Intestinal Anti-inflammatory Effects of Outer Membrane Vesicles from Escherichia coli Nissle 1917 in DSS-Experimental Colitis in Mice

Escherichia coli Nissle 1917 (EcN) is a probiotic strain with proven efficacy in inducing and maintaining remission of ulcerative colitis. However, the microbial factors that mediate these beneficial effects are not fully known. Gram-negative bacteria release outer membrane vesicles (OMVs) as a direct pathway for delivering selected bacterial proteins and active compounds to the host. In fact, vesicles released by gut microbiota are emerging as key players in signaling processes in the intestinal mucosa. In the present study, the dextran sodium sulfate (DSS)-induced colitis mouse model was used to investigate the potential of EcN OMVs to ameliorate mucosal injury and inflammation in the gut. The experimental protocol involved pre-treatment with OMVs for 10 days before DSS intake, and a 5-day recovery period. Oral administration of purified EcN OMVs (5 μg/day) significantly reduced DSS-induced weight loss and ameliorated clinical symptoms and histological scores. OMVs treatment counteracted altered expression of cytokines and markers of intestinal barrier function. This study shows for the first time that EcN OMVs can mediate the anti-inflammatory and barrier protection effects previously reported for this probiotic in experimental colitis. Remarkably, translation of probiotics to human healthcare requires knowledge of the molecular mechanisms involved in probiotic–host interactions. Thus, OMVs, as a non-replicative bacterial form, could be explored as a new probiotic-derived therapeutic approach, with even lower risk of adverse events than probiotic administration.his work was funded by the “Ministerio de Economía y Competitividad”, Spain (Grants AGL2012-34985, AGL2016-79113-R (AEI/FEDER, UE) and AGL2015-67995-C3-3-R, all co-financed with European Commission ERDF funds), by the Generalitat de Catalunya, AGAUR (Grant 2014SGR1017), and by the Junta de Andalucía (Grant CTS-164). The CIBEREHD is funded by the Instituto de Salud Carlos III. M-JF acknowledges her FPI fellowship from the Spanish Ministry of Economy and Competitiveness

Study of Melatonin as Preventive Agent of Gastrointestinal Damage Induced by Sodium Diclofenac

: Safety profile of nonsteroidal anti‐inflammatory drugs (NSAIDs) has been widely studied and both therapeutic and side effects at the gastric and cardiovascular level have been generally associated with the inhibitory effect of isoform 1 (COX‐1) and 2 (COX‐2) cyclooxygenase enzymes. Now there are evidences of the involvement of multiple cellular pathways in the NSAIDs‐mediated‐gastrointestinal (GI) damage related to enterocyte redox state. In a previous review we summarized the key role of melatonin (MLT), as an antioxidant, in the inhibition of inflammation pathways mediated by oxidative stress in several diseases, which makes us wonder if MLT could minimize GI NSAIDs side effects. So, the aim of this work is to study the effect of MLT as preventive agent of GI injury caused by NSAIDs. With this objective sodium diclofenac (SD) was administered alone and together with MLT in two experimental models, ex vivo studies in pig intestine, using Franz cells, and in vivo studies in mice where stomach and intestine were studied. The histological evaluation of pig intestine samples showed that SD induced the villi alteration, which was prevented by MLT. In vivo experiments showed that SD altered the mice stomach mucosa and induced tissue damage that was prevented by MLT. The evaluation by quantitative reverse transcription PCR (RT‐qPCR) of two biochemical markers, COX‐2 and iNOS, showed an increase of both molecules in less injured tissues, suggesting that MLT promotes tissue healing by improving redox state and by increasing iNOS/NO that under non‐oxidative condition is responsible for the maintenance of GI‐epithelium integrity, increasing blood flow and promoting angiogenesis and that in presence of MLT, COX‐2 may be responsible for wound healing in enterocyte. Therefore, we found that MLT may be a preventive agent of GI damages induced by NSAIDs. Keywords: melatonin; NSAIDs; gastric injuries; antioxidan

Desarrollo de aprendizajes activos en primeros cursos universitarios: Workshop cónica.

Como ya es conocido, los profesores de Matemáticas utilizamos los ejemplos como recursos de aprendizaje para enseñar algún contenido matemático concreto, de modo que las generalizaciones y abstracciones sean más fácilmente entendidas por los alumnos, pasando de lo concreto a lo abstracto, como otra forma de enseñar y practicar en Matemáticas. Esta metodología de trabajo se ve potenciada por el uso de dispositivos móviles llamados mobile-learning (m-learning) o educación móvil (educación-m), en español. Siguiendo esta línea de trabajo, se ha realizado el workshop de cónicas que se presenta en este artículo, empleando estas nuevas tecnologías (TIC) y con el objetivo de desarrollar aprendizajes activos en Geometría a través de la resolución de problemas en los primeros cursos de Grado en las ingenierías. ABSTRACT: As it is already known, math teachers, use examples as learning resources, to teach some specific math contents, so that generalizations and abstractions are more easily understood by students, from concrete to abstract, as another way of Mathematics teaching and training. This methodology is enhanced by the use of mobile devices, called mobile-learning (m-learning) o “educación móvil” (educación-m), in Spanish. Following this strategy, the workshop of conic sections shown in this paper has been carried out, using these new technologies (ICT) and in order to develop active learning in Geometry through problem-solving at the first years of engineering degrees

Outer membrane vesicles from probiotic and commensal Escherichia coli activate NOD1-mediated immune responses in intestinal epithelial cells

Gut microbiota plays a critical role in maintaining human intestinal homeostasis and host health. Bacterial extracellular vesicles are key players in bacteria-host communication, as they allow delivery of effector molecules into the host cells. Outer membrane vesicles (OMVs) released by Gram-negative bacteria carry many ligands of pattern recognition receptors that are key components of innate immunity. NOD1 and NOD2 cytosolic receptors specifically recognize peptidoglycans present within the bacterial cell wall. These intracellular immune receptors are essential in host defense against bacterial infections and in the regulation of inflammatory responses. Recent contributions show that NODs are also fundamental to maintain intestinal homeostasis and microbiota balance. Peptidoglycan from non-invasive pathogens is delivered to cytosolic NODs through OMVs, which are internalized via endocytosis. Whether this pathway could be used by microbiota to activate NOD receptors remains unexplored. Here, we report that OMVs isolated from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 activate NOD1 signaling pathways in intestinal epithelial cells. NOD1 silencing and RIP2 inhibition significantly abolished OMV-mediated activation of NF-κB and subsequent IL-6 and IL-8 expression. Confocal fluorescence microscopy analysis confirmed that endocytosed OMVs colocalize with NOD1, trigger the formation of NOD1 aggregates, and promote NOD1 association with early endosomes. This study shows for the first time the activation of NOD1-signaling pathways by extracellular vesicles released by gut microbiota. Keywords: gut microbiota, Escherichia coli Nissle 1917, NF-κB activation, bacterial extracellular vesicles, NOD

Development of Pranoprofen Loaded Nanostructured Lipid Carriers to Improve Its Release and Therapeutic Efficacy in Skin Inflammatory Disorders

Abstract: Pranoprofen (PF)-loaded nanostructured lipid carriers (NLCs), prepared using a high-pressure homogenization method, have been optimized and characterized to improve the biopharmaceutical profile of the drug. The optimized PF-NLCs exhibited physicochemical characteristics and morphological properties that were suitable for dermal application. Stability assays revealed good physical stability, and the release behavior of PF from these NLCs showed a sustained release pattern. Cell viability results revealed no toxicity. Ex vivo human skin permeation studies in Franz diffusion cells were performed to determine the influence of different skin penetration enhancers (pyrrolidone, decanol, octanoic acid, nonane, menthone, squalene, linoleic acid, and cineol) on skin penetration and retention of PF, being the highest dermal retention in the presence of linoleic acid. The selected formulations of NLCs exhibited a high retained amount of PF in the skin and no systemic effects. In vivo mice anti-inflammatory efficacy studies showed a significant reduction in dermal oedema. NLCs containing linoleic acid presented better anti-inflammatory efficacy by decreasing the production of interleukins in keratinocytes and monocytes. The biomechanical properties of skin revealed an occlusive effect and no hydration power. No signs of skin irritancy in vivo were detected. According to these results, dermal PF-NLCs could be an effective system for the delivery and controlled release of PF, improving its dermal retention, with reduced dermal oedema as a possible effect of this drug KEYWORDS: anti-inflammatory activity; linoleic acid; nanostructured lipid carriers; penetration enhancers; pranoprofen; skin deliver

Identificación de áreas de recarga a través de un modelo hidrológico en la región del Arco Seco de Panamá

Las variaciones del ciclo hidrológico en los mecanismos de recarga de las aguas subterráneas están poco estudiadas en los países tropicales como Panamá. En las últimas décadas, regiones del país como el Arco Seco, han enfrentado sequías y problemas de suministro principalmente debido a alteraciones antropogénicas y efectos del cambio climático. Igualmente, las áreas de recarga de aguas subterráneas no están bien definidas y ha existido una explotación desordenada de las aguas subterráneas produciendo un aumento de extracciones, afectando su calidad y cantidad. Esta investigación tiene como objetivo comprender las interacciones de variables hidrometeorológicas a través de la aplicación del modelo hidrológico SWAT+ para identificar áreas de recarga en la cuenca del río La Villa. Datos meteorológicos genéricos, e información de la Cuenca se combinan para generar Unidades de Respuesta Hidrológica para proporcionar un análisis espacial y físico de los mecanismos de recarga en la región. Los resultados presentan que la mayoría de las áreas de recarga en la zona de estudio están cambiando su uso de suelo, utilizándolo en mayor medida para la agricultura y la ganadería. Estas actividades tienen un impacto en la cubierta forestal y alteran significativamente la capacidad de infiltración del suelo y, por lo tanto, afectan la recarga de aguas subterráneas. A través de esta investigación se busca potenciar la gestión de los recursos hídricos y las políticas de uso de la tierra para garantizar un uso sostenible de las zonas de recarga de aguas subterráneas en la región

Characterization and In Vivo Anti-Inflammatory Efficacy of Copal (Dacryodes peruviana (Loes.) H.J. Lam) Essential Oil

Essential oils are natural aromatic substances that contain complex mixtures of many volatile compounds frequently used in pharmaceutical and cosmetic industries. Dacryodes peruviana (Loes.) H.J. Lam is a native species from Ecuador whose anti-inflammatory activity has not been previously reported, thus the aim of this study was to evaluate the anti-inflammatory activity of D. peruviana essential oil. To that end, essential oil from D. peruviana fruits was isolated by hydrodistillation and characterized physically and chemically. The tolerance of the essential oil was analyzed by cytotoxicity studies using human keratinocytes. The anti-inflammatory activity was evaluated by an arachidonic acid-induced edema model in mouse ear. The predominant compounds in D. peruviana essential oil were α-phellandrene, limonene, and α-pinene, with the three compounds reaching approximately 83% of the total composition. Tolerance studies showed high biocompatibility of this essential oil with human keratinocytes. In vivo studies demonstrated a moisturizing effect and an alleviation of several events occurred during the inflammatory process after topical treatment with D. peruviana essential oil such as decline in skin edema; reduction in leukocytic infiltrate; and decrease in inflammatory cytokines TNFα, IL-8, IL-17A, and IL-23. Therefore, this essential oil could be an attractive treatment for skin inflammation

Activation of immune and defense responses in the intestinal mucosa by outer membrane vesicles of commensal and probiotic Escherichia coli strains

The influence of microbiota in human health is well established. Imbalances in microbiome structure have been linked to several diseases. Modulation of microbiota composition through probiotic therapy is an attempt to harness the beneficial effects of commensal microbiota. Although there is wide knowledge of the responses induced by gut microbiota, the microbial factors that mediate these effects are not fully known. Gram-negative bacteria release outer membrane vesicles (OMVs) as a delivery mechanism of microbial factors, having an important role in intercellular communication. Here we investigated whether OMVs from the probiotic Escherichia coli strain Nissle 1917 or the commensal E. coli strain ECOR12 trigger immune responses in various cellular models: (i) peripheral blood mononuclear cells (PBMCs) as a model of intestinal barrier disruption, (ii) apical stimulation of Caco-2/PMBCs co-culture as a model of intact intestinal mucosa, and (iii) colonic mucosa explants as an ex vivo model. Stimulations with bacterial lysates were also performed for comparison. Whereas OMVs and lysates activated expression and secretion of several cytokines and chemokines in PBMCs, only OMVs induced basolateral secretion and mRNA upregulation of these mediators in the co-culture model. We provide evidence that OMVs are internalized in polarized Caco-2 cells, and that activated epithelial cells elicit a response in the underlying immunocompetent cells. The OMVs effects were corroborated in the ex vivo model. This experimental study shows that OMVs are an effective strategy used by beneficial gut bacteria to communicate with and modulate host responses, activating signaling events through the intestinal epithelial barrier.

Intestinal Anti-inflammatory Effects of Outer Membrane Vesicles from Escherichia coli Nissle 1917 in DSS-Experimental Colitis in Mice

Escherichia coli Nissle 1917 (EcN) is a probiotic strain with proven efficacy in inducing and maintaining remission of ulcerative colitis. However, the microbial factors that mediate these beneficial effects are not fully known. Gram-negative bacteria release outer membrane vesicles (OMVs) as a direct pathway for delivering selected bacterial proteins and active compounds to the host. In fact, vesicles released by gut microbiota are emerging as key players in signaling processes in the intestinal mucosa. In the present study, the dextran sodium sulfate (DSS)-induced colitis mouse model was used to investigate the potential of EcN OMVs to ameliorate mucosal injury and inflammation in the gut. The experimental protocol involved pre-treatment with OMVs for 10 days before DSS intake, and a 5-day recovery period. Oral administration of purified EcN OMVs (5 mg/day) significantly reduced DSS-induced weight loss and ameliorated clinical symptoms and histological scores. OMVs treatment counteracted altered expression of cytokines and markers of intestinal barrier function. This study shows for the first time that EcN OMVs can mediate the anti-inflammatory and barrier protection effects previously reported for this probiotic in experimental colitis. Remarkably, translation of probiotics to human healthcare requires knowledge of the molecular mechanisms involved in probiotic-host interactions. Thus, OMVs, as a non-replicative bacterial form, could be explored as a new probiotic-derived therapeutic approach, with even lower risk of adverse events than probiotic administration. Keywords: probiotic, Escherichia coli Nissle 1917, outer membrane vesicles, DS

Apremilast Microemulsion as Topical Therapy for Local Inflammation: Design, Characterization and Efficacy Evaluation

Apremilast (APR) is a selective phosphodiesterase 4 inhibitor administered orally in the treatment of moderate-to-severe plaque psoriasis and active psoriatic arthritis. The low solubility and permeability of this drug hinder its dermal administration. The purpose of this study was to design and characterize an apremilast-loaded microemulsion (APR-ME) as topical therapy for local skin inflammation. Its composition was determined using pseudo-ternary diagrams. Physical, chemical and biopharmaceutical characterization were performed. Stability of this formulation was studied during 90 days. Tolerability of APR-ME was evaluated in healthy volunteers while its anti-inflammatory potential was studied using in vitro and in vivo models. A homogeneous formulation with Newtonian behavior and droplets of nanometric size and spherical shape was obtained. APR-ME released the incorporated drug following a first-order kinetic and facilitated drug retention into the skin, ensuring a local effect. Anti-inflammatory potential was observed for its ability to decrease the production of IL-6 and IL-8 in the in vitro model. This effect was confirmed in the in vivo model histologically by reduction in infiltration of inflammatory cells and immunologically by decrease of inflammatory cytokines IL-8, IL-17A and TNFα. Consequently, these results suggest that this formulation could be used as an attractive topical treatment for skin inflammation