Microbial hydrogen economy alleviates colitis by reprogramming colonocyte metabolism and reinforcing intestinal barrier.

With the rapid development and high therapeutic efficiency and biosafety of gas-involving theranostics, hydrogen medicine has been particularly outstanding because hydrogen gas (H2), a microbial-derived gas, has potent anti-oxidative, anti-apoptotic, and anti-inflammatory activities in many disease models. Studies have suggested that H2-enriched saline/water alleviates colitis in murine models; however, the underlying mechanism remains poorly understood. Despite evidence demonstrating the importance of the microbial hydrogen economy, which reflects the balance between H2-producing (hydrogenogenic) and H2-utilizing (hydrogenotrophic) microbes in maintaining colonic mucosal ecosystems, minimal efforts have been exerted to manipulate relevant H2-microbe interactions for colonic health. Consistent with previous studies, we found that administration of hydrogen-rich saline (HS) ameliorated dextran sulfate sodium-induced acute colitis in a mouse model. Furthermore, we demonstrated that HS administration can increase the abundance of intestinal-specific short-chain fatty acid (SCFA)-producing bacteria and SCFA production, thereby activating the intracellular butyrate sensor peroxisome proliferator-activated receptor γ signaling and decreasing the epithelial expression of Nos2, consequently promoting the recovery of the colonic anaerobic environment. Our results also indicated that HS administration ameliorated disrupted intestinal barrier functions by modulating specific mucosa-associated mucolytic bacteria, leading to substantial inhibition of opportunistic pathogenic Escherichia coli expansion as well as a significant increase in the expression of interepithelial tight junction proteins and a decrease in intestinal barrier permeability in mice with colitis. Exogenous H2 reprograms colonocyte metabolism by regulating the H2-gut microbiota-SCFAs axis and strengthens the intestinal barrier by modulating specific mucosa-associated mucolytic bacteria, wherein improved microbial hydrogen economy alleviates colitis.

Immunomodulatory assessment of Portulaca oleracea L. extract in a mouse model of colitis.

Ulcerative colitis (UC) is a gastrointestinal inflammatory disease with a multifactorial pathophysiology. This study aims to investigate the immunomodulatory effect of Portulaca oleracea leaf ethanolic extract (POE) on acetic acid (AA)-induced UC in mice. Experimental animals received oral doses of POE (200 mg/kg for 7 days) after an induction of colitis by intrarectal AA administration. In mice with AA-induced UC treated with POE, the results revealed a significant modulation in body weight and colon length. Moreover, treatment with POE downregulated the interleukin 1, 6, and 17, tumor necrosis factor-alpha, gamma interferon, and nuclear factor-kappa B levels compared with the colitis group. Furthermore, POE markedly inhibited histological damage, decreased myeloperoxidase activity and reduced fecal calprotectin level compared with the colitis group. These data are consistent with the reduction in total bacterial content in the colon. Taken together, treatment with POE may reduce colonic inflammation by alleviating the immune response and inhibiting the severity of colitis. The HPLC analysis of POE resulted in the identification of seven medicinal compounds comprising two phenolic acids (ferulic and caffeic acids) and five flavonoids (kaempferol, quercetin, rutin, narenginin and hesperidin). Subsequent analysis of POE by GC-MS revealed ten phytocomponents; the major percentages were hexadecenoic acid, methyl ester (29.8119%), α-linolenic acid (25.8431%), 16-octadecenoic acid, methyl ester (15.1578%) and α-tocopherol (10.7848%). Delta-lactams and alkanes were the minor components. Such natural plant-derived substances and their probable synergistic action appear to contribute to a promising therapeutic protocol for colitis.

1H NMR-based metabonomic assessment of probiotic effects in a colitis mouse model.

Metabolic profiling of the fecal extracts of male mice was carried out to assess the effects of probiotics on colonic inflammation using (1)H NMR spectroscopy coupled with multivariate data analysis. The control group (n = 5) was administered phosphate buffered saline for 14 days. Acute colitis was induced with dextran sulfate sodium (DSS) for 7 days following administration of phosphate buffered saline for 7 days (DSS-treated group, n = 5). LAB + DSS-treated group (n = 5) was administered lactic acid bacteria (LAB) daily for 7 days followed by treatment with DSS for 7 days to investigate protective effect of LAB against DSS-inducible colitis. Histological damage, myeloperoxidase activity, and malondialdehyde content of colon tissue were reduced, whereas colon length increased in LAB + DSS-treated mice compared to those in DSS-treated mice. DSS treatment was associated with fecal excretion of amino acids, short chain fatty acids, and nucleotides, revealing significant decreases of threonine, alanine, glutamate, glutamine, aspartate, lysine, glycine, butyrate, uracil, and hypoxanthine together with increases of monosaccharides, glucose, and trimethylamine in the feces of mice with DSS-induced colitis. Increased levels of acetate, butyrate, and glutamine and decreased levels of trimethylamine were found in the feces of LAB + DSS-treated mice compared to DSS-treated mice alone. The increased short chain fatty acids levels in the feces of mice fed with LAB indicate that the probiotics have protective effects against DSS-induced colitis via modulation of the gut microbiota. This work highlights the possibility for alternative approach of metabonomics in feces for assessing the probiotic effect in an animal model of inflammatory bowel disease.

Metabolic assessment of gradual development of moderate experimental colitis in IL-10 deficient mice.

Evidence has linked genetic predisposition and environmental exposures to the worldwide pandemic of inflammatory bowel diseases (IBD), but underlying biochemical events remain largely undefined. Here, we studied the gradual development of colitis in Interleukin 10 deficient mice using a combination of (i) histopathological analysis of intestinal sections, (ii) metabolic profiling of blood plasma, and (iii) measurement of plasma inflammatory biomarkers. Data integration using chemometric tools, including Independent Component Analysis, provided a new strategy for measuring and mapping the metabolic effects associated with the development of intestinal inflammation at the age of 1, 8, 16, and 24 weeks. Chronic inflammation appeared at 8 weeks and onward, and was associated with altered cecum and colon morphologies and increased inflammatory cell infiltration into the mucosa and the submucosa. Blood plasma profiles provided additional evidence of loss of energy homeostasis, impaired metabolism of lipoproteins and glycosylated proteins. In particular, IL-10-/-mice were characterized by decreased levels of VLDL and increased concentrations of LDL and polyunsaturated fatty acids, which are related to the etiology of IBD. Moreover, higher levels of lactate, pyruvate, citrate and lowered glucose suggested increased fatty acid oxidation and glycolysis, while higher levels of free amino acids reflected muscle atrophy, breakdown of proteins and interconversions of amino acids to produce energy. These integrated system investigations demonstrate the potential of metabonomics for investigating the mechanistic basis of IBD, and it will provide novel avenues for management of IBD.

Immunohistochemical assessment of mucosal cytokine profile in acetic acid experimental colitis.

UNLABELLED: Experimental colitis induced by acetic acid has been used extensively as a model for intestinal inflammatory disease. Colonic tissue lesions of intestinal inflammatory disease patients seem to be related to the increased local production of pro-inflammatory cytokines (IL-1, IL-6, TNF-alpha, and IFN-gamma). PURPOSE: To assess the cytokine expression pattern identified through immunohistochemistry in colonic mucosa after experimental colitis induced by acetic acid and establish the relationship between this pattern and the presence of macroscopic lesions. MATERIALS AND METHODS: Adult male Wistar rats (n = 39) were divided at random into 4 groups: NC45 and NC24 (control without colitis; sacrificed at 45 minutes and 24 hours, respectively); and WC45 and WC24 (with experimental colitis induced by acetic acid; sacrificed at 45 minutes and 24 hours, respectively). Macroscopic and microscopic alterations in colonic tissue were evaluated, and cytokine expression was assessed through immunohistochemistry. RESULTS: After 24 hours, IL-1 expression was greater in the groups with colitis when compared to the groups without colitis. IL-4 expression was higher in the WC45 group. There was an increase in both INF-gamma and IL-6 related to the presence of necrosis of the colonic mucosa in the groups with colitis for both periods evaluated. CONCLUSION: The immunohistochemical technique was efficient for the analysis of various cytokine expressions in the colonic tissue. There was an increase in the IL-1 pro-inflammatory cytokines as well as in IL-6 and IFN-gamma associated with the presence of colonic necrosis. Experimental colitis induced by acetic acid is a useful model for the development of studies assessing the role of cytokines in the inflammation of mucosa as well as anti-cytokine therapies.

Immunohistochemical assessment of mucosal cytokine profile in acetic acid experimental colitis

O modelo de colite experimental induzida por ácido acético (CEAA) vem sendo extensamente utilizado em estudos sobre doenças inflamatórias intestinais (DII). Lesões no tecido colônico em portadores de DII parecem estar relacionados à produção local aumentada de citocinas pró-inflamatórias (IL-1, IL-6, TNF-a e IFN-g). OBJETIVO: Avaliar o padrão de expressão de citocinas identificadas por imunohistoquímica em tecido colônico após CEAA e relacioná-lo à presença de lesões macroscópicas. MATERIAL E MÉTODOS: Ratos machos Wistar adultos (n=39) foram submetidos ou não à CEAA e sacrificados para retirada do tecido colônico em dois períodos distintos, perfazendo 4 grupos aleatórios: SC45 e SC24 (sem colite; sacrifício 45 minutos e 24 horas, respectivamente); CC45 e CC24 (com colite; sacrifício 45 minutos e 24 horas, respectivamente). Avaliaram-se alterações macro e microscópicas do cólon e sua expressão de citocinas foi avaliada por imunohistoquímica. RESULTADOS: Após 24 horas, a expressão de IL-1 foi maior no grupo com colite, em relação ao sem colite. IL-4 foi mais expressa no grupo CC45. Houve aumento de INF-g e IL-6, relacionados à presença de necrose da mucosa colônica, nos grupos com colite, em ambos os períodos avaliados. CONCLUSÃO: A técnica de imunohistoquímica foi eficiente para a análise da expressão de citocinas na mucosa colônica. Houve aumento da expressão das citocinas pró-inflamatórias IL-1 e de IL-6 e IFN-g associado à presença de necrose colônica. A CEAA é um bom modelo para o desenvolvimento de estudos destinados a avaliar o papel das citocinas na inflamação da mucosa e terapias anti-citocinas.

Colitis increases albumin synthesis at the expense of muscle protein synthesis in macronutrient-restricted piglets.

Our aim was to examine the effect of acute inflammation localized in the colon and early macronutrient restriction on protein synthesis in a piglet model. In a 2 x 2 factorial design, piglets (n = 32) were fed an adequate or macronutrient-restricted diet with or without dextran sulfate-induced colitis for 7 d. The stable isotope tracer L-[5,5,5-(2)H(3)]leucine was infused to determine protein kinetics at the whole-body level and synthesis of tissue and plasma proteins. In the well-nourished state, colitis did not affect weight gain or protein kinetics except for an increase in albumin synthesis (P < 0.05). Macronutrient restriction alone caused a general slowing of protein metabolism including decreased weight gain (P < 0.0004), whole-body protein turnover (P < 0.0001), and liver (P < 0.01) and plasma protein (P < 0.03) synthesis. However, in the presence of macronutrient restriction, colitis compromised weight gain further (P < 0.02) and decreased muscle protein synthesis (P < 0.05) due to a redistribution of protein metabolism that supported enhanced synthesis of plasma proteins. The increased contribution of plasma protein synthesis to whole-body protein turnover was attributable mainly to increased synthesis of albumin (P < 0.006). Concentrations of plasma proteins were unaffected despite dramatic changes in their synthesis rates, thereby underestimating the effects of malnutrition and colitis on protein metabolism. Increased synthesis of plasma proteins, particularly the negative acute phase reactant albumin, compromises weight gain and muscle protein synthesis only when macronutrient intake is inadequate, underscoring the role of adequate nutrition in preventing growth impairment and muscle wasting in acute inflammation. These results suggest that the hypoalbuminemia of inflammatory bowel disease should not be attributed to decreased synthesis.

Antioxidant effects of aminosalicylates and potential new drugs for inflammatory bowel disease: assessment in cell-free systems and inflamed human colorectal biopsies.

BACKGROUND: The therapeutic efficacy of 5-aminosalicylic acid in inflammatory bowel disease may be related to its antioxidant properties. AIM: To compare in vitro the antioxidant effects of conventional drugs (5-aminosalicylic acid, corticosteroids, metronidazole), with new aminosalicylates (4-aminosalicylic acid, balsalazide) and other potential therapies (ascorbate, N-acetylcysteine, glutathione, verapamil). METHODS: Compounds were assessed for efficacy in reducing the in vitro production of reactive oxygen species by cell-free systems (using xanthine/xanthine oxidase, with or without myeloperoxidase) and by colorectal biopsies from patients with ulcerative colitis using luminol-amplified chemiluminescence. RESULTS: 5-aminosalicylic acid and balsalazide were more potent antioxidants than 4-aminosalicylic acid or N-acetyl-5-aminosalicylic acid in cell-free systems. 5-aminosalicylic acid (20 mM) and balsalazide (20 mM) inhibited rectal biopsy chemiluminescence by 93% and 100%, respectively, compared with only 59% inhibition by 4-aminosalicylic acid (20 mM). Hydrocortisone, metronidazole and verapamil had no significant effect on chemiluminescence in any system. Ascorbate (20 mM) inhibited chemiluminescence by 100% in cell-free systems and by 60% in rectal biopsies. N-acetyl cysteine (10 mM), and both oxidized and reduced glutathione (10 mM), completely inhibited chemiluminescence in cell-free systems, but not with rectal biopsies. CONCLUSIONS: The antioxidant effects of compounds varies between cell-free systems and inflamed colorectal biopsies. The effect of drugs on the chemiluminescence produced by these two assay systems is useful for screening potentially new antioxidant treatments for inflammatory bowel disease. Ascorbate seems worth further study as a novel therapy.

[Energy expenditure of patients with Crohn's disease. Course study during hospitalization]. / Consumo de energía en pacientes con enfermedad de Crohn. Estudio evolutivo durante la hospitalización.

Weight loss and protein energy malnutrition are frequent in Crohn's disease. The increase of resting energy expenditure has been pointed out as the cause of these findings. In this study we report eleven patients with Crohn's disease, six women and five men hospitalized with active Crohn's disease. In three patients the disease involved the small bowel, in five the large bowel and three shared large and small bowel involvement. At the beginning of the hospitalization the activity disease index of Van Hees was 196 +/- 52, range: 132-265. The resting energy expenditure was eleven percent higher than that, of a healthy population (p: n.s). During hospitalization the energy expenditure decreased weekly with statistically significant difference. No relation has been observed between the activity index of Van Hees, and any of the energy parameters studied. Patients with body weight lower 90% of ideal weight, had an increased resting energy expenditure when that was expressed in kcal/kg of weight (p = 0.003). Fever was the sole parameter analyzed with statistically significant relation with consumption of oxygen: with the index of oxygen consumption (p = 0.03) and with the percentage of resting energy expenditure (p = 0.006). In summary, the REE in active Crohn's disease is higher than that of healthy population, although without statistically significance. The REE tends to normalization coinciding with the decreasing of inflammatory activity. Increased energy expenditure has been detected in weight loss patients.

Faecal alpha-1-antitrypsin and excretion of 111indium granulocytes in assessment of disease activity in chronic inflammatory bowel diseases.

Intestinal protein loss in chronic inflammatory bowel diseases may be easily determined by measurement of alpha-1-antitrypsin (alpha 1-AT) stool concentration and alpha 1-AT clearance. Both parameters were significantly raised in 36 and 34 patients respectively with chronic inflammatory bowel diseases, compared with eight patients with non-inflammatory bowel diseases, or 19 healthy volunteers. There was wide range of overlap between active and inactive inflammatory disease. Contrary to serum alpha 1-AT, faecal excretion and clearance of alpha 1-AT did not correlate with ESR, serum-albumin, orosomucoid, and two indices of disease activity. A comparison of alpha 1-AT faecal excretion and clearance with the faecal excretion of 111In labelled granulocytes in 27 patients with chronic inflammatory bowel diseases, showed no correlation between the intestinal protein loss and this highly specific marker of intestinal inflammation. Enteric protein loss expressed by faecal excretion and clearance of alpha 1-AT does not depend on mucosal inflammation only, but may be influenced by other factors.