Animals ; Codonopsis ; Colitis, Ulcerative/drug therapy* ; Colon ; Male ; NF-kappa B ; Rats ; Rats, Wistar ; Saponins/pharmacology* ; Trinitrobenzenesulfonic Acid

OBJECTIVE: To investigate the protective effect of procyanidin B2 (PCB2) on the intestinal barrier and against enteritis in mice with trinitrobenzene sulphonic acid (TNBS)-induced colitis and explore the possible mechanism. METHODS: A mouse model of TNBS-induced colitis was established in male Balb/c mice aged 6-8 weeks. The successfully established mouse models were randomly divided into PCB2 treatment group (=10) and model group (=10) and were treated with daily intragastric administration of PCB2 (100 mg/kg, 0.2 mL) and 0.2 mL normal saline, respectively. After 4 weeks, the disease symptoms, intestinal inflammation, intestinal mucosal cell barrier function and the changes in PI3K/AKT signaling were evaluated using HE staining, immunofluorescence assay and Western blotting. RESULTS: The disease activity index of the mice was significantly lower and the mean body weight was significantly greater in PCB2 group than in the model group in the 3rd and 4th weeks of intervention ( < 0.05). The levels of colonic inflammation and intestinal mucosal inflammatory mediators IL-1β and TNF-α were significantly lower while IL-10 was significantly higher in PCB2 group than in the model group ( < 0.05). Compared with those in the model group, the mice in PCB2 treatment group showed a significantly lower positive rate of bacterial translocation in the mesenteric lymph nodes and a lower thiocyanate-dextran permeability of the intestinal mucosa ( < 0.05). Western blotting showed that PCB2 treatment significantly increased the expressions of claudin-1 and ZO-1 ( < 0.05) and significantly lowered the expression levels of p-PI3K and p-AKT in the intestinal mucosa as compared with those in the model group ( < 0.05). CONCLUSIONS PCB2 suppresses intestinal inflammation and protects intestinal mucosal functions and structural integrity by inhibiting intestinal PI3K/AKT signaling pathway, suggesting the potential of PCB2 as a new drug for Crohn's disease.
Animals ; Biflavonoids ; Catechin ; Colitis ; chemically induced ; Colon ; Enteritis ; Intestinal Mucosa ; Male ; Mice ; Phosphatidylinositol 3-Kinases ; Proanthocyanidins ; Trinitrobenzenesulfonic Acid

BACKGROUND/AIMS: Inflammation-induced alterations in smooth muscle contractility may be due to the effects on smooth muscle itself, neurotransmitters or enteric nerves. In dextran sulfate sodium-induced colitic rat, the delay in colonic transit was caused by decreased activity and production of neuronal nitric oxide synthase (nNOS) in the myenteric plexus of the distal colon. The aim of this study was to investigate the relationship between the delay in colonic transit and the distribution of inducible NOS (iNOS) and nNOS immunoreactive cells in the myenteric plexus of trinitrobenzene sulfonic acid (TNBS)-induced colitic guinea pig. METHODS: Sacrificed and their colonic tissues of forty-five TNBS-induced colitic guinea pigs were used to measure the colonic transit, and analyzed by immunohistochemistry. RESULTS: Colonic transit was delayed significantly at 3, 7 and 14 days after administration of TNBS. In control, nNOS immunoreactivity was present in the mucosa, submucosa, lamina propria, and ganglion cells of the myenteric plexus, while after TNBS treatment, reduced nNOS cells were found. However, the number of nNOS ganglion cells in the myenteric plexus was similar to those seen in controls. After administration of TNBS, iNOS immunoreactivity was increased in the mucosa and submucosa, but the number of iNOS positive ganglion cells in the myenteric plexus was not changed compared to control. CONCLUSIONS: It is suggested that in TNBS-induced guinea pig colitis, delayed colonic transit is not associated with the expression of nNOS nor iNOS in the myenteric plexus.
Animals ; Colitis/chemically induced/enzymology/*physiopathology ; Colon/*enzymology/innervation ; English Abstract ; *Gastrointestinal Transit ; Guinea Pigs ; Male ; Myenteric Plexus/*enzymology ; Nitric-Oxide Synthase/*metabolism ; Trinitrobenzenesulfonic Acid

OBJECTIVETo explore the efficacy of adipose-derived mesenchymal stem cells (ADMSCs) in a murine model of inflammatory bowel disease, and its potential mechanism.

METHODSMurine colitis mouse model of Crohn's disease(CD) was created by trinitrobenzene sulfonic acid(TNBS)-induced colitis. Seventy-five 6-8 weeks female BALB/c mice were randomly divided into 3 groups: control group, TNBS group and ADMSC group. To verify the therapeutic effect of ADMSC, real-time PCR and immunohistochemical staining were performed to measure inflammatory cytokines levels in colon tissues. The 10-day survival statuses were recorded after the infusion of ADMSCs.

RESULTSIntraperitoneal injection of ADMSCs alleviated the clinical and histopathologic severity of intestinal inflammation, and increased survival(60% vs. 30%, P<0.05) in the TNBS-induced mouse model of CD. Compared with TNBS group, proinflammatory cytokines, including TNF-α, IL-12 and VEGF of ADMSC group were significantly reduced, with significant increase of IL-10 expression.

CONCLUSIONADMSCs can effectively repair the injury of colonitis through down-regulation of proinflammatory cytokines TNF-α, IL-12 and VEGF expression, and up-regulation of anti-inflammatory cytokine IL-10 expression, which may be a potential new alternative of cell-based therapy for CD.

Adipocytes ; Animals ; Colitis ; Crohn Disease ; Cytokines ; Disease Models, Animal ; Down-Regulation ; Female ; Inflammatory Bowel Diseases ; Mesenchymal Stromal Cells ; Mice ; Mice, Inbred BALB C ; Trinitrobenzenesulfonic Acid ; Up-Regulation

OBJECTIVETo investigate the role of mast cells and gut hormones and their interactions in TNBS-induced ulcerative colitis.

METHODSRat models of ulcerative colitis were established by a single intracolonic injection of 100 mg/kg TNBS (in 0.3 ml 50% ethanol). At 0, 6, 11, 16, 21 days after TNBS injection, the rats were sacrificed to determine the count of the mast cells. Histamine level in the whole blood, and the levels of histamine, substance P (SP), vasoactive intestinal peptide (VIP), and somatostatin (SS) in the distal colons were measured by fluorimetry or radioimmune assay. Immunofluorescence double staining was used to observe the relationship of the mast cells with SP, VIP, and SS positive nerve fibers.

RESULTSOn day 6 after TNBS injection, obvious ulcers occurred in the distal colon of the rats with significantly increased histamine level in the whole blood (P<0.05) but significantly decreased colonic histamine levels (P<0.05). The histamine levels in the whole blood and distal colon gradually recovered the normal levels. The mast cells significantly increased on day 16 (P<0.05) and maintained the high level till day 21. The distribution of mast cells was altered after TNBS injection, and the cells were found to aggregate in the myenteric region. SP levels in the distal colon significantly increased on day 11 (P<0.05) and maintained the high level till day 21. Immunofluorescence double staining revealed numerous mast cells close to the SP- and VIP-positive nerve fibers at different time points after TNBS injection. VIP positivity and the number of VIP-positive nerve fibers in the myenteric region were markedly increased, but no mast cells were observed in association with SP- and VIP-positive nerve fibers. The distribution of MC was not found to associate with the SS-positive nerve fibers.

CONCLUSIONThe mast cells and histamine released by them, as well as parasecretion of SP and VIP, participate in tissue damage by TNBS-induced colitis. Bidirectional neuroimmunomodulation of the mast cells, SP and VIP have important effect on the development of TNBS-induced colitis.

Animals ; Colitis, Ulcerative ; chemically induced ; metabolism ; pathology ; Disease Models, Animal ; Male ; Mast Cells ; secretion ; Rats ; Rats, Sprague-Dawley ; Substance P ; metabolism ; Trinitrobenzenesulfonic Acid ; toxicity ; Vasoactive Intestinal Peptide ; metabolism

BACKGROUND/AIMS: Saccharomyces boulardii has been reported to be beneficial in the treatment of inflammatory bowel disease. The aim of this work was to evaluate the effect of S. boulardii in a mice model of 2,4,6-trinitrobencene sulfonic acid (TNBS) induced colitis and analyze the expression of genes in S. boulardii treated mice by microarray. METHODS: BALB/c mice received TNBS or TNBS and S. boulardii treatment for 4 days. Microarray was performed on total mRNA form colon, and histologic evaluation was also performed. RESULTS: In mice treated with S. boulardii, the histological appearance and mortality rate were significantly restored compared with rats receiving only TNBS. Among 330 genes which were altered by both S. boulardii and TNBS (>2 folds), 193 genes were down-regulated by S. boulardii in microarray. Most of genes which were down-regulated by S. bouardii were functionally classified as inflammatory and immune response related genes. CONCLUSIONS: S. boulardii may reduce colonic inflammation along with regulation of inflammatory and immune responsive genes in TNBS-induced colitis.
Animals ; Colitis/chemically induced/genetics/*therapy ; Colon/metabolism/pathology ; Gene Expression Profiling ; Mice ; Mice, Inbred BALB C ; Oligonucleotide Array Sequence Analysis ; *Probiotics ; *Saccharomyces ; Trinitrobenzenesulfonic Acid

OBJECTIVES: Liver disease is the most common extra-intestinal manifestation of ulcerative colitis (UC), but the underlying pathogenesis is still not clarified. It is well accepted that the occurrence of UC-related liver disease has close correlation with immune activation, intestinal bacterial liver translocation, inflammatory cytokine storm, and the disturbance of bile acid circulation. The occurrence of UC-related liver disease makes the therapy difficult, therefor study on the pathogenesis of UC-related liver injury is of great significance for its prevention and treatment. Glutathione (GSH) shows multiple physiological activities, such as free radical scavenging, detoxification metabolism and immune defense. The synthesis and the oxidation-reduction all contribute to GSH antioxidant function. It is reported that the deficiency in hepatic GSH antioxidant function participates in multiple liver diseases, but whether it participates in the pathogenesis of UC-related liver injury is still not clear. This study aims to investigate the feature and underlying mechanism of GSH synthesis and oxidation-reduction function during the development of UC, which will provide useful information for the pathogenesis study on UC-related liver injury. METHODS: UC model was induced by 2,4,6-trinitrobenzenesulfonic acid (TNBS)-ethanol solution (5 mg/0.8 mL per rat, 50% ethanol) via intra-colonic administration in rats, and the samples of serum, liver, and colon tissue of rats were collected at the 3rd, 5th, and 7th days post TNBS. The severity degree of colitis was evaluated by measuring the disease activity index, colonic myeloperoxidase activity, and histopathological score, and the degree of liver injury was evaluated by histopathological score and the serum content of alanine aminotransferase. Spearman correlation analysis was also conducted between the degree of colonic lesions and index of hepatic histopathological score as well as serum aspartate aminotransferase level to clarify the correlation between liver injury and colitis. To evaluate the hepatic antioxidant function of GSH in UC rats, hepatic GSH content, enzyme activity of GSH peroxidase (GSH-Px), and GSH reductase (GR) were determined in rats at the 3rd, 5th, and 7th days post TNBS, and the protein expressions of glutamine cysteine ligase (GCL), GSH synthase, GSH-Px, and GR in the liver of UC rats were also examined by Western blotting. RESULTS: Compared with the control, the disease activity index, colonic myeloperoxidase activity, and histopathological score were all significantly increased at the 3rd, 5th, and 7th days post TNBS (all P<0.01), the serum aspartate aminotransferase level and hepatic histopathologic score were also obviously elevated at the 7th day post TNBS (all P<0.05). There was a significant positive correlation between the degree of liver injury and the severity of colonic lesions (P=0.000 1). Moreover, compared with the control, hepatic GSH content and the activity of GSH-Px and GR were all significantly decreased at the 3rd and 5th days post TNBS (P<0.05 or P<0.01), and the protein expressions of GCL, GSH-Px, and GR were all obviously down-regulated at the 3rd, 5th, and 7th days post TNBS (P<0.05 or P<0.01). CONCLUSIONS There is a significant positive correlation between the degree of liver injury and the severity of colonic lesions, and the occurrence of reduced hepatic GSH synthesis and decreased GSH reduction function is obviously earlier than that of the liver injury in UC rats. The reduced hepatic expression of enzymes that responsible for GSH synthesis and reduction may contribute to the deficiency of GSH synthesis and oxidation-reduction function, indicating that the deficiency in GSH antioxidant function may participate in the pathogenesis of UC related liver injury.
Animals ; Antioxidants ; Aspartate Aminotransferases ; Colitis/chemically induced* ; Colitis, Ulcerative/metabolism* ; Colon/pathology* ; Glutathione/biosynthesis* ; Liver/metabolism* ; Peroxidase/metabolism* ; Rats ; Trinitrobenzenesulfonic Acid

OBJECTIVE: To investigate the effects of N-methyl-D-aspartate receptor (NMDAR) in the spinal dorsal horn in visceral hypersensitivity in rats with colonic inflammation. METHODS: Seventy adult male Sprague-Dawley (SD) rats were randomly divided into the experimental group and the control group. Colonic inflammation was induced in the experimental rats by intraluminal administration of trinitrobenzenesulfonic acid (TNBS). Saline was administered intraluminally in the control rats. After 3, 7, 14, and 28 days of administration, abdominal contractions induced by inflation of a balloon colonically inserted were recorded in rats by implanting electrodes in the abdominal striated muscles. Immunohistochemistry method was used to study the expression of NMDAR1 and NMDAR2A/B in lumbarsacral spinal cord after inflammation. RESULTS: Colonic distension evoked a significant increase of abdominal contractions after 3, 7 and 14 days of TNBS administration. After 28 days of TNBS administration, abdominal contractions were still significantly increased in 2 TNBS-treated rats compared with the control rats. After 7 and 14 days of TNBS administration, NMDAR1 and NMDAR2A/B-immunoreactive cells were significantly increased compared with the control group (P <0.05). Twenty-eight days after TNBS administration, the number of NMDAR1-IR and NMDAR2A/B-IR neurons was still significantly increased in 4 TNBS-treated rats compared with the saline-treated rats (P < 0.05). CONCLUSION NMDAR was involved in the transmission of visceral nociceptive stimuli. After the remission of colonic inflammation, increased expression of NMDAR1 and NMDAR2A/B in the spinal dorsal horn may induce persistent neuronal hyperactivity, which results in visceral hypersensitivity.
Animals ; Colitis ; chemically induced ; metabolism ; Male ; Posterior Horn Cells ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, N-Methyl-D-Aspartate ; biosynthesis ; genetics ; Trinitrobenzenesulfonic Acid

OBJECTIVE: To investigate the anti-inflammatory activity of Radix Panacis quinguefolii root extract (RPQE) and its therapeutic effects on inflammatory bowel disease (IBD). METHODS: The 72-hour post-fertilization zebrafish was used to generate the local and systematic inflammation models through tail-amputation and lipopolysaccharide (LPS)-induction (100 µ g/mL), respectively. The Tg(zlyz:EGFP) zebrafish was induced with 75 µ g/mL 2,4,6-trinitrobenzene sulfonic acid (TNBS) for establishing the IBD model. The tail-amputated, LPS-, and TNBS-induced models were subjected to RPQE (ethanol fraction, 10-20 µ g/mL) administration for 12 and 24 h, respectively. Anti-inflammatory activity of RPQE was evaluated by detecting migration and aggregation of leukocytes and expression of inflammation-related genes. Meanwhile, TNBS-induced fish were immersed in 0.2% (W/V) calcein for 1.5 h and RPQE for 12 h before photographing to analyze the intestinal efflux efficiency (IEE). Moreover, the expression of inflammation-related genes in these fish was detected by quantitative polymerase chain reaction. RESULTS: Subject to RPQE administration, the migration and aggregation of leukocytes were significantly alleviated in 3 zebrafish models (P<0.01). Herein, RPQE ameliorated TNBS-induced IBD with respect to a significantly reduced number of leukocytes, improved IEE, and inhibited gene expression of pro-inflammatory factors (P<0.05 or P<0.01). CONCLUSION RPQE exhibited therapeutic effects on IBD by inhibiting inflammation.
Animals ; Zebrafish ; Lipopolysaccharides ; Disease Models, Animal ; Inflammatory Bowel Diseases/metabolism* ; Inflammation/drug therapy* ; Anti-Inflammatory Agents/therapeutic use* ; Trinitrobenzenesulfonic Acid/adverse effects* ; Colitis/drug therapy*

BACKGROUND/AIMS: Bone marrow-derived cells (BMDC) contribute to tissue maintenance under many kinds of pathologic conditions. We carried out a study to see how BMDC play a role in the treatment of experimental murine colitis. METHODS: We divided the animals into 3 groups and treated them with 50% ethanol (control group), 2,4,6-trinitrobenzene sulfinic acid colitis (TNBS group), and TNBS+bone marrow transplant (BMT group). To induce colitis, TNBS (5.0 mg/mouse) dissolved in 50% ethanol was injected into anus weekly for two weeks. Bone marrow transplantations were performed using bone marrow of male transgenic mouse (donor) with green fluoresence protein (GFP) into female wild type mouse (recipient) three weeks before TNBS instillation. All animals were sacrificed, and colons were extracted one week after the last TNBS instillation. We measured microscopic scores of mucosal injury and investigated the GFP expression for bone marrow engraftment. The immunostaining of vimentin and alpha-smooth muscle actin (alpha-SMA) for myofibroblasts was performed. RESULTS: The score of mucosal injury in the TNBS group was much more severe than those in control, and reduced significantly by BMT (p<0.05). GFP-positive cells were almost deposited in pericryptal niche of BMT group but not at all in both control and TNBS group. Most of myofibroblasts stained with both vimentin and SMA also infiltrated into pericryptal niche. But, the number of myofibroblasts stained with vimentin and SMA in both control and TNBS group was smaller than that in BMT group. CONCLUSIONS: BMDC deposited on pericryptal niche might have a significant role in repairing acute experimental murine colitis.
Actins/metabolism ; Acute Disease ; Animals ; *Bone Marrow Transplantation ; Colitis/chemically induced/pathology/*surgery ; Female ; Fibroblasts/cytology ; Intestinal Mucosa/cytology ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Transgenic ; Transplantation, Homologous ; Trinitrobenzenesulfonic Acid/*toxicity ; Vimentin/metabolism