Edge-based relative entropy as a sensitive indicator of critical transitions in biological systems.

BACKGROUND: Disease progression in biosystems is not always a steady process but is occasionally abrupt. It is important but challenging to signal critical transitions in complex biosystems. METHODS: In this study, based on the theoretical framework of dynamic network biomarkers (DNBs), we propose a model-free method, edge-based relative entropy (ERE), to identify temporal key biomolecular associations/networks that may serve as DNBs and detect early-warning signals of the drastic state transition during disease progression in complex biological systems. Specifically, by combining gene‒gene interaction (edge) information with the relative entropy, the ERE method converts gene expression values into network entropy values, quantifying the dynamic change in a biomolecular network and indicating the qualitative shift in the system state. RESULTS: The proposed method was validated using simulated data and real biological datasets of complex diseases. The applications show that for certain diseases, the ERE method helps to reveal so-called "dark genes" that are non-differentially expressed but with high ERE values and of essential importance in both gene regulation and prognosis. CONCLUSIONS: The proposed method effectively identified the critical transition states of complex diseases at the network level. Our study not only identified the critical transition states of various cancers but also provided two types of new prognostic biomarkers, positive and negative edge biomarkers, for further practical application. The method in this study therefore has great potential in personalized disease diagnosis.

Polyphenol-Rich Extract from 'Limoncella' Apple Variety Ameliorates Dinitrobenzene Sulfonic Acid-Induced Colitis and Linked Liver Damage.

Inflammatory bowel conditions can involve nearly all organ systems and induce pathological processes through increased oxidative stress, lipid peroxidation and disruption of the immune response. Patients with inflammatory bowel disease (IBD) are at high risk of having extra-intestinal manifestations, for example, in the hepatobiliary system. In 30% of patients with IBD, the blood values of liver enzymes, such as AST and ALT, are increased. Moreover, treatments for inflammatory bowel diseases may cause liver toxicity. Apple polyphenol extracts are widely acknowledged for their potential antioxidant effects, which help prevent damage from oxidative stress, reduce inflammation, provide protection to the liver, and enhance lipid metabolism. The aim of this study was to investigate whether the polyphenol apple extract from Malus domestica cv. 'Limoncella' (LAPE) may be an effective intervention for the treatment of IBD-induced hepatotoxicity. The LAPE was administrated in vivo by oral gavage (3-300 mg/kg) once a day for 3 consecutive days, starting 24 h after the induction of dinitro-benzenesulfonic acid (DNBS) colitis in mice. The results showed that LAPE significantly attenuated histological bowel injury, myeloperoxidase activity, tumor necrosis factor and interleukin (IL-1ß) expressions. Furthermore, LAPE significantly improved the serum lipid peroxidation and liver injury in DNBS-induced colitis, as well as reduced the nuclear transcription factor-kappaB activation. In conclusion, these results suggest that LAPE, through its antioxidant and anti-inflammatory properties, could prevent liver damage induced by inflammatory bowel disease.

Chemopreventive and immunomodulatory effects of phenolic-rich extract of Commiphora leptophloeos against inflammatory bowel disease: Preclinical evidence.

ETHNOPHARMACOLOGY RELEVANCE: Commiphora leptophloeos (Mart.) J.B. Gillet (Burseraceae) is a medicinal plant native to Brazil, popularly known as "imburana". Homemade leaf decoction and maceration were used to treat general inflammatory problems in the Brazilian Northeast population. Our previous research confirmed the anti-inflammatory activity of the C. leptophloeos hydroalcoholic leaf extract. AIM OF THE STUDY: Inflammatory bowel disease (IBD) is a chronic and relapsing inflammatory disorder of the gut with no ideal treatment to maintain the remissive status. This work aimed to characterize the phytochemical composition and physicochemical properties of the C. leptophloeos hydroalcoholic leaf extract and its efficacy in chemopreventive and immunomodulatory responses in inflammatory bowel disease in non-clinical models. MATERIALS AND METHODS: Mass spectrometry and physicochemical tests determined the phytochemical profile and physicochemical characteristics of the Commiphora leptophloeos (CL) extract. The chemopreventive and immunomodulatory effects of CL extract (50 and 125 µg/mL) were evaluated in vitro in the RAW 264.7 lipopolysaccharide (LPS) induced cell assay and in vivo in the model of intestinal inflammation induced by 2,4-Dinitrobenzenesulfonic acid (DNBS) in mice when they were treated with CL extract by intragastric gavage (i.g.) at doses of 300, 400 and 500 mg/kg. RESULTS: Phytochemical annotation of CL extract showed a complex phenolic composition, characterized as phenolic acids and flavonoids, and satisfactory physicochemical characteristics. In addition, CL extract maintained the viability of RAW macrophages, reduced ROS and NO production, and negatively regulated COX-2, iNOS, TNF-α, IL-1ß, IL-6, and IL-17 (p < 0.05). In the intestinal inflammation model, CL extract was able to downregulate NF-κB p65/COX-2, mTOR, iNOS, IL-17, decrease levels of malondialdehyde and myeloperoxidase and cytokines TNF-α, IL-1ß and IL-6 (p < 0.05). CONCLUSION: Based on these findings, CL extract reduced inflammatory responses by down-regulating pro-inflammatory markers in macrophages induced by LPS and DNBS-induced colitis in mice through NF-κB p65/COX-2 signaling. CL leaf extract requires further investigation as a candidate for treating inflammatory bowel disease.

A Tumor-Targeting Dual-Stimuli-Activatable Photodynamic Molecular Beacon for Precise Photodynamic Therapy.

A multifunctional photodynamic molecular beacon (PMB) has been designed and synthesized which contains an epidermal growth factor receptor (EGFR)-targeting cyclic peptide and a trimeric phthalocyanine skeleton in which the three zinc(II) phthalocyanine units are each substituted with a glutathione (GSH)-responsive 2,4-dinitrobenzenesulfonate (DNBS) quencher and are linked via two cathepsin B-cleavable GFLG peptide chains. This tailor-made conjugate is fully quenched in the native form due to the photoinduced electron transfer effect of the DNBS moieties and the self-quenching of the phthalocyanine units. It can target the EGFR overexpressed in cancer cells, and after receptor-mediated endocytosis, it can be activated selectively by the co-existence of intracellular GSH and cathepsin B, both of which are also overproduced in cancer cells, in terms of fluorescence emission and singlet oxygen generation. The cell-selective behavior of this PMB has been demonstrated using a range of cancer cells with different expression levels of EGFR, while the stimuli-responsive properties have been studied both in vitro and in various aqueous media. The overall results show that this advanced PMB, which exhibits several levels of control of the tumor specificity, is a promising photosensitizer for precise antitumoral photodynamic therapy.

Ethanolic extract from Lepidium virginicum L. ameliorates DNBS-induced colitis in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Lepidium virginicum L. (Brassicaceae) is a plant widely used in traditional Mexican medicine as an expectorant, diuretic, and as a remedy to treat diarrhea and dysentery, infection-derived gastroenteritis. However, there is no scientific study that validates its clinical use as an anti-inflammatory in the intestine. AIM OF THE STUDY: This study aimed to investigate the anti-inflammatory properties of the ethanolic extract of Lepidium virginicum L. (ELv) in an animal model of inflammatory bowel disease (IBD)-like colitis. MATERIALS AND METHODS: The 2,4-dinitrobenzene sulfonic acid (DNBS) animal model of IBD was used. Colitis was induced by intrarectal instillation of 200 mg/kg of DNBS dissolved vehicle, 50% ethanol. Control rats only received the vehicle. Six hours posterior to DNBS administration, ELv (3, 30, or 100 mg/kg) was administered daily by gavage or intraperitoneal injection. The onset and course of the inflammatory response were monitored by assessing weight loss, stool consistency, and fecal blood. Colonic damage was evaluated by colon weight/length ratio, histopathology, colonic myeloperoxidase (MPO) activity, and gene expression of proinflammatory cytokines, including tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1ß), chemokine C-X-C motif ligand 1 (CXCL-1), and interleukin-6 (IL-6). RESULTS: Rats treated with DNBS displayed significant weight loss, diarrhea, fecal blood, colon shortening, a significant increase in immune cell infiltration and MPO activity, as well as increased proinflammatory cytokine expression. Intraperitoneal administration of ELv significantly reduced colon inflammation, whereas oral treatment proved to be ineffective. In fact, intraperitoneal ELv significantly attenuated the clinical manifestations of colitis, immune cell infiltration, MPO activity, and pro-inflammatory (CXCL-1, TNF-α, and IL-1ß) gene expression in a dose-dependent manner. CONCLUSION: Traditional medicine has employed ELv as a remedy for common infection-derived gastrointestinal symptoms; however, we hereby present the first published study validating its anti-inflammatory properties in the mitigation of DNBS-induced colitis.

Preclinical Development of FA5, a Novel AMP-Activated Protein Kinase (AMPK) Activator as an Innovative Drug for the Management of Bowel Inflammation.

Acadesine (ACA), a pharmacological activator of AMP-activated protein kinase (AMPK), showed a promising beneficial effect in a mouse model of colitis, indicating this drug as an alternative tool to manage IBDs. However, ACA displays some pharmacodynamic limitations precluding its therapeutical applications. Our study was aimed at evaluating the in vitro and in vivo effects of FA-5 (a novel direct AMPK activator synthesized in our laboratories) in an experimental model of colitis in rats. A set of experiments evaluated the ability of FA5 to activate AMPK and to compare the efficacy of FA5 with ACA in an experimental model of colitis. The effects of FA-5, ACA, or dexamethasone were tested in rats with 2,4-dinitrobenzenesulfonic acid (DNBS)-induced colitis to assess systemic and tissue inflammatory parameters. In in vitro experiments, FA5 induced phosphorylation, and thus the activation, of AMPK, contextually to the activation of SIRT-1. In vivo, FA5 counteracted the increase in spleen weight, improved the colon length, ameliorated macroscopic damage score, and reduced TNF and MDA tissue levels in DNBS-treated rats. Of note, FA-5 displayed an increased anti-inflammatory efficacy as compared with ACA. The novel AMPK activator FA-5 displays an improved anti-inflammatory efficacy representing a promising pharmacological tool against bowel inflammation.

Extra virgin olive oil and related by-products (Olea europaea L.) as natural sources of phenolic compounds for abdominal pain relief in gastrointestinal disorders in rats.

Management of abdominal pain, a common symptom of IBDs and IBS, is still a clinical problem. Extra virgin olive oil (EVOO), a main component of the Mediterranean diet, shows positive effects on chronic inflammation in IBDs. In this study, the effect of the oral administration of EVOO (3 mL) and two olive milling by-products, DPA (300 mg kg-1) and DRF (300 mg kg-1), on preventing the development of abdominal pain in a DNBS-induced colitis model in rats was evaluated. The doses were chosen with the aim of simulating a plausible daily intake in humans. DPA and EVOO treatments significantly reduced the abdominal viscero-motor response to colon-rectal distension at 2 and 3 mL of balloon distension volume, both 7 and 14 days after the DNBS-injection. DRF showed efficacy in the reduction of visceral hypersensitivity only with 3 mL balloon inflation. In awake animals, DPA and DRF reduced pain perception (evaluated as abdominal withdrawal reflex) with all balloon distension volumes, while EVOO was effective only with higher distension volumes. Fourteen days after the DNBS-injection, all samples reduced the macroscopic intestinal damage (quantified as the macroscopic damage score) also showing, at the microscopic level, a reduction of the inflammatory infiltrate (quantified by hematoxylin and eosin analysis), fibrosis (highlighted by picrosirius red staining), the increase in mast cells and their degranulation (analyzed by triptase immunohistochemistry). This is the first report on the promotion of abdominal pain relief in a rat model obtained administering EVOO and two derived by-products. Our results suggest a protective role of phenol-rich EVOO and milling by-products, which may be proposed as food ingredients for novel functional foods.

Colon-Targeted Delivery Facilitates the Therapeutic Switching of Sofalcone, a Gastroprotective Agent, to an Anticolitic Drug via Nrf2 Activation.

We investigated if the therapeutic switching of sofalcone (SFC), a gastroprotective agent, to an anticolitic agent is feasible using colon-targeted drug delivery. SFC can activate the anti-inflammatory nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-hemeoxygenase-1 (HO-1) pathway in human colon epithelial cells and murine macrophages. For the efficient treatment of colitis, SFC was coupled with acidic amino acids to yield SFC-aspartic acid (SFC-AA) and SFC-glutamic acid, and their colon targetability and therapeutic effects were assessed as an anticolitic agent in a 2,4-dinitrobenezenesulfonic acid-induced rat colitis model. The SFC derivatives were decoupled up to 72% in the cecal contents but remained stable in the small intestinal contents. Oral gavage of SFC-AA (oral SFC-AA, equivalent to 1.67 mg/kg of SFC) delivered SFC (maximal cecal concentration: 57.36 µM) to the cecum, while no SFC was detected with oral gavage of SFC (oral SFC, 1.67 mg/kg). Moreover, oral SFC-AA (equivalent to 10 mg/kg of SFC) did not afford detectable concentration of SFC in the blood but detected up to 4.64 µM with oral SFC (10 mg/kg), indicating efficient colonic delivery and limited systemic absorption of SFC upon oral SFC-AA. Oral SFC-AA ameliorated colonic damage and inflammation in rat colitis with elevating colonic levels of HO-1 and nuclear Nrf2 protein, and the anticolitic effects of SFC-AA were significantly undermined by an HO-1 inhibitor. At an equivalent dose of SFC, oral SFC-AA but not oral SFC increased colonic HO-1 and nuclear Nrf2 levels, and oral SFC-AA was more effective than oral SFC in treating rat colitis. Moreover, oral SFC-AA was as effective against colitis as oral sulfasalazine being used for the treatment of inflammatory bowel disease. In conclusion, colon-targeted delivery of SFC facilitated the therapeutic switching of the drug to an anticolitic drug via Nrf2 activation.

Preventive effects of guanosine on intestinal inflammation in 2, 4-dinitrobenzene sulfonic acid (DNBS)-induced colitis in rats.

BACKGROUND: Guanosine, a guanine-based purine, is an extracellular signaling molecule exerting anti-inflammatory and antioxidative effects in several in vivo and in vitro injury models. We aimed to investigate its protective effects on 2, 4-dinitrobenzene sulfonic acid (DNBS)-induced colitis in rat. METHODS: Rats were divided into five groups and colitis was induced by intracolonic instillation of DNBS (15 mg/rat). Guanosine (4 or 8 mg/kg) was administered for 6 days i.p. starting the day of the colitis induction. Body weight loss, stool consistency, colon weight/length, histological analysis, myeloperoxidase activity (MPO) and pro-inflammatory cytokine levels were assessed. Immunoblotting of nuclear factor-κB (NF-κB) p65 protein levels and detection of oxidative and nitrosative stress markers were also performed. RESULTS: Guanosine, in a dose-dependent manner, significantly ameliorated the severity of DNBS-induced colitis, reducing body weight loss and diarrhea incidence, preventing the DNBS-induced macroscopic and microscopic damage to the colonic mucosa, and the MPO increase. Guanosine treatment also lowered interleukin-1ß, interleukin-6, and tumor necrosis factor-α mRNA levels. Importantly, guanosine in DNBS rats down-regulated the expression of NF-κB p65 and the levels of reactive oxygen species and nitrite. CONCLUSIONS: In conclusion, guanosine exerts beneficial effects in DNBS-induced colitis in rats, through modulation of colonic inflammation, downregulating of NFκB-mediated signaling.

Endoscopic Transplantation of Mesenchymal Stem Cell Sheets in Experimental Colitis in Rats.

Owing to the recent progress in regenerative medicine technology, clinical trials that harnessed the regeneration and immune modulation potentiality of stem cells for treating IBD have shown promising results. We investigated the feasibility and utility of intraluminal endoscopic transplantation of rat MSC sheets in murine models of experimental colitis for targeted delivery of stem cells to lesions. We isolated adipose-derived mesenchymal stem cells (AD-MSC) and bone marrow-derived mesenchymal stem cells (BM-MSC) from EGFP-transgenic rats and fabricated the cells in sheet forms using temperature-responsive culture dishes. The MSC sheets were endoscopically transplanted to the inflamed area in electrocoagulation and DNBS colitis model. The effect of the transplantation was verified using endoscopic scoring and histological analysis. In the electrocoagulation model, the AD-MSC group showed significantly decreased ulcer size in the transplanted regions. In the DNBS colitis model, the AD-MSC group showed decreased inflammation and colitis in the transplanted regions. Histologic analysis showed that the MSC sheets had successfully attached to the inflamed mucosa in both the electrocoagulation and DNBS colitis model. Our results show that endoscopic transplantation of MSC sheets could be a new effective mode of stem cell therapy for IBD treatment.

The Role of Interleukin-19 in Contact Hypersensitivity.

Interleukin (IL)-19 is a member of the IL-10 family of interleukins and is an immuno-modulatory cytokine produced by the main macrophages. The gastrointestinal tissues of IL-19 knockout mice show exacerbated experimental colitis mediated by the innate immune system and T cells. There is an increasing focus on the interaction and relationship of IL-19 with the function of T cells. Contact hypersensitivity (CHS) is T cell-mediated cutaneous inflammation. Therefore, we asked whether IL-19 causes CHS. We investigated the immunological role of IL-19 in CHS induced by 1-fluoro-2,4-dinitrofluorobenzene as a hapten. IL-19 was highly expressed in skin exposed to the hapten, and ear swelling was increased in IL-19 knockout mice. The exacerbation of the CHS response in IL-19 knockout mice correlated with increased levels of IL-17 and IL-6, but no alterations were noted in the production of interferon (IFN)γ and IL-4 in the T cells of the lymph nodes. In addition to the effect on T cell response, IL-19 knockout mice increased production of inflammatory cytokines. These results show that IL-19 suppressed hapten-dependent skin inflammation in the elicitation phase of CHS.

The antioxidant glutathione protects against enteric neuron death in situ, but its depletion is protective during colitis.

Enteric glia play an important neuroprotective role in the enteric nervous system (ENS) by producing neuroprotective compounds such as the antioxidant reduced glutathione (GSH). The specific cellular pathways that regulate glial production of GSH and how these pathways are altered during, or contribute to, neuroinflammation in situ and in vivo are not fully understood. We investigated this issue using immunohistochemistry to localize GSH synthesis enzymes within the myenteric plexus and tested how the inhibition of GSH synthesis with the selective inhibitor l-buthionine sulfoximine impacts neuronal survival and inflammation. Both enteric glia and neurons express the cellular machinery necessary for GSH synthesis. Furthermore, glial GSH synthesis is necessary for neuronal survival in isolated preparations of myenteric plexus. In vivo depletion of GSH does not induce colitis but alters myenteric plexus neuronal phenotype and survival. Importantly, global depletion of glutathione is protective against some macroscopic and microscopic measures of colonic inflammation. Together, our data highlight the heterogeneous roles of GSH in the myenteric plexus of the ENS and during gastrointestinal inflammation. NEW & NOTEWORTHY Our results show that both enteric glia and neurons express the cellular machinery necessary for glutathione (GSH) synthesis and that glial GSH synthesis is necessary for neuronal survival in isolated enteric nervous system (ENS) preparations. In vivo depletion of GSH with the selective inhibitor l-buthionine sulfoximine is not sufficient to induce inflammation but does alter neuronal neurochemical composition and survival. Together, our data highlight novel heterogeneous roles for GSH in the ENS and during gastrointestinal inflammation.

Intestinal anti-inflammatory effects of goat whey on DNBS-induced colitis in mice.

This study evaluated the intestinal anti-inflammatory effects of goat whey in a mouse model of colitis induced by 2,4-dinitrobenzenesulfonic acid that resembles human IBD. At a concentration of 4 g/kg/day, the goat whey improved the symptoms of intestinal inflammation, namely by decreasing the disease activity index, colonic weight/length, and leukocyte infiltration. Moreover, goat whey inhibited NF-κB p65 and p38 MAPK signaling pathways and consequently down-regulated the gene expression of various proinflammatory markers such as IL-1ß, IL-6, IL-17, TNF-α, iNOS, MMP-9, ICAM-1. Also, goat whey increased the expression of proteins such as mucins, occludin proteins and cytokine signalling suppressors. The immunomodulatory properties of goat whey were also evaluated in vitro using the murine macrophage cell line Raw 264 and CMT-93 cells derived from mouse rectum carcinomas. The results revealed the ability of goat whey to inhibit the production of NO and reduce IL-6 production in LPS-stimulated cells. In conclusion, goat whey exhibited anti-inflammatory effects in the DNBS model of intestinal inflammation, and these observations were confirmed by its immunomodulatory properties in vitro. Together, our results indicate that goat whey could have applications for the treatment of IBD.

Intestinal Epithelial Cell-Specific Deletion of PLD2 Alleviates DSS-Induced Colitis by Regulating Occludin.

Ulcerative colitis is a multi-factorial disease involving a dysregulated immune response. Disruptions to the intestinal epithelial barrier and translocation of bacteria, resulting in inflammation, are common in colitis. The mechanisms underlying epithelial barrier dysfunction or regulation of tight junction proteins during disease progression of colitis have not been clearly elucidated. Increase in phospholipase D (PLD) activity is associated with disease severity in colitis animal models. However, the role of PLD2 in the maintenance of intestinal barrier integrity remains elusive. We have generated intestinal-specific Pld2 knockout mice (Pld2 IEC-KO) to investigate the mechanism of intestinal epithelial PLD2 in colitis. We show that the knockout of Pld2 confers protection against dextran sodium sulphate (DSS)-induced colitis in mice. Treatment with DSS induced the expression of PLD2 and downregulated occludin in colon epithelial cells. PLD2 was shown to mediate phosphorylation of occludin and induce its proteasomal degradation in a c-Src kinase-dependent pathway. Additionally, we have shown that treatment with an inhibitor of PLD2 can rescue mice from DSS-induced colitis. To our knowledge, this is the first report showing that PLD2 is pivotal in the regulation of the integrity of epithelial tight junctions and occludin turn over, thereby implicating it in the pathogenesis of colitis.

Adelmidrol, a Palmitoylethanolamide Analogue, as a New Pharmacological Treatment for the Management of Inflammatory Bowel Disease.

Leukocyte infiltration, improved levels of intercellular adhesion molecule 1 (ICAM-1), and oxidative stress in the colon are the principal factors in inflammatory bowel disease. The goal of the current study was to explore the effects of adelmidrol, an analog of the anti-inflammatory fatty acid amide signaling molecule palmitoylethanolamide, in mice subjected to experimental colitis. Additionally, to clarify whether the protective action of adelmidrol is dependent on the activation of peroxisome proliferator-activated receptors (PPARs), we investigated the effects of a PPARγ antagonist, GW9662, on adelmidrol action. Adelmidrol (10 mg/kg daily, o.s.) was tested in a murine experimental model of colitis induced by intracolonic administration of dinitrobenzene sulfonic acid. Nuclear factor-κB translocation, cyclooxygenase-2, and phosphoextracellular signal-regulated kinase, as well as tumor necrosis factor-α and interleukin-1ß, were significantly increased in colon tissues after dinitrobenzene sulfonic acid administration. Immunohistochemical staining for ICAM-1, P-selectin, nitrotyrosine, and poly(ADP)ribose showed a positive staining in the inflamed colon. Treatment with adelmidrol decreased diarrhea, body weight loss, and myeloperoxidase activity. Adelmidrol treatment, moreover, reduced nuclear factor-κB translocation, cyclooxygenase-2, and phosphoextracellular signal-regulated kinase expression; proinflammatory cytokine release; and the incidence of nitrotyrosine and poly(ADP)ribose in the colon. It also decreased the upregulation of ICAM-1 and P-selectin. Adelmidrol treatment produced a reduction of Bax and an intensification of Bcl-2 expression. This study clearly demonstrates that adelmidrol exerts important anti-inflammatory effects that are partly dependent on PPARγ, suggesting that this molecule may represent a new pharmacologic approach for inflammatory bowel disease treatment.

Intestinal anti-inflammatory effects of total alkaloid extract from Fumaria capreolata in the DNBS model of mice colitis and intestinal epithelial CMT93 cells.

BACKGROUND: Fumaria capreolata L. (Papaveraceae) is a botanical drug used in North Africa for its gastro-intestinal and anti-inflammatory properties. It is characterized for the presence of several alkaloids that could be responsible for some of its effects, including an immunomodulatory activity. PURPOSE: To test in vivo the intestinal anti-inflammatory properties of the total alkaloid fraction extracted from the aerial parts of F. capreolata (AFC), and to evaluate its effects on an intestinal epithelial cell line. STUDY DESIGN AND METHODS: AFC was chemically characterized by liquid chromatography coupled to diode array detection and high resolution mass spectrometry. Different doses of AFC (25, 50 and 100mg/kg) were assayed in the DNBS model of experimental colitis in mice, and the colonic damage was evaluated both histologically and biochemically. In addition, in vitro experiments were performed with this alkaloid fraction on the mouse intestinal epithelial cell line CMT93 stimulated with LPS. RESULTS: The chemical analysis of AFC revealed the presence of 23 alkaloids, being the most abundants stylopine, protopine and coptisine. Oral administration of AFC produced a significant inhibition of the release and the expression of IL-6 and TNF-α in the colonic tissue. It also suppressed in vivo the transcription of other pro-inflammatory mediators such as IL-1ß, iNOS, IL-12 and IL-17. Furthermore, AFC showed an immunomodulatory effect in vitro since it was able to inhibit the mRNA expression of IL-6, TNF-α and ICAM-1. Moreover, the beneficial effect of AFC in the colitic mice could also be associated with the normalization of the expression of MUC-2 and ZO-1, which are important for the intestinal epithelial integrity. CONCLUSION: The present study suggests that AFC, containing 1.3% of stylopine and 0.9% of protopine, significantly exerted intestinal anti-inflammatory effects in an experimental model of mouse colitis. This fact could be related to a modulation of the intestinal immune response and a restoration of the intestinal epithelial function.

Identification of novel anti-inflammatory probiotic strains isolated from pulque.

Probiotics are live microorganisms which when administered in adequate amounts, confer health benefits on the host. Their use is more and more widespread for both prevention and treatment of diseases, including traveler's diarrhea and inflammatory bowel diseases (IBDs). In this work, we isolated and characterized novel candidate probiotic strains from pulque (xaxtle), a traditional Mexican alcoholic fermented beverage. A total of 14 strains were obtained from xaxtle samples isolated from three different Mexican regions. Species identification was performed by biochemical methods and 16S rRNA gene targeted PCR. The isolates belonged to the Lactobacillus plantarum, Lactobacillus paracasei, Lactobacillus brevis, and Lactobacillus composti phylogenetic groups, with L. brevis being the most dominant group. Bacteria were tested for lysozyme, low pH, and bile acid resistance. Moreover, the strains were tested for adherence to human intestinal epithelial cells and screened for their immunomodulatory properties using a cellular model. Selected bacterial strains with anti-inflammatory properties were then tested in vivo in a dinitro-benzene sulfonic acid (DNBS)-induced chronic colitis mouse model, and weight loss, gut permeability, and cytokine profiles were measured as readouts of inflammation. One of the selected strains, Lactobacillus sanfranciscensis LBH1068, improved mice health as observed by a reduction of weight loss, significant decreases in gut permeability, and cytokine modulation. Altogether, our results highlighted the potential of lactobacilli isolated from pulque and in particular the strain L. sanfranciscensis LBH1068 as a novel probiotic to treat IBD.

Dimethyl Fumarate Reduces Inflammatory Responses in Experimental Colitis.

BACKGROUND AND AIMS: Fumaric acid esters have been proven to be effective for the systemic treatment of psoriasis and multiple sclerosis. We aimed to develop a new treatment for colitis. METHODS: We investigated the effect of dimethylfumarate [DMF, 10-30-100mg/kg] on an experimental model of colitis induced by dinitrobenzene sulphuric acid [DNBS]. We also evaluated the therapeutic activity of 7 weeks' treatment with DMF [30mg/kg] on 9-week-old IL-10KO mice that spontaneously develop a T helper-1 [Th1]-dependent chronic enterocolitis after birth, that is fully established at 8-10 weeks of age. The mechanism of this pharmacological potential of DMF [10 µM] was investigated in colonic epithelial cell monolayers [Caco-2] exposed to H2O2. The barrier function was evaluated by the tight junction proteins. RESULTS: The treatment with DMF significantly reduced the degree of haemorrhagic diarrhoea and weight loss caused by administration of DNBS. DMF [30 and 100mg/kg] also caused a substantial reduction in the degree of colon injury, in the rise in myeloperoxidase [MPO] activity, and in the increase in tumour necrosis factor [TNF]-α expression, as well as in the up-regulation of ICAM-1 caused by DNBS in the colon. Molecular studies demonstrated that DMF impaired NF-κB signalling via reduced p65 nuclear translocalisation. DMF induced a stronger antioxidant response as evidenced by a higher expression of Mn-superoxide dismutase. Moreover, DMF protected human intestinal epithelial cells against H2O2-induced barrier dysfunction, restoring ZO-1 occludin expression, via the HO-1 pathway. CONCLUSIONS: DMF treatment reduces the degree of colitis caused by DNBS. We propose that DMF treatment may be useful in the treatment of inflammatory bowel disease.

Vasoactive intestinal polypeptide promotes intestinal barrier homeostasis and protection against colitis in mice.

Inflammatory bowel disease is a chronic gastrointestinal inflammatory disorder associated with changes in neuropeptide expression and function, including vasoactive intestinal peptide (VIP). VIP regulates intestinal vasomotor and secretomotor function and motility; however, VIP's role in development and maintenance of colonic epithelial barrier homeostasis is unclear. Using VIP deficient (VIPKO) mice, we investigated VIP's role in epithelial barrier homeostasis, and susceptibility to colitis. Colonic crypt morphology and epithelial barrier homeostasis were assessed in wildtype (WT) and VIPKO mice, at baseline. Colitic responses were evaluated following dinitrobenzene sulfonic acid (DNBS) or dextran-sodium sulfate (DSS) exposure. Mice were also treated with exogenous VIP. At baseline, VIPKO mice exhibited distorted colonic crypts, defects in epithelial cell proliferation and migration, increased apoptosis, and altered permeability. VIPKO mice also displayed reduced goblet cell numbers, and reduced expression of secreted goblet cell factors mucin 2 and trefoil factor 3. These changes were associated with reduced expression of caudal type homeobox 2 (Cdx2), a master regulator of intestinal function and homeostasis. DNBS and DSS-induced colitis were more severe in VIPKO than WT mice. VIP treatment rescued the phenotype, protecting VIPKO mice against DSS colitis, with results comparable to WT mice. In conclusion, VIP plays a crucial role in the development and maintenance of colonic epithelial barrier integrity under physiological conditions and promotes epithelial repair and homeostasis during colitis.

Effect of chenodeoxycholic acid and sodium hydrogen sulfide in dinitro benzene sulfonic acid (DNBS)--Induced ulcerative colitis in rats.

BACKGROUND: Ulcerative colitis is a chronic inflammatory condition in which the inflammatory response confined to the colon. There is a need to explore the new targets for UC such as Farnesoid X receptor and hydrogen sulfide pathway. METHODS: Wistar rats of either sex (200-250 g) were used. 2,4-Dinitrobenzene sulfonic acid (DNBS) (25mg/rat) given by rectal route into the colon to induced symptoms of ulcerative colitis. Chenodeoxycholic acid (CDCA) (10 and 20mg/kg) and sodium hydrogen sulfide (NaHS) (10 and 30 µmol/kg) and a inhibitor of cystathionine-γ-lyase enzyme (CSE) i.e. dl-propargylglycine (10mg/kg) treatment given along with 2,4-dinitrobenzene sulfonic acid. The disease activity index was assessed by daily change in body weight and rectal bleed score and change in length of colon. Oxidative stress markers (reduced glutathione, malondialdehyde (MDA), nitrite, and catalase and myeloperoxidase enzyme activity), serum glutamic-oxaloacetic transaminase (SGOT) and serum glutamic pyruvic transaminase (SGPT) levels in blood serum, and cardiac hemodynamic were performed on last day. RESULTS: The administration of DNBS intra-rectally in rats produced loss of body weight and bloody diarrhea with significant increase in oxidative stress markers in the colon. CDCA (10 and 20mg/kg) and NaHS (10 and 30 µmol/kg) significantly attenuated DNBS-induced UC in rats. The combination of CDCA (10mg/kg) and NaHS (10 µmol/kg) showed synergetic effect whereas; dl-propargylglycine reversed the protective effect of CDCA. CONCLUSION: The observed beneficial effects following CDCA may be due to its action through activation of CSE enzyme which leads to hydrogen sulfide generation.