Abnormal regulation of Na,K-ATPase in Glucose Intolerant Rats.

Introduction: Glucose is the most important physiological insulin secretagogue. However, the mechanisms underlying glucose-induced insulin release are not fully understood. The role of electrogenic systems such as ionic pumps, to these events remains essentially uninvestigated. Na,K-ATPase, responsible for maintaining Na+ and K+ gradients across the plasma membrane and generates a net outward current, thus changes in its activity may contribute to the early ionic events regulating insulin secretion (Therien and Blostein, 2000). Objective: The aim of this work was to evaluate the regulation of Na,K-ATPase activity by glucose in intact -cells of normal and glucose intolerant (GI) rats and its putative contribution to the regulation of insulin secretion. Material and Methods: Pancreatic -cells, from normal or control or GI rats, were isolated and cultured (48h). Cell batches were pre-incubated (30min) with 2mM glucose to reach basal. Afterwards cells were challenged with glucose in the interval 0-11mM for 60min, for dose-dependence evaluation, or with 8mM glucose for 5-120min, for time-dependence evaluation. ATPase activity was assessed in intact cells by colorimetric quantification of Pi formed in 30min. Na,K-ATPase activity was calculated by the difference between the activities obtained in the absence and in presence the of 1mM ouabain (Costa et al., 2009). Results: In β-cells from normal rats, glucose induced a bimodal regulation of Na,K-ATPase. In the absence of glucose, Na,K-ATPase activity was 0.056±0.015 U/mg. Stimulation with 2mM glucose induced an increase of Na,K-ATPase activity of ~4 fold whereas for [glucose] above 2mM it was observed a significant inhibition of Na,K-ATPase activity (0.061±0.013, 0.080±0.009 and 0.064±0.005 U/mg for 5.6, 8.4 and 11mM glucose, respectively, compared to 0.188±0.035 U/mg observed in 2mM G; n=3-8). β-cells from GI rats does not present this profile; in the absence of glucose, Na,K-ATPase activity was 0.202±0.036 U/mg and no significant differences from this value were observed with the other glucose concentration tested. Addicionally, in β-cells from normal rats, glucose (8mM) induced a time-dependent inhibition, with a biphasic profile, of Na,K-ATPase - it was observed a decrease in the pump activity between 0 and 20min stimulation where it reached a minimum value (77%). For incubation periods over 20min, the pump activity slowly and partially recovered (54%, 55% and 52%, for 30, 60 and 120min, respectively; n=7). In β-cells from GI animals, an less accentuated decrease of Na,K-ATPase activity between 0 ans 20min was also observed (34%), and is not observed further recover in activity. Conclusions: This work demonstrates there Na,K-ATPase is strictly regulated by glucose in pancreatic β-cell. This regulation is unpaired in GI animals. Na,K-ATPase contribution to glucose-induced ionic events and insulin secretion might be relevant and must be explored as a possible therapeutic target in TD2 . 1. Therien AG, Blostein R (2000) Mechanisms of sodium pump regulation. Am J Physiol Cell Physiol 279:C541-C566 2. Costa AR, Real J, Antunes CM, Cruz-Morais J (2009) A new approach for determination of Na,K-ATPase activity: application to intact pancreatic beta-cells. In Vitro Cell Dev Biol Ani

Implication of AMPK in glucose-evoked modulation of Na,K-ATPase

Background and aims: Na,K-ATPase is an integral membrane protein that maintains the gradients of Na+ and K+, using the energy of ATP hydrolysis, maintaining the ionic gradients that allow electrical activity to occur. It has been demonstrated that, in pancreatic β-cells, Na,K-ATPase is regulated by glucose and that this phenomenon is impaired in glucose intolerant subjects. However, the mechanism underlying glucose-induced modulation of Na,K-ATPase is still unclear. The AMP-activated protein kinase (AMPK) is a molecular key player in energy homeostasis, providing exquisite sensitivity to small changes in intracellular AMP levels and thus to intracellular [ATP]/[ADP] ratio, that is known to activate protein regulatory pathways. Since in pancreatic β-cell, glucose has marked effects on oxidative metabolism and total intracellular ATP and AMP levels, the involvement of AMPK in the cascade of events regulating Na,K-ATPase regulation in pancreatic β-cells was postulated. The aim of this work was to evaluate the putative role of AMPK in the glucose-evoked regulation of Na,K-ATPase activity in the pancreatic β-cell. Materials and methods: Pancreatic -cells from normal (control) or glucose-intolerant Wistar rats (GIR) were isolated and cultured (48h). Cell batches were pre-incubated (30min) with 2.1mM glucose to reach basal activity. Afterwards cells were challenged to 8.4mM glucose for 20min, in the presence or absence of AMPK agonists (AICAR) and antagonists (compound C; CC). ATPase activity was assessed in intact cells by colorimetric quantification of Pi formed in 30min. Na,K-ATPase activity was calculated by the difference between the activities obtained in the absence and in presence the of 1mM ouabain. Results: In basal conditions the activity of Na,K-ATPase from normal and GIR pancreatic β-cell was similar (0.184±0.030 and 0.186±0.020 molPi/min/mgProt, respectively). Challenging the control β-cells with glucose 8.4mM evoked a 62% reduction of Na,K-ATPase activity whereas in GIR β-cells a significantly lower inhibition (40%) was observed. The addition of AICAR 1mM abolished glucose-induced Na,K-ATPase inhibition (0,166±0.011 molPi/min/mg). In control β-cell, the addition of CC 10 μM had no effect on glucose-induced inhibition of Na,K-ATPase. In the contrary, in GIR β-cells it significantly potentiated glucose-evoked inhibition of Na,K-ATPase reaching values similar to that observed in the controls (66%). Conclusions: The AMPK agonist AICAR counteracts the inhibitory action of glucose on Na,K-ATPase of control β-cells whereas CC amplified the glucose-induced inhibition of Na,K-ATPase in GIR β-cells. These results suggest that AMPK plays a central role in the cascade of events underlying glucose-induced modulation of Na,K-ATPase and that the defect must be upstream of AMPK. Finally, abnormal glucose-induced regulation of Na,K-ATPase occurs prior to overt type 2 diabetes and might be a feature in the disease development

Introduction

Este documento é uma tradução do livro publicado em português, “Pobreza e Paz nos PALOP”, Sextante Editora (2009), ISBN 978-989-676-007-6As the end result of a multidisciplinary investigation project conducted in Africa, this publication about Poverty and Peace in the Portuguese-speaking African Countries1 has the purpose, on one hand, of presenting the main conclusions of the studies conducted in these countries and, on the other, to provide a collection of guidelines for future research relevant to the comprehension of the combination of both phenomena

Reproductive biology of Megabalanus azoricus (Pilsbry), the Azorean Barnacle

Copyright © 2007, Taylor & Francis.The reproductive biology of Megabalanus azoricus (Pilsbry), the commercially exploited barnacle in the Azores, was studied in an attempt to provide the scientific basis for the sustainable management of this heavily exploited regional marine resource. Both the Ospar Commission and WWF have expressed concern for this species, considering it at risk and in urgent need for scientific study. Barnacles were collected every month from shallow water (.3 m depth) around São Miguel Island from October 2004 to September 2005. Individuals were measured and gonads processed for histology and analysed with a stereological method. M. azoricus has a hermaphroditic reproductive system with separate gonads and it was possible to describe the various stages of gametic maturation in both. Data on fertility were also obtained by egg counts and calculation of the Gonadosomatic index (GSI). Throughout the year, some specimens of M. azoricus can be found that rare mature, but two reproductive peaks were observed, one in January and a smaller one in July. A strong positive correlation between GSI and environmental factors such as photoperiod and water temperature was observed.CIRN (Research Centre in Natural Resources), Universidade dos Açores

The percepetion of unlisted companies on gender quotas law

Mestrado Bolonha em FinançasThe lack of women representation on boards has recently become an important issue that needs to be addressed due to the benefits generated by gender diversity in boards’ structure. As a result of increased interest in this topic, academic investigations have arisen and, more recently, increasing regulatory pressure has emerged on the percentage of women in top management positions. In Portugal, in 2017, Law nº 62/2017, known as the “Gender Quotas Law”, was released. It aimed a more balanced representation of women and men in management bodies by imposing a minimum of 33% of the least represented gender in management bodies. At this time, in Portugal, this Law is only applied to companies listed on the stock exchange and, as this Law can most likely be adopted also for companies not listed on the stock exchange, this study purposes to understand the point of view of Portuguese non-listed companies, in this case specific of Portuguese Public Limited Companies, about the implementation of this Law, by inquiring them on different aspects inherent to Gender Quotas Law. The results obtained revel that Portuguese Public Limited Companies agree on the conception of measures to promote gender equality, but not in the form of law. They also believe that women are competent enough to assume top management positions and that Gender Quotas Law are important for risk assessment in the decision-making process, for the company's overall performance, for financial reporting, for the company's growth and expansion and for the company's sustainability long-term. It is essential to eliminate the injustices and gender inequalities that still exist in the labor market in Portugal and this Law is another step in that direction, hence it is important to understand the companies' point of view on this issue.info:eu-repo/semantics/publishedVersio

Educational Process Mining based on Moodle courses: a review of literature

With the prevalence of E-Learning, it is important to analyze how students progress in this environment. These systems collect data about the students’ learning path, and Process Mining (PM) can provide a detailed model of this path. Based on the analysis of ten Educational Process Mining (EPM) case studies involving Moodle event logs, this article aims to contribute a literature review on EPM’s research. Beyond a theoretical introduction to PM and its implications for educational data, the review concludes on what PM tools and techniques are used, as well as the challenges faced in practice. The technical options include software, process discovery algorithms and representation models. These results aim to create a list of available options for future EPM endeavors, in addition to a list of issues to consider in future research involving Moodle

The K-framed quadtrees approach for path planning through a known environment

One of the most important tasks for a mobile robot is to navigate in an environment. The path planning is required to design the trajectory that generates useful motions from the original to the desired position. There are several methodologies to perform the path planning. In this paper, a new method of approximate cells decomposition, called K-Framed Quadtrees is present, to which the algorithm A ⋆ is applied to determine trajectories between two points. To validate the new approach, we made a comparative analysis between the present method, the grid decomposition, quadtree decomposition and framed quadtree decomposition. Results and implementation specifications of the four methods are presented.Project ”TEC4Growth - Pervasive Intelligence, Enhancers and Proofs of Concept with Industrial Impact/NORTE-01-0145-FEDER-000020” is financed by the North Portugal Regional Operational. Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, and through the European Regional Development Fund (ERDF). This work is also financed by the ERDF – European Regional Development Fund through the Operational Programme for Competitiveness and Internationalisation - COMPETE 2020 Programme within project POCI-01-0145-FEDER-006961, and by National Funds through the FCT – Fundação para a Ciência e a Tecnologia (Portuguese Foundation for Science and Technology) as part of project UID/EEA/50014/2013.info:eu-repo/semantics/publishedVersio

Produção de emulsões tipo Pickering por processos de alta e baixa energia

Orientadores: Rosiane Lopes da Cunha, David A. WeitzTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos, Harvard UniversityResumo: Partículas de quitosana (Ch) e celulose (nanofibras de celulose; CNFs e cristais de celulose; CCrys) foram utilizadas como estabilizantes de emulsões Pickering óleo em água (O/A) produzidas por técnicas de alta (ultrassom e homogeneizador a alta pressão) e baixa energia (dispositivos de microfluídica). As características das partículas e emulsões foram avaliadas frente às variáveis de processo de emulsificação e condições gastrointestinais. Em uma primeira etapa, Ch foram produzidas e caracterizadas quanto aos efeitos da variação da potência aplicada e tempo de processamento em ultrassom sobre a estabilidade destas partículas e das emulsões. Os intensos efeitos de cavitação gerados com altas potências de ultrassom foram capazes de quebrar as partículas de quitosana e as gotas de óleo em menores tamanhos promovendo a formação de uma rede tridimensional entre gotas. Além disso, a alta estabilidade das emulsões foi associada à redução da tensão interfacial entre o óleo e a água e aumento da hidrofobicidade das partículas. Os efeitos das condições de emulsificação usando ultrassom e homogeneizador a alta pressão sobre as propriedades das CNFs obtidas da casca da banana e das emulsões Pickering foram avaliados na segunda etapa deste projeto. O fenômeno de coalescência foi observado nas emulsões produzidas no homogeneizador a alta pressão, enquanto que, a floculação das gotas ocorreu naquelas obtidas em ultrassom de alta intensidade. Nesta última, a estabilidade à coalescência foi associada com o ligeiro aumento da viscosidade da emulsão e rompimento das CNFs durante o processo de ultrassonicação. Em uma terceira etapa, um protocolo de digestibilidade in vitro foi utilizado para elucidar o papel das partículas (Ch, CNFs and CCrys) na taxa de digestão lipídica das emulsões tipo Pickering. A alta carga positiva das emulsões estabilizadas por quitosana levou à desagregação das gotas após a etapa gástrica, o que favoreceu a maior digestão lipídica na etapa intestinal. Por outro lado, a emulsão estabilizada com CNFs apresentou menor digestão lipídica e a forte aderência das partículas de CCrys na interface das gotas tornou-as resistentes ao deslocamento por componentes tensoativos. Na última etapa, a formação e propriedades de emulsões estabilizadas por CCrys foi estudada usando um dispositivo microcapilar. Gotas de óleo altamente monodispersas e emulsões estáveis ao longo do tempo foram produzidas a partir do balanço entre o tempo de geração das gotas e adsorção das partículas. Por outro lado, grandes gotas de óleo e emulsões menos estáveis foram obtidas após a ocorrência de eventos de coalescência dentro do microcanal ou devido ao aumento da viscosidade da fase dispersa. A abordagem microfluídica contribuiu para uma melhor compreensão das condições dinâmicas de estabilização das emulsões Pickering dentro e fora dos microcanais revelando eventos discretos que geralmente estão escondidos em emulsões produzidas por métodos convencionais de emulsificaçãoAbstract: High-energy (ultrasound and high-pressure homogenizer) and low-energy (microfluidic devices) processes were used to obtain oil-in-water (O/W) Pickering emulsions stabilized by food-grade particles (chitosan nanoparticles; Ch, cellulose nanofibers; CNFs and cellulose crystals; CCrys). In the first step, the effect of time and ultrasound power on physicochemical properties of Ch and O/W Pickering emulsions were evaluated using an ultrasonic device. The surface activity of chitosan particles was evidenced with the reduction of interfacial tension between oil-water phases. The emulsion stability mechanism by deprotonated chitosan particles was also associated to an increase of the particle hydrophobicity and the formation of a droplet network structure due to the intense effects of cavitation generated at higher ultrasonication power. The effects of the emulsification conditions using ultrasound and high-pressure homogenization on the properties of the CNFs obtained from the banana peel and Pickering emulsions were evaluated in the second step. Coalescence phenomenon was observed in the emulsions produced using high-pressure homogenizer, whereas droplets flocculation occurred in emulsions processed by ultrasound. In the latter, coalescence stability was associated with effects of cavitation forces acting on the CNFs breakup. In the third step, an in vitro digestibility protocol was used to elucidate the role of different emulsifying polysaccharides particles (Ch, CNFs and CCrys) on the lipid digestion rate of oil-in-water Pickering emulsions. The highly positive charge of the emulsions stabilized by chitosan led to the disaggregation of droplets after the gastric step, which favored a more intense lipid digestion in the intestinal step. On the other hand, Tween 80, CCrys and CNFs were able to inhibit lipid digestion and no changes on droplet mean size were observed following intestinal step. CNFs-stabilized emulsion showed the lowest lipid digestion, whereas the strong adherence of the CCrys particles onto the droplet interface became them resistant to displacement by surface-active components. In the last step, the formation and stability of oil droplets in the presence of cellulose nanocrystals was studied using a microcapillary device. Monodisperse oil droplets and stable emulsions over time were produced from a balance between droplets generation time and particles adsorption. On the other hand, large oil droplets and less stable emulsions were obtained after coalescence events inside the microchannel or due to the increase on the dispersed phase viscosity. Microfluidic approach contributed to a better understanding of the dynamic conditions of stabilization inside and outside the microchannels revealing discrete events that usually are hidden in emulsions produced by conventional emulsification methodsDoutoradoEngenharia de AlimentosDoutora em Engenharia de Alimentos140710/2015-9; 204109/2017-5CNP

Chitosan/carrageenan nanoparticles: effect of cross-linking with tripolyphosphate

Chitosan/carrageenan/tripolyphosphate nanoparticles were prepared by polyelectrolyte complexation/ionic gelation, the latter compound acting as cross-linker. The incorporation of the three components in the nanoparticle matrix was assessed by analytical techniques (FTIR, XPS and TOF-SIMS). Using chitosan/carrageenan nanoparticles as control, the effect of the cross-linker in the particles properties was studied. A decrease in size (from 450-500 nm to 150-300 nm) and in zeta potential (from +75 - +85 mV to +50 - +60 mV), and an increase in production yield (from 15-20% to 25-35%), and in stability (from one week to up to 9 months) were observed. Also, a correlation between positive to negative charge ratios in the formulations and the above characteristics was established. The small size and high positive surface charge make the developed chitosan/carrageenan/tripolyphosphate nanoparticles potential tools for an application in mucosal delivery of macromolecules