Contribution of NO synthases to neutrophil infiltration in the gastric mucosal lesions in rats with water immersion restraint stress

AbstractA decrease in constitutive NO synthase (cNOS) activity and an increase in inducible NO synthase (iNOS) activity occurred with an increase in myeloperoxidase (MPO) activity, an index of neutrophil infiltration, in the gastric mucosa of rats with water immersion restraint (WIR) stress. This increase in gastric mucosal MPO activity was enhanced by pretreatment with NG-monomethyl l-arginine, a non-selective NOS inhibitor, but was prevented with maintenance of gastric mucosal cNOS activity by pretreatment with aminoguanidine, a selective iNOS inhibitor. The MPO activity was negatively correlated with the cNOS activity in all WIR-stressed rats used (r=−0.723). These results suggest that a decrease in cNOS activity could contribute to an increase in neutrophil infiltration in the gastric mucosa of WIR-stressed rats

Calcium Pyruvate Exerts Beneficial Effects in an Experimental Model of Irritable Bowel Disease Induced by DCA in Rats

Pyruvate is a normal constituent of the body that participates in carbohydrate metabolism and functions as a scavenger of free radicals. Calcium pyruvate monohydrate (CPM) is a more stable derivative that has proved its anti-inflammatory effect in experimental colitis, among other disorders, and that could also be considered a source of calcium. Thus, it would be useful for the treatment of diseases with an inflammatory component and a high prevalence of osteoporosis like the irritable bowel syndrome (IBS). The aim of the present study is to evaluate the effects of CPM in a rat model of chronic post-inflammatory visceral pain induced by deoxycholic acid (DCA) that resembles IBS. Rats were administered DCA for three days intracolonically and then treated daily with CPM (40 and 100 mg/kg) or gabapentin (70 mg/kg) (positive control) by oral gavage for 17 days. The treatments reduced the visceral hypersensitivity measured by response to colorectal distension and referred pain. DCA induced changes in the colonic immune response characterized by increased expression of the cytokine Il-1b and the inducible enzyme Cox-2, which was reduced by the treatments. DCA also decreased the gut expression of the mucins Muc-2 and Muc-3, which was normalized by CPM, whereas gabapentin only increased significantly Muc-3. Moreover, DCA increased the expression of Tlr3, which was decreased to basal levels by all the treatments. However, the serotonin receptor Htr-4, which was also elevated, was not affected by any of the treatments, indicating no effect through this signalling pathway. In conclusion, CPM ameliorated the visceral hypersensitivity and the referred pain caused by DCA, being as effective as the control drug. Furthermore, it improved the immune status of the animals, which could contribute to the visceral analgesia and the regeneration of the intestinal epithelial barrier integrity.This work was supported by the Junta de Andalucía (CTS 164) and by the Spanish Ministry of Economy and Competitiveness (AGL2015-67995-C3-3-R) with funds from the European Union. The CIBER-EHD is funded by the Instituto de Salud Carlos III

The Antioxidant Properties of Salvia verbenaca Extract Contribute to Its Intestinal Antiinflammatory Effects in Experimental Colitis in Rats

La enfermedad inflamatoria intestinal (EII) es una inflamación gastrointestinal crónica con fluctuaciones de síntomas impredecibles. Si bien no existe una cura eficaz para la EII, varios tratamientos tienen como objetivo controlar los síntomas y mejorar la calidad de vida de las personas afectadas. En los últimos años, ha habido un interés creciente en los beneficios potenciales de ciertas plantas y hierbas naturales en el tratamiento de la EII. En este sentido, este estudio tuvo como objetivo evaluar los efectos inmunomoduladores y antiinflamatorios de un extracto bien caracterizado de Salvia verbenaca (S. verbenaca) en un modelo experimental de colitis en ratas. Curiosamente, la administración diaria de S. verbenaca (10 y 25 mg/kg) alivió eficazmente los síntomas de la colitis, como lo demuestra la reducción de la relación peso/longitud y el daño colónico. Además, redujo los marcadores de estrés oxidativo (MPO y GSH), disminuyó la expresión de citocinas proinflamatorias (Il-6, Il-12a, Il-1β, Il-23, Icam-1, Mcp-1, Cinc-1) y conservó la integridad de la barrera intestinal (Villin, Muc-2, Muc-3). Estos efectos sugieren que el extracto de S. verbenaca podría representar un potencial candidato complementario para tratar los trastornos gastrointestinales. Sus acciones beneficiosas pueden estar relacionadas con sus propiedades antioxidantes, así como con la regulación negativa de la respuesta inmune, lo que puede resultar en la mejora de la barrera epitelial del intestino.Inflammatory bowel disease (IBD) is a chronic gastrointestinal inflammation with unpredictable symptom fluctuations. While there is no effective cure for IBD, various treatments aim to manage symptoms and improve the quality of life for affected individuals. In recent years, there has been growing interest in the potential benefits of certain natural plants and herbs in the management of IBD. In this regard, this study aimed to evaluate the immunomodulatory and anti-inflammatory effects of a well-characterized extract of Salvia verbenaca (S. verbenaca) in an experimental model of colitis in rats. Interestingly, the daily administration of S. verbenaca (10 and 25 mg/kg) effectively alleviated colitis symptoms, as evidenced by reduced weight/length ratio and colonic damage. Moreover, it reduced oxidative stress markers (MPO and GSH), decreased pro-inflammatory cytokine expression (Il-6, Il-12a, Il-1β, Il-23, Icam-1, Mcp-1, Cinc-1), and preserved the integrity of the intestinal barrier (Villin, Muc-2, Muc-3). These effects suggest S. verbenaca extract could represent a potential complementary candidate to treat gastrointestinal disorders. Its beneficial actions can be related to its antioxidant properties as well as the downregulation of the immune response, which can result in the improvement in the intestine epithelial barrier.Junta de Andalucia (AGR-6826)Junta de Andalucia (CTS-164)Ministerio de Economía y Competitividad (SAF2011-29648)Instituto de Salud Carlos III (pFIS (FI20/00159)Instituo de Saud Carlos III (Miguel Servet CP22/00153

Limosilactobacillus fermentum CECT5716: Mechanisms and Therapeutic Insights

This work was supported by the Junta de Andalucía (CTS 164) and Instituto de Salud Carlos III (PI19/01058) with funds from the European Union.M.J. Rodríguez-Sojo is a predoctoral fellow from University of Granada (“Programa de Doctorado en Biomedicina”); A.J. Ruiz-Malagón is a predoctoral fellow from Formación de Profesorado Universitario Program (“Programa de Doctorado en Medicina Clínica y Salud Pública”), and A. Rodríguez-Nogales is a postdoctoral fellow of Instituto de Salud Carlos III (Miguel Servet Program).Probiotics microorganisms exert their health-associated activities through some of the following general actions: competitive exclusion, enhancement of intestinal barrier function, production of bacteriocins, improvement of altered microbiota, and modulation of the immune response. Among them, Limosilactobacillus fermentum CECT5716 has become one of the most promising probiotics and it has been described to possess potential beneficial effects on inflammatory processes and immunological alterations. Different studies, preclinical and clinical trials, have evidenced its anti-inflammatory and immunomodulatory properties and elucidated the precise mechanisms of action involved in its beneficial effects. Therefore, the aim of this review is to provide an updated overview of the effect on host health, mechanisms, and future therapeutic approaches.Junta de Andalucia CTS 164Instituto de Salud Carlos III European Commission PI19/01058European Commissio

Relationship between Changes in Microbiota and Liver Steatosis Induced by High-Fat Feeding-A Review of Rodent Models

Several studies have observed that gut microbiota can play a critical role in nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH) development. The gut microbiota is influenced by different environmental factors, which include diet. The aim of the present review is to summarize the information provided in the literature concerning the impact of changes in gut microbiota on the effects which dietary fat has on liver steatosis in rodent models. Most studies in which high-fat feeding has induced steatosis have reported reduced microbiota diversity, regardless of the percentage of energy provided by fat. At the phylum level, an increase in Firmicutes and a reduction in Bacteroidetes is commonly found, although widely diverging results have been described at class, order, family, and genus levels, likely due to differences in experimental design. Unfortunately, this fact makes it difficult to reach clear conclusions concerning the specific microbiota patterns associated with this feeding pattern. With regard to the relationship between high-fat feeding-induced changes in liver and microbiota composition, although several mechanisms such as alteration of gut integrity and increased permeability, inflammation, and metabolite production have been proposed, more scientific evidence is needed to address this issue and thus further studies are needed.This research was funded by MINECO (AGL-2015-65719-R-MINECO/FEDER, UE) and Instituto de Salud Carlos III (CIBERobn, CB12/03/30007

Spontaneous Luteolysis in the Cyclic Golden Hamster: Evidence for Mediation Through Inflammation and Apoptosis

Mammalian corpora lutea (CL) are transient ovarian glands which function to support pregnancy by secretion of progesterone (P). Sustained P secretion during pregnancy in the hamster requires gonadotrophic support, i.e., follicle stimulating hormone (FSH) and prolactin (Prl). Infertile cycles require CL regression. In luteolysis, CL first regress functionally (FL) and then structurally (SL). FL permits the next estrous. Estrous occurs every fourth day in the hamster such that CL form, function briefly, and then regress rapidly and completely. With only one generation of CL present in each ovary at any one time, the hamster is an ideal model for studying luteolysis. FL, begins on day three as P secretion plummets, and is followed immediately by SL as neutrophils —cellular markers of inflammation— invade the CL. SL proceeds via apoptosis and is not reversible with exogenous FSH-Prl. Apoptosis is synonymous with physiological cell death, and, as a rule, evokes no inflammation. In fact, it may have evolved to prevent inflammation and the excessive tissue damage which often accompanies pathologic cell death. Since apoptosis and inflammation coexist in luteolysis, experiments are designed in the present study to help explain this paradox. Light and transmission electron microscopic observations of ovarian sections chronicled cyclic luteal morphology. Enzyme histochemistry was conducted on ovarian sections for lysosomal nonspecific esterase activity. Luteal and nonluteal ovarian samples were probed for neutrophilic myeloperoxidase by immunoblot. Microscopic observations revealed that some luteal cell organelles atrophy in FL prior to the apoptosis seen in SL. Nonspecific esterase staining showed a dramatic increase in autolytic lysosomal activity during SL and follicular atresia. Immunoblots showed that myeloperoxidase is present at the onset of SL. The findings indicate that luteolysis is a dynamic process which may require a tightly regulated acute inflammatory response for rapid completion. Neutrophils may play a role in the abrupt onset of cell death required for SL. It is possible that in hamster luteolysis the damaging sequalae of chronic inflammation is prevented by apoptosis in which viable luteal cells are transformed into phagocytes that clear the gland of apoptotic cellular debris

Analysis of antioxidants mechanisms in the antiulcerogenic activity of Anacardium humile St. Hil. (Anacardiaceae)

Orientador: Alba Regina Monteiro Souza BritoDissertação (mestrado) - Universidade Estadual de Campinas, Instituto de BiologiaResumo: Anacardium humile, popularmente conhecida como cajuzinho-do-cerrado, é utilizada na medicina tradicional para o tratamento de diversas inflamações. Extratos e frações de suas folhas apresentam compostos fenólicos e significativa atividade anti-úlcera. Nesse trabalho, avaliou-se possíveis mecanismos antioxidantes na gastroproteção conferida pela fração acetato de etila das folhas de A. humile (AHFAc). AHFAc é rica em compostos fenólicos, há mais de 30% de fenólicos solúveis nessa fração, segundo resultados do ensaio de Folin-Ciocalteu. AHFAc apresenta atividade redutora em ensaio de redução de 1,1-difenil-1-picril-hidrazila (DPPH), o que representa ação sequestradora de radicais livres em potencial. Foram registrados o perfil cromatográfico de AHFAc, em cromatografia líquida de alta eficiência e o espectro de massas com ionização por electrospray, que indicam a presença de ácido gálico e/ou derivados, quercetina, kaempferol e amentoflavona (um biflavonóide).A atividade farmacológica de AHFAc foi analisada em modelos de indução de úlcera gástrica por isquemia e reperfusão e por etanol absoluto. Ratos Unib: WH foram tratados com o veículo Tween 80® 12% (10 ml.kg-1) ou AHFAc (25, 50 e 100 mg.kg-1) e submetidos à isquemia (30 minutos) e reperfusão (60 minutos) do estômago ou à administração de 1 ml de etanol absoluto. Um grupo SHAM foi formado por animais não tratados, expostos aos procedimentos experimentais, mas sem efetiva indução de úlcera. Após a realização dos modelos de indução de úlcera gástrica, o estômago dos animais foi removido, a área de lesão ulcerativa determinada, uma porção do estômago foi fixada para análises histológicas (colorações hematoxilina-eosina e ácido periódico de Schiff) e imunohistoquímicas (reação para mieloperoxidase e superóxido dismutase) e o restante da porção glandular do estômago foi raspado e homogeneizado para ensaios bioquímicos. Foram dosados os níveis de grupamentos sulfidrila (G-SH), fragmentação de DNA, atividade das enzimas mieloperoxidase (MPO), glutationa peroxidase (GPx), glutationa redutase (GR) e superóxido dismutase (SOD). O pré-tratamento com AHFAc (50 mg.kg-1) apresenta efeito gastroprotetor nos dois modelos experimentais de úlcera gástrica analisados, com manutenção da integridade da mucosa e tendência de aumento dos níveis de muco. AHFAc evitou o aumento da atividade de MPO na mucosa gástrica de ratos submetidos aos dois modelos experimentais investigados, o que indica menor infiltração de neutrófilos no estômago dos animais tratados com a fração. Além disso, a administração de AHFAc também foi eficaz em manter os níveis normais de G-SH e fragmentação de DNA bem como da atividade da SOD e GPx na mucosa gástrica exposta à ação lesiva do etanol absoluto. A ação antiulcerogênica de AHFAc deve envolver atividade antioxidante, garantida pela sua composição fenólica, aliada a outros mecanismos de ação. Uma vez que não foi observada modulação na atividade das enzimas antioxidantes, e por apresentar capacidade redutora do DPPH, a atividade antioxidante de AHFAc provavelmente envolve sequestro de radicais livres.Abstract: Anacardium humile, popularly known as "cajuzinho-do-cerrado", is used in traditional medicine for the treatment of various inflammatory diseases. Extracts and fractions of leaves of A. humile present phenolic compounds and significant antiulcer activity. In this study, we evaluated the possible antioxidant mechanisms in the antiulcerogenic activity conferred by the ethyl acetate fraction of leaves of A. humile (AHFAc). AHFAc is rich in phenolic compounds; there are more than 30% of soluble phenolic compounds in this fraction, according to results from the Folin Ciocalteu assay. AHFAc shows antioxidant activity in the 1, 1-diphenyl-1-picryl picrylhydrazyl (DPPH) assay, which represents a potential free radical scavenging action. There have been recorded the chromatographic profile AHFAc in high performance liquid chromatography and the mass spectrum by electrospray ionization mass spectrometry, which indicate the presence of gallic acid and derivatives, quercetin, kaempferol and amentoflavone (biflavonoid).The pharmacological activity of AHFAc was examined in ischemia and reperfusion induced and absolute ethanol-induced gastric ulcer models. Unib: WH rats were treated with the vehicle Tween 80 12% (10 ml.kg-1) or AHFAc (25, 50 and 100 mg.kg-1) and underwent ischemia (30 minutes) and reperfusion (60 minutes) of stomach or the administration of 1 ml of absolute ethanol. SHAM group was formed by untreated animals exposed to experimental procedures, but without effective induction of ulcer. After experimental models, the stomach of the animals was removed, the ulcerative lesion area (ULA) determined, a portion of the stomach was fixed for histological analysis (hematoxylin-eosin staining and periodic acid Schiff staining) and immunohistochemistry analysis (myeloperoxidase and superoxide dismutase), the remaining portion of the glandular stomach was scraped and homogenized for biochemical assays. We measured the levels of sulfhydryl groups (G-SH), DNA fragmentation, activity of myeloperoxidase (MPO), glutathione peroxidase (GPx), glutathione reductase (GR) and superoxide dismutase (SOD) enzymes. The pre-treatment with AHFAc (50 mg.kg-1) showed gastroprotective effect in both experimental models of gastric ulcer, while maintaining the integrity of the mucosa and a tendency for increased levels of mucus. The administration of AHFAc was also effective in maintaining normal levels of G-SH and fragmentation of DNA, SOD and GPx activities in the gastric mucosa exposed to the harmful effect of absolute ethanol. In addition, AHFAc prevented the increase of MPO activity in gastric mucosa of rats subjected to the experimental models investigated, indicating less infiltration of neutrophils in the stomach of animals treated with the fraction. The anti-ulcer action of AHFAc must involve antioxidant activity, due to its phenolic composition, combined with other mechanisms of action. Since there was no modulation in the activity of antioxidant enzymes, and for presenting reduction capacity of DPPH, the antioxidant activity of AHFAc probably involves scavenger activity of free radicals.MestradoFisiologiaMestre em Biologia Funcional e Molecula

Neutrophils and vascular reactivity in ischaemia/reperfusion

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