1,721,099 research outputs found
Gene expression and stress response in sea urchin embryos with skeleton defects caused by magnesium deprivation.
No full textEchinoderms have an extensive endoskeleton composed of magnesian calcite, a form of calcium carbonate that contains small amounts of magnesium carbonate and occluded matrix proteins [1]. In the frame of the Biomintec European Project focused on the understanding of basic biomineralization mechanisms for the design of novel strategies in nano-biotechnology, we studied the effects of magnesium deprivation on Arbacia lixula sea urchin embryo development. Embryos were morphologically monitored, evaluating developmental abnormalities at different endpoints (3, 6, 24, 48 and 72 hours). In parallel, the spatial transcriptional levels of a skeleton matrix protein (msp130) and the protein synthesis levels of a signaling protein (p38MAPk) were detected by in situ hybridization (ISH) and Western Blotting (WB). The morphological analysis evidenced a general delay in development soon after the morula stage (6 hours), the absence of biomineral deposition after 24 hours and severe skeleton malformations after 48-72 hours. We observed the ectopic localization of primary mesenchyme cells (PMCs), the only cells in the embryo directing skeleton formation, by immuno-fluorescence with a FITC-conjugated lectin (WGA) able to bind specifically the PMCs. By ISH we detected the msp130 mRNA correctly in PMCs, although it was found persistently expressed even at 48h after fertilization, unlike control embryos where the messenger is down-regulated at this stage. At 48 hours we found high levels of p38MAPk by WB, if compared to control embryos; preliminary experiments showed hsp70 levels similar to controls. Further experiments will be performed to characterize the role of magnesium in the biomineralization process: we have now a toolkit of probes for various skeletogenic-specific genes (msp130, SM30, SM50, p16, p19) [2, 3] that will be used to study their embryonic temporal and spatial expression profiles. The internal calcium content and eventually the calcium carbonate polymorph forms [4] occurring in embryonic spicules in Mg-deprived embryos will be studied in comparison to control embryo.
References
[1] Matranga V, Bonaventura R, Costa C, Karakostis K, Pinsino A, Russo R, Zito F. Echinoderms as blueprints for biocalcification: regulation of skeletogenic genes and matrices. Prog Mol Subcell Biol (2011) 52:225-48.
[2] Pinsino A, Roccheri MC, Costa C, Matranga V. Manganese interferes with calcium, perturbs ERK signaling, and produces embryos with no skeleton. Toxicol Sci (2011) 123:217-230.
[3] Costa C, Karakostis K, Zito F, Matranga V. Phylogenetic analysis and expression patterns of p16 and p19 in Paracentrotus lividus embryos. Dev Genes Evol (2012) 222:245-51.
[4] Raz S, Hamilton PC, Wilt FH, Weiner S, Addadi L. The transient phase of amorphous calcium carbonate in sea urchin larval spicules: the involvement of proteins and magnesium ions in its formation and stabilization. Adv Mater (2003) 13:480-486
Regolazione genica e biomineralizzazione in embrioni di riccio di mare.
No full textGli echinodermi possiedono un esteso endoscheletro composto da calcite di magnesio, una forma di carbonato di calcio che contiene piccole quantità di carbonato di magnesio e di proteine occluse nella matrice [1]. Nell’ambito del progetto europeo del 7FP Biomintec 2008-2012 (http://www.biomintec.de/), focalizzato sui meccanismi molecolari che stanno alla base dei processi di biomineralizzazione e volto alla futura applicazione di nuovi biomateriali, abbiamo studiato gli effetti della deprivazione di magnesio sullo sviluppo dell’embrione di riccio di mare Arbacia lixula. Gli embrioni sono stati analizzati morfologicamente, valutando l’impatto sullo sviluppo a vari stadi (3, 6, 24, 48 e 72 ore). Tramite ibridazione in situ (ISH) e Western Blotting (WB) sono stati valutati: la localizzazione del trascritto (mRNA) di una proteina della matrice scheletrica (msp130); i livelli della proteina p38MAPk, nota per la sua implicazione nella scheletogenesi embrionale. L’osservazione morfologica mediante microscopia ottica ha messo in evidenza un generale rallentamento dello sviluppo, visibile già allo stadio di morula (6 ore), l’assenza di formazione minerale dopo 24 ore, e gravi malformazioni scheletriche dopo 48-72 ore. Tramite l’uso di marcatori molecolari (WGA-FITC) abbiamo osservato mediante microscopia a fluorescenza la localizzazione ectopica delle cellule del mesenchima primario (PMCs), le uniche cellule deputate alla formazione dello scheletro. Inoltre, mediante ISH abbiamo valutato la trascrizione del gene PMC-specifico msp130, dimostrando la presenza di alti livelli del trascritto in tutte le PMCs anche a 48 ore di sviluppo, a differenza di quanto accade negli embrioni controllo in cui solo una parte delle PMCs sono marcate e per un tempo minore. A livello proteico, mediante WB abbiamo dimostrato un aumento nei livelli di p38MAPk rispetto agli embrioni controllo. Risultati preliminari indicano livelli proteici costanti di hsp70. Esperimenti in corso sono volti all’analisi dei profili di espressione spazio-temporale di alcuni geni implicati nella scheletogenesi, tra cui msp130, SM30, SM50, p16, p19, VEGF, VEGFR [1-3], mediante l’uso delle specifiche sonde molecolari prodotte nel nostro laboratorio. Ulteriori esperimenti saranno necessari per accertare il ruolo del magnesio nel complesso processo di biomineralizzazione, attraverso la quantificazione del contenuto interno di calcio e la determinazione delle forme polimorfiche di carbonato di calcio presenti nelle spicole degli embrioni coltivati in assenza di magnesio in confronto agli embrioni controllo [4].
Alcuni dei risultati sono parte della tesi di laurea magistrale in Biologia Cellulare e Molecolare di Chiara Martino, di cui è stato relatore la Chiar.ma Prof.ssa Maria Carmela Roccheri del Dipartimento di Scienze e Tecnologiche Biologiche, Chimiche e Farmaceutiche dell’Università degli Studi di Palermo.
Bibliografia
[1] Matranga V, Bonaventura R, Costa C, Karakostis K, Pinsino A, Russo R, Zito F. (2011) Echinoderms as blueprints for biocalcification: regulation of skeletogenic genes and matrices. Prog Mol Subcell Biol 52:225-48.
[2] Pinsino A, Roccheri MC, Costa C, Matranga V. (2011) Manganese interferes with calcium, perturbs ERK signaling, and produces embryos with no skeleton. Toxicol Sci 123:217-230.
[3] Costa C, Karakostis K, Zito F, Matranga V. (2012) Phylogenetic analysis and expression patterns of p16 and p19 in Paracentrotus lividus embryos. Dev Genes Evol 222:245-51.
[4] Raz S, Hamilton PC, Wilt FH, Weiner S, Addadi L. (2003) The transient phase of amorphous calcium carbonate in sea urchin larval spicules: the involvement of proteins and magnesium ions in its formation and stabilization. Adv Mater 13:480-486
Recensione a G. Ruffino, "Variazione diatopica in Sicilia. Cartografia elementare"
No full textSi recensisce il voume di G. Ruffino, “Variazione diatopiaca in Sicilia. Cartografia elementare”, Centro di studi filologici e linguistici sicilani, Palermo 2018. Il saggio ripropone, con minimi aggiornamenti, il fondamentale studio sull’assetto linguistico della Sicilia che lo stesso Autore aveva presentato – con il titolo di “Isoglosse siciliane” – in occasione del congresso su “Tre millenni di storia linguistica della Sicilia, svoltosi a Palermo nel 1983 (atti curati da Adriana Quattordio Moreschini, Giardini Editori e stampatori, Pisa 1984)
L’informatica nella ricerca geolinguistica. L’esperienza siciliana
No full textLa ricerca dialettologica ha risentito molto di alcuni momenti dello sviluppo tecnologico e, successivamente, di quello informatico, non soltanto sul piano del rinnovamento metodologico, ma anche su alcuni aspetti teorici di questa disciplina. Insieme ad alcune considerazioni di ordine generale, l’implicazione dell’informatica nella ricerca dialettologica, e in quella geolinguistica in particolare, sarà valutata e discussa attraverso il piano operativo di uno specifico progetto di ricerca geo(socio)linguistico: quello dell’Atlante Linguistico della Sicilia (ALS).Dialectological research has been widely affected by technological advances to some extent and, as a result, by computer sciences as well. This has had an impact not only on the level of methodological renewals, but also on the level of a theoretical rethinking of the discipline itself. In light of some general considerations, the implications of information technology in dialectological research, and, in particular, in geolinguistic research, will be evaluated and discussed by taking into account the operational plan relating to a specific geo(socio)-linguistic research project, namely, the Atlante Linguistico della Sicilia (ALS)
Toxic effects of engineered nanoparticles in the marine environment: model organisms and molecular approaches
No full textEngineered nanoparticles (ENPs) have been produced by nano-biotech companies in recent decades to generate innovative goods in various fields, including agriculture, electronics, biomedicine, manufacturing, pharmaceuticals and cosmetics. The nano-scale size of the particles can confer novel and significantly improved physical, chemical and biological properties to scientific phenomena and processes. As their applications to science and technology expand, the need to understand the putative noxious effects of ENPs on humans and ecosystems is becoming increasingly important. ENPs are emerging as a new class of pollutants with eco-toxicological impacts on marine ecosystems because the particles can end up in waterways and reach the sea. Recent laboratory studies in invertebrates and fishes suggest that exposure to ENPs could have harmful effects. Because there is not much data available for gauging the effects of ENPs on marine wildlife, the ultimate ecotoxicological impacts of chronic exposure to ENPs should be investigated further using laboratory tests and field studies. We propose the use of model organisms to understand the molecular pathways involved in the mechanisms that may be affected by exposure to ENPs. Sensitive and innovative molecular methods will provide information regarding the hazards of ENPs that may exist in the marine environment. Model organisms that have not been conventionally used for risk assessment and the development of eco-toxicogenomic approaches will result in an improved understanding of the mechanistic modes of action of contaminating ENPs in the marine environment. © 2012 Elsevier Ltd
Cellular, biochemical and molecular effects of cadmium on marine invertebrates: focus on Paracentrotus lividus sea urchin development
No full textCadmium is a heavy metal that is toxic for living organisms even at low
concentrations. The presence in the environment of this metal has grown because of its
large employment in some industrial and agricultural activities. Although heavy metals
are terrestrially produced, they flow into the sea through effluents and sewage or are
directly discharged from industries placed on the seawater front. In addition to its release
into costal waters, cadmium fallout, following atmospheric events, contributes to the
pollution of marine ecosystems. It should be considered that cadmium concentrations
determined in the field vary widely according to different seawater latitudes and depths
and can be strongly influenced by freshwater discharges from heavily polluted rivers.
Cadmium does not have any biological role and, since it cannot be degraded, it is
irreversibly accumulated into cells, interacting with cellular components and molecular
targets. Experimental evidence suggests that the metal crosses the plasma membrane as a
bivalent ion, exerting an agonistic role against calcium ionic channels. The metal has
been associated with blockage of oxidative phosphorylation, glutathione depletion and
antioxidant enzymatic activity inhibition, production of ROS, DNA damage, and
inhibition of relative repair mechanisms, a general reduction of protein synthesis coupled
with an increase in stress proteins. For many years, the effects of various pollutants onaquatic invertebrates have been studied. Most of these studies examined the
developmental defects due to the presence, in rearing media, of a specific toxicant such as
cadmium. Considering the capacity of sea urchin embryos and larvae to accumulate
contaminants during development, this developmental model offers an excellent
opportunity to investigate the possible adaptive response of cells exposed to cadmium
during differentiation. In this review, we will examine the cellular, biochemical and
molecular effects produced by cadmium ions on Paracentrotus lividus sea urchin
embryos. Specifically we will consider the following: i) Cd2+ accumulation during
embryonic development; ii) stress protein synthesis and accumulation (HSPs) and
metallothioneins; and iii) induction of apoptosis and related pathways
Sea urchin embryos as an in vivo model for the assessment of manganese toxicity: developmental and stress response effects.
No full textManganese (Mn), one of the most abundant metals in nature present in rocks, soil and water, is also found in soft bottom sediments of the oceans. It represents a trace element that is accumulated and utilized by all forms of life and plays multiple roles ranging from bone mineralization to cellular protection. Although Mn is an essential nutrient, exposure of cells/organisms to high levels of Mn cause toxicity. In the marine environment, increased concentrations of bio-available Mn often result from anthropogenic activities, and consequently, Mn represents a new important factor in environmental contamination. Emission of Mn into the marine environment occurs from metallurgic and chemical industries including municipal wastewater discharges, sewage sludge, mining and mineral processing, emissions from alloy, steel, and iron production. Thus, a sharp Mn solubility gradient exists along the oxic/anoxic interfaces in such water columns. During hypoxia, that in many costal areas occurs as a result of eutrophication, Mn is released in a divalent ionic form (Mn2+) and reaches high concentrations in bottom waters.
In this study, we investigated the effects of manganese chloride (MnCl2) on embryos of the sea urchin Paracentrotus lividus used as a model of Mn-induced toxicity. Embryos were continuously exposed to different MnCl2 concentrations from fertilization to the pluteus stage and examined after 6,12, 20, 24, 48, 72 hours. We found a severe dose-dependent inhibition of embryonic development, with consequent specific malformations. The EC50 value was determined in 7,7 mg/l, defined as the concentration causing 50% abnormal embryo development. These effects were rapidly reversed upon Mn washout, except after 40 hours of treatment. Major developmental defects consisted in the absence or reduced elongation of skeletal rods (spicules), suggesting a key role for Mn in embryonic skeleton development. Mn accumulation was determined in exposed embryos by AAS analysis and compared to physiological calcium (Ca) concentration found in the same specimens. We found that the Mn is accumulated into embryos in a time and dose-dependent manner, with a drastic increase 24 hrs post-fertilization. In contrast, Ca concentration is reduced in an opposite proportional way, consistent with its poor detection in primary mesenchyme cells, observed by in vivo labelling with cell permeable fluorescent calcein. A direct correlation has been observed between malformations, accumulation of Mn ions and the regulation of key stress proteins that provide the major protection against stressors. We found that Mn exposure caused an increase in HSC70 and HSC60 levels, but not in HSC90 levels and did not induce synthesis of the HSPs inducible forms. Analysis by 2D gel electrophoresis showed different patterns of protein spots in control and exposed embryos highlighting qualitative protein expression differences in response to Mn exposure. No increase in apoptosis in comparison to controls was measured by the TUNEL assay, as well as no reactive oxygen species (ROS) production was found by a fluorescent detection assay on live embryos. To conclude, developmental and stress response effects observed could be explained as a Mn toxicity linked to the accumulation and bio-concentration of Mn into the embryos tissues and a possible competition between Mn and Ca for membrane transport sites or other specific molecular targets
Manganese-exposed embryos as blueprints to study signaling pathways involved in development
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