Prokineticin-2 Is Highly Expressed in Colonic Mucosa of Early Parkinson's Disease Patients.

BACKGROUND: Elevated levels of prokineticin-2 (PK2), regarded as a protein involved in modulating immune/inflammatory responses, have been detected in the substantia nigra, serum, and olfactory neurons of Parkinson's disease (PD) patients. Of note, emerging evidence suggests that gut alterations, including dysbiosis and enteric inflammation, play a role in PD via the gut-brain axis. OBJECTIVES: Our goal was to investigate the expression of PK2 in colonic biopsies of PD patients. METHODS: Mucosal biopsies from the descending colon were obtained in 11 PD patients and five asymptomatic subjects. Biopsy samples were processed for PK2 immunofluorescence and western blot. RESULTS: We revealed an increased PK2 expression in colonic mucosa from PD patients in the early stages compared to controls. In addition, we found that PK2 was expressed by activated enteric glial cells and macrophages. CONCLUSIONS: PK2 is highly expressed within neurogenic/inflammatory cells of colonic mucosa from early PD patients, suggesting a potential role of PK2 in gut inflammation, especially in the early stages of PD. © 2024 International Parkinson and Movement Disorder Society.

Pathological Remodeling of the Gut Barrier as a Prodromal Event of High-Fat Diet-Induced Obesity.

Intestinal barrier alterations represent a primum movens in obesity and related intestinal dysfunctions. However, whether gut barrier remodeling represents prodromal events in obesity before weight gain, metabolic alterations, and systemic inflammation remains unclear. Herein, we examined morphologic changes in the gut barrier in a mouse model of high-fat diet (HFD) since the earliest phases of diet assumption. C57BL/6J mice were fed with standard diet (SD) or HFD for 1, 2, 4, or 8 weeks. Remodeling of intestinal epithelial barrier, inflammatory infiltrate, and collagen deposition in the colonic wall was assessed by histochemistry and immunofluorescence analysis. Obese mice displayed increased body and epididymal fat weight along with increased plasma resistin, IL-1ß, and IL-6 levels after 8 weeks of HFD. Starting from 1 week of HFD, mice displayed (1) a decreased claudin-1 expression in lining epithelial cells, (2) an altered mucus in goblet cells, (3) an increase in proliferating epithelial cells in colonic crypts, (4) eosinophil infiltration along with an increase in vascular P-selectin, and (5) deposition of collagen fibers. HFD intake is associated with morphologic changes in the large bowel at mucosal and submucosal levels. In particular, the main changes include alterations in the mucous layer and intestinal epithelial barrier integrity and activation of mucosal defense-enhanced fibrotic deposition. These changes represent early events occurring before the development of obesity condition that could contribute to compromising the intestinal mucosal barrier and functions, opening the way for systemic dissemination.

Intestinal histomorphological and molecular alterations in patients with Parkinson's disease.

BACKGROUND AND PURPOSE: Changes in gut microbiota composition, enteric inflammation, impairments of the intestinal epithelial barrier and neuroplastic changes in the enteric nervous system have been reported in Parkinson's disease (PD) patients and could contribute to the onset of both neurological and gastrointestinal symptoms. However, their mutual interplay has rarely been investigated. This study evaluated, in an integrated manner, changes in faecal microbiota composition, morphofunctional alterations of colonic mucosal barrier and changes of inflammatory markers in blood and stools of PD patients. METHODS: Nineteen PD patients and nineteen asymptomatic subjects were enrolled. Blood lipopolysaccharide binding protein (LBP, marker of altered intestinal permeability) and interleukin-1ß (IL-1ß) levels, as well as stool IL-1ß and tumour necrosis factor (TNF) levels, were evaluated. Gut microbiota analysis was performed. Epithelial mucins, collagen fibres, claudin-1 and S100-positive glial cells as markers of an impairment of the intestinal barrier, mucosal remodelling and enteric glial activation were evaluated on colonic mucosal specimens collected during colonoscopy. RESULTS: Faecal microbiota analysis revealed a significant difference in the α-diversity in PD patients compared to controls, while no differences were found in the ß-diversity. Compared to controls, PD patients showed significant chenags in plasma LBP levels, as well as faecal TNF and IL-1ß levels. The histological analysis showed a decrease in epithelial neutral mucins and claudin-1 expression and an increased expression of acidic mucins, collagen fibres and S100-positive glial cells. CONCLUSIONS: Parkinson's disease patients are characterized by enteric inflammation and increased intestinal epithelial barrier permeability, as well as colonic mucosal barrier remodelling, associated with changes in gut microbiota composition.

Pathological remodelling of colonic wall following dopaminergic nigrostriatal neurodegeneration.

BACKGROUND AND AIM: Patients with Parkinson's disease (PD) are often characterized by functional gastrointestinal disorders. Such disturbances can occur at all stages of PD and precede the typical motor symptoms of the disease by many years. However, the morphological alterations associated with intestinal disturbances in PD are undetermined. This study examined the remodelling of colonic wall in 6-hydroxydopamine (6-OHDA)-induced PD rats. METHODS: 8 weeks after 6-OHDA injection animals were sacrificed. Inflammatory infiltrates, collagen deposition and remodelling of intestinal epithelial barrier and tunica muscularis in the colonic wall were assessed by histochemistry, immunohistochemistry, immunofluorescence and western blot analysis. RESULTS: 6-OHDA rats displayed significant alterations of colonic tissues as compared with controls. Signs of mild inflammation (eosinophil infiltration) and a transmural deposition of collagen fibres were observed. Superficial colonic layers were characterized by severe morphological alterations. In particular, lining epithelial cells displayed a reduced claudin-1 and transmembrane 16A/Anoctamin 1 (TMEM16A/ANO1) expression; goblet cells increased their mucin expression; colonic crypts were characterized by an increase in proliferating epithelial cells; the density of S100-positive glial cells and vimentin-positive fibroblast-like cells was increased as well. Several changes were found in the tunica muscularis: downregulation of α-smooth muscle actin/desmin expression and increased proliferation of smooth muscle cells; increased vimentin expression and proliferative phenotype in myenteric ganglia; reduction of interstitial cells of Cajal (ICCs) density. CONCLUSIONS: A pathological remodelling occurs in the colon of 6-OHDA rats. The main changes include: enhanced fibrotic deposition; alterations of the epithelial barrier; activation of mucosal defense; reduction of ICCs. These results indicate that central nigrostriatal denervation is associated with histological changes in the large bowel at mucosal, submucosal and muscular level. These alterations might represent morphological correlates of digestive symptoms in PD.

Prodromal Intestinal Events in Alzheimer's Disease (AD): Colonic Dysmotility and Inflammation Are Associated with Enteric AD-Related Protein Deposition.

Increasing evidence suggests that intestinal dysfunctions may represent early events in Alzheimer's disease and contribute to brain pathology. This study examined the relationship between onset of cognitive impairment and colonic dysfunctions in a spontaneous AD model before the full development of brain pathology. SAMP8 mice underwent Morris water maze and assessment of faecal output at four, six and eight months of age. In vitro colonic motility was examined. Faecal and colonic Aß, tau proteins, α-synuclein and IL-1ß were assessed by ELISA. Colonic citrate synthase activity was assessed by spectrophotometry. Colonic NLRP3, caspase-1 and ASC expression were evaluated by Western blotting. Colonic eosinophil density and claudin-1 expression were evaluated by immunohistochemistry. The effect of Aß on NLRP3 signalling and mitochondrial function was tested in cultured cells. Cognitive impairment and decreased faecal output occurred in SAMP8 mice from six months. When compared with SAMR1, SAMP8 animals displayed: (1) impaired in vitro colonic contractions; (2) increased enteric AD-related proteins, IL-1ß, active-caspase-1 expression and eosinophil density; and (3) decreased citrate synthase activity and claudin-1 expression. In THP-1 cells, Aß promoted IL-1ß release, which was abrogated upon incubation with caspase-1 inhibitor or in ASC-/- cells. Aß decreased mitochondrial function in THP-1 cells. In SAMP8, enteric AD-related proteins deposition, inflammation and impaired colonic excitatory neurotransmission, occurring before the full brain pathology development, could contribute to bowel dysmotility and represent prodromal events in AD.

Interplay between colonic inflammation and tachykininergic pathways in the onset of colonic dysmotility in a mouse model of diet-induced obesity.

BACKGROUND: The murine model of high fat diet (HFD)-induced obesity is characterized by an increment of intestinal permeability, secondary to an impairment of mucosal epithelial barrier and enteric inflammation, followed by morphofunctional rearrangement of the enteric nervous system. The present study investigated the involvement of abdominal macrophages in the mechanisms underlying the development of enteric dysmotility associated with obesity. METHODS: Wild type C57BL/6J mice were fed with HFD (60% kcal from fat) or normocaloric diet (NCD, 18% kcal from fat) for 8 weeks. Groups of mice fed with NCD or HFD were treated with clodronate encapsulated into liposomes to deplete abdominal macrophages. Tachykininergic contractions, elicited by electrical stimulation or exogenous substance P (SP), were recorded in vitro from longitudinal muscle colonic preparations. Substance P distribution was examined by confocal immunohistochemistry. The density of macrophages in the colonic wall was examined by immunohistochemical analysis. Malondialdehyde (MDA, colorimetric assay) and IL-1ß (ELISA assay) levels were also evaluated. RESULTS: MDA and IL-1ß levels were increased in colonic tissues from HFD-treated animals. In colonic preparations, electrically evoked tachykininergic contractions were enhanced in HFD mice. Immunohistochemistry displayed an increase in substance P immunoreactivity in myenteric ganglia, as well as in the muscular layers of colonic cryosections from obese mice. Macrophage depletion in HFD mice was associated with a significant reduction of colonic inflammation. In addition, the decrease in macrophage density attenuated the morphofunctional alterations of tachykininergic pathways observed in obese mice. CONCLUSION: Obesity elicited by HFD determines a condition of colonic inflammation, followed by a marked rearrangement of motor excitatory tachykininergic enteric nerves. Macrophage depletion counteracted the morphofunctional changes of colonic neuromuscular compartment, suggesting a critical role for these immune cells in the onset of enteric dysmotility associated with obesity.

Aging Modulates the Influence of Arginase on Endothelial Dysfunction in Obesity.

Objective- Arginase can reduce NO availability. In this study, we explored arginase as a determinant of endothelial dysfunction in small arteries from obese patients and its relationship with aging and microvascular remodeling. Approach and Results- Small arteries were dissected after subcutaneous fat biopsies and evaluated on a pressurized micromyograph. Endothelium-dependent vasodilation was assessed by acetylcholine, repeated under L-NAME ( N G-nitro-L-arginine-methyl ester), N(ω)-hydroxy-nor-l-arginine (arginase inhibitor) and gp91ds-tat (NADPH [nicotinamide adenine dinucleotide phosphate oxidase] oxidase inhibitor) in vessels from young (age <30 years) control and obese and old (>30 years) control and obese subjects. Media-lumen ratio and amount of vascular wall fibrosis were used as markers of vascular remodeling. Amount of vascular superoxide anions and NO production were determined with immunofluorescence, whereas arginase expression was quantified using Western blot and quantitative polymerase chain reaction. Obese and older age groups had lower vascular NO, as well as higher media-lumen ratio, wall fibrosis, intravascular superoxide, and blunted inhibitory effect of L-NAME on acetylcholine versus controls and younger age groups. N(ω)-hydroxy-nor-l-arginine restored the acetylcholine-induced vasodilation in young and, to a lesser extent, in old obese subjects. This effect was abolished by addition of L-NAME. Gp91ds-tat increased the vasodilatory response to N(ω)-hydroxy-nor-l-arginine in old obese. Superoxide anions and arginase I/II levels were higher in the vascular wall of obese versus controls. Conclusions- Arginase contributes to microvascular endothelial dysfunction in obesity. Its impact is reduced by aging because of higher levels of vascular oxidative stress. Obesity is accompanied by accelerated microvascular remodeling, the extent of which is related to the amount of arginase in the vascular wall.

Colonic motor dysfunctions in a mouse model of high-fat diet-induced obesity: an involvement of A2B adenosine receptors.

Adenosine A2B receptors (A2BR) regulate several enteric functions. However, their implication in the pathophysiology of intestinal dysmotility associated with high-fat diet (HFD)-induced obesity has not been elucidated. We investigated the expression of A2BR in mouse colon and their role in the mechanisms underlying the development of enteric dysmotility associated with obesity. Wild-type C57BL/6J mice were fed with HFD (60% kcal from fat) or normocaloric diet (NCD; 18% kcal from fat) for 8 weeks. Colonic A2BR localization was examined by immunofluorescence. The role of A2BR in the control of colonic motility was examined in functional experiments on longitudinal muscle preparations (LMPs). In NCD mice, A2BR were predominantly located in myenteric neurons; in HFD animals, their expression increased throughout the neuromuscular layer. Functionally, the A2BR antagonist MRS1754 enhanced electrically induced NK1-mediated tachykininergic contractions in LMPs from HFD mice, while it was less effective in tissues from NCD mice. The A2B receptor agonist BAY 60-6583 decreased colonic tachykininergic contractions in LMPs, with higher efficacy in preparations from obese mice. Both A2BR ligands did not affect contractions elicited by exogenous substance P. Obesity is related with a condition of colonic inflammation, leading to an increase of A2BR expression. A2BR, modulating the activity of excitatory tachykininergic nerves, participate to the enteric dysmotility associated with obesity.

Alteration of colonic excitatory tachykininergic motility and enteric inflammation following dopaminergic nigrostriatal neurodegeneration.

BACKGROUND: Parkinson's disease (PD) is frequently associated with gastrointestinal (GI) symptoms, including constipation and defecatory dysfunctions. The mechanisms underlying such disorders are still largely unknown, although the occurrence of a bowel inflammatory condition has been hypothesized. This study examined the impact of central dopaminergic degeneration, induced by intranigral injection of 6-hydroxydopamine (6-OHDA), on distal colonic excitatory tachykininergic motility in rats. METHODS: Animals were euthanized 4 and 8 weeks after 6-OHDA injection. Tachykininergic contractions, elicited by electrical stimulation or exogenous substance P (SP), were recorded in vitro from longitudinal muscle colonic preparations. SP, tachykininergic NK1 receptor, and glial fibrillary acidic protein (GFAP) expression, as well as the density of eosinophils and mast cells in the colonic wall, were examined by immunohistochemical analysis. Malondialdehyde (MDA, colorimetric assay), TNF, and IL-1ß (ELISA assay) levels were also examined. The polarization of peritoneal macrophages was evaluated by real-time PCR. RESULTS: In colonic preparations, electrically and SP-evoked tachykininergic contractions were increased in 6-OHDA rats. Immunohistochemistry displayed an increase in SP and GFAP levels in the myenteric plexus, as well as NK1 receptor expression in the colonic muscle layer of 6-OHDA rats. MDA, TNF, and IL-1ß levels were increased also in colonic tissues from 6-OHDA rats. In 6-OHDA rats, the number of eosinophils and mast cells was increased as compared with control animals, and peritoneal macrophages polarized towards a pro-inflammatory phenotype. CONCLUSIONS: The results indicate that the induction of central nigrostriatal dopaminergic degeneration is followed by bowel inflammation associated with increased oxidative stress, increase in pro-inflammatory cytokine levels, activation of enteric glia and inflammatory cells, and enhancement of colonic excitatory tachykininergic motility.

Enteric Dysfunctions in Experimental Parkinson's Disease: Alterations of Excitatory Cholinergic Neurotransmission Regulating Colonic Motility in Rats.

Parkinson's disease is frequently associated with gastrointestinal symptoms, mostly represented by constipation and defecatory dysfunctions. This study examined the impact of central dopaminergic denervation, induced by injection of 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle, on distal colonic excitatory cholinergic neuromotor activity in rats. Animals were euthanized 4 and 8 weeks after 6-OHDA injection. In vivo colonic transit was evaluated by radiologic assay. Electrically induced and carbachol-induced cholinergic contractions were recorded in vitro from longitudinal and circular muscle colonic preparations, whereas acetylcholine levels were assayed in the incubation media. Choline acetyltransferase (ChAT), HuC/D (pan-neuronal marker), muscarinic M2 and M3 receptors were assessed by immunohistochemistry or western blot assay. As compared with control rats, at week 4, 6-OHDA-treated animals displayed the following changes: decreased in vivo colonic transit rate, impaired electrically evoked neurogenic cholinergic contractions, enhanced carbachol-induced contractions, decreased basal and electrically stimulated acetylcholine release from colonic tissues, decreased ChAT immunopositivity in the neuromuscular layer, unchanged density of HuC/D immunoreactive myenteric neurons, and increased expression of colonic muscarinic M2 and M3 receptors. The majority of such alterations were also detected at week 8 post 6-OHDA injection. These findings indicate that central nigrostriatal dopaminergic denervation is associated with an impaired excitatory neurotransmission characterized by a loss of myenteric neuronal ChAT positivity and decrease in acetylcholine release, resulting in a dysregulated smooth muscle motor activity, which likely contributes to the concomitant decrease in colonic transit rate.

An integrated assessment of histopathological changes of the enteric neuromuscular compartment in experimental colitis.

Bowel inflammatory fibrosis has been largely investigated, but an integrated assessment of remodelling in inflamed colon is lacking. This study evaluated tissue and cellular changes occurring in colonic wall upon induction of colitis, with a focus on neuromuscular compartment. Colitis was elicited in rats by 2,4-dinitrobenzenesulfonic acid (DNBS). After 6 and 21 days, the following parameters were assessed on paraffin sections from colonic samples: tissue injury and inflammatory infiltration by histology; collagen and elastic fibres by histochemistry; HuC/D, glial fibrillar acidic protein (GFAP), proliferating cell nuclear antigen (PCNA), nestin, substance P (SP), von Willebrand factor, c-Kit and transmembrane 16A/Anoctamin1 (TMEM16A/ANO1) by immunohistochemistry. TMEM16A/ANO1 was also examined in isolated colonic smooth muscle cells (ICSMCs). On day 6, inflammatory alterations and fibrosis were present in DNBS-treated rats; colonic wall thickening and fibrotic remodelling were evident on day 21. Colitis was associated with both an increase in collagen fibres and a decrease in elastic fibres. Moreover, the neuromuscular compartment of inflamed colon displayed a significant decrease in neuron density and increase in GFAP/PCNA-positive glia of myenteric ganglia, enhanced expression of neural SP, blood vessel remodelling, reduced c-Kit- and TMEM16A/ANO1-positive interstitial cells of Cajal (ICCs), as well as an increase in TMEM16A/ANO1 expression in muscle tissues and ICSMCs. The present findings provide an integrated view of the inflammatory and fibrotic processes occurring in the colonic neuromuscular compartment of rats with DNBS-induced colitis. These morphological alterations may represent a suitable basis for understanding early pathophysiological events related to bowel inflammatory fibrosis.

Enteric glial NLRP3 inflammasome contributes to gut mucosal barrier alterations in a mouse model of diet-induced obesity.

AIM: In the present study, we investigated the involvement of NLRP3 inflammasome in the intestinal epithelial barrier (IEB) changes associated with obesity, and its role in the interplay between enteric glia and intestinal epithelial cells (IECs). METHODS: Wild-type C57BL/6J and NLRP3-KO (-/-) mice were fed with high-fat diet (HFD) or standard diet for 8 weeks. Colonic IEB integrity and inflammasome activation were assessed. Immunolocalization of colonic mucosal GFAP- and NLRP3-positive cells along with in vitro coculture experiments with enteric glial cells (EGCs) and IECs allowed to investigate the potential link between altered IEB, enteric gliosis, and NLRP3 activation. RESULTS: HFD mice showed increased body weight, altered IEB integrity, increased GFAP-positive glial cells, and NLRP3 inflammasome hyperactivation. HFD-NLRP3-/- mice showed a lower increase in body weight, an improvement in IEB integrity and an absence of enteric gliosis. Coculture experiments showed that palmitate and lipopolysaccharide contribute to IEB damage and promote enteric gliosis with consequent hyperactivation of enteric glial NLRP3/caspase-1/IL-1ß signaling. Enteric glial-derived IL-1ß release exacerbates the IEB alterations. Such an effect was abrogated upon incubation with anakinra (IL-1ß receptor antagonist) and with conditioned medium derived from silenced-NLRP3 glial cells. CONCLUSION: HFD intake elicits mucosal enteric gliotic processes characterized by a hyperactivation of NLRP3/caspase-1/IL-1ß signaling pathway, that contributes to further exacerbate the disruption of intestinal mucosal barrier integrity. However, we cannot rule out the contribution of NLRP3 inflammasome activation from other cells, such as immune cells, in IEB alterations associated with obesity. Overall, our results suggest that enteric glial NLRP3 inflammasome might represent an interesting molecular target for the development of novel pharmacological approaches aimed at managing the enteric inflammation and intestinal mucosal dysfunctions associated with obesity.

Histopathology in gastrointestinal neuromuscular diseases: methodological and ontological issues.

Gastrointestinal neuromuscular diseases (GINMDs) comprise a heterogenous group of chronic conditions associated with impaired gut motility. These gastrointestinal (GI) disorders, differing for etiopathogenic mechanisms, pathologic lesions, and region of gut involvement, represent a relevant matter for public health, because they are very common, can be disabling, and determine major social and economic burdens. GINMDs are presumed or proven to arise as a result of a dysfunctioning GI neuromuscular apparatus, which includes myenteric ganglia (neurons and glial cells), interstitial cells of Cajal and smooth muscle cells. Despite the presence of symptoms related to gut dysmotility in the clinical phenotype of these patients, in the diagnostic setting scarce attention is usually paid to the morphologic pattern of the GI neuromuscular apparatus. It is also objectively difficult to collect full-thickness gut tissue samples from patients with GINMDs, because their disease, which can be only functional in nature, may not justify invasive diagnostic procedures as a first-line approach. As a consequence, whenever available, bioptic gut specimens, retrieved from these patients, must be regarded as a unique chance for obtaining relevant diagnostic information. On the basis of these arguments, there is an urgent need of standardized and validated histopathologic methods, aiming at overcoming the discrepancies affecting current approaches, which usually lead to conflicting definitions of normality and hamper the identification of disease-specific pathologic patterns. This review article intends to address current methodological and ontological issues in the histopathologic diagnosis of GINMDs, to foster the debate on how to discriminate normal morphology from abnormalities.

Role of cyclooxygenase pathways in bowel fibrotic remodelling in a murine model of experimental colitis.

OBJECTIVE: Gut fibrosis occurs under chronic inflammation. This study examined the effects of different cyclooxygenase (COX) inhibitors on fibrosis in the inflamed colon. METHODS: Colitis was induced by 2,4-dinitrobenzenesulfonic acid (DNBS) in albino male Sprague-Dawley rats. After 6, 12 and 18 days, macroscopic and microscopic damage, collagen and elastic fibre content were examined. At day 6, pro-fibrotic factors (collagen I and III, hydroxyproline, fibronectin, matrix metalloproteinase-2 and -9), transforming growth factor-beta (TGF-ß) signalling [TGF-ß, Ras homolog gene family member A (RhoA), phosphorylated small mother against decapentaplegic (pSMAD)-2 and -6] and peristalsis were assessed, and the effects of indomethacin, SC-560 or celecoxib were tested. KEY FINDINGS: Six days after DNBS administration, significant histopathological signs of fibrotic remodelling were observed in rats. At day 6, pro-fibrotic factors were up-regulated and colonic peristalsis was altered. COX inhibitors reversed the histochemical, molecular and functional changes in the fibrotic colon. COX inhibition reduced TGF-ß expression, SMAD2 phosphorylation and RhoA, and increased SMAD6 expression. CONCLUSIONS: Colonic fibrosis is associated with altered bowel motility and induction of profibrotic factors driven by TGF-ß signalling. COX-1 and COX-2 inhibition counteracts this fibrotic remodelling by the modulation of TGF-ß/SMAD signalling, mainly via SMAD6 induction and reduction in SMAD2 phosphorylation.

Enteric Glia and Brain Astroglia: Complex Communication in Health and Disease along the Gut-Brain Axis.

Several studies have provided interesting evidence about the role of the bidirectional communication between the gut and brain in the onset and development of several pathologic conditions, including inflammatory bowel diseases (IBDs), neurodegenerative diseases, and related comorbidities. Indeed, patients with IBD can experience neurologic disorders, including depression and cognitive impairment, besides typical intestinal symptoms. In parallel, patients with neurodegenerative disease, such as Parkinson disease and Alzheimer disease, are often characterized by the occurrence of functional gastrointestinal disorders. In this context, enteric glial cells and brain astrocytes are emerging as pivotal players in the initiation/maintenance of neuroinflammatory responses, which appear to contribute to the alterations of intestinal and neurologic functions observed in patients with IBD and neurodegenerative disorders. The present review was conceived to provide a comprehensive and critical overview of the available knowledge on the morphologic, molecular, and functional changes occurring in the enteric glia and brain astroglia in IBDs and neurologic disorders. In addition, our intent is to identify whether such alterations could represent a common denominator involved in the onset of comorbidities associated with the aforementioned disorders. This might help to identify putative targets useful to develop novel pharmacologic approaches for the therapeutic management of such disturbances.

Dietary Supplementation with the Probiotic SF68 Reinforces Intestinal Epithelial Barrier in Obese Mice by Improving Butyrate Bioavailability.

SCOPE: Modifications in intestinal microbiota and its metabolites, the short-chain fatty acids (SCFA) are main factors altering intestinal epithelial barrier integrity and eliciting the onset of a meta-inflammation observed in obesity. The present study is aimed at evaluating the efficacy of Enterococcus faecium (SF68) administration in counteracting the impairment of gut barrier and enteric inflammation in a model of diet-induced obesity, characterizing the molecular mechanisms underlying such beneficial effects. METHODS AND RESULTS: Male C57BL/6J mice, fed with standard diet (SD) or high-fat diet (HFD), are treated with SF68 (108  CFU day-1 ). After 8 weeks, plasma interleukin (IL)-1ß and lipopolysaccharide binding protein (LBP) are measured, analysis of fecal microbiota composition and butyrate content as well as intestinal malondialdehyde, myeloperoxidase, mucins, tight junction protein, and butyrate transporter expression are investigated. After 8 weeks, SF68 administration counteracts the body weight gain in HFD mice, reducing plasma IL-1ß and LBP. In parallel, SF68 treatment acts against the intestinal inflammation in HFD-fed animals and improves the intestinal barrier integrity and functionality in obese mice via the increase in tight junction protein and intestinal butyrate transporter (sodium-coupled monocarboxylate transporter 1 ) expression. CONCLUSIONS: Supplementation with SF68 reduces intestinal inflammation and reinforces the enteric epithelial barrier in obese mice, improving the transport and utilization of butyrate.

Immunohistochemical analysis of myenteric ganglia and interstitial cells of Cajal in ulcerative colitis.

Ulcerative colitis (UC) is an inflammatory bowel disease with alterations of colonic motility, which influence clinical symptoms. Although morpho-functional abnormalities in the enteric nervous system have been suggested, in UC patients scarce attention has been paid to possible changes in the cells that control colonic motility, including myenteric neurons, glial cells and interstitial cells of Cajal (ICC). This study evaluated the neural-glial components of myenteric ganglia and ICC in the colonic neuromuscular compartment of UC patients by quantitative immunohistochemical analysis. Full-thickness archival samples of the left colon were collected from 10 patients with UC (5 males, 5 females; age range 45-62 years) who underwent elective bowel resection. The colonic neuromuscular compartment was evaluated immunohistochemically in paraffin cross-sections. The distribution and number of neurons, glial cells and ICC were assessed by anti-HuC/D, -S100ß and -c-Kit antibodies, respectively. Data were compared with findings on archival samples of normal left colon from 10 sex- and age-matched control patients, who underwent surgery for uncomplicated colon cancer. Compared to controls, patients with UC showed: (i) reduced density of myenteric HuC/D(+) neurons and S100ß(+) glial cells, with a loss over 61% and 38%, respectively, and increased glial cell/neuron ratio; (ii) ICC decrease in the whole neuromuscular compartment. The quantitative variations of myenteric neuro-glial cells and ICC indicate considerable alterations of the colonic neuromuscular compartment in the setting of mucosal inflammation associated with UC, and provide a morphological basis for better understanding the motor abnormalities often observed in UC patients.

Nonsteroidal anti-inflammatory drug-activated gene-1 plays a role in the impairing effects of cyclooxygenase inhibitors on gastric ulcer healing.

Nonsteroidal anti-inflammatory drugs (NSAIDs) can impair gastric ulcer healing. This study investigates the involvement of NSAID-activated gene-1 (NAG-1) in ulcer repair impairment by cyclooxygenase (COX) inhibitors. Gastric ulcers were induced in rats by acetic acid. Four days later, animals received daily intragastric indomethacin (nonselective COX-1/COX-2 inhibitor; 1 mg/kg), 5-(4-chlorophenyl)-1-(4-methoxyphenyl)-3-trifluoromethylpyrazole (SC-560) (selective COX-1 inhibitor; 2.5 mg/kg), (5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulfonyl) phenyl-2(5H)-furanone (DFU) (selective COX-2 inhibitor; 5 mg/kg), celecoxib (selective COX-2 inhibitor; 1 mg/kg), and valdecoxib (selective COX-2 inhibitor; 1 mg/kg), for 1, 3, or 7 days. Ulcerated tissues were processed to assess: 1) COX-1, COX-2, NAG-1, proliferating cell nuclear antigen (PCNA), and activated caspase-3 expression; 2) ulcer area; and 3) prostaglandin E(2) (PGE(2)) levels. COX-1 expression in ulcerated tissues was decreased, whereas COX-2 expression was enhanced. Ulcer healing was delayed by indomethacin, DFU, and SC-560, but not by celecoxib and valdecoxib. Ulcer PGE(2) levels were decreased by SC-560, DFU, celecoxib, valdecoxib, and indomethacin. NAG-1 was overexpressed in ulcerated tissues and further enhanced by indomethacin, DFU, and SC-560, but not by celecoxib or valdecoxib. PCNA expression in ulcerated areas was reduced by indomethacin, but not by the other test drugs. The expression of activated caspase-3 in ulcers was increased and enhanced further by indomethacin, DFU, and SC-560, but not by celecoxib and valdecoxib. These findings indicate that: 1) COX inhibitors exert differential impairing effects on gastric ulcer healing, through mechanisms unrelated to the inhibition of COX isoforms and prostaglandin production; and 2) NAG-1 induction, followed by activation of proapoptotic pathways, can contribute to the impairing effects of COX inhibitors on ulcer healing.

From the intestinal mucosal barrier to the enteric neuromuscular compartment: an integrated overview on the morphological changes in Parkinson's disease.

Gastrointestinal dysfunctions represent the most common non-motor symptoms in Parkinson's disease (PD). Of note, changes in gut microbiota, impairments of intestinal epithelial barrier (IEB), bowel inflammation and neuroplastic rearrangements of the enteric nervous system (ENS) could be involved in the pathophysiology of the intestinal disturbances in PD. In this context, although several review articles have pooled together evidence on the alterations of enteric bacteria-neuro-immune network in PD, a revision of the literature on the specific morphological changes occurring in the intestinal mucosal barrier, the ENS and enteric muscular layers in PD, is lacking. The present review provides a complete appraisal of the available knowledge on the morphological alterations of intestinal mucosal barrier, with particular focus on IEB, ENS and enteric muscular layers in PD. In particular, our intent was to critically discuss whether, based on evidence from translational studies and pre-clinical models, morphological changes in the intestinal barrier and enteric neuromuscular compartment contribute to the pathophysiology of intestinal dysfunctions occurring in PD.

NLRP3 at the crossroads between immune/inflammatory responses and enteric neuroplastic remodelling in a mouse model of diet-induced obesity.

BACKGROUND AND PURPOSE: Enteric neurogenic/inflammation contributes to bowel dysmotility in obesity. We examined the role of NLRP3 in colonic neuromuscular dysfunctions in mice with high-fat diet (HFD)-induced obesity. EXPERIMENTAL APPROACH: Wild-type C57BL/6J and NLRP3-KO (Nlrp3-/- ) mice were fed with HFD or standard diet for 8 weeks. The activation of inflammasome pathways in colonic tissues from obese mice was assessed. The role of NLRP3 in in vivo colonic transit and in vitro tachykininergic contractions and substance P distribution was evaluated. The effect of substance P on NLRP3 signalling was tested in cultured cells. KEY RESULTS: HFD mice displayed increased body and epididymal fat weight, cholesterol levels, plasma resistin levels and plasma and colonic IL-1ß levels, colonic inflammasome adaptor protein apoptosis-associated speck-like protein containing caspase-recruitment domain (ASC) and caspase-1 mRNA expression and ASC immunopositivity in macrophages. Colonic tachykininergic contractions were enhanced in HFD mice. HFD NLRP3-/- mice developed lower increase in body and epididymal fat weight, cholesterol levels, systemic and bowel inflammation. In HFD Nlrp3-/- mice, the functional alterations of tachykinergic pathways and faecal output were normalized. In THP-1 cells, substance P promoted IL-1ß release. This effect was inhibited upon incubation with caspase-1 inhibitor or NK1 antagonist and not observed in ASC-/- cells. CONCLUSION AND IMPLICATIONS: In obesity, NLRP3 regulates an interplay between the shaping of enteric immune/inflammatory responses and the activation of substance P/NK1 pathways underlying the onset of colonic dysmotility. Identifying NLRP3 as a therapeutic target for the treatment of bowel symptoms related to obesity.