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International Journal of Biological... Mar 2024Oxidative stress-induced enteric neuropathy is an important factor in slow transit constipation (STC). Cistanche deserticola crude polysaccharides (CDCP) are natural...
Oxidative stress-induced enteric neuropathy is an important factor in slow transit constipation (STC). Cistanche deserticola crude polysaccharides (CDCP) are natural antioxidants with various biological activities. We prepared CDCP through water-extract and alcohol-precipitation methods. The structural characteristics of CDCP were analyzed by infrared spectroscopy and methylation analysis. The results showed that CDCP was primarily composed of (1 → 4)-linked glucans with minor amounts of pectic polysaccharides. Different doses of CDCP (100, 200, and 400 mg/kg) were administered to loperamide-induced STC mice to explore the therapeutic effects of CDCP. Compared with the untreated group, CDCP treatment significantly improved constipation symptoms, relevant gut-regulating peptides levels, colonic pathological damage, and colonic myenteric nerons injury. CDCP enhanced the antioxidant capacity by decreasing Malondialdehyde (MDA) content, increasing Superoxide Dismutase (SOD) activity and Reduced Glutathione (GSH) content. CDCP significantly reduced oxidative stress-induced injury by preserving mitochondrial function in the colonic myenteric plexus. Furthermore, the neuroprotective effects of CDCP might be associated with the Nrf2/Keap1 pathway. Thus, our findings first revealed the potential of CDCP to protect the colonic myenteric plexus against oxidative stress-induced damage in STC, establishing CDCP as promising candidates for natural medicine in the clinical management of STC.
Topics: Mice; Animals; Cistanche; Neuroprotective Agents; Kelch-Like ECH-Associated Protein 1; NF-E2-Related Factor 2; Constipation; Oxidative Stress; Antioxidants; Polysaccharides
PubMed: 38246435
DOI: 10.1016/j.ijbiomac.2024.129527 -
Science China. Life Sciences Feb 2024Weaning piglets usually suffer from severe diarrhea (commonly known as postweaning diarrhea, PWD) along with intestinal motility disorder. Intestinal peristalsis is...
Weaning piglets usually suffer from severe diarrhea (commonly known as postweaning diarrhea, PWD) along with intestinal motility disorder. Intestinal peristalsis is mainly regulated by the longitudinal muscle-myenteric plexus (LM-MP). To understand the relationship between intestinal LM-MP function and the development of PWD, we compared the intestinal electrical activity, and the transcriptional profile of the LM-MP between 21-day-old piglets (just weaned, n=7) and 24-day-old piglets (suffered the most severe weaning stress, n=7). The results showed that 24-day-old piglets exhibited different degrees of diarrhea. A significant increase in the slow-wave frequency in the ileum and colon was observed in 24-day-old piglets, while c-kit expression in the intestinal LM-MPs was significantly decreased, indicating that PWD caused by elevated slow-wave frequency may be associated with loss of c-kit. The real-time quantitative PCR (RT-qPCR) and enzyme-linked immunosorbent assay (ELISA) showed that intestinal LM-MPs in 24-day-old piglets may undergo inflammation and oxidative stress. Significant increases in 8-hydroxy-2'-deoxyguanosine and decreases in thioredoxin suggest that weaning may lead to DNA damage in the LM-MP of 24-day-old piglets. In addition, activating transcription factor 3 was significantly upregulated, indicating nerve damage in the LM-MP of 24-day-old piglets. The transcriptomic results showed that most of the differentially expressed genes in the ileal LM-MP after weaning were downregulated and closely related to the cell cycle process. Subsequent RT-qPCR analysis showed that the relative expression of p21 was upregulated, while the expression of cyclin A2, cyclin B1, and proliferating cell nuclear antigen was downregulated in the ileal and colonic LM-MP of 24-day-old piglets, suggesting that weaning may inhibit cell proliferation and cause G1/S cell cycle arrest in ileal and colonic LM-MP. In conclusion, weaning may lead to cell cycle arrest by causing DNA damage in the LM-MP, impairing intestinal motility regulation, and ultimately leading to diarrhea in piglets.
Topics: Animals; Swine; Myenteric Plexus; Weaning; Intestines; Diarrhea; Muscles; Intestinal Mucosa
PubMed: 37824029
DOI: 10.1007/s11427-022-2391-x -
Cell and Tissue Research Jan 2024Nothobranchius furzeri is emerging as an exciting vertebrate organism in the field of biomedicine, developmental biology and ecotoxicology research. Its short generation...
Nothobranchius furzeri is emerging as an exciting vertebrate organism in the field of biomedicine, developmental biology and ecotoxicology research. Its short generation time, compressed lifespan and accelerated ageing make it a versatile model for longitudinal studies with high traceability. Although in recent years the use of this model has increased enormously, there is still little information on the anatomy, morphology and histology of its main organs. In this paper, we present a description of the digestive system of N. furzeri, with emphasis on the intestine. We note that the general architecture of the intestinal tissue is shared with other vertebrates, and includes a folding mucosa, an outer muscle layer and a myenteric plexus. By immunohistochemical analysis, we reveal that the mucosa harbours the same type of epithelial cells observed in mammals, including enterocytes, goblet cells and enteroendocrine cells, and that the myenteric neurons express neurotransmitters common to other species, such as serotonin, substance P and tyrosine hydroxylase. In addition, we detect the presence of a proliferative compartment at the base of the intestinal folds. The description of the normal intestinal morphology provided here constitutes a baseline information to contrast with tissue alterations in future lines of research assessing pathologies, ageing-related diseases or damage caused by toxic agents.
Topics: Animals; Aging; Intestines; Mammals
PubMed: 38015266
DOI: 10.1007/s00441-023-03845-8 -
European Journal of Pharmacology Dec 2023Gastric lesions have several aetiologies, among which stress is the most prominent. Therefore, identification of new therapies to prevent stress is of considerable...
Gastric lesions have several aetiologies, among which stress is the most prominent. Therefore, identification of new therapies to prevent stress is of considerable importance. Alpha-ketoglutarate (α-kg) several beneficial effects and has shown promise in combating oxidative stress, inflammation, and premature aging. Thus, this study aimed to evaluate the protective effect of α-kg in a gastric damage model by water-immersion restraint stress (WIRS). Pretreatment with α-kg decreased stress-related histopathological scores of tissue oedema, cell loss, and inflammatory infiltration. The α-kg restored the percentage of type III collagen fibres. Mucin levels were preserved as well as the structure and area of the myenteric plexus ganglia were preserved after pretreatment with α-kg. Myeloperoxidase (MPO) levels and the expression of pro-inflammatory cytokines (TNF-α and IL-1β) were also reduced following α-kg pretreatment. Decreased levels of glutathione (GSH) in the stress group were restored by α-kg. The omeprazole group was used as standard drug e also demonstrated improve on some parameters after the exposition to WIRS as inflammatory indexes, GSH and mucin. Through this, was possible to observe that α-kg can protect the gastric mucosa exposed to WIRS, preserve tissue architecture, reduce direct damage to the mucosa and inflammatory factors, stimulate the production of type III collagen and mucin, preserve the myenteric plexus ganglia, and maintain antioxidant potential. Due to, we indicate that α-kg has protective activity of the gastric mucosa, demonstrating its ability to prevent damage associated with gastric lesions caused by stress.
Topics: Mice; Animals; Ketoglutaric Acids; Stomach Ulcer; Collagen Type III; Immersion; Gastric Mucosa; Glutathione; Mucins; Water; Restraint, Physical
PubMed: 37871764
DOI: 10.1016/j.ejphar.2023.176118 -
Cellular and Molecular Gastroenterology... 2024The presence of myenteric plexitis in the proximal resection margins is a predictive factor of early postoperative recurrence in Crohn's disease. To decipher the...
BACKGROUND & AIMS
The presence of myenteric plexitis in the proximal resection margins is a predictive factor of early postoperative recurrence in Crohn's disease. To decipher the mechanisms leading to their formation, T-cell interactions with enteric neural cells were studied in vitro and in vivo.
METHODS
T cells close to myenteric neural cells were retrospectively quantified in ileocolonic resections from 9 control subjects with cancer and 20 patients with Crohn's disease. The mechanisms involved in T-cell adhesion were then investigated in co-cultures of T lymphocytes with enteric glial cells (glia). Finally, the implication of adhesion molecules in the development of plexitis and colitis was studied in vitro but also in vivo in Winnie mice.
RESULTS
The mean number of T cells close to glia, but not neurons, was significantly higher in the myenteric ganglia of relapsing patients with Crohn's disease (2.42 ± 0.5) as compared with controls (0.36 ± 0.08, P = .0007). Co-culture experiments showed that exposure to proinflammatory cytokines enhanced T-cell adhesion to glia and increased intercellular adhesion molecule-1 (ICAM-1) expression in glia. We next demonstrated that T-cell adhesion to glia was inhibited by an anti-ICAM-1 antibody. Finally, using the Winnie mouse model of colitis, we showed that the blockage of ICAM-1/lymphocyte function-associated antigen-1 (LFA-1) with lifitegrast reduced colitis severity and decreased T-cell infiltration in the myenteric plexus.
CONCLUSIONS
Our present work argues for a role of glia-T-cell interaction in the development of myenteric plexitis through the adhesion molecules ICAM-1/LFA-1 and suggests that deciphering the functional consequences of glia-T-cell interaction is important to understand the mechanisms implicated in the development and recurrence of Crohn's disease.
Topics: Humans; Crohn Disease; Intercellular Adhesion Molecule-1; Neuroglia; Animals; T-Lymphocytes; Male; Coculture Techniques; Cell Adhesion; Female; Adult; Myenteric Plexus; Mice; Middle Aged; Retrospective Studies; Aged
PubMed: 38428588
DOI: 10.1016/j.jcmgh.2024.02.016 -
British Journal of Pharmacology Feb 2024Glial cell-derived neurotrophic factor (GDNF) maintains gut homeostasis. Dopamine promotes GDNF release in astrocytes. We investigated the regulation by dopamine of...
BACKGROUND AND PURPOSE
Glial cell-derived neurotrophic factor (GDNF) maintains gut homeostasis. Dopamine promotes GDNF release in astrocytes. We investigated the regulation by dopamine of colonic GDNF secretion.
EXPERIMENTAL APPROACH
D receptor knockout (D R ) mice, adeno-associated viral 9-short hairpin RNA carrying D receptor (AAV9-shD R)-treated mice, 6-hydroxydopamine treated (6-OHDA) rats and primary enteric glial cells (EGCs) culture were used. Incubation fluid from colonic submucosal plexus and longitudinal muscle myenteric plexus were collected for GDNF and ACh measurements.
KEY RESULTS
D receptor-immunoreactivity (IR), but not D receptor-IR, was observed on EGCs. Both D receptor-IR and D receptor-IR were co-localized on cholinergic neurons. Low concentrations of dopamine induced colonic GDNF secretion in a concentration-dependent manner, which was mimicked by the D receptor agonist SKF38393, inhibited by TTX and atropine and eliminated in D R mice. SKF38393-induced colonic ACh release was absent in D R mice. High concentrations of dopamine suppressed colonic GDNF secretion, which was mimicked by the D receptor agonist quinpirole, and absent in AAV-shD R-treated mice. Quinpirole decreased GDNF secretion by reducing intracellular Ca levels in primary cultured EGCs. Carbachol ( ACh analogue) promoted the release of GDNF. Quinpirole inhibited colonic ACh release, which was eliminated in the AAV9-shD R-treated mice. 6-OHDA treated rats with low ACh and high dopamine content showed decreased GDNF content and increased mucosal permeability in the colon.
CONCLUSION AND IMPLICATIONS
Low concentrations of dopamine promote colonic GDNF secretion via D receptors on cholinergic neurons, whereas high concentrations of dopamine inhibit GDNF secretion via D receptors on EGCs and/or cholinergic neurons.
Topics: Rats; Mice; Animals; Dopamine; Glial Cell Line-Derived Neurotrophic Factor; Quinpirole; Oxidopamine; 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; Receptors, Dopamine D1; Receptors, Dopamine D2; Cholinergic Agents
PubMed: 37614042
DOI: 10.1111/bph.16226 -
Cell and Tissue Research Jan 2024The pig is an important translational model for studying intestinal physiology and disorders for its many homologies with humans, including the organization of the...
The pig is an important translational model for studying intestinal physiology and disorders for its many homologies with humans, including the organization of the enteric nervous system (ENS), the major regulator of gastrointestinal functions. This study focused on the quantification and neurochemical characterization of substance P (SP) neurons in the pig ascending (AC) and descending colon (DC) in wholemount preparations of the inner submucosal plexus (ISP), outer submucosal plexus (OSP), and myenteric plexus (MP). We used antibodies for the pan-neuronal marker HuCD, and choline acetyltransferase (ChAT) and neuronal nitric oxide synthase (nNOS), markers for excitatory and inhibitory transmitters, for multiple labeling immunofluorescence and high-resolution confocal microscopy. The highest density of SP immunoreactive (IR) neurons was in the ISP (222/mm in the AC, 166/mm in the DC), where they make up about a third of HuCD-IR neurons, compared to the OSP and MP (19-22% and 13-17%, respectively, P < 0.001-0.0001). HuCD/SP/ChAT-IR neurons (up to 23%) were overall more abundant than HuCD/SP/nNOS-IR neurons (< 10%). Most SP-IR neurons contained ChAT-IR (62-85%), whereas 18-38% contained nNOS-IR with the highest peak in the OSP. A subpopulation of SP-IR neurons contains both ChAT- and nNOS-IR with the highest peak in the OSP and ISP of DC (33-36%) and the lowest in the ISP of AC (< 10%, P < 0.001). SP-IR varicose fibers were abundant in the ganglia. This study shows that SP-IR neurons are functionally distinct with variable proportions in different plexuses in the AC and DC reflecting diverse functions of specific colonic regions.
Topics: Humans; Swine; Animals; Myenteric Plexus; Submucous Plexus; Substance P; Neurons; Colon; Choline O-Acetyltransferase
PubMed: 37982872
DOI: 10.1007/s00441-023-03842-x -
Journal of Neurogastroenterology and... Oct 2023Multiple sclerosis (MS) is an inflammatory disease characterized by the demyelination of primarily the central nervous system. Diffuse esophageal spasm (DES) and...
BACKGROUND/AIMS
Multiple sclerosis (MS) is an inflammatory disease characterized by the demyelination of primarily the central nervous system. Diffuse esophageal spasm (DES) and achalasia are both disorders of esophageal peristalsis which cause clinical symptoms of dysphagia. Mechanisms involving dysfunction of the pre- and post-ganglionic nerve fibers of the myenteric plexus have been proposed. We sought to determine whether MS confers an increased risk of developing achalasia or DES.
METHODS
Cohort analysis was done using the Explorys database. Univariate logistic regression was performed to determine the odds MS confers to each motility disorder studied. Comparison of proportions of dysautonomia comorbidities was performed among the cohorts. Patients with a prior diagnosis of diabetes mellitus, chronic Chagas' disease, opioid use, or CREST syndrome were excluded from the study.
RESULTS
Odds of MS patients developing achalasia or DES were (OR, 2.09; 95% CI, 1.73-2.52; < 0.001) and (OR, 3.15; 95% CI, 2.89-3.42; < 0.001), respectively. In the MS/achalasia cohort, 27.27%, 18.18%, 9.09%, and 45.45% patients had urinary incontinence, gastroparesis, impotence, and insomnia, respectively. In the MS/DES cohort, 35.19%, 11.11%, 3.70%, and 55.56% had these symptoms. In MS patients without motility disorders, 12.64%, 0.79%, 2.21%, and 21.85% had these symptoms.
CONCLUSIONS
Patients with MS have higher odds of developing achalasia or DES compared to patients without MS. MS patients with achalasia or DES have higher rates of dysautonomia comorbidities. This suggests that these patients have a more severe disease phenotype in regards to the extent of neuronal degradation and demyelination causing the autonomic dysfunction.
PubMed: 37528077
DOI: 10.5056/jnm22173 -
Gastroenterology Mar 2024Hirschsprung's disease is defined by the absence of the enteric nervous system (ENS) from the distal bowel. Primary treatment is "pull-through" surgery to remove bowel...
BACKGROUND & AIMS
Hirschsprung's disease is defined by the absence of the enteric nervous system (ENS) from the distal bowel. Primary treatment is "pull-through" surgery to remove bowel that lacks ENS, with reanastomosis of "normal" bowel near the anal verge. Problems after pull-through are common, and some may be due to retained hypoganglionic bowel (ie, low ENS density). Testing this hypothesis has been difficult because counting enteric neurons in tissue sections is unreliable, even for experts. Tissue clearing and 3-dimensional imaging provide better data about ENS structure than sectioning.
METHODS
Regions from 11 human colons and 1 ileal specimen resected during Hirschsprung's disease pull-through surgery were cleared, stained with antibodies to visualize the ENS, and imaged by confocal microscopy. Control distal colon from people with no known bowel problems were similarly cleared, stained, and imaged.
RESULTS
Quantitative analyses of human colon, ranging from 3 days to 60 years old, suggest age-dependent changes in the myenteric plexus area, ENS ganglion area, percentage of myenteric plexus occupied by ganglia, neurons/mm, and neuron Feret's diameter. Neuron counting using 3-dimensional images was highly reproducible. High ENS density in neonatal colon allowed reliable neuron counts using 500-μm × 500-μm regions (36-fold smaller than in adults). Hirschsprung's samples varied 8-fold in proximal margin enteric neuron density and had diverse ENS architecture in resected bowel.
CONCLUSIONS
Tissue clearing and 3-dimensional imaging provide more reliable information about ENS structure than tissue sections. ENS structure changes during childhood. Three-dimensional ENS anatomy may provide new insight into human bowel motility disorders, including Hirschsprung's disease.
PubMed: 38494035
DOI: 10.1053/j.gastro.2024.02.045 -
Frontiers in Neuroscience 2023The central and peripheral nervous systems provide cholinergic innervation in the colon. The ability to assess their neuroanatomical distinctions is still a challenge....
INTRODUCTION
The central and peripheral nervous systems provide cholinergic innervation in the colon. The ability to assess their neuroanatomical distinctions is still a challenge. The pig is regarded as a relevant translational model due to the close similarity of its enteric nervous system (ENS) with that of human. Opioid-induced constipation is one of the most common side effects of opioid therapy.
METHODS
We developed an approach to differentiate the central and peripheral cholinergic innervation of the pig colon using double immunolabeling with a novel mouse anti-human peripheral type of choline acetyltransferase (hpChAT) antibody combined with a rabbit anti-common type of ChAT (cChAT) antibody, a reliable marker of cholinergic neurons in the central nervous system. We examined their spatial configurations in 3D images of the ENS generated from CLARITY-cleared colonic segments. The density was quantitated computationally using Imaris 9.7. We assessed changes in the distal colon induced by daily oral treatment for 4 weeks with the μ opioid receptor agonist, loperamide (0.4 or 3 mg/kg).
RESULTS
The double labeling showed strong cChAT immunoreactive (ir) fibers in the cervical vagus nerve and neuronal somata and fibers in the ventral horn of the sacral (S2) cord while hpChAT immunoreactivity was visualized only in the ENS but not in the vagus or sacral neural structures indicating the selectivity of these two antibodies. In the colonic myenteric plexus, dense hpChAT-ir neurons and fibers and varicose cChAT-ir fibers surrounding hpChAT-ir neurons were simultaneously visualized in 3D. The density of cChAT-ir varicose fibers in the outer submucosal plexus of both males and females were higher in the transverse and distal colon than in the proximal colon and in the myenteric plexus compared to the outer submucosal plexus and there was no cChAT innervation in the inner submucosal plexus. The density of hpChAT in the ENS showed no segmental or plexus differences in both sexes. Loperamide at the highest dose significantly decreased the density hpChAT-ir fibers + somata in the myenteric plexus of the distal colon.
DISCUSSION
These data showed the distinct density of central cholinergic innervation between myenteric and submucosal plexuses among colonic segments and the localization of cChAT-ir fibers around peripheral hpChAT neurons in 3D. The reduction of cholinergic myenteric innervation by chronic opiate treatment points to target altered prokinetic cholinergic pathway to counteract opiate constipation.
PubMed: 37650102
DOI: 10.3389/fnins.2023.1204233