TNBS-treated Animals Research Articles

P-Cymene and Rosmarinic Acid Ameliorate TNBS-Induced Intestinal Inflammation Upkeeping ZO-1 and MUC-2: Role of Antioxidant System and Immunomodulation

p-Cymene (p-C) and rosmarinic acid (RA) are secondary metabolites that are present in medicinal herbs and Mediterranean spices that have promising anti-inflammatory properties. This study aimed to evaluate their intestinal anti-inflammatory activity in the trinitrobenzene sulphonic acid (TNBS)-induced colitis model in rats. p-C and RA (25–200 mg/kg) oral administration reduced the macroscopic lesion score, ulcerative area, intestinal weight/length ratio, and diarrheal index in TNBS-treated animals. Both compounds (200 mg/kg) decreased malondialdehyde (MDA) and myeloperoxidase (MPO), restored glutathione (GSH) levels, and enhanced fluorescence intensity of superoxide dismutase (SOD). They also decreased interleukin (IL)-1β and tumor necrosis factor (TNF)-α, and maintained IL-10 basal levels. Furthermore, they modulated T cell populations (cluster of differentiation (CD)4+, CD8+, or CD3+CD4+CD25+) analyzed from the spleen, mesenteric lymph nodes, and colon samples, and also decreased cyclooxigenase 2 (COX-2), interferon (IFN)-γ, inducible nitric oxide synthase (iNOS), and nuclear transcription factor kappa B subunit p65 (NFκB-p65) mRNA transcription, but only p-C interfered in the suppressor of cytokine signaling 3 (SOCS3) expression in inflamed colons. An increase in gene expression and positive cells immunostained for mucin type 2 (MUC-2) and zonula occludens 1 (ZO-1) was observed. Altogether, these results indicate intestinal anti-inflammatory activity of p-C and RA involving the cytoprotection of the intestinal barrier, maintaining the mucus layer, and preserving communicating junctions, as well as through modulation of the antioxidant and immunomodulatory systems.

Bacteriocin biosynthesis contributes to the anti-inflammatory capacities of probiotic Lactobacillus plantarum.

Plantaricin EF (PlnEF) is a class IIb bacteriocin produced by Lactobacillus plantarum. We compared L. plantarum NCIMB8826 and LM0419, a plnEFI deletion mutant of that strain lacking plnEF and the gene for the cognate immunity protein plnI, in a 2,4,6-trinitrobenzenesulfonic acid (TNBS) induced mouse model of acute inflammatory bowel disease. Mice fed either L. plantarum NCIMB8826 or LM0419 were not protected against TNBS according to either disease activity or histology (Ameho) scores. Mice consuming NCIMB8826 exhibited intermediate (non-significant) levels of colonic tumour necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) that ranged between the TNBS-treated animals and healthy controls. By comparison, TNF-α and IL-6 quantities were elevated in mice given L. plantarum LM0419 and equivalent to mice given TNBS alone. Both strains survived digestive tract transit in equal numbers and did not result in global changes to the bacterial composition in the intestine according to 16S rRNA gene sequencing either prior to or after TNBS administration. Examination of intestinal taxa showed that mice consuming wild-type L. plantarum, but not LM0419 contained lower proportions of Mucispirillum (Deferribacteres phylum) in the faeces prior to TNBS administration and Parabacteroides (Bacteroidetes phylum) in the caecum after disease induction. Parabacteroides also positively correlated with disease activity and histology scores. These findings suggest a role for PlnEFI production by L. plantarum in benefiting digestive tract health.

The Toll-like receptor 4 antagonist TAK-242 protects against chronic pancreatitis in rats.

Chronic pancreatitis is a progressive disease characterized by irreversible morphological changes to the pancreas, typically causing pain and permanent loss of function. It is a poorly understood disease with the pathogenesis remaining unclear. The authors' previous data demonstrated that the inhibition of Toll-like receptor 4 (TLR4) using TLR4 antagonist kinase (TAK)-242 attenuates taurocholate-induced oxidative stress via the regulation of mitochondrial function in the pancreatic acinar cells of mice. In the present study, the effect of TAK-242 on trinitrobenzene sulfonic acid (TNBS)-induced chronic pancreatitis was investigated in rats. The results revealed that TAK-242 attenuated the severity of chronic pancreatic injury, and regulated extracellular matrix secretion and cellular immunity. In addition, TAK-242 treatment significantly decreased cell apoptosis, as evidenced by the reduction in Terminal deoxynucleotidyl transferase dUTP nick end labeling-positive cells in pancreas tissue sections, and also promoted cell proliferation in TNBS-treated animals. Furthermore, the results of the calibrated von Frey filament assay demonstrated that TAK-242 could prevent the pancreatitis-induced referred abdominal hypersensitivity. In summary, TAK-242 exhibits protective effects against TNBS-induced chronic pancreatitis and may be a potential therapeutic strategy for the treatment of patients with chronic pancreatitis.

Encenicline, an 7 Nicotinic Acetylcholine Receptor Partial Agonist, Reduces Immune Cell Infiltration in the Colon and Improves Experimental Colitis in Mice

The α7 pentamer nicotinic acetylcholine receptors (nAChRs) are a target in transduction of anti-inflammatory signals from the central nervous system to the gastrointestinal (GI) tract. The aim of this study was to investigate the anti-inflammatory action of the novel α7 nAChR partial agonist encenicline and to determine the mechanism underlying its activity. Anti-inflammatory activity of encenicline was evaluated using trinitrobenzenesulfonic acid (TNBS)- and dextran sulfate sodium (DSS)-induced models of colitis. Macroscopic score, ulcer score, colon length and thickness, as well as myeloperoxidase (MPO) activity were recorded. Immunohistochemistry (IHC) was used to measure the infiltration of immune cells in the colon. Furthermore, we employed flow cytometry to determine the effect of encenicline on frequencies of FoxP3(+) and interleukin (IL)-17A(+) T cells in the mouse colon. Encenicline attenuated TNBS- and DSS-induced colitis in mice via α7 nAChRs, as indicated by significantly reduced macroscopic parameters and MPO activity. Treatment with encenicline significantly reduced the infiltration of macrophages, neutrophils, and B cells in the colon of TNBS-treated animals, as indicated by IHC. In the TNBS model encenicline reduced the frequency of FoxP3(+) IL-17A(+) T cells in the colon. In the DSS-model treatment encenicline increased the frequency of FoxP3(+) T cells and reduced IL-17A(+) T cells. Stimulation of α7 nAChR with partial agonist encenicline alleviates colitis via alteration of the number and/or activation status of the immune cells in the gut, emphasizing a potential role of α7 nAChRs as a target for anticolitic drugs.

Toward a better understanding of the central consequences of intestinal inflammation.

Inflammatory bowel diseases (IBDs), which include Crohn's disease and ulcerative colitis, are inflammatory diseases of the gastrointestinal tract. Quality of life for IBD patients is negatively affected by associated pain and gastrointestinal dysfunction, but also by serious behavioral symptoms that include depression, anxiety, fatigue, and cognitive dysfunction. Because these behavioral comorbidities are poorly understood, we have investigated them in a rat model of IBD caused by infusion of a hapten (trinitrobenzene sulfonic acid (TNBS)) into the lower colon. TNBS colitis has many similarities to Crohn's disease, and we have found that it is associated with changes in central nervous system function. TNBS-treated animals have lowered seizure thresholds, which resolve following remission, and hippocampal slices from such animals display increased excitability. There are significant changes in excitatory, AMPA receptor-mediated transmission, in part due to increased numbers of AMPA receptors lacking the GluR2 subunit. Long-term potentiation and depression are reduced in colitic animals, and the synaptic alterations are reversed if microglial activation and tumor necrosis factor α synthesis within the brain are blocked.

Protective effect of simvastatin and rosuvastatin on trinitrobenzene sulfonic acid-induced colitis in rats.

Objective:Statins have anti-inflammatory effects that are not directly related to their cholesterol lowering activity. This study was carried out to evaluate the effect of simvastatin or rosuvastatin on the extent of colonic mucosal damage and on the inflammatory response in trinitrobenzene sulfonic acid (TNBS)-induced ulcerative colitis.Materials and Methods:Ulcerative colitis was induced by single intrarectal injection of 120 mg/kg TNBS. Test groups were treated with simvastatin (10 mg/kg, p.o.) or rosuvastatin (10 mg/kg, p.o.). Colonic mucosal inflammation was evaluated clinically, biochemically, and histologically.Result:Disease activity index score in TNBS-treated rats, as determined by weight loss, stool consistency, fecal occult blood, were significantly lowers in simvastatin or rosuvastatin-treated rats than TNBS-treated animals. Simvastatin or rosuvastatin counteracted the reduction in colon length, decreased colon weight, neutrophil accumulation, and tumor necrosis factor-alpha level in TNBS-induced colitis. Simvastatin and rosuvastatin also inhibited the increase in oxidative stress levels after TNBS administration.Conclusions:These results suggest that simvastatin and rosuvastatin significantly ameliorate experimental colitis in rats, and these effects could be explained by their anti-inflammatory and antioxidant activity.

Experimental ileitis alters prostaglandin biosynthesis in mesenteric lymphatic and blood vessels

Prostaglandins are important mediators responsible for many changes that occur during the inflammatory response. Specifically, in inflammatory bowel disease (IBD), prostaglandins are key players in maintenance of blood flow and mucosal defense. In blood vessels, prostaglandins modulate and inhibit transmigration. In lymphatic vessels, on the other hand, prostaglandin E2 (PGE2) and prostacyclin (PGI2) have been shown to potently inhibit lymphatic contractility. Inhibition of lymphatic contractility could impair proper tissue fluid drainage during inflammation, consequently leading to the submucosal oedema observed in IBD. Alterations in production of PGE2 and PGI2 during inflammation could have severe implications on lymphatic and vascular functions within the small intestine. Using the 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced ileitis guinea pig and rat models, we assessed by quantitative PCR changes in mRNA transcript of enzymes and receptors involved in the production and actions of prostaglandins in mesenteric lymphatic and blood vessels as well as in the affected ileum. Furthermore, we also assessed lymphatic tissue levels of PGE2 and PGI2 during inflammation. We observed significant changes in lymphatic mRNA expression of COX-1, COX-2, MPGES-1, PGIS, EP4 and IP and increases in PGE2 and PGI2 in tissues of TNBS-treated animals. Changes in mRNA in blood vessels from TNBS-treated animals included differences in COX-1, COX-2, MPGES-1, PGIS, EP1, EP2 and IP expression. Prostaglandin metabolites are differentially regulated in both lymphatic and blood vessels during intestinal inflammation.

Thrombin Mediates Vagal Apoptosis and Dysfunction in Inflammatory Bowel Disease

BackgroundIn inflammatory bowel disease, autonomic dysfunction contributes to symptoms, morbidity, and health care resource utilization. Efferent vagal neurons, which provide the primary parasympathetic input to the gastrointestinal tract, are housed in the dorsal motor nucleus of the vagus (DMV) in the brainstem. This study seeks to characterize the effects of IBD on DMV neuronal survival and function. MethodsTNBS (picrylsulfonic acid) was administered by enema to induce colitis in rats. Brain sections through the DMV were examined for neuronal apoptosis using TUNEL labeling, and for glial cell activation by immunofluorescence. Prothrombin production was evaluated via quantitative RT-PCR from DMV tissue, as well as by double immunofluorescence in DMV sections. To investigate the effects of thrombin in the DMV, thrombin or thrombin and an antagonist to its receptor were administered into the fourth ventricle via a stereotactically placed cannula. DMV sections were then examined for apoptosis by TUNEL assay. To evaluate the effect of thrombin on DMV neuronal function, we examined calcium signaling in primary DMV neuron cultures following exposure to thrombin and other neurotransmitters. ResultsTNBS colitis is associated with significantly increased rates of DMV neuronal apoptosis, affecting 12.7 % of DMV neurons in animals with colitis, compared to 3.4 % in controls. There was a corresponding increase in DMV neuron activated caspase-3 immunoreactivity (14.8 vs. 2.6 % of DMV neurons). TNBS-treated animals also demonstrated significantly increased DMV astrocyte and microglial immunoreactivity, indicating glial cell activation. DMV prothrombin production was significantly increased in TNBS colitis, with a close anatomic relationship between prothrombin and microglia. Direct DMV exposure to thrombin replicated the apoptosis and activation of caspase-3 seen in TNBS colitis; these effects were prevented by coadministration of the PAR-1 inhibitor FR171113. Cultured DMV neurons exhibited impaired calcium signaling in response to neurotransmitters following exposure to thrombin. Glutamate-induced calcium transients decreased by 59 %, and those triggered by GABA were reduced by 61 %. PAR-1 antagonism prevented these thrombin-induced changes in calcium signaling. ConclusionsIBD is associated with DMV microglial activation and production of prothrombin. Thrombin in the DMV causes vagal neuron apoptosis and decreased sensitivity to neurotransmitters.

Neuron-glial communication mediated by TNF-α and glial activation in dorsal root ganglia in visceral inflammatory hypersensitivity

Communication between neurons and glia in the dorsal root ganglia (DRG) and the central nervous system is critical for nociception. Both glial activation and proinflammatory cytokine induction underlie this communication. We investigated whether satellite glial cell (SGC) and tumor necrosis factor-α (TNF-α) activation in DRG participates in a 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced rat model of visceral hyperalgesia. In TNBS-treated rats, TNF-α expression increased in DRG and was colocalized to SGCs enveloping a given neuron. These SGCs were activated as visualized under electron microscopy: they had more elongated processes projecting into the connective tissue space and more gap junctions. When nerves attached to DRG (L6-S1) were stimulated with a series of electrical stimulations, TNF-α were released from DRG in TNBS-treated animals compared with controls. Using a current clamp, we noted that exogenous TNF-α (2.5 ng/ml) increased DRG neuron activity, and visceral pain behavioral responses were reversed by intrathecal administration of anti-TNF-α (10 μg·kg(-1)·day(-1)). Based on our findings, TNF-α and SGC activation in neuron-glial communication are critical in inflammatory visceral hyperalgesia.

Involvement of the NO-cGMP-KATP channel pathway in the mesenteric lymphatic pump dysfunction observed in the guinea pig model of TNBS-induced ileitis

Mesenteric lymphatic vessels actively transport lymph, immune cells, fat, and other macromolecules from the intestine via a rhythmical contraction-relaxation process called lymphatic pumping. We have previously demonstrated that mesenteric lymphatic pumping was compromised in the guinea pig model of 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced ileitis, corroborating clinical and experimental observations of a dilated and/or obstructed phenotype of these vessels in inflammatory bowel disease. Many mediators released during the inflammatory process have been shown to alter lymphatic contractile activity. Among them, nitric oxide (NO), an inflammatory mediator abundantly released during intestinal inflammation, decreases the frequency of lymphatic contractions through activation of ATP-sensitive potassium (K(ATP)) channels. The objective of this study was to investigate the role of NO and K(ATP) channels in the lymphatic dysfunction observed in the guinea pig model of TNBS-induced ileitis. Using quantitative real-time PCR, we demonstrated that expression of Kir6.1, SUR2B, and inducible NO synthase (iNOS) mRNAs was significantly upregulated in TNBS-treated animals. Pharmacological studies performed on isolated, luminally perfused mesenteric lymphatic vessels showed that the K(ATP) channels blocker glibenclamide, the selective iNOS inhibitor 1400W, and the guanylyl cyclase inhibitor ODQ (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one) significantly improved lymphatic pumping in quiescent lymphatic vessels from TNBS-treated animals. Membrane potential measurement with intracellular microelectrodes revealed that vessels from TNBS-treated animals were hyperpolarized compared with their sham counterpart and that the hyperpolarization was significantly attenuated in the presence of glibenclamide and ODQ. Our findings suggest that NO and K(ATP) play a major role in the lymphatic contractile dysfunction that occurred as a consequence of the intestinal inflammation caused by TNBS.

Colitis is associated with a loss of intestinofugal neurons

Intestinofugal neurons sense and receive information regarding mechanical distension of the bowel and transmit this information to postganglionic sympathetic neurons in the prevertebral ganglia. Previous studies have demonstrated that trinitrobenzene sulfonic acid (TNBS)-induced colitis is associated with a loss of myenteric neurons that occurs within the first 12 h following the inflammatory insult. The purpose of this study was to test the hypothesis that intestinofugal neurons are among the myenteric neurons lost during TNBS-induced colitis. The retrograde tracing dye Fast Blue was used to label intestinofugal neurons, and immunohistochemical staining for the RNA-binding proteins HuC/D was used to count all myenteric neurons. Ongoing synaptic input to neurons in the guinea pig inferior mesenteric ganglion (IMG) was recorded via conventional intracellular electrophysiology. In control preparations, intestinofugal neurons account for 0.25% of myenteric neurons. In the distal colon of TNBS-treated animals, the proportion of intestinofugal neurons was reduced to 0.05% (an 80% reduction) within the region of inflammation where 20-25% of myenteric neurons were lost. Neither intestinofugal neurons specifically nor myenteric neurons were reduced in more proximal uninflamed regions. There is a reduction in the frequency of ongoing synaptic potentials in visceromotor neurons of the IMG at 12 and 24 h and 6 and 56 days after TNBS. Collectively, the results of this study suggest that intestinofugal neurons are among the myenteric neurons lost during inflammation and may be selectively targeted. Because intestinofugal neurons are a major driver of sympathetic output to the gut, the loss of intestinofugal neurons may have a profound pathophysiological significance.

Altered response to hydrogen sulphide during experimental colitis in rats

Hydrogen sulphide (H2 S) is produced in the intestine by sulphate-reducing bacteria and during metabolism of L-cysteine within the mucosa. This gasotransmitter induces anion secretion by stimulating enteric neurons and by a direct effect on epithelial cells. As H2 S is discussed to exert both pro- and anti-inflammatory actions, we aimed to investigate the role of H2 S during experimental colitis by comparing the effects of blockade of H2 S-forming endogenous enzymes with the effect of a S-reduced diet to diminish microbial production of H2 S. Rectal application of trinitrobenzenesulfonic acid (TNBS) was used to induce chronic colitis. The level of inflammation was assessed macroscopically and histologically. In Ussing chamber experiments, colonic specimens from TNBS-treated animals exhibited a higher tissue conductance, that is, a higher epithelial permeability, and a slightly reduced basal short-circuit current (a measure of net ion transport) in relation to non-inflamed control tissue. Analgetic treatment with flupirtine, a central antinociceptive analgetic, did not interfere with the induction of the inflammatory response so that all animals were treated with flupirtine to reduce pain and distress during the development of colitis. The secretory response evoked by an exogenous H2 S donor, NaHS, was significantly decreased after induction of colitis, whereas the response to Ca(2+) - or cAMP-dependent secretagogues was unaltered. This downregulation was not observed in the colitis group fed on a S-reduced diet. The decreased NaHS response indicates a desensitization of the tissue by inflammation, which might be explained by an upregulation of colonic H2 S production as described in some models of inflammation.

Mo1844 Selective Agonists of Somatostatin Receptors Type 1 or 2 Prevent Visceral Hyperalgesia Induced by Activation of Peripheral CRF1 Receptors in Mice

G A A b st ra ct s brought a new understanding on the mechanisms of chronic visceral pain. The present study hypothesizes that inflammation of visceral organs cause glial activation, leading to the release of pro-inflammatory cytokines, resulting in hyperalgesia. Therefore, the inhibition of microglial activation can attenuate visceral hyperalgesia. We also hypothesize that glialspecific purinergic P2X7 receptors play a role in glial-neuron cross talk and blocking of these receptors might produce an analgesic effect. Methods: Colonic inflammation was induced inmale Sprague-Dawley rats (n=12) by intracolonic administration of either zymosan (25mg/ml, 0.5ml) or tri-nitrobenzenesulphonic acid (TNBS). Visceral hyperalgesia was determined by measuring the viscero-motor response (VMR) to graded colorectal distension (CRD) prior, 3hr (for zymosan) and 7 days (for TNBS) after induction of colonic inflammation. The effects of minocycline, (a broad-spectrum tetracycline antibiotic and a microglial inhibitor) and brilliant blue G (BBG; a selective P2X7 antagonist) were tested on VMRs to CRD in preand post-inflamed conditions. Electrophysiological studies involving extracellular single-unit recordings were made from CRD-sensitive pelvic nerve afferent (PNA) fibers in naive non-inflamed and TNBS-treated animals and the effect of minocycline (50mg/kg, i.v.) on the responses of PNAs to graded CRD was studied 30 and 60mins following drug administration. Results: Colonic inflammation produced visceral hyperalgesia to CRD. Administration of minocycline (50mg/kg, i.p.) significantly (p<0.05) attenuated the visceral hyperalgesia in both zymosanand TNBS-treated rats, but not in naive non-inflamed rats. Similarly, selective blocking of P2X7 receptors by BBG (50mg/kg i.p.) attenuated VMRs to CRD in TNBS-treated rats, but not in naive rats. In electrophysiology experiments, minocycline significantly (p<0.05) attenuated mechanotransduction of CRD-sensitive PNA (n=6) to CRD, but had no effect on responses of CRD-sensitive PNA (n=4) from naive non-inflamed rats. Conclusions: Activated glial cells plays an important role in initiating and maintaining visceral hyperalgesia in a rat model of post-inflammatory pain. Blocking the function of glial cells by minocycline or by blocking the glial-specific P2X7 receptor can attenuate visceral hyperalgesia. This effect is partly via peripheral actions of these drugs, which might prove to be a vital target for the treatment of visceral hyperalgesia.

Probiotics Modulate Intestinal Expression of Nuclear Receptor and Provide Counter-Regulatory Signals to Inflammation-Driven Adipose Tissue Activation

BackgroundAdipocytes from mesenteric white adipose tissue amplify the inflammatory response and participate in inflammation-driven immune dysfunction in Crohn's disease by releasing proinflammatory mediators. Peroxisome proliferator-activated receptors (PPAR)-α and -γ, pregnane x receptor (PXR), farnesoid x receptor (FXR) and liver x-receptor (LXR) are ligand-activated nuclear receptor that provide counter-regulatory signals to dysregulated immunity and modulates adipose tissue.AimsTo investigate the expression and function of nuclear receptors in intestinal and adipose tissues in a rodent model of colitis and mesenteric fat from Crohn's patients and to investigate their modulation by probiotics.MethodsColitis was induced by TNBS administration. Mice were administered vehicle or VSL#3, daily for 10 days. Abdominal fat explants obtained at surgery from five Crohn's disease patients and five patients with colon cancer were cultured with VSL#3 medium.ResultsProbiotic administration attenuated development of signs and symptoms of colitis, reduced colonic expression of TNFα, IL-6 and IFNγ and reserved colonic downregulation of PPARγ, PXR and FXR caused by TNBS. Mesenteric fat depots isolated from TNBS-treated animals had increased expression of inflammatory mediators along with PPARγ, FXR, leptin and adiponectin. These changes were prevented by VSL#3. Creeping fat and mesenteric adipose tissue from Crohn's patients showed a differential expression of PPARγ and FXR with both tissue expressing high levels of leptin. Exposure of these tissues to VSL#3 medium abrogates leptin release.ConclusionsMesenteric adipose tissue from rodent colitis and Crohn's disease is metabolically active and shows inflammation-driven regulation of PPARγ, FXR and leptin. Probiotics correct the inflammation-driven metabolic dysfunction.

The effects of glucagon-like peptide 2 on enteric neurons in intestinal inflammation

Intestinal inflammation alters the structure and function of the enteric nervous system (ENS). Glucagon-like peptide 2 (GLP-2) reduces intestinal inflammation and has trophic effects on isolated neurons. This study examined the effects of GLP-2 treatment on the submucosal plexus of rat colon in the trinitrobenzene sulfonic acid (TNBS) model of colitis. After administration of TNBS or saline/ethanol for controls, animals were allocated to treatment with GLP-2 (50 μg kg⁻¹ day⁻¹, s.c.) or sham injection of vehicle, twice daily. Animals were monitored, following clinical parameters, and killed on day 5. The number of neuronal cell bodies per ganglion was quantified using immunohistochemistry on submucosal whole mount preparations, with further characterization of specific subpopulations using antibodies against vasoactive intestinal polypeptide (VIP), neuronal nitric oxide synthase (nNOS), and enteric glial cells with glial fibrillary acid protein and S100. Glucagon-like peptide 2 treatment was associated with a significant amelioration of weight loss, and reduced neutrophil infiltration and microscopic colitis scores in the TNBS animals. Inflammation resulted in a loss of enteric neurons in submucosal ganglia; GLP-2 treatment restored the enteric neuronal populations to normal. In control, non-inflamed animals, GLP-2 treatment increased the number of VIP expressing neurons per ganglion; in TNBS-treated animals, GLP-2 prevented an inflammation-induced reduction in the numbers of VIP expressing neurons per ganglion. Glucagon-like peptide 2 did not change the numbers of nNOS neurons or enteric glial cells in either the control, or inflamed state. These findings show that GLP-2 increased the number of VIP expressing neurons in normal animals, and prevents the inflammation-induced loss of neurons in the colonic submucosal ganglia, with an increase in the proportion of VIP expressing neurons. They suggest that GLP-2 may have a role in protecting or regulating the circuitry of the ENS under basal and inflamed states.

Antinociceptive effects of melatonin in a rat model of post-inflammatory visceral hyperalgesia: A centrally mediated process

Previous reports suggest that melatonin may play an important role in visceral nociception and neurogenic inflammation. We aimed to examine the role of melatonin on visceral hypersensitivity and to explore the site of action using a rat model of post-inflammatory visceral hyperalgesia. In all rats, a baseline viscero-motor response (VMR) to graded colorectal distension (CRD; 10–60 mmHg) was recorded prior and 1 week following tri-nitrobenzenesulfonic acid (TNBS) induced colonic inflammation. Melatonin (30, 45 or 60 mg/kg, ip) was given 20 min before testing the VMR in naïve and TNBS-treated rats. Extracellular single-unit recordings were made from CRD-sensitive pelvic nerve afferent (PNA) fibers and lumbosacral (LS) spinal neurons in TNBS-treated animals. The effect of melatonin (60 mg/kg) was examined on responses of PNAs and spinal neurons to graded CRD. In separate experiments, luzindole (non-specific MT 1/MT 2 receptor antagonist) or naltrexone (non-specific opiod receptor antagonist) was injected prior to melatonin. Following TNBS, there was a significant increase in the VMR to CRD compared to baseline. This increase was attenuated by melatonin (60 mg/kg) at pressures >20 mmHg. The same dose of melatonin had no effect on the VMR in naïve animals. In TNBS-treated rats, melatonin significantly attenuated the responses of CRD-sensitive spinal neurons to CRD, but had no effect in spinal transected rats or PNA fibers. Both luzindole and naltrexone blocked melatonin’s effect on the VMR and LS spinal neurons. Results indicate melatonin’s antinociceptive effects are not via a peripheral site of action but rather a supra-spinal process linked to the central opioidergic system.

Efficacy of a novel sphingosine kinase inhibitor in experimental Crohn’s disease

Activation of sphingosine kinase (SK) is a key response to many inflammatory processes. The present studies test the hypothesis that an orally available SK inhibitor, ABC294640, would be effective in rodent models of Crohn's disease. Trinitrobenzene sulfonic acid (TNBS) was administered rectally to mice and rats. Rats were treated with ABC294640 orally alone or in combination with olsalazine and disease progression was monitored. For both rodent species, treatment with ABC294640 attenuated disease progression. Colon samples from the ABC294640-treated animals had improved histology and cytokine parameters when compared with vehicle-treated animals. The expression of SK was similarly increased in TNBS-treated animals and in human colon tissue specimens from inflammatory bowel disease patients relative to normal, control patients. Sphingosine kinase may be a critical mediator of colonic damage during intestinal inflammation, and pharmacologic inhibitors of this enzyme may prove useful in the treatment of Crohn's disease.

Effects of ethanol extract of ficus bengalensis (bark) on inflammatory bowel disease

Objective:The present study was designed to evaluate the effects of ethanol extract of Ficus bengalensis Linn. bark (AEFB) on inflammatory bowel disease (IBD).Materials and Methods:Effects of AEFB were studied on 2,4,6-trinitrobenzenesulfonic acid (TNBS, 0.25 ml 120 mg/ml in 50% ethanol intrarectally, on first day only) induced IBD in rats. The effects of co-administration of prednisolone (2 mg/kg) and AEFB (250, 500 mg/kg) for 21 days were evaluated. Animals sacrificed at end of the experiment and various histopathological parameters like colon mucosal damage index (CMDI) and disease activity index (DAI) were assessed. In the colon homogenate malondialdehyde (MDA), myeloperoxidase (MPO), superoxide dismutase (SOD), and nitric oxide (NO) levels and in mesentery % mast cell protection was also measured.Results:Rats treated with only TNBS showed more score of CMDI and DAI, higher MDA, NO, MPO, and lower SOD activity as compared to the control group. Treatment with AEFB significantly declined both indices scores and decreased the MPO, MDA, NO, and increased the SOD activity. AEFB also increased the % mast cell protection compared to alone TNBS-treated animals.Conclusion:In our study, we found that AEFB has a significant protective effect in the IBD in rats that is comparable to that of prednisolone and may be because of the presence of flavonoids, terpenoids, and phenolic compounds.

Differential adipokine response in genetically predisposed lean and obese rats during inflammation: a role in modulating experimental colitis?

The relationship between a predisposition to obesity and the development of colitis is not well understood. Our aim was to characterize the adipokine response and the extent of colitis in diet-induced obese (DIO) rats. DIO and control, diet-resistant (DR) animals were administered either saline or trinitrobenzene sulfonic acid (TNBS) to induce colitis. Macroscopic damage scores and myeloperoxidase (MPO) activity were measured to determine the extent of inflammation. Trunk blood was collected for the analysis of plasminogen activator inhibitor-1 (PAI-1) as well as leptin, ghrelin, and adiponectin. Colonic epithelial physiology was assessed using Ussing chambers. DIO rats had a modestly increased circulating PAI-1 before TNBS treatment; however, during colitis, DR animals had more than a fourfold increase in circulating PAI-1 compared with DIO rats. Circulating leptin was higher in DIO rats compared with DR animals, in the inflamed and noninflamed states. These changes in TNBS-induced adipokine profile were accompanied by decreased macroscopic tissue damage score in DIO animals compared with DR tissues. Furthermore, TNBS-treated DR animals lost significantly more weight than DIO rats during active inflammation. Colonic epithelial physiology was comparable between groups, as was MPO activity. The factors contributing to the decreased colonic damage are almost certainly multifold, driven by both genetic and environmental factors, of which adipokines are likely to play a part given the increasing body of evidence for their role in modulating intestinal inflammation.

A novel murine model for chronic inflammatory alveolar bone loss

Chronic inflammatory bowel disease (IBD) demonstrates some similarities to the dysregulated chronic immunoinflammatory lesion of periodontitis. Trinitrobenzene sulphonic acid (TNBS) and dextran sodium sulphate (DSS) administered to rodents have been shown to elicit inflammatory responses that undermine the integrity of the gut epithelium in a similar manner to IBD in humans. The objective of this study was to evaluate the ability of these chemicals to elicit periodontal inflammation as a novel model for alveolar bone loss. Mice were treated by oral application of TNBS twice a week, or with DSS in the diet over a period of 18 weeks. Alveolar bone loss was assessed on the defleshed skull using morphometric measures for area of bone resorption. The TNBS-treated animals tolerated oral administration with no clinical symptoms and gained weight at a similar rate to normal control animals. In contrast, DSS exerted a systemic response, including shortening of colonic tissue and liver enzyme changes. Both TNBS and DSS caused a localized action on periodontal tissues, with alveolar bone loss observed in both maxilla and mandibles, with progression in a time-dependent manner. Bone loss was detected as early as week 7, with more severe periodontitis increasing over the 18 weeks (p < 0.001). Young (7-month-old) and old (12-month-old) mice with severe combined immunodeficiency were treated with TNBS for a period of 7 weeks and did not develop significant bone loss. These data show that oral administration of TNBS or DSS provokes alveolar bone loss in concert with the autochthonous oral microbiota.