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Microbiome Oct 2021The intestinal microbiota plays an important role in regulating gastrointestinal (GI) physiology in part through interactions with the enteric nervous system (ENS)....
BACKGROUND
The intestinal microbiota plays an important role in regulating gastrointestinal (GI) physiology in part through interactions with the enteric nervous system (ENS). Alterations in the gut microbiome frequently occur together with disturbances in enteric neural control in pathophysiological conditions. However, the mechanisms by which the microbiota regulates GI function and the structure of the ENS are incompletely understood. Using a mouse model of antibiotic (Abx)-induced bacterial depletion, we sought to determine the molecular mechanisms of microbial regulation of intestinal function and the integrity of the ENS. Spontaneous reconstitution of the Abx-depleted microbiota was used to assess the plasticity of structure and function of the GI tract and ENS. Microbiota-dependent molecular mechanisms of ENS neuronal survival and neurogenesis were also assessed.
RESULTS
Adult male and female Abx-treated mice exhibited alterations in GI structure and function, including a longer small intestine, slower transit time, increased carbachol-stimulated ion secretion, and increased intestinal permeability. These alterations were accompanied by the loss of enteric neurons in the ileum and proximal colon in both submucosal and myenteric plexuses. A reduction in the number of enteric glia was only observed in the ileal myenteric plexus. Recovery of the microbiota restored intestinal function and stimulated enteric neurogenesis leading to increases in the number of enteric glia and neurons. Lipopolysaccharide (LPS) supplementation enhanced neuronal survival alongside bacterial depletion, but had no effect on neuronal recovery once the Abx-induced neuronal loss was established. In contrast, short-chain fatty acids (SCFA) were able to restore neuronal numbers after Abx-induced neuronal loss, demonstrating that SCFA stimulate enteric neurogenesis in vivo.
CONCLUSIONS
Our results demonstrate a role for the gut microbiota in regulating the structure and function of the GI tract in a sex-independent manner. Moreover, the microbiota is essential for the maintenance of ENS integrity, by regulating enteric neuronal survival and promoting neurogenesis. Molecular determinants of the microbiota, LPS and SCFA, regulate enteric neuronal survival, while SCFA also stimulates neurogenesis. Our data reveal new insights into the role of the gut microbiota that could lead to therapeutic developments for the treatment of enteric neuropathies. Video abstract.
Topics: Animals; Enteric Nervous System; Female; Gastrointestinal Microbiome; Intestine, Small; Male; Mice; Neuroglia; Neurons
PubMed: 34702353
DOI: 10.1186/s40168-021-01165-z -
Archives of Pathology & Laboratory... May 2022Intestinal neuronal dysplasia type B (IND B) is a complex entity involving the enteric nervous system, clinically manifested with constipation in infancy. Diagnosis has...
CONTEXT.—
Intestinal neuronal dysplasia type B (IND B) is a complex entity involving the enteric nervous system, clinically manifested with constipation in infancy. Diagnosis has been established by histopathologic analysis of rectal biopsies. However, the criteria for the diagnosis have been questioned and modified, hindering diagnostic practice.
OBJECTIVE.—
To analyze the applicability of PTEN immunohistochemistry in the diagnosis of IND B and to compare with control cases and cases of Hirschsprung disease (HD).
DESIGN.—
PTEN immunohistochemical expression was analyzed in colorectal samples from 29 cases of IND B and compared with 4 control cases and 6 cases of HD. The pattern of PTEN immunoexpression was analyzed in glial cells of the submucosal and myenteric nerve plexuses and in neural fibrils of the muscularis propria using a scoring system.
RESULTS.—
Marked reduction or absence of PTEN expression was observed in glial cells of the submucosal nerve plexuses in all cases of the IND B group and in the myenteric nerve plexuses in 28 of 29 cases (96.5%). Lack of PTEN expression was detected in neural fibrils within the muscularis propria in 21 of 29 cases (72%) of the IND B group. PTEN expression was positive in the same neural structures of the control and HD groups.
CONCLUSIONS.—
PTEN immunohistochemistry may be a valuable tool in the diagnostic evaluation of IND B. Lack of or reduction of PTEN expression in neural fibrils within the muscularis propria suggests that involvement of the neuromuscular junction may be a key event in the pathogenesis of the motility disturbance occurring in IND B.
Topics: Humans; Immunohistochemistry; Enteric Nervous System; Myenteric Plexus; Hirschsprung Disease; Constipation; PTEN Phosphohydrolase
PubMed: 35943858
DOI: 10.5858/arpa.2021-0424-OA -
Nature Neuroscience Jan 2021Autonomous regulation of the intestine requires the combined activity of functionally distinct neurons of the enteric nervous system (ENS). However, the variety of...
Autonomous regulation of the intestine requires the combined activity of functionally distinct neurons of the enteric nervous system (ENS). However, the variety of enteric neuron types and how they emerge during development remain largely unknown. Here, we define a molecular taxonomy of 12 enteric neuron classes within the myenteric plexus of the mouse small intestine using single-cell RNA sequencing. We present cell-cell communication features and histochemical markers for motor neurons, sensory neurons and interneurons, together with transgenic tools for class-specific targeting. Transcriptome analysis of the embryonic ENS uncovers a novel principle of neuronal diversification, where two neuron classes arise through a binary neurogenic branching and all other identities emerge through subsequent postmitotic differentiation. We identify generic and class-specific transcriptional regulators and functionally connect Pbx3 to a postmitotic fate transition. Our results offer a conceptual and molecular resource for dissecting ENS circuits and predicting key regulators for directed differentiation of distinct enteric neuron classes.
Topics: Animals; Cell Communication; Enteric Nervous System; Homeodomain Proteins; Interneurons; Mice; Mice, Inbred C57BL; Motor Neurons; Myenteric Plexus; Neurons; Proto-Oncogene Proteins; RNA; Sensory Receptor Cells; Sequence Analysis, RNA; Single-Cell Analysis; Transcriptome
PubMed: 33288908
DOI: 10.1038/s41593-020-00736-x -
Nature Medicine Jan 2017The enteric nervous system (ENS) of the gastrointestinal tract controls many diverse functions, including motility and epithelial permeability. Perturbations in ENS...
The enteric nervous system (ENS) of the gastrointestinal tract controls many diverse functions, including motility and epithelial permeability. Perturbations in ENS development or function are common, yet there is no human model for studying ENS-intestinal biology and disease. We used a tissue-engineering approach with embryonic and induced pluripotent stem cells (PSCs) to generate human intestinal tissue containing a functional ENS. We recapitulated normal intestinal ENS development by combining human-PSC-derived neural crest cells (NCCs) and developing human intestinal organoids (HIOs). NCCs recombined with HIOs in vitro migrated into the mesenchyme, differentiated into neurons and glial cells and showed neuronal activity, as measured by rhythmic waves of calcium transients. ENS-containing HIOs grown in vivo formed neuroglial structures similar to a myenteric and submucosal plexus, had functional interstitial cells of Cajal and had an electromechanical coupling that regulated waves of propagating contraction. Finally, we used this system to investigate the cellular and molecular basis for Hirschsprung's disease caused by a mutation in the gene PHOX2B. This is, to the best of our knowledge, the first demonstration of human-PSC-derived intestinal tissue with a functional ENS and how this system can be used to study motility disorders of the human gastrointestinal tract.
Topics: Animals; Calcium; Cell Line; Chick Embryo; Enteric Nervous System; Gastrointestinal Motility; Hirschsprung Disease; Homeodomain Proteins; Humans; Immunohistochemistry; In Vitro Techniques; Induced Pluripotent Stem Cells; Interstitial Cells of Cajal; Intestines; Mice; Mice, SCID; Microscopy, Confocal; Models, Biological; Mutation; Myenteric Plexus; Neural Crest; Neurogenesis; Neuroglia; Neurons; Organoids; Permeability; Real-Time Polymerase Chain Reaction; Submucous Plexus; Tissue Engineering; Transcription Factors
PubMed: 27869805
DOI: 10.1038/nm.4233 -
The American Journal of Surgical... Aug 2021Congenital myenteric hypoganglionosis is a rare developmental disorder characterized clinically by severe and persistent neonatal intestinal pseudoobstruction. The...
Congenital myenteric hypoganglionosis is a rare developmental disorder characterized clinically by severe and persistent neonatal intestinal pseudoobstruction. The diagnosis is established by the prevalence of small myenteric ganglia composed of closely spaced ganglion cells with sparse surrounding neuropil. In practice, the diagnosis entails familiarity with the normal appearance of myenteric ganglia in young infants and the ability to confidently recognize significant deviations in ganglion size and morphology. We review clinical, histologic, and immunohistochemical findings from 12 patients with congenital myenteric hypoganglionosis in comparison with similar data from age-matched controls and clearly delineate the diagnostic features of the condition. Practical guidelines are provided to assist surgical pathologists, who are likely to encounter this condition only infrequently. The diagnosis typically requires full-thickness intestinal biopsy as the abnormality is confined to the myenteric plexus in many patients. Immunohistochemistry for Hu C/D may be used to confirm hypoganglionosis. Reduced staining for calretinin and NeuN implicates a selective deficiency of intrinsic primary afferent neurons in this disease.
Topics: Child; Child, Preschool; Colonic Diseases; Digestive System Abnormalities; Female; Humans; Infant; Intestinal Pseudo-Obstruction; Male; Myenteric Plexus; Neurons
PubMed: 33492848
DOI: 10.1097/PAS.0000000000001670 -
Gastroenterology Jul 2020The enteric nervous system (ENS) exists in close proximity to luminal bacteria. Intestinal microbes regulate ENS development, but little is known about their effects on...
BACKGROUND & AIMS
The enteric nervous system (ENS) exists in close proximity to luminal bacteria. Intestinal microbes regulate ENS development, but little is known about their effects on adult enteric neurons. We investigated whether intestinal bacteria or their products affect the adult ENS via toll-like receptors (TLRs) in mice.
METHODS
We performed studies with conventional C57/BL6, germ-free C57/BL6, Nestin-creER:tdTomato, Nestin-GFP, and ChAT-cre:tdTomato. Mice were given drinking water with ampicillin or without (controls). Germ-free mice were given drinking water with TLR2 agonist or without (controls). Some mice were given a blocking antibody against TLR2 or a TLR4 inhibitor. We performed whole gut transit, bead latency, and geometric center studies. Feces were collected and analyzed by 16S ribosomal RNA gene sequencing. Longitudinal muscle myenteric plexus (LMMP) tissues were collected, analyzed by immunohistochemistry, and levels of nitric oxide were measured. Cells were isolated from colonic LMMP of Nestin-creER:tdTomato mice and incubated with agonists of TLR2 (receptor for gram-positive bacteria), TLR4 (receptor for gram-negative bacteria), or distilled water (control) and analyzed by flow cytometry.
RESULTS
Stool from mice given ampicillin had altered composition of gut microbiota with reduced abundance of gram-positive bacteria and increased abundance of gram-negative bacteria, compared with mice given only water. Mice given ampicillin had reduced colon motility compared with mice given only water, and their colonic LMMP had reduced numbers of nitrergic neurons, reduced neuronal nitric oxide synthase production, and reduced colonic neurogenesis. Numbers of colonic myenteric neurons increased after mice were switched from ampicillin to plain water, with increased markers of neurogenesis. Nestin-positive enteric neural precursor cells expressed TLR2 and TLR4. In cells isolated from the colonic LMMP, incubation with the TLR2 agonist increased the percentage of neurons originating from enteric neural precursor cells to approximately 10%, compared with approximately 0.01% in cells incubated with the TLR4 agonist or distilled water. Mice given an antibody against TLR2 had prolonged whole gut transit times; their colonic LMMP had reduced total neurons and a smaller proportion of nitrergic neurons per ganglion, and reduced markers of neurogenesis compared with mice given saline. Colonic LMMP of mice given the TLR4 inhibitor did not have reduced markers of neurogenesis. Colonic LMMP of germ-free mice given TLR2 agonist had increased neuronal numbers compared with control germ-free mice.
CONCLUSIONS
In the adult mouse colon, TLR2 promotes colonic neurogenesis, regulated by intestinal bacteria. Our findings indicate that colonic microbiota help maintain the adult ENS via a specific signaling pathway. Pharmacologic and probiotic approaches directed towards specific TLR2 signaling processes might be developed for treatment of colonic motility disorders related to use of antibiotics or other factors.
Topics: Adult; Ampicillin; Animals; Cells, Cultured; Colon; Disease Models, Animal; Dysbiosis; Enteric Nervous System; Gastrointestinal Microbiome; Gastrointestinal Motility; Germ-Free Life; Humans; Male; Mice; Mice, Transgenic; Myenteric Plexus; Nestin; Neurogenesis; Nitrergic Neurons; Nitric Oxide; Primary Cell Culture; Toll-Like Receptor 2; Toll-Like Receptor 4
PubMed: 32234538
DOI: 10.1053/j.gastro.2020.03.050 -
Journal of Crohn's & Colitis Jan 2024Pain is a cardinal symptom in inflammatory bowel disease [IBD]. An important structure in the transduction of pain signalling is the myenteric plexus [MP]. Nevertheless,...
BACKGROUND AND AIMS
Pain is a cardinal symptom in inflammatory bowel disease [IBD]. An important structure in the transduction of pain signalling is the myenteric plexus [MP]. Nevertheless, IBD-associated infiltration of the MP by immune cells lacks in-depth characterisation. Herein, we decipher intra- and periganglionic immune cell infiltrations in Crohn´s disease [CD] and ulcerative colitis [UC] and provide a comparison with murine models of colitis.
METHODS
Full wall specimens of surgical colon resections served to examine immune cell populations by either conventional immuno-histochemistry or immunofluorescence followed by either bright field or confocal microscopy. Results were compared with equivalent examinations in various murine models of intestinal inflammation.
RESULTS
Whereas the MP morphology was not significantly altered in IBD, we identified intraganglionic IBD-specific B cell- and monocyte-dominant cell infiltrations in CD. In contrast, UC-MPs were infiltrated by CD8+ T cells and revealed a higher extent of ganglionic cell apoptosis. With regard to the murine models of intestinal inflammation, the chronic dextran sulphate sodium [DSS]-induced colitis model reflected CD [and to a lesser extent UC] best, as it also showed increased monocytic infiltration as well as a modest B cell and CD8+ T cell infiltration.
CONCLUSIONS
In CD, MPs were infiltrated by B cells and monocytes. In UC, mostly CD8+ cytotoxic T cells were found. The chronic DSS-induced colitis in the mouse model reflected best the MP-immune cell infiltrations representative for IBD.
Topics: Animals; Mice; Colitis, Ulcerative; Crohn Disease; Myenteric Plexus; Inflammatory Bowel Diseases; Colitis; Neurotransmitter Agents; Pain; Inflammation
PubMed: 37565754
DOI: 10.1093/ecco-jcc/jjad122 -
Annals of the New York Academy of... Dec 2020Achalasia is a rare motility disorder with incomplete relaxation of the lower esophageal sphincter and ineffective contractions of the esophageal body. It has been... (Review)
Review
Achalasia is a rare motility disorder with incomplete relaxation of the lower esophageal sphincter and ineffective contractions of the esophageal body. It has been hypothesized that achalasia does not result from only one pathway but rather involves a combination of infectious, autoimmune, and familial etiological components. On the basis of other observations, a novel hypothesis suggests that a muscular form of eosinophilic esophagitis is involved in the pathophysiology of achalasia in some patients. This appears to progressively diminish the myenteric plexus at stage III, gradually destroy it at stage II, and finally eliminate it at stage I, the most advanced and final stage of achalasia. Although high-resolution manometry has identified these three different types of achalasia, another subset of patients with a normal-appearing sphincter relaxation has been proposed. Provocative maneuvers, such as the rapid drinking challenge, have recently been demonstrated to improve diagnosis in certain borderline patients, but have to be studied in more detail. However, whether the different types of achalasia will have a long-term impact on tailored therapies is still a matter of debate. Additionally, novel aspects of the standard timed barium swallow appear to be an important adjunct of diagnosis, as it has been shown to have a diagnostic as well as a predictive value.
Topics: Autoimmunity; Deglutition; Eosinophilic Esophagitis; Esophageal Achalasia; Esophageal Sphincter, Lower; Humans; Male; Manometry; Myenteric Plexus
PubMed: 33140485
DOI: 10.1111/nyas.14510