Funcionalidad del aceite de oliva virgen extra en la artritis reumatoide experimental

Falta palabras clavesIntroduction Rheumatoid arthritis (RA) is a chronic systemic inflammatory disease characterized by inflammation of the synovial membrane and progressive destruction of the articular cartilage and bone. (Salgado and Maneiro 2014). RA patients exhibit an inflammatory chronic condition, which usually affects symmetrically arthrodial and small joints of hands and feet Global prevalence of RA has been estimated to be around 0.5-1.0% of adults in developed countries with a large variation across regions and approximately three-times more common in the female gender. The disease may begin at any age, but around 80% of all patients initiate the disease between the ages of 35 and 50 years (Rudan et al. 2015). Although the specific triggers and exact mechanisms of tissue damage in RA is still unknown, an increase in inflammatory mediators as well as a dysregulation of the immune system with uncontrolled T cell activity, play a remarkable role in its pathogenesis. Pharmacological treatment in AR including nonsteroidal anti-inflammatory drugs (NSAIDs), glucocorticoids, disease-modifying antirheumatic drugs (DMARDs) and biological agents have improved the signs and symptoms of RA, but these drugs are only effective in a fraction of patients and have other limitations including a high cost, the requirement for parenteral administration and important side effects. Therefore, new therapeutic strategies are under investigation including nutritional therapy. The beneficial effects of the Mediterranean diet have been proven not only in cardiovascular diseases but also in diabetes, obesity, arthritis and cancer (Cardeno, Sanchez-Hidalgo, and Alarcon-de-la-Lastra 2013). Evidence points out that Mediterranean diet decreases both pain and disease activity leading to better outcomes, and decreasing the doses of anti-inflammatory drugs, which exhibit important secondary effects (Smedslund et al. 2010). Olive Oil is the characteristic culinary fat of the Mediterranean area being described as a key bio-active food (Puertollano et al. 2010). Extra virgin olive oil (EVOO) is obtained from the fruit of the olive tree (Olea europea L.) solely by mechanical or other physical means under conditions that do not alter its natural composition. Traditionally the beneficial effects of EVOO have been ascribed to its monounsaturated fatty acid (MUFA) (Bermudez et al. 2011). However, a wide range of evidence indicates that many of the beneficial effects of EVOO intake are due to its minor highly bioactive components (about 1–2 % of oil weight) (Alarcon de la Lastra et al. 2001) Among them, phenolic compounds such as hydroxytyrosol, tyrosol and oleuropein have shown anti-inflammatory and antioxidant effects (Omar et al. 2010). Current experimental studies support a beneficial role of polyphenols from EVOO in several inflammatory diseases, including RA (Martinez-Dominguez, de la Puerta, and Ruiz-Gutierrez 2001; Impellizzeri et al. 2011; Gong et al. 2009). Although EVOO has demonstrated anti-inflammatory effects, it has not reported so much evidence of its possible immunomodulatory effects. Therefore, the objectives of this thesis were: 1. To determinate the possible protective effect of dietary extra virgin olive oil (EVOO) in collagen-induced arthritis (CIA) in DBA 1/J mice, an experimental model of RA and explore the biochemical routes and possibly intracellular signalling pathways. 2. To evaluate the oral polyphenolic extract from EVOO treatment in murine collageninduced arthritis and study the biochemical routes and signalling pathway involved. 3. To investigate the effects of hydroxytyrosol (HTy) or hydroxytyrosol acetate (HTy- Ac), polyphenolics compounds from EVOO enriched-diets in experimental arthritis model in mice and elucidate the molecular mechanisms and signalling pathways involved. Results and Discussion 1. Dietary extra-virgin olive oil prevents inflammatory response and cartilage matrix degradation in murine collagen‑induced arthritis. (Rosillo et al. Eur J Nutr. 2015 In press) Three-week-old male DBA-1/J mice were randomized into four experimental groups: (1) Sham sunflower diet (SO-Sham) group received a diet elaborated with a marketable sunflower oil; (2) CIA sunflower diet (SO-CIA) group received a diet elaborated with a marketable sunflower oil; (3) Sham EVOO diet (EVOO-Sham) group were fed with a diet made with a marketable EVOO picual variety and (4) CIA EVOO diet (EVOO-CIA) group were fed with a diet made with a marketable EVOO picual variety. After 6 weeks, arthritis was induced by type II collagen. Experiments followed a protocol approved by the Animal Ethics Committee of the University of Seville, and all experiments were in accordance with the recommendations of the European Union regarding animal experimentation (Directive of the European Counsel 2010/630/EU) Our results revealed, that EVOO, as the lipid component of the diet, effectively exhibited preventive and therapeutic effects in the development of inflammatory arthritis and joint damage in CIA arthritic mice in comparison with those CIA mice fed with SO. This effect was correlated to an improved arthritis score, a minor inflammatory cells infiltration into articular tissues, reduced exudation into the synovial space, synovial hyperplasia and cartilage erosion. Overexpression of pro-inflammatory cytokines, such as IL-1β, TNF-α and IL-17 may activate osteoclasts and macrophages and recruit leukocytes in inflamed joints. Besides, it is well-known that IL-17 is able to induce the release of IL-8 and IL-6, and plays a remarkable role in the additive/synergistic effects induced by TNF-α and IL-1β (Jeong et al. 2004). Our results indicate that animals fed with EVOO diet showed a significant reduction in IL-1β, TNF-α and IL-17 proinflammatory cytokines levels in paw homogenates. Cartilage oligomeric matrix protein (COMP) is a matrix protein with a great potential as a biological marker of cartilage metabolism in arthritis (Saxne and Heinegard 1992). In addition, COMP is a putative substrate for metalloproteinases (MMPs). Particularly, MMP-3 is a proteinase secreted by synovial fibroblasts and chondrocytes and its activity results in degradation of aggrecan core protein, cartilage link protein, fibronectin and collagen. Our data showed that the production of both cartilage (COMP) and synovial (MMP-3) biomarkers was significantly inhibited by dietary EVOO treatment in CIA mice. Abnormal signalling pathways play an important role in the inflammatory process and can lead to a dysregulation of the inflammatory response being crucial in RA pathogenesis. Nuclear factor κB (NF-κB) is a crucial transcriptional activator for the expression of multiple proinflammatory genes involved in the microenvironment of the arthritic joints, playing an important role in the development of RA. (Morel and Berenbaum 2004; Okamoto et al. 2010). Our results suggested that dietary EVOO treatment suppressed NF-κB activation in CIA-induced arthritic mice. Similarly, the mitogen-activated protein kinase (MAPK) family also plays critical roles in RA pathogenesis (Han et al. 2001; Suzuki et al. 2000) regulating cytokine production, and activating the janus kinase-signal transducer and activator of transcription (JAK-STAT) signalling pathway through STAT-3 phosphorylation (Aaronson and Horvath 2002). Our results demonstrated that EVOO diet intake reduced significantly both MAPKs and STAT-3 activation. On the other hand, nuclear factor E2-related factor 2 (Nrf2) plays a central role for expression of heme oxigenase 1 (HO-1), antioxidant enzyme. Our data showed that HO-1 could represent a potential molecular target susceptible to EVOO modulation, since dietary EVOO treatment strongly augmented Nrf2 and HO-1 protein expression conferring a role of HO-1 in the beneficial effects of EVOO in this murine model of chronic inflammation. Altogether, our results confirm, for the first time, that EVOO intake dramatically attenuated the progression and severity of arthritis in CIA DBA/1 J mice through Nrf2/HO-1 upregulation and NF-κB, MAPKs and JAK-STAT signalling pathway inhibition, decreasing the inflammatory cascade induced by CIA.Premio Extraordinario de Doctorado U

Extra-virgin olive oil and its phenolic extract prevent inflammatory response and joint damage in murine experimental arthritis

he consumption of EVOO in Mediterranean countries has shown beneficial effects. A wide range of evidence indicates that the phenolic compounds present in EVOO are endowed with anti-inflammatory properties. In this work, we evaluated the effects of dietary EVOO and treatment with its phenolic extract (PE) in a model of RA, the collagen-induced arthritis (CIA) in mice. On day 0, DBA-1/J mice were immunized with bovine collagen type II (CII). On day 21, the mice received a booster injection. We have demonstrated that EVOO and its PE decreases joint edema, cell migration, cartilage degradation and bone erosion. Our data indicate that dietary EVOO and PE treatment inhibit JNK, p38 and signal transducer and STAT-3. In addition, both EVOO and PE decrease NF-κB translocation leading to the down-regulation of the arthritic process. These results support the interest of natural diet components in the development of therapeutic products for arthritic conditions.El aceite de oliva extra virgen y su fracción polifenólica previenen la respuesta inflamatoria y el daño articular en un modelo de artritis experimental en murino. El consumo de Aceite de oliva virgen extra (AOVE) en los países mediterráneos ha demostrado tener efectos beneficiosos. Una amplia gama de pruebas indica que los compuestos fenólicos presentes en el AOVE tienen propiedades anti-inflamatorias. En este trabajo, se evaluaron los efectos de AOVE y el tratamiento en dieta de su fracción polifenólica (FP) en un modelo de la artritis reumatoide inducida por colágeno (CIA) en ratones. En el día 0, los ratones DBA-1/J se inmunizaron con colágeno bovino tipo II (CII). En el día 21, los ratones recibieron una inyección de refuerzo. Hemos demostrado que el AOVE y su FP disminuyen conjuntamente el edema, la migración celular, la degradación del cartílago y erosión ósea. Nuestros datos indican que la dieta con AOVE y el tratamiento con FP inhiben JNK, p38 y el transductor de señal y activador de la transcripción 3 (STAT-3). Además, tanto el AOVE como la FP disminuyen la translocación NF- κB que conduce a la mejora del proceso artrítico. Estos resultados apoyan el interés de una dieta con componentes naturales y el desarrollo de productos terapéuticos para desordenes artríticos.Ministerio de Economia y Competitividad AGL 2011-26949Junta de Andalucía P-10AGR-660

Protective effect of ellagic acid, a natural polyphenolic compound, in a murine model of Crohn’s disease

Current epidemiological and experimental studies support a beneficial role of dietary polyphenols in several gastrointestinal diseases, including inflammatory bowel disease. The aim of this study was to gain a better understanding of the effects of a naturally occurring polyphenol, ellagic acid, present in some fruits such as pomegranate, raspberries and nuts among others, in an experimental murine model of Crohn's disease by intra-colonic administration of TNBS in rats. Analysis of the lesions were carried out by macroscopic and histological technics. Inflammation response was assessed by histology and myeloperoxidase activity. iNOS and COX-2 are upregulated by MAPKs and NF-κB nuclear transcription factor in intestinal epithelial cells thus, we determined the expression of iNOS, COX-2 and the involvement of the p38, JNK, ERK1/2 MAPKs and NF-κB signalling in the protective effect of EA by western blotting. Oral administration of EA (10–20 mg/kg) diminished the severity and extension of the intestinal injuries induced by TNBS although there was no observed a significant dose–response. In addition, EA increased mucus production in goblet cells in colon mucosa, decreased neutrophil infiltration and pro-inflammatory proteins COX-2 and iNOS overexpression. Also EA was capable of reducing the activation of p38, JNK and ERK1/2 MAPKs, preventing the inhibitory protein IκB-degradation and inducing an inhibition of the nuclear translocation level of p65 in colonic mucosa. In conclusion, EA reduces the damage in a rat model of Crohn's disease, alleviates the oxidative events and returns pro-inflammatory proteins expression to basal levels probably through MAPKs and NF-κB signalling pathways.Junta de Andalucía y Ministerio de Ciencia e Innovación de España AGL 2008-0247

An opinion on the regulation of bone marrow adipose tissue by dietary fatty acids

Obesity has a significant impact on predisposition to various diseases and also affects the viabil­ity and choice of haematopoietic stem cells (HSCs) to favour myeloid cell production and/or turnover, all of which are extremely important for the functioning of immune system. As the production of blood cells and mobilization of HSCs and their progeny are regulated, at least in part, by multifaceted interactions through signals that come from the bone marrow (BM) microenvironment, it does not seem astonishing to assume that circumstances that cause alterations in BM structure will unavoidably cause alterations in mesenchymal cells such as adipocytes and lineages from HSCs. The existence of adipose tissue in BM or marrow fat (BMAT) is well known, although its origin, expansion, and functions are poorly understood. Inspired by other studies showing the potential role for olive oil and omega-3 long chain polyunsaturated fatty acids (omega-3 PUFAs) on BM health, and by our own preliminary findings showing the effects of monounsaturated (olive oil) but not saturated (milk cream) dietary fats to contain neutrophils and CD14high monocytes in BM during postprandial periods in healthy volunteers, herein we asked whether dietary fats (saturated fatty acids, SFAs, monounsatu­rated fatty acids, MUFAs, and omega-3 PUFAs) may be a candidate lifestyle factor to modulate the expansion, composition, and function of BMAT, the infiltration of adipose tissue macrophages (ATMs) in BMAT and the mobilization of HSCs and mature myeloid cells from BM during high-fat-induced obesity in mice. This is the first time that the interplay between different dietary fatty acids, obesity, and BM is addressed.Spanish Ministry of Science, Innovation and Universities AGL2016-80852-

Amoxicillin and Clarithromycin Mucoadhesive Delivery System for Helicobacter pylori Infection in a Mouse Model: Characterization, Pharmacokinetics, and Efficacy

Helicobacter pylori is the main pathogen responsible for gastric ulcers and a predisposing factor of stomach cancer. Although current treatment is usually successful, it requires high doses and frequent administration. An innovative mucoadhesive system (Mucolast®) loaded with amoxicillin and clarithromycin is proposed to improve the efficacy of treatment against H. pylori. The drug product was optimized based on its viscoelastic properties to obtain long-term stability of the vehicle. The drug release mechanisms were different for both antibiotics based on their solubilization status. A systemic and stomach pharmacokinetic profile was obtained after three different doses were administered to mice, obtaining similar systemic exposure levels but an increase in drug concentration in the stomach. The efficacy results in mice infected with H. pylori also demonstrated the superiority of the antibiotics when administered in Mucolast®, as shown by the bacterial count in stomach tissue and under histopathological and biochemical evaluation. The proposed treatment was efficacious and safe and is presented as a realistic alternative to current treatment options to improve patient compliance and to reduce bacterial resistance.Ministerio de Ciencia e Innovación. Gobierno de España (programa RETOS)-RTC-2015-4437-

MUFAs in High-Fat Diets Protect against Obesity-Induced Bias of Hematopoietic Cell Lineages

Scope: The role of dietary fatty acids in the generation of bone marrow (BM) immune cells and their trafficking to extramedullary compartments in the obesity is not yet fully understood. Methods and Results: C57BL/6J mice are randomly assigned to isocaloric high-fat diets (HFDs) formulate with dietary fats rich in saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs) or MUFAs fortified with eicosapentaenoic and docosahexaenoic acids for 20 weeks, followed by profiling of the obese metabolic phenotype and immunophenotypic features of immune cells in blood, spleen, and BM. All HFDs induce an obese phenotype, but it becomes largely less disruptive after the HFDs are enriched in MUFAs, which also induce signs of granulopoiesis and an expansion of long-term hematopoietic stem and granulocyte-macrophage progenitor cells in BM. In contrast, a HFD enriched in SFAs disturbs the fitness of medullary lymphocytes and promotes monopoiesis in favor of pro-inflammatory activated subsets. Conclusion: The reshaping of the fatty acid pools with MUFAs from the diet serves to manipulate the generation and trafficking of immune cells that are biased during obesity. These findings reveal a novel strategy by which dietary MUFAs may be instrumental in combating HFD-induced dysfunctional immune systems.Ministerio de Ciencia, Innovación y Universidades AGL2016-80852-

CD4+ and CD8+ T-cell responses in bone marrow to fatty acids in high-fat diets

Obesity is associated with disruptions in the adaptive immune system; however, dietary fatty acids in high-fat diets (HFDs) that induce obesity have consequences that are currently unclear regarding T-cell maintenance in bone marrow (BM). C57BL/6J mice were randomly assigned to isocaloric HFDs formulated with dietary fats rich in saturated fatty acids (SFAs), monounsaturated fatty acids (MUFAs), or MUFAs supplemented with eicosapentaenoic and docosahexaenoic acids for 20 weeks, followed by an analysis of the immunophenotypic feature of lymphocytes (CD3+) T and their subsets CD4+ and CD8+ T cells in spleen and BM, identification of fatty acids in BM extracellular fluid and analysis of the correspondence between fatty acids with the frequency of T-cell subsets in BM. Splenic CD3+ T cells were reduced irrespective of HFDs. In BM, CD3+ T cells were reduced after HFD-SFAs, while CD4+ T cells were increased after HFDs enriched in MUFAs and CD8+ T cells were reduced irrespective of HFDs. In BM extracellular fluid, the content of palmitic and myristic acids increased after HFD-SFAs and that of oleic acid increased after HFDs enriched in MUFAs. There was a statistical correspondence between HFD-induced changes in fatty acids in BM extracellular fluid and HFD-induced changes in the frequency of CD3+ and CD4+ T cells in BM. These findings reveal an undervalued critical role for dietary fatty acids in the selective acquisition of T-cell subsets in BM, highlighting that oleic acid existing in the surroundings of T-cell niches during HFD-induced obesity could be instrumental in the maintenance of CD4+ T cells.Fondo Social Europeo y Universidad de Sevilla-VPPI-USConsejo Superior de Investigaciones Científicas (CSIC)/Juan de la Cierva-FJCI-2017-3313

Aplicación de actividades colaborativas en los seminarios de Farmacología y Farmacoterapia III y Farmacia Clínica

En la búsqueda de soluciones para combatir el bajo nivel de motivación en los alumnos universitarios, encontramos la aplicación de técnicas de innovación docente, donde uno de los objetivos es la dinamización de las clases en las que el alumno tome mayor protagonismo. Hay que destacar la metodología de aprendizaje basado en problemas (ABP), donde el docente propone un caso o situación problema, y son los alumnos mediante la investigación y la reflexión los que deben resolverlo, fomentando de esta manera el aprendizaje significativo. En este sentido, pretendemos realizar una actividad colaborativa para logar un aprendizaje significativo de los alumnos, mediante la metodología ABP en las clases de seminario de la asignatura “Farmacología y Farmacoterapia III y Farmacia Clínica”. Los alumnos en pequeños grupos trabajarán en la resolución de los casos propuestos por el profesor, expondrán dichos resultados y finalmente serán evaluados, mediante preguntas elaboradas por el profesor con la herramienta Wooclap.Looking for solutions to combat the low level of motivation in university students, we find the application of teaching innovation techniques, where one of these objectives is the revitalization of classes in which the student takes a greater role. We highlight the problem-based learning methodology (PBL), where the teacher proposes a problem case or situation, and the student, through research and reflection, must solve it, thus promoting meaningful learning. In this way, we are going to carry out a collaborative activity to realize a significant learning of the students, through the PBL methodology in the seminar classes of the subject "Pharmacology and Pharmacotherapy III and Clinical Pharmacy". The students in small groups will work on solving the cases proposed by the teacher, they will present said results, and finally they will be evaluated, through questions prepared by the teacher with the Wooclap tool

Dietary squalene supplementation improvesDSS-induced acute colitis by downregulating p38 MAPKand NFkB signaling pathways

Scope:Squalene is a polyunsaturated triterpene, which has exhibited anticancer and antioxidantactivities among others. We investigated dietary squalene supplementation effect on an acutecolitis model induced by dextran sulfate sodium (DSS) in C57BL/6 mice.Methods and results:Mice were fed from weaning with squalene at 0.02% and 0.1%. After4 weeks, mice were exposed to 3% DSS for 5 days developing acute colitis. After DSS removal(5 days), colons were histological and biochemically processed. Our results showed that dietarysqualene treatment exerts anti-inflammatory action in DSS-induced acute colitis. Western blotrevealed that squalene downregulated COX-2 (where COX is cyclooxygenase) and induciblenitric oxide synthase system by inhibition of mitogen-activated protein kinase p38 and thenuclear factor-kappa B signaling pathways, preventing an increase in the cytokines levels.Under our experimental conditions, STAT3 and FOXP3 (where FOXP3 is forkhead box P3)were not modified and the transcriptional regulation of antioxidant and/or detoxifying enzymes,Nrf2 (where Nrf2 is nuclear factor (erythroid-derived 2)-like 2), was reduced in DSS-inducedcolitis. However, any change could be observed after squalene supplementation.Conclusion:Squalene was able to improve the oxidative events and returned proinflammatoryproteins expression to basal levels probably through p38 mitogen-activated protein kinase andnuclear factor-kappa B signaling pathways. However, supplementary studies are needed inorder to provide a basis for developing a new dietary supplementation strategy

Dietary oleocanthal supplementation prevents inflammation and oxidative stress in collagen-induced arthritis in mice

Oleocanthal (OLE), a characteristic and exclusive secoiridoid of Oleoaceae family, is mainly found in extra virgin olive oil (EVOO). Previous studies have reported its antioxidant, anti-inflammatory, antimicrobial, anticancer and neuroprotective effects. Since the pathogenesis of rheumatoid arthritis (RA) involves inflammatory and oxidative components, this study was designed to evaluate the preventive role of dietary OLE-supplemented effects in collagen-induced arthritis (CIA) murine model. Animals were fed with a preventive OLE-enriched dietary during 6 weeks previous to CIA induction and until the end of experiment time. At day 43 after first immunization, mice were sacrificed: blood was recollected and paws were histological and biochemically processed. Dietary OLE prevented bone, joint and cartilage rheumatic affections induced by collagen. Levels of circulatory matrix metalloproteinase (MMP)-3 and pro-inflammatory cytokines (IL-6, IL-1β, TNF-α, IL-17, IFN-γ) were significantly decreased in secoiridoid fed animals. Besides, dietary OLE was able to diminish COX-2, mPGES-1 and iNOS protein expressions and, also, PGE2 levels. The mechanisms underlying these protective effects could be related to Nrf-2/HO-1 axis activation and the inhibition of relevant signaling pathways including JAK-STAT, MAPKs and NF-κB, thus controlling the production of inflammatory and oxidative mediators. Overall, our results exhibit preliminary evidences about OLE, as a novel dietary tool for the prevention of autoimmune and inflammatory disorders, such as RA.Ministerio de Economía y Competitividad AG-2017-89342-PJunta de Andalucía CTS259, FQM13