utilised as a substrate to produce SCFA by gut microbiota, would increase colonic 5-HT content in mice. Male C57BL/6J mice were fed a purified diet and water supplemented with 4 % (w/v) 1-kestose (KES) for 2 weeks. Colonic 5-HT content and enterochromaffin (EC) cell numbers were lower in mice supplemented with KES than those without supplementation, while monoamine oxidase A activity and mRNA levels of tryptophan hydroxylase 1 (Tph1), chromogranin A (Chga), Slc6a4 and monoamine oxidase A (Maoa) genes in the colonic mucosa, serum 5-HT concentration and total 5-HT content in the colonic contents did not differ between groups. Caecal acetate concentration and Bifidobacterium pseudolongum population were higher in KES-supplemented mice. Similar trends were observed in mice supplemented with other indigestible saccharides, that is, fructo-oligosaccharides, inulin and raffinose. Intragastric administration of live B. pseudolongum (108 colony-forming units/d) for 2 weeks reduced colonic 5-HT content and EC cell numbers. These results suggest that changes in synthesis, reuptake, catabolism and overflow of 5-HT in the colonic mucosa are not involved in the reduction of colonic 5-HT content by dietary indigestible saccharides in mice. We propose that gut microbes including B. pseudolongum could contribute to the reduction of 5-HT content in the colonic mucosa via diminishing EC cells.

Serotonin Deficiency Is Associated With Delayed Gastric Emptying

Gastrointestinal (GI) motility is regulated by serotonin (5-hydroxytryptamine [5-HT]), which is primarily produced by enterochromaffin (EC) cells in the GI tract. However, the precise roles of EC cell-derived 5-HT in regulating gastric motility remain a major point of conjecture. Using a novel transgenic mouse line, we investigated the distribution of EC cells and the pathophysiologic roles of 5-HT deficiency in gastric motility in mice and humans. We developed an inducible, EC cell-specific Tph1CreERT2/+ mouse, which was used to generate a reporter mouse line, Tph1-tdTom, and an EC cell-depleted line, Tph1-DTA. We examined EC cell distribution, morphology, and subpopulations in reporter mice. GI motility was measured invivo and exvivo in EC cell-depleted mice. Additionally, we evaluated 5-HT content in biopsy and plasma specimens from patients with idiopathic gastroparesis (IG). Tph1-tdTom mice showed EC cells that were heterogeneously distributed throughout the GI tract with the greatest abundance in the antrum and proximal colon. Two subpopulations of EC cells were identified in the gut: self-renewal cells located at the base of the crypt and mature cells observed in the villi. Tph1-DTA mice displayed delayed gastric emptying, total GI transit, and colonic transit. These gut motility alterations were reversed by exogenous provision of 5-HT. Patients with IG had a significant reduction of antral EC cell numbers and 5-HT content, which negatively correlated with gastric emptying rate. The Tph1CreERT2/+ mouse provides a powerful tool to study the functional roles of EC cells in the GI tract. Our findings suggest a new pathophysiologic mechanism of 5-HT deficiency in IG.

Enterochromaffin Cell-Enriched Monolayer Platform for Assaying Serotonin Release from Human Primary Intestinal Cells.

Enteroendocrine (EE) cells within the intestinal epithelium produce a range of hormones that have key roles in modulating satiety and feeding behavior in humans. The regulation of hormone release from EE cells as a potential therapeutic strategy to treat metabolic disorders is highly sought after by the pharmaceutical industry. However, functional studies are limited by the scarcity of EE cells (or surrogates) in both in vivo and in vitro systems. Enterochromaffin (EC) cells are a subtype of EE cells that produce serotonin (5HT). Here, we explored simple strategies to enrich EC cells in in vitro monolayer systems derived from human primary intestinal stem cells. During differentiation of the monolayers, the EC cell lineage was significantly altered by both the culture method [air-liquid interface (ALI) vs submerged] and the presence of vasoactive intestinal peptide (VIP). Compared with traditional submerged cultures without VIP, VIP-assisted ALI culture significantly boosted the number of EC cells and their 5HT secretion by up to 430 and 390%, respectively. The method also increased the numbers of other subtypes of EE cells such as L cells. Additionally, this method generated monolayers with enhanced barrier integrity, so that directional (basal or apical) 5HT secretion was measurable. For all donor tissues, the enriched EC cells improved the signal-to-background ratio and reliability of 5HT release assays. The enhancement in the 5HT secretion behavior was consistent over time from a single donor, but significant variation in the amount of secreted 5HT was present among tissues derived from five different donors. To demonstrate the utility of the EC-enriched monolayer system, 13 types of pungent food ingredients were screened for their ability to stimulate 5HT secretion. Curcumin found in the spice turmeric derived from the Curcuma longa plant was found to be the most potent secretagogue. This EC-enriched cell monolayer platform can provide a valuable analytical tool for the high-throughput screening of nutrients and gut microbial components that alter the secretion of 5HT.

Acute stress disrupts intestinal homeostasis via GDNF-RET.

ObjectivesEnterochromaffin (EC) cells have been associated with functional gastrointestinal disorders such as IBS. Recently, we found that glial cell‐derived neurotrophic factor (GDNF)‐rearranged during transfection (RET) localized in EC cells in human colonic epithelia. Here, we examine the role of GDNF‐RET in the pathophysiology of diarrhoea‐predominant irritable bowel syndrome (IBS‐D).Materials and MethodsGDNF was assessed by ELISA and immunohistochemistry in biopsies from IBS‐D patients and healthy controls. Stress was induced by using a wrap‐restraint stress (WRS) procedure to serve as an acute stress‐induced IBS model. The function of GDNF‐RET axis to intestinal stem cell (ISC) homeostasis, and EC cell numbers were assessed in vivo and in vitro.ResultsGDNF‐RET was expressed in EC cells in human colon. GDNF was significantly increased in IBS‐D patients. WRS mice showed increased GDNF‐RET levels in colon. WRS induced visceral hypersensitivity by expanding of ISC and differentiation of EC cell via GDNF‐RET. Furthermore, GDNF‐treated mice recapitulated the phenotype of WRS mice. In vitro, GDNF treatment amplified Wnt signal and increased serotonin levels in colonic organoids in a dose‐dependent manner.ConclusionsWe identified GDNF‐RET was presented in colonic epithelium of patients with IBS‐D. GDNF‐RET played important roles in regulating ISC and EC cell differentiation. Our findings, thus, provide RET inhibitor as new therapeutic targets for treatment of patients with IBS‐D.

Diet differentially regulates enterochromaffin cell serotonin content, density and nutrient sensitivity in the mouse small and large intestine.

Enterochromaffin (EC) cells are specialized enteroendocrine cells lining the gastrointestinal (GI) tract and the source of almost all serotonin (5-hydroxytryptamine; 5-HT) in the body. Gut-derived 5-HT has a plethora of physiological roles, including regulation of gastrointestinal motility, and has been implicated as a driver of obesity and metabolic disease. This is due to 5-HT influencing key metabolic processes, such as hepatic gluconeogenesis, adipose tissue lipolysis and hindering thermogenic capacity. Increased circulating 5-HT occurs in humans with obesity and type 2 diabetes. However, despite the known metabolic roles of gut-derived 5-HT, the mechanisms underlying the cellular-level change in EC cells under obesogenic conditions remains unknown. We use a mouse model of diet-induced obesity (DIO) to identify the regional changes that occur in primary EC cells from the duodenum and colon. Transcriptional changes in the nutrient sensing profile of primary EC cells were assessed, and responses to nutrient stimuli in culture were determined by 5-HT ELISA. We find that obesogenic conditions affect EC cells in a region-dependent manner. Duodenal EC cells from DIO mice have impaired sugar sensing even in the presence of increased 5-HT content per cell, while colonic EC cell numbers are significantly increased, but have unaltered nutrient sensing capacity. Our findings from this study add novel insights into the mechanisms by which functional changes to EC cells occur at a cellular level, which may contribute to the altered circulating 5-HT seen with obesity and metabolic disease, and associated gastrointestinal disorders.

Dietary Proteins Regulate Serotonin Biosynthesis and Catabolism by Specific Gut Microbes.

More than 90% of serotonin is produced in the intestine. Previous studies have shown that different protein diets significantly affect serum serotonin levels. Here, the colonic microbiota and intestinal serotonin were measured to elaborate how protein diets affect serotonin production in a mouse model. The emulsion-type sausage protein and cooked pork protein diets increased the mRNA levels of tryptophan hydroxylase 1 (Tph1) and monoamine oxidase A (Maoa) and serotonin level as well but reduced the number of enterochromaffin cells. However, the soy protein diet increased the number of enterochromaffin cells and Tph1 mRNA level but decreased the Maoa mRNA level and the serotonin content. Specific gut microbes that responded to dietary changes and affected the content of short-chain fatty acids were significantly related to serotonin-associated biomarkers. These results suggest that dietary proteins may regulate serotonin biosynthesis and catabolism by altering specific gut microbes.

Human‐derived Bifidobacterium dentium Metabolites Modulate the Mammalian Serotonergic System

BackgroundRecent work has identified an essential role for the gut microbiota in regulating host‐derived serotonin (5‐hydroxytryptamine; 5‐HT) from enterochromaffin cells. Studies using germ‐free mice have revealed impaired 5‐HT levels and signaling in the absence of microbes. While it is known that addition of microbial communities to germ‐free mice restores 5‐HT levels, information on which microbes directly influence 5‐HT remains unknown. Commensal Bifidobacterium species produce acetate, a short chain fatty acid known to stimulate 5‐HT release in vitro. We hypothesize that B. dentium, a model Bifidobacterium species, secretes acetate and modulates the serotonergic system.Methods & ResultsAdult germ‐free mice were mono‐associated with acetate producing B. dentium, propionate producing Bacteroides ovatus, or heat killed B. dentium. Mass spectrometry of cecal and fecal samples revealed elevated levels of acetate in B. dentium mono‐associated mice and increased ileal, colonic, fecal and serum 5‐HT compared to all other groups. No increases in acetate or 5‐HT were observed in heat‐killed B. dentium or B. ovatus treated mice compared to germ‐free controls, as assessed by mass spec, immunostaining, and qPCR. Using mouse and human enteroids, we observed that addition of B. dentium metabolites and acetate stimulated release of 5‐HT in a dose‐dependent manner. Depletion of acetate in B. dentium secreted products reduced the amount of 5‐HT released from enteroids. We also confirmed our findings using a novel NGN3 over‐expression system in human enteroids, which increased the number of enterochromaffin cells. In addition to 5‐HT alterations, we found increased expression of 5‐HT receptors 2a and 4 in B. dentium mono‐associated mouse ileum and colon. RNA in situ hybridization revealed a similar increase in 5‐HT receptor 2a in the hippocampus of B. dentium treated mice compared to germ‐free controls. Functionally, live B. dentium modulated species typical behavior previously shown to be linked to 5‐HT receptor 2a.ConclusionCollectively, these data indicate that B. dentium, and the bacterial metabolite acetate, regulate key components of the serotonergic system.B. dentium colonizes the intestinal mucus layer (left) and it’s metabolites stimulate serotonin release in mouse organoids (right).Figure 1

TLR2 Plays a Pivotal Role in Mediating Mucosal Serotonin Production in the Gut.

Serotonin (5-hydroxytryptamine [5-HT]) is a key enteric signaling molecule that mediates various physiological processes in the gut. Enterochromaffin (EC) cells in the mucosal layer of the gut are the main source of 5-HT in the body and are situated in close proximity to the gut microbiota. In this study, we identify a pivotal role of TLR2 in 5-HT production in the gut. Antibiotic treatment reduces EC cell numbers and 5-HT levels in naive C57BL/6 mice, which is associated with downregulation of TLR2 expression but not TLR1 or TLR4. TLR2-deficient (Tlr2 -/-) and Myd88 -/- mice express lower EC cell numbers and 5-HT levels, whereas treatment with TLR2/1 agonist upregulates 5-HT production in irradiated C57BL/6 mice, which are reconstituted with Tlr2 -/- bone marrow cells, and in germ-free mice. Human EC cell line (BON-1 cells) release higher 5-HT upon TLR2/1 agonist via NF-κB pathway. Tlr2 -/- mice and anti-TLR2 Ab-treated mice infected with enteric parasite, Trichuris muris, exhibited attenuated 5-HT production, compared with infected wild-type mice. Moreover, excretory-secretory products from T. muris induce higher 5-HT production in BON-1 cells via TLR2 in a dose-dependent manner, whereby the effect of excretory-secretory products is abrogated by TLR2 antagonist. These findings not only suggest an important role of TLR2 in mucosal 5-HT production in the gut by resident microbiota as well as by a nematode parasite but also provide, to our knowledge, novel information on the potential benefits of targeting TLR2 in various gut disorders that exhibit aberrant 5-HT signaling.

Sugar Responses of Human Enterochromaffin Cells Depend on Gut Region, Sex, and Body Mass.

Gut-derived serotonin (5-HT) is released from enterochromaffin (EC) cells in response to nutrient cues, and acts to slow gastric emptying and modulate gastric motility. Rodent studies also evidence a role for gut-derived 5-HT in the control of hepatic glucose production, lipolysis and thermogenesis, and in mediating diet-induced obesity. EC cell number and 5-HT content is increased in the small intestine of obese rodents and human, however, it is unknown whether EC cells respond directly to glucose in humans, and whether their capacity to release 5-HT is perturbed in obesity. We therefore investigated 5-HT release from human duodenal and colonic EC cells in response to glucose, sucrose, fructose and α-glucoside (αMG) in relation to body mass index (BMI). EC cells released 5-HT only in response to 100 and 300 mM glucose (duodenum) and 300 mM glucose (colon), independently of osmolarity. Duodenal, but not colonic, EC cells also released 5-HT in response to sucrose and αMG, but did not respond to fructose. 5-HT content was similar in all EC cells in males, and colonic EC cells in females, but 3 to 4-fold higher in duodenal EC cells from overweight females (p < 0.05 compared to lean, obese). Glucose-evoked 5-HT release was 3-fold higher in the duodenum of overweight females (p < 0.05, compared to obese), but absent here in overweight males. Our data demonstrate that primary human EC cells respond directly to dietary glucose cues, with regional differences in selectivity for other sugars. Augmented glucose-evoked 5-HT release from duodenal EC is a feature of overweight females, and may be an early determinant of obesity.

Characterization of Serotonin Signaling Components in Patients with Inflammatory Bowel Disease.

BackgroundTryptophan hydroxylase (TPH)1 catalyzes the biosynthesis of serotonin (5-hydroxytrptamine; 5-HT) in enterochromaffin (EC) cells, the predominant source of gut 5-HT. Secreted 5-HT regulates various gut functions through diverse 5-HT receptor (5-HTR) families, and 5-HT transporter (5-HTT) sequesters its activity via uptake into surrounding cells. In inflammatory bowel disease (IBD) mucosal 5-HT signaling is altered, including upregulated EC cell numbers and 5-HT levels. We examined key mucosal 5-HT signaling components and blood 5-HT levels and, as part of a pilot study, investigated the association between 5-HTT gene-linked polymorphic region (5HTTLPR) and Crohn’s disease (CD).MethodsIn the context of inflammation, colonic expressions of TPH1, 5-HTT and 5-HTRs were studied in CD patients (n=15) and healthy controls (HC; n=10) using quantitative polymerase chain reaction (qPCR). We also investigated 5HTTLPR in 40 CD patients and HC utilizing PCR and measured platelet-poor plasma (PPP) and plasma 5-HT concentrations.ResultsCompared with HC, inflammation in CD patients was associated with elevated TPH1, 5-HTR3, 5-HTR4, 5-HTR7 and downregulated 5-HTT expressions. In our second cohort of participants, significantly higher PPP and plasma 5-HT levels and higher S-genotype (L/S+S/S) than L/L genotype were observed in CD patients compared with HC.ConclusionOur results suggest that augmented mucosal 5-HT signaling and specific 5-HTTLPR genotype–associated decreased efficiency in 5-HT reuptake, the latter through increased 5-HT availability, may contribute to inflammation in CD patients. These findings revealed important information on various components of 5-HT signaling in intestinal inflammation which may ultimately lead to effective strategies targeting this pathway in IBD.