Gli echinodermi come modelli per lo studio di Distruttori Endocrini: un approccio integrato

Gli echinodermi rappresentano dei validi modelli sperimentali nell\u2019ambito dell\u2019ecotossicologia marina. Essi infatti sono organismi bentonici a diretto contatto con potenziali fonti di xenobiotici e offrono un ampio range di processi biologici utili per questo tipo di approccio. In questo lavoro sono presentati i risultati pi\uf9 significativi ottenuti nel nostro laboratorio utilizzando questi organismi come modelli per lo studio degli effetti di potenziali Distruttori Endocrini (ED). Sono stati presi in esame due diverse specie di echinodermi, il riccio di mare Paracentrotus lividus e il crinoide Antedon mediterranea, e due diversi aspetti della loro fisiologia, la biologia riproduttiva e lo sviluppo rigenerativo, rispettivamente. Esemplari adulti di entrambe le specie sono stati esposti a diversi ED, potenzialmente in grado di interferire con gli ormoni sessuali. Le concentrazioni utilizzate erano comparabili a quelle ambientali. La ricerca si \ue8 basata su un approccio multidisciplinare che ha previsto analisi morfologiche, chimiche e biochimiche. I risultati ottenuti hanno evidenziato la capacit\ue0 di questi composti di interferire con diversi parametri quali la crescita rigenerativa, il pattern istologico e il diametro delle uova prodotte, sebbene lo specifico meccanismo d\u2019azione sia ancora da chiarire. Infatti, nel caso di P. lividus, la somministrazione diretta di ormoni sessuali, quali l\u2019estradiolo, non produce effetti significativi sulla biologia riproduttiva dell\u2019animale. Nel complesso i risultati ottenuti sottolineano sia l\u2019importanza di utilizzare modelli alternativi nei test ecotossicologici sia la necessit\ue0 di approfondire maggiormente la fisiologia di base degli organismi modello

New insight into mutable collagenous tissue : work in progress and applied perspectives in Paracentrotus lividus

The mechanically adaptable connective tissue of echinoderms (Mutable Collagenous Tissue, MCT), which can undergo drastic nervously-mediated changes in stiffness, tensile strength and viscosity, represents a promising model for biomaterial design and biomedical applications. MCT could be a source of inspiration for new composite materials whose molecular interactions and structural conformation can be changed in response to external stimuli. MCT is composed of collagen fibrils comparable to those of mammals plus other fibrillar structures, proteoglycans and glycoproteins. According to literature, the extracellular matrix of holothurians includes at least two glycoproteins, stiparin and tensilin, that can modulate the aggregation of collagen fibrils and their capacity for reciprocal sliding. This contribution presents the latest results of a detailed analysis of MCT components in Paracentrotus lividus: focusing on biochemical characterization of the fibrillar components (extraction, purification and quantification) and biomolecular analysis of the glycoprotein components. The final aims will be to confirm the presence and the role of these glycoproteins in echinoids and to manipulate simpler components in order to produce a composite with mutable mechanical properties. In the long term, MCT could provide inspiration for biomimetic materials and offer great potential for economically relevant biotechnological and clinical applications that require the controlled and reversible plasticization and/or stiffening of connective tissue

Echinoderms are valid deuterostome marine invertebrate models to study repair phase events after arm injury

Echinoderms are often subjected to traumatic amputations that damage or remove whole body parts i.e. arms. After such severe injuries, the repair phase must be effective with rapid emergency reaction and re-epithelialisation as well finely regulated extracellular matrix (ECM) remodelling to ensure subsequent arm regeneration. Here, we used the brittle star Amphiura filiformis (Ophiuroidea) and the starfish Echinaster sepositus (Asteroidea) as valid deuterostome marine invertebrate models to study similarities and differences in the repair phase phenomena of these two echinoderm species and discuss them in comparison with those of animals with limited regenerative abilities (i.e. mammals). To achieve this goal, we used an integrated approach based on both microscopy and molecular analyses. We showed that in both echinoderm models, immediately after injury, emergency reaction and re-epithelialisation are extremely rapid and more efficient than those displayed by mammals. The remodelling and the formation of the ECM, mainly collagen, is ensured by delayed activation of ECM genes and protein deposition and, together with absence of fibrosis (i.e. over-deposition of ECM), seem to be advantageous for regeneration-competent animals in comparison to mammals. Overall, we found that the echinoderm species here studied show comparable repair events. The differences between regeneration-competent and non-competent animals suggest that rapid wound closure and delayed ECM deposition are necessary to ensure an effective regeneration of whole lost body parts. Further molecular and functional analyses must be performed to confirm this hypothesis

Ultrastructural and biochemical characterization of mechanically adaptable collagenous structures in the edible sea urchin Paracentrotus lividus

The viscoelastic properties of vertebrate connective tissues rarely undergo significant changes withinphysiological timescales, the only major exception being the reversible destiffening of the mammalianuterine cervix at the end of pregnancy. In contrast to this, the connective tissues of echinoderms (seaurchins, starfish, sea cucumbers, etc.) can switch reversibly between stiff and compliant conditions intimescales of around a second to minutes. Elucidation of the molecular mechanism underlying suchmutability has implications for the zoological, ecological and evolutionary field. Important informationcould also arise for veterinary and biomedical sciences, particularly regarding the pathological plasti-cization or stiffening of connective tissue structures. In the present investigation we analyzed aspectsof the ultrastructure and biochemistry in two representative models, the compass depressor ligamentand the peristomial membrane of the edible sea urchin Paracentrotus lividus, compared in three differentmechanical states. The results provide further evidence that the mechanical adaptability of echinodermconnective tissues does not necessarily imply changes in the collagen fibrils themselves. The higher gly-cosaminoglycan (GAG) content registered in the peristomial membrane with respect to the compassdepressor ligament suggests a diverse role of these molecules in the two mutable collagenous tissues.The possible involvement of GAG in the mutability phenomenon will need further clarification. Duringthe shift from a compliant to a standard condition, significant changes in GAG content were detected onlyin the compass depressor ligament. Similarities in terms of ultrastructure (collagen fibrillar assembling)and biochemistry (two alpha chains) were found between the two models and mammalian collagen.Nevertheless, differences in collagen immunoreactivity, alpha chain migration on SDS-PAGE and BLASTalignment highlighted the uniqueness of sea urchin collagen with respect to mammalian collagen

Regeneration in Echinoderms: repair, regrowth, cloning

Regenerative potential is expressed to a maximum extent in echinoderms. It is a common phenomenon in all the classes, extensively employed to reconstruct external appendages and internal organs often subjected to amputation, self-induced or traumatic, rapidly followed by complete successful re-growth of the lost parts. Regeneration has been studied in adult individuals as well as in larvae. In armed echinoderms, regeneration of arms is obviously frequent: in many cases, the detached body fragments can undergo phenomena of partial or total regeneration independently of the donor animal, and, in a few cases (asteroids), the individual autotomised arms can even regenerate to produce new complete adults, offering superb examples of cloning strategies. In the species examined so far most results throw light on aspects related to wound healing, growth, morphogenesis and differentiation, even though in most cases many crucial questions remain unanswered. The present paper provides an overview of the current understanding of the phenomenon and covers the main biological aspects of regeneration giving an idea of the \u201cstate of the art\u201d across the phylum in terms of experimental approaches and representative models

Z-line and supercontraction in the hydraulic muscular systems of insect larvae

An electron microscopic study has been made on the body wall muscles from Lepidoptera (Noctua pronuba, Philosamia cynthia) and Diptera larvae (Tipula tipula), whose locomotion is based on a hydraulic muscle antagonism. All the fibers examined had interrupted Z\u2010bands, formed of single dense bodies, irregularly placed around the sarcomere; myosin filaments could pass through the spaces between one Z\u2010body and the next, making it possible for the sarcomere to shorten its resting length by up to 60%. In supercontracted muscles, both the thick and the thin filaments pass through the Z\u2010line; since these actin filaments are very long, they can overlap and interact with the myosin filaments of the adjacent sarcomeres. The muscles examined can be placed in the category of supercontracting muscles and can be considered a type of striated muscle, with morphological and functional characteristics similar to those of helical or smooth muscle

Regenerative response and Endocrine Disrupters in crinoid Echinoderms: an old experimental model, a new ecotoxicological test

The regenerative phenomena that reproduce developmental processes in adult organisms and are regulated by endocrine and neurohumoral mechanisms can provide new sensitive tests for monitoring the effects of exposure to anthropogenic chemicals such as endocrine disrupter (ED) contaminants. These pollutants in fact can be bioaccumulated by the organisms, causing dysfunctions in steroid hormone production/metabolism and activities and inducing dramatic effects on reproductive competence, development and growth in many animals, man included. Current research is exploring the effects of exposure to different classes of compounds well known for their ED activity, such as polychlorinated biphenyls (PCBs), nonylphenols and organotins, on regenerative potential of echinoderms, a relatively unexplored and promising applied approach which offers the unique chance to study physiological developmental processes in adult animals. The selected test species is the crinoid Antedon mediterranea, which represents a valuable experimental model for investigation into the regenerative process from the macroscopic to the molecular level. The present study employs an integrated approach which combines exposure experiments, chemical analysis and biological analysis utilizing classical methods of light (LM) and electron (TEM and SEM) microscopy and immunocytochemistry. The experiments were carried out on experimentally induced arm regenerations in controlled conditions with exposure concentrations comparable to those of moderately polluted coastal zones in order to reproduce common conditions of exposure to environmental contaminants. The results of the exposure tests were analysed in terms of effects at the whole organism, at the tissue and cellular level, and possible sites of action of EDs. Our results show that prolonged exposure to these compounds significantly affects the regenerative mechanisms by inducing appreciable anomalies in terms of regeneration times, overall growth, general morphology and histological and cellular pattern. A concentration/effect relationship could be found for all substances. Interestingly, contrasting results in terms of inhibition or acceleration of regeneration phenomenon were obtained for different chemicals