Fructooligosaccharides exert intestinal anti-inflammatory activity in the CD4+ CD62L+ T cell transfer model of colitis in C57BL/6J mice.

PURPOSE: Fructooligosaccharides (FOS) are used as functional foods due to their prebiotic effects. Intestinal anti-inflammatory activity has been established in most, but not all, studies in animal models of colitis, using mainly chemically induced inflammation. Our goal was to test the effect of FOS (degree of polymerization 2-8) in the chronic, lymphocyte-driven CD4+ CD62L+ T cell transfer model of colitis. METHODS: Colitis was induced by transfer of CD4+ CD62L+ T cells to C57BL/6J Rag1(-/-) mice. FOS (75 mg day(-1)) was administered by gavage as a post-treatment. Three groups were established: non-colitic (NC), colitic control (C, CD4+ CD62L+ transferred mice treated with vehicle) and colitic+FOS (C+FOS, similar but treated with FOS). Mice were killed after 13 days. RESULTS: Treatment of mice with FOS ameliorated colitis, as evidenced by an increase in body weight, a lesser myeloperoxidase and alkaline phosphatase activities, a lower secretion of proinflammatory cytokines by mesenteric lymph node cells ex vivo (IFN-γ, IL-17, and TNF-α), and a higher colonic expression of occludin (C+FOS vs. C, p < 0.05). Increased relative abundance of lactic acid bacteria was observed in FOS-treated mice (p < 0.05). CONCLUSIONS: FOS exert intestinal anti-inflammatory activity in T lymphocyte-dependent colitis, suggesting it may be useful in the management of inflammatory bowel disease in appropriate conditions.

A new therapeutic association to manage relapsing experimental colitis: Doxycycline plus Saccharomyces boulardii.

Immunomodulatory antibiotics have been proposed for the treatment of multifactorial conditions such as inflammatory bowel disease. Probiotics are able to attenuate intestinal inflammation, being considered as safe when chronically administered. The aim of the study was to evaluate the anti-inflammatory effects of doxycycline, a tetracycline with immunomodulatory properties, alone and in association with the probiotic Saccharomyces boulardii CNCMI-745. Doxycycline was assayed both in vitro (Caco-2 epithelial cells and RAW 264.7 macrophages) and in vivo, in the trinitrobenzenesulfonic acid (TNBS) model of rat colitis and the dextran sodium sulfate (DSS) model of mouse colitis. In addition, the anti-inflammatory effect of the association of doxycycline and the probiotic was evaluated in vitro and in vivo in a DSS model of reactivated colitis in mice. Doxycycline displayed immunomodulatory activity in vitro, reducing IL-8 production by intestinal epithelial cells and nitric oxide by macrophages. Doxycycline administration to TNBS-colitic rats (5, 10 and 25 mg/kg) ameliorated the intestinal inflammatory process, being its efficacy comparable to that previously showed by minocycline. Doxycycline treatment was also effective in reducing acute intestinal inflammation in the DSS model of mouse colitis. The association of doxycycline and S. boulardii helped managing colitis in a reactivated model of colitis, by reducing intestinal inflammation and accelerating the recovery and attenuating the relapse. This was evidenced by a reduced disease activity index, colonic tissue damage and expression of inflammatory mediators. This study confirms the intestinal anti-inflammatory activity of doxycycline and supports the potential use of its therapeutic association with S. boulardii for the treatment of inflammatory bowel diseases, in which doxycycline is used to induce remission and long term probiotic administration helps to prevent the relapses.

Intestinal anti-inflammatory activity of apigenin K in two rat colitis models induced by trinitrobenzenesulfonic acid and dextran sulphate sodium.

Flavonoids are polyphenolic compounds that are widespread in nature, and consumed as part of the human diet in significant amounts. The aim of the present study was to test the intestinal anti-inflammatory activity of apigenin K, a soluble form of apigenin, in two models of rat colitis, namely the trinitrobenzenesulfonic acid (TNBS) model and the dextran sulphate sodium (DSS) model. Apigenin K (1, 3 and 10 mg/kg; by the oral route; n 4-6 per group) was administered as a pre-treatment to rats with TNBS and DSS colitis, and colonic status was checked by macroscopic and biochemical examination. Apigenin K pre-treatment resulted in the amelioration of morphological signs and biochemical markers in the TNBS model. The results demonstrated a reduction in the inflamed area, as well as lower values of score and colonic weight:length ratio compared with the TNBS group. Myeloperoxidase (MPO) activity was reduced by 30 % (P< 0·05). Moreover, apigenin K pre-treatment ameliorated morphological signs and biochemical markers in the DSS model. Thus, macroscopic damage was significantly reduced and the colonic weight:length ratio was lowered by approximately 10 %, while colonic MPO and alkaline phosphatase activities were decreased by 35 and 21 %, respectively (P< 0·05). Apigenin K pre-treatment also tended to normalise the expression of a number of colonic inflammatory markers (e.g. TNF-α, transforming growth factor-ß, IL-6, intercellular adhesion molecule 1 or chemokine (C-C motif) ligand 2). In conclusion, apigenin K is found to have anti-inflammatory effects in two preclinical models of inflammatory bowel disease.

Active hexose correlated compound exerts therapeutic effects in lymphocyte driven colitis.

Active hexose correlated compound (AHCC) is a commercial extract of Basidiomycetes fungi enriched in oligosaccharides that is used as a human nutritional supplement for various purposes in humans. Our aim was to study the anti-inflammatory effect of AHCC in the CD4+ CD62L(+) T cell transfer model of colitis, considered one of the closest to the human disease. Colitis was induced by transfer of CD4(+) CD62L(+) T cells to recombination activating gene 1(-/-) mice. AHCC (75 mg/d) was administered by gavage as a post-treatment. Three groups were established: noncolitic, colitic (CD4(+) CD62L(+) transferred mice treated with vehicle), and AHCC (colitic treated with AHCC). AHCC improved colitis, as evidenced by a 24% lower colonic myeloperoxidase and a 21% lower alkaline phosphatase activity. In addition, a decreased secretion of proinflammatory genes assessed by RT-qPCR was observed, particularly TNF-α and IL-1ß. Ex vivo mesenteric lymph node cells obtained from AHCC treated mice exhibited a fully normalized production of IL-6, IL-17, and IL-10 (p < 0.05). Also, AHCC treated mice exhibited decreased STAT4 and IκB-α phosphorylation in splenic CD4(+) cells. Our data provide validation of AHCC colonic anti-inflammatory activity in a chronic, T cell driven model of inflammatory bowel disease.

Prebiotic oligosaccharides directly modulate proinflammatory cytokine production in monocytes via activation of TLR4.

SCOPE: Prebiotic oligosaccharides are currently used in a variety of clinical settings for their effects on intestinal microbiota. Here, we have examined the direct, microbiota independent, effects of prebiotics on monocytes and T lymphocytes in vitro. METHODS AND RESULTS: Prebiotics generally evoked cytokine secretion (TNF-α, IL-6, and IL-10) by mouse splenocytes but inhibited LPS -induced IFN-γ and IL-17 release. Inulin was found to enhance LPS-induced IL-10 secretion. Splenocytes from TLR4(-/-) (where TLR is Toll-like receptor) mice showed a markedly depressed response. Conversely, in both basal and LPS-stimulated conditions, prebiotic inhibition of IFN-γ levels was preserved. These results suggested a predominant effect on monocytes via TLR4 ligation and possible inhibition of T cells. Hence, we studied the modulation of primary rat monocytes and T lymphocytes, focusing on fructooligosaccharides (FOS) and inulin. In monocytes, FOS and inulin induced TNF-α, growth-regulated oncogene α, and IL-10, but not IL-1ß release. The NF-κB inhibitor Bay 11-7082 fully prevented these effects. Pharmacological evidence also indicated a significant involvement of mitogen-activated protein kinase and phosphatidylinositol-3-kinase. There was little effect on T cells. FOS and inulin also generally increased TNF-α, IL-1ß, and IL-10, but not IL-8, in human peripheral blood monocytes. CONCLUSION: We conclude that prebiotics may act as TLR4 ligands or as indirect TLR4 modulators to upregulate cytokine secretion in monocytes.

Nondigestible oligosaccharides exert nonprebiotic effects on intestinal epithelial cells enhancing the immune response via activation of TLR4-NFκB.

SCOPE: Prebiotic effects of non absorbable glucids depend mainly on digestion by the colonic microbiota. Our aim was to assess nonprebiotic, direct effects of 4 prebiotics, namely fructooligosaccharides, inulin, galactooligosaccharides, and goat's milk oligosaccharides on intestinal epithelial cells. METHODS AND RESULTS: Prebiotics were tested in intestinal epithelial cell 18 (IEC18), HT29, and Caco-2 cells. Cytokine secretion was measured by ELISA and modulated with pharmacological probes and gene silencing. Prebiotics induced the production of growth-related oncogene, (GROα), monocyte chemoattractant protein 1 (MCP-1), and macrophage inflammatory protein 2 (MIP2) in IEC18 cells, with an efficacy that was 50-80% that of LPS. Prebiotics did not change RANTES expression, which was robustly induced by LPS in IEC18 cells. Cytokine secretion was suppressed by Bay11-7082, an inhibitor of IκB-α phosphorylation. The response was markedly decreased by Myd88 or TLR4 gene knockdown. Prebiotics also elicited cytokine production in HT29 but not in Caco-2 cells, consistent with reduced and vestigial expression of TLR4 in these cell lines, respectively. Prebiotic-induced MCP-1 secretion was reduced also in colonic explants from TLR4 KO mice compared with the controls. CONCLUSIONS: We conclude that prebiotics are TLR4 ligands in intestinal epithelial cells and that this may be a relevant mechanism for their in vivo effects.

Intestinal anti-inflammatory activity of hydroalcoholic extracts of Phlomis purpurea L. and Phlomis lychnitis L. in the trinitrobenzenesulphonic acid model of rat colitis.

ETHNOPHARMACOLOGICAL RELEVANCE: Different species from genus Phlomis, frequently native from the the eastern Mediterranean zone, have been used in traditional medicine as an anti-inflammatory remedy. Among other constituents, they contain polyphenols that show antioxidant properties, which are interesting for the treatment of inflammatory pathologies associated with oxidative stress in humans, such as inflammatory bowel disease (IBD). The aim of this study was to evaluate the intestinal anti-inflammatoy effect of hydroalcoholic extracts of Phlomis lychnitis and P. purpurea in the trinitrobenzenesulphonic acid (TNBS) model of rat colitis, a well characterized experimental model with some resemblance to human IBD. MATERIALS AND METHODS: Hydroalcoholic extracts of both plants were characterized by determining their polyphenolic content and then assayed in the TNBS model of rat colitis. For this purpose, female Wistar rats were assigned to seven groups (n=10): healthy control, untreated TNBS-colitis and five TNBS- colitis groups treated with Phlomis lychnitis (10 and 20mg/kg), P. purpurea (10 and 25mg/kg) and sulphasalazine (200mg/kg), as a positive control. Treatments started the same day of TNBS colitis induction, and rats were sacrificed one week later. Colonic inflammation was evaluated both histologically and biochemically. RESULTS: The histological (macroscopic and microscopic) analysis of colonic samples revealed that both extracts showed an anti-inflammatory effect, which was confirmed biochemically by a decreased colonic MPO activity, a maker of neutrophil infiltration, an increased colonic glutathione content, which counteracts the oxidative status associated with the inflammatory process, and a down-regulated iNOS expression. However, only the extract of P. purpurea reduced the expression of the proinflammatory cytokines IL-1ß and IL-17, the chemokines CINC-1 and MCP-1, as well as the adhesion molecule ICAM-1, ameliorating the altered immune response associated with the colonic inflammation. Furthermore, both P. lychnitis and P. purpurea extracts were able to significantly increase the expression of markers of epithelial integrity such as MUC-2, MUC-3 and villin, thus revealing an improvement in the altered colonic permeability that characterizes colonic inflammation. CONCLUSIONS: Both extracts showed intestinal anti-inflammatory activity in the TNBS model of rat colitis, thus confirming their traditional use in digestive inflammatory complaints. In addition to their antioxidant properties, other mechanisms can contribute to this beneficial effect, like an improvement in the intestine epithelial barrier and a downregulation of the immune response.

The nutritional supplement Active Hexose Correlated Compound (AHCC) has direct immunomodulatory actions on intestinal epithelial cells and macrophages involving TLR/MyD88 and NF-κB/MAPK activation.

Active Hexose Correlated Compound (AHCC) is an immunostimulatory nutritional supplement. AHCC effects and mechanism of action on intestinal epithelial cells or monocytes are poorly described. AHCC was added to the culture medium of intestinal epithelial cells (IEC18 and HT29 cells) and monocytes (THP-1 cells) and assessed the secretion of proinflammatory cytokines by ELISA. Inhibitors of NFκB and MAPKs were used to study signal transduction pathways while TLR4 and MyD88 were silenced in IEC18 cells using shRNA. It was found that AHCC induced GROα and MCP1 secretion in IEC18 and IL-8 in HT29 cells. These effects depended on NFκB activation, and partly on MAPKs activation and on the presence of MyD88 and TLR4. In THP-1 cells AHCC evoked IL-8, IL-1ß and TNF-α secretion. The induction of IL-8 depended on JNK and NFκB activation. Therefore, AHCC exerts immunostimulatory effects on intestinal epithelial cells and monocytes involving TLR4/MyD88 and NFκB/MAPK signal transduction pathways.

Active hexose-correlated compound and Bifidobacterium longum BB536 exert symbiotic effects in experimental colitis.

PURPOSE: Active hexose-correlated compound (AHCC) is a commercial extract obtained from Basidiomycetes under controlled conditions, yielding a 74 % content in oligosaccharides, especially α-glucans. AHCC has a number of therapeutic effects, including intestinal anti-inflammatory activity. Bifidobacterium longum BB536 is a probiotic with potential health-promoting effect at the gut level. The purpose of the present study was to evaluate the possibility of synergism between AHCC, which is believed to act as a prebiotic, and B. longum BB536. METHODS: We used the trinitrobenzene sulfonic acid model (TNBS) of colitis in rats. AHCC (100 or 500 mg kg(-1)) and B. longum BB536 (5 × 10(6) CFU rat(-1) day(-1)) were administered together or separately for 7 days prior to colitis induction and then for another 7 days and compared with control (noncolitic) and TNBS rats. RESULTS: The results show that both treatments had intestinal anti-inflammatory activity separately, which was enhanced when used in combination, as shown by changes in body weight gain, colonic weight to length ratio, myeloperoxydase activity and iNOS expression. Interestingly, the association of AHCC 100 mg kg(-1) + B. longum BB536 showed the highest anti-inflammatory activity. CONCLUSIONS: Our data provide a preclinical experimental basis for the synergistic effect of AHCC and B. longum BB536 on inflammatory bowel disease.

Dose-dependent antiinflammatory effect of ursodeoxycholic acid in experimental colitis.

The denomination of inflammatory bowel disease comprises a group of chronic inflammatory diseases of the digestive tract, ulcerative colitis and Crohn's disease being the most important conditions. Bile acids may play a role both in etiology and pharmacology of this disease. Thus, although deoxycholic acid is regarded as a proinflammatory agent ursodeoxycholic acid, which is currently being used to treat certain types of cholestasis and primary biliary cirrhosis, because of their choleretic, cytoprotective and immunomodulatory effects, it has been reported to exert an anti-inflammatory activity. We aim to confirm and characterize the intestinal antiinflammatory activity of ursodeoxycholic acid. The experimental model trinitrobenzenesulfonic acid (TNBS)-induced colitis in rats has been used. Animal status was characterized by a number of macroscopic and biochemical parameters. Oral administration of ursodeoxycholic acid was able to ameliorate experimental colonic inflammation. This occurred only at a relatively high dose (50 mg/kg day), whereas ursodeoxycholic acid was without significant effect at doses of 10 and 25 mg/kg day. The therapeutic effect was evidenced, among others, by a higher body weight recovery, a diminished affected to total mucosal area and lower alkaline phosphatase activity in treated vs. control (TNBS treated) animals. These results indicate that, at the appropriate dose, ursodeoxycholic acid is a potentially useful drug to reduce intestinal inflammation and could be envisaged to be incorporated in the treatment of inflammatory bowel diseases.

The immunomodulatory properties of viable Lactobacillus salivarius ssp. salivarius CECT5713 are not restricted to the large intestine.

PURPOSE: The aim of this study was to better characterise the biological effects of Lactobacillus salivarius ssp. salivarius CECT5713, a probiotic with immunomodulatory properties. METHODS: Live or dead probiotic was assayed in the TNBS model of rat colitis to determine whether viability was a requisite to exert the beneficial effects. In vitro studies were also performed in Caco-2 cells to evaluate its effects on epithelial cell recovery and IL-8 production. Finally, the probiotic was assayed in the LPS model of septic shock in mice to establish its effects when there is an altered systemic immune response. RESULTS: The viability of the probiotic was required for its anti-inflammatory activity. The probiotic inhibited IL-8 production in stimulated Caco-2 cells and facilitated the recovery of damaged intestinal epithelium. In LPS-treated mice, the probiotic inhibited the production of TNFα in plasma and lungs and increased the hepatic glutathione content. These effects were associated with an improvement in the altered production of the T-cell cytokines in splenocytes, by reducing IL-2 and IL-5 and by increasing IL-10. Finally, it reduced the increased plasma IgG production in LPS-treated mice. CONCLUSION: The anti-inflammatory effects of viable L. salivarius ssp. salivarius CECT5713 are not restricted to the gastrointestinal tract.

DNFB-DNS hapten-induced colitis in mice should not be considered a model of inflammatory bowel disease.

BACKGROUND: The dinitrofluorobenzene/dinitrosulfonic acid (DNFB/DNS) model was originally described as an experimental model of intestinal inflammation resembling human ulcerative colitis (UC). Due to the absence of acceptable UC experimental models for pharmacological preclinical assays, here we examine the immune response induced in this model. METHODS: Balb/c mice were sensitized by skin application of DNFB on day 1, followed by an intrarectal challenge with DNS on day 5. We further expanded this model by administering a second DNS challenge on day 15. The features of colonic inflammation and immune response were evaluated. RESULTS: The changes observed in colonic tissue corresponded, in comparison to the trinitrobenzene sulfonic acid (TNBS) colitis model, to a mild mucosal effect in the colon, which spontaneously resolved in less than 5 days. Furthermore, the second hapten challenge did not exacerbate the inflammatory response. In contrast to other studies, we did not observe any clear involvement of tumor necrosis factor alpha (TNF-α) or other Th1 cytokines during the initial inflammatory response; however, we found that a more Th2-humoral response appeared to mediate the first contact with the hapten. An increased humoral response was detected during the second challenge, although an increased Th1/Th17-cytokine expression profile was also simultaneously observed. CONCLUSIONS: On the basis of these results, although the DNFB/DNS model can display some features found in human UC, it should be considered as a model for the study of the intestinal hypersensitivity seen, for example, during food allergy or irritable bowel syndrome but not intestinal inflammation per se.

Antihypertensive effects of peroxisome proliferator-activated receptor-ß activation in spontaneously hypertensive rats.

Activation of nuclear hormone receptor peroxisome proliferator-activated receptor ß/δ (PPARß) has been shown to improve insulin resistance and plasma high-density lipoprotein levels, but nothing is known about its effects in genetic hypertension. We studied whether the PPARß agonist GW0742 might exert antihypertensive effects in spontaneously hypertensive rats (SHRs). The rats were divided into 4 groups, Wistar Kyoto rat-control, Wistar Kyoto rat-treated (GW0742, 5 mg · kg(-1) · day(-1) by oral gavage), SHR-control, and SHR-treated, and followed for 5 weeks. GW0742 induced a progressive reduction in systolic arterial blood pressure and heart rate in SHRs and reduced the mesenteric arterial remodeling, the increased aortic vasoconstriction to angiotensin II, and the endothelial dysfunction characteristic of SHRs. These effects were accompanied by a significant increase in endothelial NO synthase activity attributed to upregulated endothelial NO synthase and downregulated caveolin 1 protein expression. Moreover, GW0742 inhibited vascular superoxide production, downregulated p22(phox) and p47(phox) proteins, decreased both basal and angiotensin II-stimulated NADPH oxidase activity, inhibited extracellular-regulated kinase 1/2 activation, and reduced the expression of the proinflammatory and proatherogenic genes, interleukin 1ß, interleukin 6, or intercellular adhesion molecule 1. None of these effects were observed in Wistar Kyoto rats. PPARß activation, both in vitro and in vivo, increased the expression of the regulators of G protein-coupled signaling proteins RGS4 and RGS5, which negatively modulated the vascular actions of angiotensin II. PPARß activation exerted antihypertensive effects, restored the vascular structure and function, and reduced the oxidative, proinflammatory, and proatherogenic status of SHRs. We propose PPARß as a new therapeutic target in hypertension.

The intestinal anti-inflammatory effect of minocycline in experimental colitis involves both its immunomodulatory and antimicrobial properties.

Some antibiotics, including minocycline, have recently been reported to display immunomodulatory properties in addition to their antimicrobial activity. The use of a compound with both immunomodulatory and antibacterial properties could be very interesting in the treatment of inflammatory bowel disease (IBD), so the aim of our study was to evaluate the anti-inflammatory effect of minocycline in several experimental models of IBD. Firstly, the immunomodulatory activity of the antibiotic was tested in vitro using Caco-2 intestinal epithelial cells and RAW 264.7 macrophages; minocycline was able to inhibit IL-8 and nitrite production, respectively. In vivo studies were performed in trinitrobenzenesulfonic acid (TNBS)-induced rat colitis and dextran sodium sulfate (DSS)-induced mouse colitis. The results revealed that minocycline exerted an intestinal anti-inflammatory effect when administered as a curative treatment in the TNBS model, modulating both immune and microbiological parameters, being confirmed in the DSS model; whereas none of the other antibiotics tested (tetracycline and metronidazole) showed anti-inflammatory effect. However, minocycline administration before the colitis induction was not able to prevent the development of the intestinal inflammation, thus showing that only its antimicrobial activity is not enough for the anti-inflammatory effect. In conclusion, minocycline displays an anti-inflammatory effect on different models of rodent colitis which could be attributed to the association of its antibacterial and immunomodulatory properties.

Tissue-nonspecific alkaline phosphatase is activated in enterocytes by oxidative stress via changes in glycosylation.

BACKGROUND: Intestinal inflammation produces an induction of alkaline phosphatase (AP) activity that is attributable in part to augmented expression, accompanied by a change in isoform, in epithelial cells. METHODS: This study focuses on induction of AP in intestinal epithelial cells in vitro. RESULTS: Treatment with the oxidants H2O2, monochloramine, or tButOOH increases AP activity in vitro in Caco-2, HT29, and IEC18 cells. We selected IEC18 cells for further testing. Basal AP activity in IEC18 cells is of the tissue-nonspecific (bone-liver-kidney) type, as indicated by Northern and Western blot analysis. Oxidative stress augments AP activity and the sensitivity of the enzyme to levamisole, homoarginine, and heat in IEC18 cells. Increased immunoreactivity to tissue-nonspecific AP antibodies suggests an isoform shift from liver to either kidney or bone type. This effect occurs without changes at the mRNA level and is sensitive to tunicamycin, an inhibitor of N-glycosylation, and neuraminidase digestion. Saponin and deoxycholate produce similar effects to oxidants. Butyrate but not proinflammatory cytokines or LPS can induce a similar effect but without toxicity. The AP increase is not prevented by modulators of the MAPK, NF-κB, calcium, and cyclic adenosine monophosphate (cAMP) pathways, and is actually enhanced by actinomycin D via higher cell stress. CONCLUSIONS: Oxidative stress causes a distinct increase in enterocyte AP activity together with cell toxicity via changes in the glycosylation of the enzyme that correspond to a shift in isotype within the tissue-nonspecific paradigm. We speculate that this may have physiological implication for gut defense.

New insights into the immunological effects of food bioactive peptides in animal models of intestinal inflammation.

Bioactive peptides have proven to be active in several conditions, including inflammatory bowel disease (IBD). This is a chronic and relapsing condition of unknown aetiology that comprises chiefly ulcerative colitis and Crohn's disease. Although there are treatments for IBD, they have frequent side effects and they are not always effective; therefore there is a need for new therapies that could alleviate this condition. Two bioactive peptides present in milk (transforming growth factor-beta (TGF-beta) and casein macropeptide, also named glycomacropeptide) have been shown to have intestinal anti-inflammatory activities. In fact, TGF-beta is currently added to formulas intended for patients with IBD, and several studies indicate that these formulas could induce clinical remission. In this paper, evidence supporting the anti-inflammatory effect of TGF-beta and bovine glycomacropeptide, as well as their mechanisms of action, is reviewed, focusing on the evidence obtained in animal models.

Di-D-fructose dianhydride-enriched caramels: effect on colon microbiota, inflammation, and tissue damage in trinitrobenzenesulfonic acid-induced colitic rats.

In the present study we describe the preparation and chemical characterization of a caramel with a high (70%) content of difructose dianhydrides (DFAs) and glycosylated derivatives (DFAs). This product was obtained by thermal activation (90 degrees C) of highly concentrated (90% w/v) aqueous D-fructose solutions using the sulfonic acid ion-exchange resin Lewatit S2328 as caramelization catalyst. DFAs represent a unique family of cyclic fructans with prebiotic properties already present in low proportions (<15%) in commercial caramel. We report the antiinflammatory activity of the new DFA-enriched caramel in the trinitrobenzenesulfonic acid (TNBS) model of rat colitis, an experimental model that resembles human inflammatory bowel disease (IBD), and compare its effects with those obtained with a commercial sucrose caramel and with linear fructooligosaccharides (FOS). For this purpose, the effects on colon tissue damage, gut microbiota, short-chain fatty acid (SCFAs) production, and different inflammatory markers were evaluated. The administration of DFA-enriched caramel to colitic rats showed intestinal antiinflammatory effect, as evidenced macroscopically by a significant reduction in the extent of the colonic damage induced by TNBS. This effect was similar to that obtained with FOS in the same experimental settings, whereas commercial caramel was devoid of any significant antiinflammatory effect. The beneficial effect was associated with the inhibition of the colonic levels of the proinflammatory cytokines, tumor necrosis factor alpha (TNF alpha) and interleukin 1beta (IL-1beta), and the reduction in colonic myeloperoxidase (MPO) activity and inducible nitric oxide synthase (iNOS) expression. The DFA-enriched caramel also promoted a more favorable intestinal microbiota, increasing lactobacilli and bifidobacteria counts as well as inducing higher concentrations of SCFAs in the luminal colonic contents. These results reinforce the concept of DFAs and glycosyl-DFAs as dietary beneficial compounds with prebiotic properties and suggest that the novel DFA-enriched caramel here reported may be an interesting candidate to be explored for the dietary treatment of human IBD.

Butyrate in vitro immune-modulatory effects might be mediated through a proliferation-related induction of apoptosis.

Survival and proliferation signals are two processes closely interrelated and finely controlled in most cell types, whose deregulation may lead to carcinogenesis. In the last decade, different studies have suggested that both cellular functions are also intimately associated with other cellular activities such as differentiation and cellular activation, especially in immune cells. The aim of this study was to evaluate the effects of the short-chain fatty acid (SCFA) butyrate on the proliferation and activation state of different cell types involved in inflammatory bowel disease. We focused on intestinal epithelial cells, macrophages and T-lymphocytes, using both primary non-transformed cultures and established cell lines. The results showed that low concentrations of butyrate inhibited the proliferation of all the immune cell types tested in this work, whereas it only induced apoptosis in activated T-lymphocytes, non-differentiated epithelial cells and macrophage cell lines, but not in differentiated epithelial cells or primary macrophages. Butyrate apoptosis induction was mediated by caspase-3/7 activation. This SCFA was only able to modify cell activation, measured as expression of inflammatory cytokines, in those cell types in which apoptosis was induced. In conclusion, our results suggest a cell type-specificity of the immune-modulatory effects of butyrate based on the proliferation/activation characteristic physiology of these processes in different cells types.

The intestinal antiinflammatory agent glycomacropeptide has immunomodulatory actions on rat splenocytes.

Bovine glycomacropeptide (GMP) is an immunologically active milk peptide that is a part of the normal human diet. GMP has therapeutic value in preclinical models of intestinal inflammation, and its mechanism may be related to effects on lymphocytes. This study focuses on the actions of GMP on rat splenocytes in vitro and in vivo. Bovine serum albumin and lactoferrin were used for comparative purposes. GMP (0.01-0.1mgmL(-1)) enhanced Concanavalin A (ConA) evoked but not basal splenocyte proliferation. At 1mgmL(-1) GMP lost this effect but augmented basal TNF-alpha secretion and also iNOS and COX2 expression. IFN-gamma, IL-2 and IL-17 were not affected by GMP in quiescent splenocytes, but IL-10 was augmented at all concentrations tested. On the other hand, GMP produced a marked inhibitory effect (70%) on IFN-gamma secretion and to a lower extent (50%) also on TNF-alpha. GMP was shown to block STAT4 but not IkappaB-alpha phosphorylation. The Treg marker Foxp3 was markedly upregulated by GMP. Bovine serum albumin had some effects on splenocyte function which were of lower magnitude and not entirely coincidental, while lactoferrin had a strong antiproliferative effect, as expected, indicating a specific effect of GMP. When administered for 3 days to normal Wistar rats, GMP reproduced the Foxp3 induction effect observed previously in vitro. This was observed in splenocytes but not in thymocytes, and only when administered by the oral rather than the intraperitoneal route. Thus our results support the hypothesis that GMP may limit intestinal inflammation acting at least in part on lymphocytes.

Lack of beneficial metabolic effects of quercetin in adult spontaneously hypertensive rats.

Insulin sensitivity is partly dependent on insulin-mediated nitric oxide (NO) release and antioxidants may decrease insulin resistance by amelioring NO bioavailability. The effects of chronic therapy with the antioxidant quercetin on blood pressure, vascular function and glucose tolerance in male spontaneously hypertensive rats (SHR), a model of genetically hypertension and insulin resistance, were analyzed. Rats were divided into four groups, WKY vehicle, WKY quercetin, SHR vehicle and SHR quercetin. Animals were daily administered by gavage for four weeks: vehicle, quercetin in vehicle (10mg/kg body weight). Blood pressure was followed by tail-cuff plethysmography. Chronic quercetin treatment reduced systolic blood pressure, and significantly reduced left ventricular (-10%) and renal (-6%) hypertrophy. However, oral glucose tolerance test, homeostatic model assessment of insulin resistance, total cholesterol and triglycerides were unaffected by quercetin in both strains of rats. It also improved the blunted aortic endothelium-dependent relaxation to acetylcholine, without affecting both endothelium-dependent relaxation to insulin and endothelium-independent relaxation to sodium nitroprusside in SHR. In WKY rats, quercetin in vitro and in vivo, impaired the relaxation to insulin. Quercetin reduced both plasma malondialdehyde levels and aortic superoxide production in SHR. Furthermore, quercetin inhibited insulin-stimulated protein kinase B (Akt)- and endothelial NO synthase (eNOS) phosphorylation. In conclusion, quercetin reduced blood pressure, left ventricular and renal hypertrophy and improved NO-dependent acetylcholine relaxation. However, and despite its antioxidant effects, quercetin was unable to improve insulin sensitivity possibly through its specific interference with the insulin signalling pathway.