Irritable Bowel Syndrome Model Research Articles

Activation of the High-Affinity Choline Transporter 1 in the Spinal Cord Relieves Stress-Induced Hyperalgesia.

The mechanism underlying irritable bowel syndrome (IBS), a common disease with hyperalgesia, remains elusive. The spinal cholinergic system is involved in pain modulation, but its role in IBS is unknown. To determine whether high-affinity choline transporter 1 (CHT1, a major determinant of the cholinergic signaling capacity), is implicated in spinal modulation of stress-induced hyperalgesia. A rat IBS model was established by water avoidance stress (WAS). Visceral sensations were detected by abdominal withdrawal reflex (AWR) and visceromotor response (VMR) to colorectal distension (CRD). Abdominal mechanical sensitivity was determined by von Frey filaments (VFFs) test. RT-PCR, Western blot, and immunostaining were performed for spinal CHT1 expression. Spinal acetylcholine (ACh) was measured by ELISA; the influence of spinal CHT1 on hyperalgesia were evaluated by intrathecal administration of MKC-231 (a choline uptake enhancer) and hemicholinium-3 (HC-3, a specific inhibitor of CHT1). Minocycline treatment was used to explore the role of spinal microglia in hyperalgesia. After 10days of WAS, AWR scores and VMR magnitude to CRD, and the number of withdrawal events in VFF test were increased. Double-labeling showed that CHT1 in the dorsal horn was expressed in most of the neurons and almost all the microglia. The CHT1 expression and ACh levels in the spinal cord and the density of CHT1-positive cell in the spinal dorsal horn were enhanced in WAS-exposed rats. HC-3 enhanced pain responses in WAS rats; MKC-231 alleviated pain in WAS rats by upregulating CHT1 expression and increasing ACh production in the spinal cord. Furthermore, microglial activation in the spinal dorsal horn promoted the stress-induced hyperalgesia, and MKC-231 achieved analgesic effects byinhibiting the spinal microglial activation. CHT1 exerts antinociceptive effects in spinal modulation of chronic stress-induced hyperalgesia by increasing ACh synthesis and suppressing microglial activation. MKC-231 has potential for treating disorders accompanied by hyperalgesia.

Effect of moxibustion on immune function homeostasis in rats with diarrhea irritable bowel syndrome based on SCF/c-kit signaling pathway

To observe the effects of moxibustion on the stem cell factor (SCF)/tyrosine kinase receptor (c-kit) signaling pathway and immune function in rats with diarrhea irritable bowel syndrome (IBS-D), and to explore the mechanism of moxibustion for IBS-D. Among 52 young rats born from 6 healthy pregnant SPF rats, 12 rats were randomly selected into the normal group, and the remaining 40 rats were treated with the three-factor combination method of maternal separation, acetic acid enema and chronic restraint stress to establish the IBS-D rat model. Thirty-six rats with successful IBS-D model were randomly divided into a model group, a moxibustion group, and a medication group, 12 rats in each group. The rats in the moxibustion group were treated with suspension moxibustion at "Tianshu" (ST 25) and "Shangjuxu" (ST 37); the rats in the medication group were treated with intragastric administration of rifaximin suspension (150 mg/kg). All the treatments were given once a day for 7 consecutive days. The body mass, loose stool rate (LSR), the minimum volume threshold when abdominal withdrawal reflex (AWR) scored 3 were measured before acetic acid enema (35 days old), after modeling (45 days old), and after intervention (53 days old). After intervention (53 days old), HE staining was used to observe the morphology of colon tissue, and spleen and thymus coefficients were measured; ELISA method was used to detect serum inflammatory factors (tumor necrosis factor a [TNF-a], interleukin [IL]-10, IL-8), T-lymphocyte subsets (CD+4, CD+8, CD+45), value of CD+4/CD+8 and immune globulin (IgA, IgG, IgM); real-time PCR method and Western blot method was used to detect the expression of SCF, c-kit mRNA and protein in colon tissue; immunofluorescence staining method were used to detect positive expression of SCF and c-kit. After intervention, compared with the normal group, in the model group, the body mass and the minimum volume threshold when AWR scored 3 were decreased (P<0.01), LSR, spleen and thymus coefficients, serum levels of TNF-α, IL-8, CD+4, CD+45, CD+4/CD+8, IgA, IgG, IgM were increased (P<0.01), serum IL-10 level and protein and mRNA expression of SCF and c-kit in colon tissue were decreased (P<0.01), and the positive expression of SCF and c-kit was decreased (P<0.01). Compared with the model group, in the moxibustion group and the medication group, the body mass and the minimum volume threshold when AWR scored 3 were increased (P<0.01, P<0.05), LSR, spleen and thymus coefficients, serum levels of TNF-α, IL-8, CD+4, CD+8, CD+45, CD+4/CD+8, IgA, IgG, IgM were decreased (P<0.01, P<0.05), serum IL-10 level and protein and mRNA expression of SCF and c-kit in colon tissue were increased (P<0.01), and the positive expression of SCF and c-kit was increased (P<0.01). Compared with the medication group, in the moxibustion group, the level of serum CD+4 was decreased (P<0.05), the value of CD+4/CD+8 was increased (P<0.01), and there was no significant difference in other indexes (P>0.05). The expression of SCF and c-kit mRNA was positively correlated with the minimum volume threshold when AWR scored 3 and IL-10 (P<0.01), and negatively correlated with remaining indexes (P<0.01, P<0.05). Moxibustion could reduce visceral hypersensitivity, improve symptoms of abdominal pain and diarrhea in IBS-D rats, and its mechanism may be related to up-regulation of the expression of SCF/c-kit signaling pathway and improvement of IBS-D immune function.

Evaluation of two laboratory model methods for diarrheal irritable bowel syndrome

BackgroundDiarrheal irritable bowel syndrome (IBS-D) is a common chronic functional gastrointestinal disorder, and the underlying pathogenic mechanism is still unclear. Animal models that mimic the pathological state of IBS-D patients were constructed to provide a reference for later drug research and model development.MethodsThe IBS-D model was induced using restraint stress and chemical stimulation (rhubarb), and rats were divided into normal control group (NC), chemically stimulated group (CS), and restraint stress group (RS). Visceral motility responses to Colorectal Balloon Dilation (CRD) were measured by Abdominal Withdrawal Reflex (AWR); evaluation of faecal properties and water content; determination of colonic tissue tight junction (TJ) mRNA expression by RT-PCR; measurement of inflammatory cytokines by ELISA; and intestinal flora and short chain fatty acids.ResultsCompared to NC group, CS and RS group rats showed increased intestinal sensitivity and Bristol stool score, significant diarrheal symptoms and weight loss. Mucin 2, ZO-1, OCLN, CLDN4 mRNA expression was reduced and the intestinal mucosal barrier function was diminished. In addition, the levels of inflammatory factors IL-1β, IL-6, IL-8, IL-10 and TNF-α increased, the abundance and diversity of intestinal flora decreased, the content of beneficial bacteria such as Bifidobacteria decreased, and SCFAs such as acetic acid, propionic acid and butyric acid decreased to different degrees. Although, no significant difference was observed for any molecular and inflammatory marker, but compared to CS group, RS group had less water in the stool, higher visceral sensitivity, and higher relative abundance of beneficial intestinal bacteria such as Actinobacteria.ConclusionIn conclusion, restraint stress combined with chemical stimulation can mimic the pathological state of diarrhoea symptoms, visceral hypersensitivity, reduced intestinal mucosal barrier permeability, immune regulatory dysfunction and dysbiosis in IBS-D patients. However, herbs with antibacterial effects such as rhubarb and senna, for example, are not suitable as the first choice for chemical stimulation, as they may lead to a decrease in harmful bacteria and an increase in beneficial bacteria in the intestinal fraction and do not perfectly mimic the imbalanced state of intestinal flora in IBS-D patients, while restraint stress may be a key factor in modelling.

Exploring the mechanism of Xingpi Capsule in diarrhea predominant-irritable bowel syndrome treatment based on multiomics technology

BackgroundXingpi Capsule (XP), a commercially available over-the-counter herbal medicine in China, plays a prominent role in treating diarrhea-predominant irritable bowel syndrome (IBS-D). Nevertheless, the potential mechanisms remain unclear. PurposeThis study aimed to investigate XP efficacy in IBS-D and elucidate the underlying molecular mechanisms. MethodsA rat IBS-D model was established by senna decoction gavage combined with restraint stress and swimming exhaustion. The changes in rat body weight and stool were recorded daily. Colon pathological changes and the number of colonic goblet cells of rats were observed by hematoxylin-eosin (HE) staining and Alcian blue plus periodic acid-Schiff (AB-PAS) staining, respectively. The expression of Occludin, a tight-junction-associated protein, was examined via immunohistochemistry. Images of colonic microvilli were obtained by TEM. Western blotting (WB) was used to analyze the protein expression of the ASK1/P38 MAPK pathway. The composition of the rat intestinal microbiota was detected by 16S rRNA sequencing. Changes in colonic metabolites were evaluated by liquid chromatography-mass spectrometry (LC-MS). Changes in colon RNA expression were assessed by RNA sequencing (RNA-Seq). The nontoxic range of hypoxanthine (HPX) was screened by Cell Counting Kit-8 (CCK8), the cell model of human colonic epithelial cells (NCM460) induced by lipopolysaccharide (LPS) was established, and the effective concentration of HPX was screened by CCK8. After transfection of pcDNA3.1-MAP3K5, Hoechst 33,342 staining, flow cytometry to detect cell apoptosis, and immunofluorescence to detect the fluorescence changes of ASK1 and ZO-1. WB detection of ASK1/P38 MAPK pathway protein expression changes. ResultsXP increased the body weight of IBS-D patients and reduced the loose stool rate, loose stool index, and Bristo score. In addition, XP mitigated colon lesions, increased the number of goblet cells and the expression of Occludin, and prevented severe distortion and effacement of the microvillous structure. Specifically, 16S rRNA gene sequence analysis showed that XP decreased the abundance of Desulfurium and Prevotella 9 at the phylum and genus levels while increasing the abundance of Bacteroides at the genus level. RNA-Seq combined with WB validation showed that XP exerted antidiarrheal effects by inhibiting the ASK1/P38 MAPK signaling pathway. Additionally, XP also increased the relative expression level of the metabolite HPX, as revealed by untargeted metabolomics analysis. Impressively, the correlation analysis between 16S rRNA sequencing and LC-MS suggested that HPX and Prevotella 9 are negatively correlated, which indicated that XP might increase the content of HPX by reducing the abundance of Prevotella 9. Meanwhile, a negative correlation between HPX and ASK1 was indicated through RNA-Seq and LC-MS, which suggested that the inhibition of ASK1 (Map3k5) may be ascribed to the increase in HPX after XP treatment. In vitro experiments have proven that HPX can alleviate LPS-induced NCM460 damage, specifically manifested as enhancing cell viability, reducing cell apoptosis, increasing ZO-1 expression, reducing the fluorescence intensity of MAP3K5 in the model group, and inhibiting the expression of ASK1/P38 MAPK pathway proteins. The protective effect of HPX was reversed after transfection with pcDNA 3.1-MAP3K5, which fully demonstrated that the protective mechanism of HPX was achieved by inhibiting MAP3K5 and its downstream pathways. ConclusionXP displayed multifaceted protection against IBS-D in rats by regulating the intestinal microbiota, increasing the relative expression level of HPX, a metabolite of the microbiota, and inhibiting the ASK1/P38 MAPK signaling pathway.

Effects of PAR2 Gene Knockout on Visceral Sensitivity, Stress Behaviors, and Colonic Electrical Activities in Irritable Bowel Syndrome.

Irritable bowel syndrome (IBS) could seriously affect the patient's health quality by its recurrence. There is a great medical and social need to explore the mechanisms and new treatment strategies of IBS. To explore the influence of protease-activated receptor 2 (PAR2) gene knockout on IBS visceral sensitivity, stress behaviors, and colonic electrical activities. The PAR2 gene knockout and IBS model rats were generated and divided into PAR2+/+ wild control (WC) group, PAR2+/+ wild IBS model (WM) group, PAR2-/- knockout control (KC) group and PAR2-/- knockout IBS model (KM) group. The stress behaviors scores, minimum rectal fluid injection capacity threshold value of abdominal withdrawal reflex (AWR) = 3, and the colonic electrical activities indexes were recorded and the experimental results were analyzed statistically. (1) PAR2 gene deletion and IBS models were successfully generated. (2) The scores of aggressive and exploratory behaviors in WM and KM groups were higher than WC and KC groups (p < 0.05). The grooming behavior scores in WC and KC groups were higher than the WM and KM groups (p < 0.05). WM group had the highest aggressive and exploratory behavior scores; WC group had the highest grooming behavior scores. (3) The minimum rectal fluid injection capacity threshold value of AWR = 3 in WM and KM groups were lower than WC and KC groups (p < 0.05); WM group had the lowest value. (4) The maximum amplitude and wave frequency of colonic fast and slow waves in WM and WC groups were greater than in the KM and KC groups, respectively (p < 0.05). The amplitude index and number of colonic contraction waves in WM and KM groups were greater than the WC and KC groups (p < 0.05). Colonic electrical activity indexes were highest in the WM group. The PAR2 gene deletion could have a beneficial effect on visceral sensitivity, stress behaviors and colonic electrical activities in rats with IBS.

Vitamin D and lactoferrin attenuate stress-induced colitis in Wistar rats via enhancing AMPK expression with inhibiting mTOR-STAT3 signaling and modulating autophagy.

Irritable bowel syndrome (IBS) is a global gastrointestinal disorder closely related to psychological stress exposure and local colonic inflammation. Herein, we investigated the effect of wrap-restraint stress (WRS) on rat behavior, on adenosine monophosphate-activated protein kinase-mammalian/mechanistic target of rapamycin-signal transducer and activator of transcription 3(AMPK-mTOR-STAT3) signaling, and autophagy in colonic mucosa. The impact of chronic administration of vitamin D3 and lactoferrin was compared.Twenty-four male Wistar rats were randomly divided into four groups. Chronic WRS protocol was applied as a rodent model of IBS. Group I: naïve animals, Group II: WRS animals, Group III: WRS-exposed and treated with vitamin D3 (500 IU/kg/day),and Group IV: WRS-exposed and treated with lactoferrin (300 mg/kg/day). In this study, we found that chronic administration of each of vitamin D3 and lactoferrin resulted in a significant increase in social interaction test, interleukin-10, AMPK, optical density of LC3B, goblet cell count and marked decrease in serum cortisol level, STAT3, inflammatory cell count, and optical density of mTOR in comparison to the WRS rats. Our findings suggest that both vitamin D3 and Lactoferrin could augment colonic autophagy through enhanced AMPK expression and inhibition of mTOR-STAT3 signaling, which offers practical insights into their clinical use in the prevention and therapy of IBS. However, lactoferrin intake as a nutritional supplement could be more helpful for stress-induced colitis treatment than vitamin D3.

Piezo2 channels expressed by colon-innervating TRPV1-lineage neurons mediate visceral mechanical hypersensitivity

Inflammatory and functional gastrointestinal disorders such as irritable bowel syndrome (IBS) and obstructive bowel disorder (OBD) underlie the most prevalent forms of visceral pain. Although visceral pain can be generally provoked by mechanical distension/stretch, the mechanisms that underlie visceral mechanosensitivity in colon-innervating visceral afferents remain elusive. Here, we show that virally mediated ablation of colon-innervating TRPV1-expressing nociceptors markedly reduces colorectal distention (CRD)-evoked visceromotor response (VMR) in mice. Selective ablation of the stretch-activated Piezo2 channels from TRPV1 lineage neurons substantially reduces mechanically evoked visceral afferent action potential firing and CRD-induced VMR under physiological conditions, as well as in mouse models of zymosan-induced IBS and partial colon obstruction (PCO). Collectively, our results demonstrate that mechanosensitive Piezo2 channels expressed by TRPV1-lineage nociceptors powerfully contribute to visceral mechanosensitivity and nociception under physiological conditions and visceral hypersensitivity under pathological conditions in mice, uncovering potential therapeutic targets for the treatment of visceral pain.

Changes in signalling from faecal neuroactive metabolites following dietary modulation of IBS pain

ObjectiveDietary therapies for irritable bowel syndrome (IBS) have received increasing interest but predicting which patients will benefit remains a challenge due to a lack of mechanistic insight. We recently found...

Effect of electroacupuncture on visceral sensitivity and colonic NGF, TrkA, TRPV1 expression in IBS-D rats

To observe the effect of electroacupuncture (EA) at "Tianshu" (ST 25) and "Shangjuxu" (ST 37) on mental state, visceral sensitivity and protein expression of nerve growth factor (NGF), tyrosine kinase receptor A (TrkA) and transient receptor potential vanilloid 1 (TRPV1) of colonic tissue in diarrhea-predominant irritable bowel syndrome (IBS-D) rats, and to explore its possible mechanism on treating IBS-D. A total of 36 male SD rats of SPF grade were randomized into a blank group, a model group, an EA group and a western medication group, 9 rats in each group. In the model group, the EA group and the western medication group, IBS-D model was established by enema of dinitrobenzene sulfonic acid (DNBS) combined with chronic restraint stress method. In the EA group, EA was applied at "Tianshu" (ST 25) and "Shangjuxu" (ST 37), with disperse-dense wave, in frequency of 2 Hz/100 Hz, 20 min each time, once a day for 7 days. In the western medication group, pinaverium bromide suspension was given by gavage (15 mg•kg-1•d-1) for 7 days. Before and after model establishment, and after intervention, the body mass, 24 h food intake and fecal water content were observed, the visceral sensitivity was detected by abdominal withdrawal reflex (AWR); after intervention, the mental state was evaluated by elevated plus maze (EPM) test, the protein expression of NGF, TrkA and TRPV1 was detected by immunohistochemistry and Western blot in the 4 groups. After model establishment, compared with the blank group, the body mass and 24 h food intake were decreased (P<0.05), first systolic latency of AWR was shortened and number of contraction wave of AWR was increased (P<0.05), and fecal water content was increased (P<0.05) in the model group, the EA group and the western medication group. After intervention, compared with the blank group, open arm residence time ratio (OT%) of EPM was decreased (P<0.05) and protein expression of NGF, TrkA, TRPV1 in colonic tissue was increased in the model group (P<0.05); compared with the model group, the body mass and 24 h food intake were increased (P<0.05), first systolic latency of AWR was lengthened and number of contraction wave of AWR was decreased (P<0.05), the fecal water content was decreased (P<0.05), OT% of EPM was increased (P<0.05), and protein expression of NGF, TrkA, TRPV1 in colonic tissue was decreased (P<0.05) in the EA group and the western medication group. Electroacupuncture at "Tianshu" (ST 25) and "Shangjuxu" (ST 37) can relieve the anxiety and depression-like behaviors in IBS-D rats, down-regulate the protein expression of NGF, TrkA, TRPV1 in colonic tissue, so as to reduce the visceral sensitivity and relieve symptoms.

YINDARA-4 relieves visceral hypersensitivity in irritable bowel syndrome rats via regulation of gut microbiota and serotonin levels

Objective: The present study aims to evaluate the in vivo efficacy of YINDARA-4 in improving the symptoms of irritable bowel syndrome (IBS) in a rat model and investigate the impact of YINDARA-4 on potential targets of IBS management, such as the serotonin level in intestinal tissues and the structure and composition of the gut microbiota. Methods: We developed an IBS rat model by combining stress from maternal separation, acetic acid administration, and restraint. We administered YINDARA-4 water extract to the IBS rat model for 10 consecutive days. The fecal water content, visceral sensitivity, gut microbiota, and serotonin levels in the colonic tissue were then analyzed and compared between the control group, IBS model group, and YINDARA-4–treated groups. Results: Treatment with YINDARA-4 reversed visceral hypersensitivity in a dose-dependent manner in the experimental rat model of IBS. The relief of visceral hypersensitivity upon treatment with YINDARA-4 involved regulation of the gut microbiota structure and composition, and normalization of elevated serotonin levels in the colon. The decrease in colonic serotonin levels with YINDARA-4 treatment might be associated with a reduction in the abundance of Helicobacter and enrichment of Butyricimonas. Conclusions: Treatment with YINDARA-4 was beneficial against visceral hypersensitivity in a rat model of IBS. The improved symptoms exhibited in IBS rats were associated with favorably altered gut microbiota and normalization of serotonin levels in the colon.

Modulation of colonic function in irritable bowel syndrome rats by electroacupuncture at ST25 and the neurobiological links between ST25 and the colon.

Irritable bowel syndrome (IBS) is a chronic functional gastrointestinal disease characterized by abdominal pain and defecation disorders. Acupuncture therapy positively affects IBS, with ST25 being the main point. However, ST25 has mostly been used in conjunction with other acupoints. This study aimed to observe the therapeutic effect of electroacupuncture at ST25 alone in IBS and the neurobiological mechanism of ST25 associated with the colon. First, we observed the effect of electroacupuncture at ST25 on the visceral pain threshold and slow-wave discharge of the colon in IBS model rats. Second, we explored the neurobiological mechanism of ST25 associated with the colon using a neural tracer technique. The results showed that (1) electroacupuncture at ST25 alone can alleviate visceral hypersensitivity and restore normal slow-wave frequency and rhythm of the colon in IBS rats; (2) there is a close neuroanatomical connection between ST25 and the colon, i.e., in the dorsal root ganglion (DRG), ST25 is similar in innervation to the colon, mainly in the T8-L1 segment, while the presence of double-labeled positive neurons is present in a part of the DRG; retrogradely labeled motor neurons associated with ST25 were observed in the anterior horn of the spinal cord, and retrogradely labeled sympathetic postganglionic neurons associated with ST25 were observed in the sympathetic nerve chain. These findings suggested that the DRGs and the dorsal horn of the spinal cord are important targets for electroacupuncture at ST25 to reduce visceral hypersensitivity in IBS rats. The sympathetic ganglia may be an important site for ST25 to regulate intestinal motility. The neurobiological mechanism of ST25 action in IBS rats should be further investigated in the future by combining related techniques, such as pseudorabies virus, optogenetics, calcium imaging, and electrophysiology.

Metagenomics and metabolomics analysis to investigate the effect of Shugan decoction on intestinal microbiota in irritable bowel syndrome rats.

The effect of Shugan Decoction (SGD) on intestinal motility and visceral hypersensitivity in Water avoid stress (WAS)-induced diarrhea predominant irritable bowel syndrome (IBS-D) model rats has been confirmed. However, the mechanisms of its action involved in the treatment of IBS-D need to be further studied. Intestinal microbiota plays an important role in maintaining intestinal homeostasis and normal physiological function. Changes in the intestinal microbiota and its metabolites are thought to participate in the pathophysiological process of IBS. This study aimed to analyze the influence of SGD on intestinal microbiota and fecal metabolites in IBS-D rats by multiple omics techniques, including metagenomic sequencing and metabolomics. We measured the intestinal motility and visceral sensitivity of three groups of rats by fecal pellets output and colorectal distension (CRD) experiment. In addition, metagenome sequencing analysis was performed to explore the changes in the number and types of intestinal microbiota in IBS-D model rats after SGD treatment. Finally, we also used untargeted metabolomic sequencing to screen the metabolites and metabolic pathways closely related to the therapeutic effect of SGD. We found that compared with the rats in the control group, the fecal pellets output of the rats in the WAS group increased and the visceral sensitivity threshold was decreased (P < 0.05). Compared with the rats in the WAS group, the fecal pellets output of the SGD group was significantly decreased, and the visceral sensitivity threshold increased (P < 0.05). Besides, compared with the rats in the WAS group, the relative abundance of Bacteroidetes increased in SGD group, while that of Firmicutes decreased at the phylum level, and at the species level, the relative abundance of Bacteroides sp. CAG:714, Lactobacillus reuteri and Bacteroides Barnesiae in SGD group increased, but that of bacterium D42-87 decreased. In addition, compared with the WAS group, several metabolic pathways were significantly changed in SGD group, including Taurine and hypotaurine metabolism, Purine metabolism, Sulfur metabolism, ABC transporters, Arginine and proline metabolism and Bile secretion. SGD can regulate specific intestinal microbiota and some metabolic pathways, which may explain its effect of alleviating visceral hypersensitivity and abnormal intestinal motility in WAS-induced IBS-D rats.

Serpin-positive Bifidobacterium breve CNCM I-5644 improves intestinal permeability in two models of irritable bowel syndrome

Probiotic supplementation can help to mitigate the pathogenesis of irritable bowel syndrome (IBS) by reinforcing the intestinal barrier, and reducing both inflammation and proteolytic activity. Here, a combination of in vitro tests was performed on 33 Bifidobacterium strains as probiotic candidates for IBS. In addition to the classical tests performed, the detection of the serine protease inhibitor (serpin) enzyme capable of decreasing the high proteolytic activity found in IBS patients was included. Three serpin-positive strains were selected: Bifidobacterium breve CNCM I-5644, Bifidobacterium longum subsp. infantis CNCM I-5645 and B. longum CNCM I-5646 for their immunomodulation properties and protection of intestinal epithelial integrity in vitro. Furthermore, we found that B. breve CNCM I-5644 strain prevented intestinal hyperpermeability by upregulating Cingulin and Tight Junction Protein 1 mRNA levels and reducing pro-inflammatory markers. The ability of CNCM I-5644 strain to restore intestinal hyperpermeability (FITC-dextran) was shown in the murine model of low-grade inflammation induced by dinitrobenzene sulfonic acid (DNBS). This effect of this strain was corroborated in a second model of IBS, the neonatal maternal separation model in mice. Altogether, these data suggest that serpin-positive B. breve CNCM I-5644 may partially prevent disorders associated with increased barrier permeability such as IBS.

Dysregulated cystathionine-β-synthase/hydrogen sulfide signaling promotes chronic stress-induced colonic hypermotility in rats.

Hydrogen sulfide (H2 S), an important endogenous gasotransmitter, is involved in the modulation of gastrointestinal motility, but whether it mediates the intestinal dysmotility in irritable bowel syndrome (IBS) is not known. This study explored the significance of cystathionine-β-synthase (CBS)/H2 S signaling in stress-induced colonic dysmotility. A rat model of IBS was established using chronic water avoidance stress (WAS). Colonic pathological alterations were detected histologically. Intestinal motility was determined by intestinal transit time (ITT) and fecal water content (FWC). Visceral sensitivity was assessed using the visceromotor response (VMR) to colorectal distension (CRD). Real-time PCR, Western blotting, and immunostaining were performed to identify the expression of CBS in the colon. The contractions of distal colon were studied in an organ bath system and H2 S content was measured by ELISA. The effects of SAM, a selective CBS activator, on colonic dysmotility were examined. MEK1 was tested as a potential upstream effector of CBS/H2 S loss. After 10 days of WAS, the ITT was decreased and FWC was increased, and the VMR magnitude in response to CRD was enhanced. The colonic CBS expression and H2 S levels were significantly declined in WAS-exposed rats, and the density of CBS-positive enteric neurons in the myenteric plexus in WAS-treated rats was lower than that in controls. SAM treatment relieved WAS-induced colonic hypermotility via increased H2 S production. AZD6244, a selective inhibitor of MEK1, partially reversed CBS downregulation and colonic hypermotility in WAS-treated rats. Decreased CBS/H2 S signaling through increased MEK1 signaling might be important in the pathogenesis of chronic stress-induced colonic hypermotility. SAM could be administered for disorders associated with intestinal hypermotility.

Repeated psychological stress, chronic vicarious social defeat stress, evokes irritable bowel syndrome-like symptoms in mice.

Increasing evidence has demonstrated that emotional states and intestinal conditions are inter-connected in so-called “brain–gut interactions.” Indeed, many psychiatric disorders are accompanied by gastrointestinal symptoms, such as the irritable bowel syndrome (IBS). However, the functional connection remains elusive, partly because there are few useful experimental animal models. Here, we focused on a highly validated animal model of stress-induced psychiatric disorders, such as depression, known as the chronic vicarious social defeat stress (cVSDS) model mice, which we prepared using exposure to repeated psychological stress, thereafter examining their intestinal conditions. In the charcoal meal test and the capsaicin-induced hyperalgesia test, cVSDS model mice showed a significantly higher intestinal transit ratio and increased visceral pain-related behaviors, respectively. These changes persisted over one month after the stress session. On the other hand, the pathological evaluations of the histological and inflammatory scores of naive and cVSDS model mice did not differ. Furthermore, keishikashakuyakuto—a kampo medicine clinically used for the treatment of IBS—normalized the intestinal motility change in cVSDS model mice. Our results indicate that cVSDS model mice present IBS-like symptoms such as chronic intestinal peristaltic changes and abdominal hyperalgesia without organic lesion. We therefore propose the cVSDS paradigm as a novel animal model of IBS with wide validity, elucidating the correlation between depressive states and intestinal abnormalities.

Electroacupuncture at Tianshu (ST25) and Zusanli (ST36) alleviates stress-induced irritable bowel syndrome in mice by modulating gut microbiota and corticotropin-releasing factor.

To investigate whether electroacupuncture (EA) at bilateral Tianshu (ST25) and Zusanli (ST36) acupoints could alleviate stress-induced irritable bowel syndrome (IBS) and evaluate its effect on gut microbiota and corticotropin-releasing factor (CRF). Thirty C57BL/6 mice were randomly divided into the normal, water avoidance stress (WAS), and WAS+EA groups (10 mice per group). An experimental model of IBS was established by exposing the animals to WAS. The mice were treated with EA at the bilateral Tianshu (ST25) and Zusanli (ST36) acupoints. The abdominal withdrawal reflex test was conducted to evaluate visceral sensitivity in IBS. Gut microbiota was analyzed using 16S rRNA sequencing and analysis. The expression of CRF was determined using immune-ofluorescence and quantitative real-time polymerase chain reaction. EA alleviated visceral hypersensitivity in a mouse model of WAS-induced IBS. It modulated the dysbiosis of gut microbiota induced by WAS. Moreover, it suppressed the WAS-induced overexpression of CRF in colon tissues. The findings of this study suggest that EA alleviated WAS-induced IBS mechanisms possibly involving the modulation of the dysbiosis of gut microbiota and suppression of CRF expression.

Abdominal Massage Improves the Symptoms of Irritable Bowel Syndrome by Regulating Mast Cells via the Trypase-PAR2-PKCε Pathway in Rats.

Background Irritable bowel syndrome (IBS) is a clinical disease mainly characterized as a syndrome of abdominal pain and discomfort, which frequently occurs in humans aged 20–50. Abdomen massage is of great medical significance for the health of the human body, including promoting intestinal peristalsis, relieving constipation, and facilitating weight loss. However, its potential benefits in alleviating IBS and the underlying mechanisms remain elusive. Methods In this study, we established an IBS model in rats to evaluate the effects of abdomen massage. Forty male Sprague Dawley (SD) rats were randomly assigned into 4 groups: the normal (control) group, IBS group, abdominal massage group, and abdominal massage + ketotifen treatment group (n = 10 rats in each group). Abdominal massage was performed once a day for 5 minutes for 14 days. On day 14, the rats were euthanized and the tissues were analyzed by transmission electron microscopy (TEM), immunohistochemistry or immunofluorescence staining, and laser confocal focus to visualize the micromorphology of the intestinal mucosa. The expression of TRPV1 and the release of trypase were determined by RT-qPCR and western blot. Results We found that compared with the control group, the mast cells in the IBS group were significantly increased and the increased MC was partially decreased by an abdominal massage with or without ketotifen treatment. We also found that TRPV1 was upregulated in the IBS group. Abdominal massage with or without ketotifen treatment could attenuate the upregulation of TRPV1 in IBS. Mechanically, results of IHC and western Blot suggested that abdominal massage reduces the sensitivity of IBS by regulating the trypase-PAR2-PKCε pathway. Conclusion Overall, our results suggested that abdominal massage produces a beneficial effect in improving the symptoms of IBS through reducing mast cell recruitment and attenuating the trypase-PAR2-PKCε pathway. Ketotifen could promote the effect of abdominal massage on IBS treatment, which can serve as a potential therapeutic strategy for IBS.

Symptoms and health experience in irritable bowel syndrome with focus on men.

Irritable bowel syndrome (IBS) is a disorder with a predominance in women; IBS in men is less studied. The present study evaluated symptoms as well as health and social experiences of men with IBS. This cross-sectional study included 293 patients with IBS (64 men) and 363 non-IBS controls (62 men). Gastrointestinal symptom diaries were filled in prospectively, and data on comorbidities and healthcare-seeking behavior were assessed by questionnaires. Men with IBS were compared with men without IBS and women with IBS. Compared with women with IBS, men with IBS had fewer contacts with the healthcare system, fewer psychiatric comorbidities, fewer sleeping problems, and less chronic pain. Urgency to defecate and nausea were less common, and stool frequency was higher in men with IBS. There was no difference between men with and without IBS in terms of educational level, satisfaction with household economy, or living with a partner. In contrast, women with IBS more often lived alone, were more often dissatisfied with household economy, and had a lower educational level than women without IBS. Men with IBS had the same proportion of full-time employment as men without IBS but in contrast, the proportion of women with IBS in full-time employment was only 34%, compared to 50% of the women without IBS. The present study improves the understanding of men's experiences of IBS and suggests that sex and gender may be integrated into the biopsychosocial model of IBS.

Ameliorating effects of electroacupuncture on the low‐grade intestinal inflammation in rat model of diarrhea‐predominant irritable bowel syndrome

We aim to investigate the effects and mechanisms of electroacupuncture (EA) at ST25 and ST37 on the intestinal low-grade inflammation (LGI) in rat model of Diarrhea-predominant irritable bowel syndrome (IBS-D). IBS-D model rats were established by acetic acid enema combined with restraint and tail clamping. Before EA intervention, they were divided into three groups: blank 1 group, blank 2 group, and IBS-D model group. Diarrhea symptoms and visceral pain sensitivity were evaluated. After constructed the model successfully, the remaining IBS-D model group rats were randomly divided into model group and EA group. Local intestinal inflammation (HE staining), changes of intestinal mucosa (occludin protein and microvascular diameter) were evaluated. Differences between two groups were compared using t-test or Mann-Whitney U-test. Differences among more than two groups were compared using one-way ANOVA or Kruskal-Wallis test. After modeling, the results of HE staining in intestinal tract of IBS-D model rats showed LGI. Compared with the model group, 4h fecal moisture content (diarrhea index) and the AWR score were decreased in the EA group. The results of HE in EA group showed that the infiltration of intestinal inflammatory cells were alleviated. Additionally, EA significantly upregulated the expression of occludin protein and partially dilated the intestinal microvascular diameter. Pearson correlation analysis showed that the symptoms of IBS-D rats were correlated with the changes of intestinal mucosa. EA may treat intestinal LGI in IBS-D rats by upregulating the expression of occludin protein and dilating the intestinal microvascular diameter.

Co-existing polysaccharides affect the systemic exposure of major bioactive ingredients in Chang-Kang-Fang, a multi-herb prescription for treatment of irritable bowel syndrome

Ethnopharmacological relevanceChang-Kang-Fang (CKF) is a traditional Chinese herbal formula used for treatment of irritable bowel syndrome (IBS) in China. Decoction is the administration form of CKF in clinical practice. Previously, CKF has been confirmed with activities of releasing pain and reversing disorders of intestinal propulsion. And alkaloids, monoglycosides, chromones were found as the main bioactive components potentially contributing to the efficacy of CKF. Polysaccharide was also a major constituent in CKF. But if and how polysaccharides influence the systemic exposure of bioactive components in CKF is unknown. Aim of the studyIn this study, we aimed to demonstrate the contribution of the co-existed polysaccharides on the systemic exposure of the major bioactive components from CKF in normal and IBS model rats. Materials and methodsAn UPLC-TQ-MS with multiple reaction monitoring (MRM) scan method was developed and validated for quantifying six major small molecular bioactive ingredients of CKF in the plasma samples, including magnoflorine (MAG), berberine (BBR), albiflorin (ALB), paeoniflorin (PAE), 5-O-methylvisamminol (5-OM) and prim-O-glucosylcimifugin (POG). The rats received CKF decoction (CKF) and CKF small molecule portion (knockout of polysaccharides, CKFSM), respectively. IBS model rats were induced by daily bondage and gavage of Sennae Folium decoction (derived from the leaf of Cassia angustifolia Vahl). The effects of the co-existing polysaccharides on the pharmacokinetic parameters of six small molecular bioactive components in normal and IBS model rats were systematically evaluated. The potential gut microbiota involved mechanisms of the effects was validated by broad-spectrum antibiotic (ABX) treatment. ResultsThe selectivity, precision, accuracy, recovery and matrix effect of the established quantification method were all within acceptable limits of biological sample. In normal rats, the co-existing polysaccharides significantly reduced the AUC(0-t) of MAG and PAE compared with CKFSM group. The Cmax and AUC(0-t) of other four compound were not influenced by co-existing polysaccharides. However, in IBS model rats, compared with CKFSM group, the Cmax and AUC(0-t) of the six ingredients significantly increased in CKF group. For CKF + ABX group, the Cmax of six ingredients decreased significantly when compared with CKF group, and the AUC(0-t) of MAG, BBR, ALB, PAE also reduced with significant differences. ConclusionsA reliable and sensitive UPLC-TQ-MS method was successfully developed and validated for evaluating influence of co-existing polysaccharides on pharmacokinetic behavior of six major small molecules components in CKF. The co-existing polysaccharides enhanced the systemic exposure of six bioactive small molecules in CKF under IBS pathological state potentially via gut microbiota involvement.