Fish models of induced osteoporosis

Osteopenia and osteoporosis are bone disorders characterized by reduced bone mineral density (BMD), altered bone microarchitecture and increased bone fragility. Because of global aging, their incidence is rapidly increasing worldwide and novel treatments that would be more efficient at preventing disease progression and at reducing the risk of bone fractures are needed. Preclinical studies are today a major bottleneck to the collection of new data and the discovery of new drugs, since they are commonly based on rodent in vivo systems that are time consuming and expensive, or in vitro systems that do not exactly recapitulate the complexity of low BMD disorders. In this regard, teleost fish, in particular zebrafish and medaka, have recently emerged as suitable alternatives to study bone formation and mineralization and to model human bone disorders. In addition to the many technical advantages that allow faster and larger studies, the availability of several fish models that efficiently mimic human osteopenia and osteoporosis phenotypes has stimulated the interest of the academia and industry toward a better understanding of the mechanisms of pathogenesis but also toward the discovery of new bone anabolic or antiresorptive compounds. This mini review recapitulates the in vivo teleost fish systems available to study low BMD disorders and highlights their applications and the recent advances in the field.UIDB/04326/2020, EAPA_151/2016/BLUEHUMANinfo:eu-repo/semantics/publishedVersio

Temporal and spatial expression patterns of pregnane X receptor and vitamin K epoxide reductase genes, two core molecular players on fish vitamin K homeostasis and skeletal development.

Vitamin K (VK) is a liposoluble vitamin known to be essential for bone metabolism by two different pathways: (i) by its role as a coenzyme in the gamma-carboxylation of some skeletal proteins (e.g. osteocalcin (OC) and matrix Gla protein (MGP); Price et al., 1998); and (ii) through its role in skeletal gene transcription via binding to the pregnane X receptor (PXR; Azuma et al., 2010)

Gene expression during regeneration of zebrafish (danio rerio) fins: relative expression levels of mineralization – related gla proteins

Most animals have the ability to regenerate epidermal injuries yet only a few can regenerate largely severed appendages that comprise several different tissues. Nowadays zebrafish is one of the most used metazoan models in regeneration studies in particular for investigation of molecular events during fin regeneration process. Fin regeneration starts through the formation of a blastema, a set of heterogeneous mesenchyma-like cells located between stump tissues and the wounded epidermis. This event, denominated epimorphic regeneration, comprises strict growth control and cell reprogramming leading to faithful restoration of the lost parts

Regular supplementation with antioxidants rescues Doxorubicin-Induced Bone deformities and mineralization delay in Zebrafish

Osteoporosis is characterized by an abnormal bone structure with low bone mass and degradation of microarchitecture. Oxidative stress induces imbalances in osteoblast and osteoclast activity, leading to bone degradation, a primary cause of secondary osteoporosis. Doxorubicin (DOX) is a widely used chemotherapy drug for treating cancer, known to induce secondary osteoporosis. The mechanism underlying DOX-induced bone loss is still not fully understood, but one of the relevant mechanisms is through a massive accumulation of reactive oxygen and nitrogen species (i.e., ROS and NOS) leading to oxidative stress. We investigated the effects of antioxidants Resveratrol and MitoTEMPO on DOX-induced bone impairment using the zebrafish model. DOX was shown to increase mortality, promote skeletal deformities, induce alterations on intestinal villi, impair growth and mineralization and significantly downregulate osteoblast differentiation markers osteocalcin 2 and osterix/sp7. Lipid peroxidation was significantly increased in DOX-supplemented groups as compared to control and antioxidants, suggesting ROS formation as one of the key factors for DOX-induced bone loss. Furthermore, DOX affected mineral contents, suggesting an altered mineral metabolism. However, upon supplementation with antioxidants, DOX-induced effects on mineral content were rescued. Our data show that supplementation with antioxidants effectively improves the overall growth and mineralization in zebrafish and counteracts DOX-induced bone anomalies.info:eu-repo/semantics/publishedVersio

Reversal of doxorubicin-Induced bone loss and mineralization by supplementation of Resveratrol and MitoTEMPO in the early development of Sparus aurata

Doxorubicin is a widely used chemotherapeutic drug known to induce bone loss. The mechanism behind doxorubicin-mediated bone loss is unclear, but oxidative stress has been suggested as a potential cause. Antioxidants that can counteract the toxic effect of doxorubicin on the bone would be helpful for the prevention of secondary osteoporosis. We used resveratrol, a natural antioxidant, and MitoTEMPO, a mitochondria-targeted antioxidant, to counteract doxorubicin-induced bone loss and mineralization on Sparus aurata larvae. Doxorubicin supplemented Microdiets increased bone deformities, decreased mineralization, and lipid peroxidation, whereas Resveratrol and MitoTEMPO supplemented microdiets improved mineralization, decreased bone deformities, and reversed the effects of doxorubicin in vivo and in vitro, using osteoblastic VSa13 cells. Partial Least-Squares Discriminant Analysis highlighted differences between groups on the distribution of skeletal anomalies and mineralization of skeleton elements. Calcium and Phosphorus content was negatively affected in the doxorubicin supplemented group. Doxorubicin reduced the mRNA expression of antioxidant genes, including catalase, glutathione peroxidase 1, superoxide dismutase 1, and hsp90 suggesting that ROS are central for Doxorubicin-induced bone loss. The mRNA expression of antioxidant genes was significantly increased on resveratrol alone or combined treatment. The length of intestinal villi was increased in response to antioxidants and reduced on doxorubicin. Antioxidant supplements effectively prevent bone deformities and mineralization defects, increase antioxidant response and reverse doxorubicin-induced effects on bone anomalies, mineralization, and oxidative stress. A combined treatment of doxorubicin and antioxidants was beneficial in fish larvae and showed the potential for use in preventing Doxorubicin-induced bone impairment.info:eu-repo/semantics/publishedVersio

Expression of Gla proteins during fish skeletal development

Senegal sole skeletal development; Skeletal malformations; Skeletal malformation in mediterranean species; Senegal sole skeletal deformities; Zebra fish as model system: skeletal development; Identification of bone cells / skeletal development; Spatial - temporal pattern of bgp expression; Single cell resolution: localization of bgp mRNA; Single cell resolution: Immunolocalization of Bgp; Single cell resolution: localization of mgp mRNA; Single cell resolution: Immunolocalization of Mgp; An in vivo system to uncover MGP and BGP function; Effects of warfarin on toadfish

Ensinar a brincar para aprender a viver

O presente Relatório de Estágio (RE) foi realizado no âmbito da Unidade Curricular (UC) da Prática de Ensino Supervisionada (PES), pertencente ao plano de estudos do Mestrado em Ensino do 1º Ciclo do Ensino Básico (CEB) e de Matemática e Ciências Naturais no 2º CEB. Neste documento encontram-se organizados pressupostos teóricos e legais que corroboram a prática docente bem como alguns momentos da PES do mestrando. No decorrer da prática, o mestrando teve contacto com duas turmas de cada ciclo (1º CEB e 2º CEB), havendo uma perspetiva constante de crescimento e desenvolvimento de estratégias e metodologias inovadoras, adequando-as às características específicas de cada um dos contextos, os quais chegaram a abranger um curto período de ensino remoto de emergência. As intervenções realizadas tinham por base um desenvolvimento exploratório de atividades digitais e gamificadas, tendo-se verificado momentos de observação, cooperação, planificação, implementação/ação e reflexão, alguns momentos de investigação. Neste documento, para além da apresentação de práticas dos contextos supramencionados, também se encontra a componente investigativa, em formato de artigo científico, a qual se associa com um estudo caso da implementação e reflexão sobre a utilização de um jogo de cartas de ciências como estratégia de motivação, incentivo e auxílio ao estudo num 5º ano de escolaridade. A mesma constitui um estudo caso do material didático, as cartas, averiguando qual o impacto das mesmas nas aprendizagens e progressões dos alunos. Durante a PES, destaca-se uma aposta num desenvolvimento de um trabalho apoiado em interesses específicos dos alunos, procurando deixá-los mais empenhados e entusiasmados, promovendo atividades que implique que os alunos sejam construtores frequentes do seu próprio conhecimento, fazendo-o através de atividades lúdicas. Deste modo, os alunos preparam-se para a vida, aprendendo com brincadeiras orientadas, mediadas e didaticamente ponderadas.This Internship Report (IR) was carried out within the scope of the Curricular Unit (CU) of Supervised Teaching Practice (STP), belonging to the study plan of the Master in Teaching of the 1st Cycle of Basic Education (CBE) and of Mathematics and Natural Sciences in the 2nd CBE. In this document are organized theoretical and legal assumptions that corroborate the teaching practice as well as some moments of the Master's STP. During the practice, the master's student had contact with two classes of each cycle (1st CBE and 2nd CBE), with a constant perspective of growth and development of innovative strategies and methodologies, adapting them to the specific characteristics of each of the contexts, one of them being held with a short period of emergency remote teaching. The interventions carried out were based on an exploratory development of digital and gamified activities, which lead to the foreseen moments of observation, cooperation, planning, implementation/action and reflection, some moments of investigation. In this document, in addition to the presentation of practices from the aforementioned contexts, there is also the investigative component, in the format of a scientific article, which is associated with a case study of the implementation and reflection of the use of a science card game as a strategy of motivation, incentive and study aid in a 5th year of school. It constitutes a case study of the didactic material, the cards, verifying the impact of the same in the learning and progression of the students. During the STP, there is a focus on the development of work based on the specific interests of students, having the objective of making them more committed and enthusiastic, promoting activities that imply that students are frequent builders of their own knowledge, doing it through playful activities. In this way, students prepare themselves for life, learning through guided, mediated and didactically weighted play

Localization of osteocalcin (BGP) during fish (Sparus aurata) development by in situ hybridization and immunohistochemistry: comparison between gene expression/protein distribution and skeletal mineralization

Osteocalcin (Bone Gla protein, BGP) is a small noncollagenous protein which is synthesized by osteoblasts and odontoblasts and is found exlusively in mineralized bony tissues. Although isolated for the first time in 1978, only recently has a function for this protein been suggested, specifically in controlling hydroxyapatite crystal growth. Appearance of osteocalcin could be linked to the presence of an hydroxyapatite-containing bony skeleton, since the protein was never found in cartilaginous fishes. Furthermore, within its primary sequence the amino acid residues known to be essential for its function are present in fish as well as in mammals, suggesting that function has been conserved over 400 million years of evolution. Taken totgether, these findings prompted us to study in detail the localization of osteocalcin gene expression in fish

New insights on vitamin K metabolism in senegalese sole (Solea senegalensis) based on ontogenetic and tissue-specific vitamin K epoxide reductase molecular data

Vitamin K (VK) is a key nutrient for several biological processes (e.g., blood clotting and bone metabolism). To fulfill VK nutritional requirements, VK action as an activator of pregnane X receptor (Pxr) signaling pathway, and as a co-factor of γ-glutamyl carboxylase enzyme, should be considered. In this regard, VK recycling through vitamin K epoxide reductases (Vkors) is essential and should be better understood. Here, the expression patterns of vitamin K epoxide reductase complex subunit 1 (vkorc1) and vkorc1 like 1 (vkorc1l1) were determined during the larval ontogeny of Senegalese sole (Solea senegalensis), and in early juveniles cultured under different physiological conditions. Full-length transcripts for ssvkorc1 and ssvkorc1l1 were determined and peptide sequences were found to be evolutionarily conserved. During larval development, expression of ssvkorc1 showed a slight increase during absence or low feed intake. Expression of ssvkorc1l1 continuously decreased until 24 h post-fertilization, and remained constant afterwards. Both ssvkors were ubiquitously expressed in adult tissues, and highest expression was found in liver for ssvkorc1, and ovary and brain for ssvkorc1l1. Expression of ssvkorc1 and ssvkorc1l1 was differentially regulated under physiological conditions related to fasting and re-feeding, but also under VK dietary supplementation and induced deficiency. The present work provides new and basic molecular clues evidencing how VK metabolism in marine fish is sensitive to nutritional and environmental conditions.FCT: UIDB/04326/2020. ALGASOLE-16-02-01-FMP-0058; RTI2018-099029-A-I00; RYC2018-025337-I.info:eu-repo/semantics/publishedVersio

Functional analysis of bone related Gla proteins from bony fish during skeletal development

Tese de Doutoramento, Biologia, Faculdade de Ciências do Mar e do Ambiente, Universidade do Algarve, 2005As proteinas dependentes da vitamina K, proteína Gla do osso (BGP ou osteocalcina) e proteína Gla da matriz (MGP) são pequenas proteinas que ligam o cálcio, tendo sido recentemente descobertas em peixe.The vitamin K dependent bone Gla protein (BGP or osteocalcin) and matrix Gla protein (MGP) are small calcium binding proteins only recently discovered in fish