Dataset Information

Sea urchin Paracentrotus lividus immune cells in culture: formulation of the appropriate harvesting and culture media and maintenance conditions.

ABSTRACT: The sea urchin is an emergent model system for studying basic and translational immunology. Here we report a new method for the harvesting and maintenance of primary immune cells isolated from adult Paracentrotus lividus, a common Mediterranean sea urchin species. This optimised method uses coelomocyte culture medium, containing a high-affinity Ca2+ chelator, as the ideal harvesting and anti-clotting vehicle and short-term culture medium (?48?h), and artificial seawater as the master medium that maintains cell survival and in vitro-ex vivo physiological homeostasis over 2?weeks. Gradually reducing the amount of anticoagulant solution in the medium and regularly replacing the medium led to improved culture viability. Access to a robust and straightforward in vitro-ex vivo system will expedite our understanding of deuterostome immunity as well as underscore the potential of sea urchin with respect to biomedicine and regulatory testing.This article has an associated First Person interview with the first author of the paper.

SUBMITTER: Pinsino A

PROVIDER: S-EPMC6451355 | biostudies-literature | 2019 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Sea urchin Paracentrotus lividus immune cells in culture: formulation of the appropriate harvesting and culture media and maintenance conditions.

Pinsino Annalisa A Alijagic Andi A

Biology open 20190305 3

The sea urchin is an emergent model system for studying basic and translational immunology. Here we report a new method for the harvesting and maintenance of primary immune cells isolated from adult Paracentrotus lividus, a common Mediterranean sea urchin species. This optimised method uses coelomocyte culture medium, containing a high-affinity Ca2+ chelator, as the ideal harvesting and anti-clotting vehicle and short-term culture medium (≤48 h), and artificial seawater as the ...[more]

PMID: 30718227

Similar Datasets

Project description:The sea urchin Paracentrotus lividus (Lamarck, 1816) is a keystone herbivore in the Mediterranean Sea due to its ability to transform macroalgal-dominated communities into barren areas characterized by increased cover of bare substrates and encrusting coralline algae, reduced biodiversity and altered ecosystem functions. P. lividus is also an excellent animal model for toxicology, physiology and biology investigations having been used for more than a century as a model for embryological studies with synchronously developing embryos which are easy to manipulate and analyze for morphological aberrations. Despite its importance for the scientific community, the complete genome is still not fully annotated. To date, only a few molecular tools are available and a few Next Generation Sequencing (NGS) studies have been performed. Here we aimed at setting-up an RNA extraction method to obtain high quality and sufficient quantity of RNA for NGS from P. lividus embryos at the pluteus stage. We compared five different RNA extraction protocols from four different pools of plutei (500, 1000, 2500 and 5000 embryos): TRIzol®, and four widely-used Silica Membrane kits, GenElute™ Mammalian Total RNA Miniprep Kit, RNAqueous® Micro Kit, RNeasy® Micro Kit and Aurum™ Total RNA Mini Kit. The quantity of RNA isolated was evaluated using NanoDrop. The quality, considering the purity, was measured as A260/A280 and A260/230 ratios. The integrity was measured by RNA Integrity Number (RIN). Our results demonstrated that the most efficient procedures were GenElute, RNeasy and Aurum, producing a sufficient quantity of RNA for NGS. The Bioanalyzer profiles and RIN values revealed that the most efficient methods guaranteeing for RNA integrity were RNeasy and Aurum combined with an initial preservation in RNAlater. This research represents the first attempt to standardize a method for high-quality RNA extraction from sea urchin embryos at the pluteus stage, providing a new resource for this established model marine organism.

Project description:Sea urchins act as a keystone herbivore in marine coastal ecosystems, regulating macrophyte density, which offers refuge for multiple species. In the Mediterranean Sea, both the sea urchin Paracentrotus lividus and fish preying on it are highly valuable target species for artisanal fisheries. As a consequence of the interactions between fish, sea urchins and macrophyte, fishing leads to trophic disorders with detrimental consequences for biodiversity and fisheries. In Sardinia (Western Mediterranean Sea), regulations for sea urchin harvesting have been in place since the mid 90s. However, given the important ecological role of P. lividus, the single-species fishery management may fail to take into account important ecosystem interactions. Hence, a deeper understanding of population dynamics, their dependance on environmental constraints and multispecies interactions may help to achieve long-term sustainable use of this resource. This work aims to highlight how sea urchin population structure varies spatially in relation to local environmental constraints and species interactions, with implications for their management. The study area (Sinis Peninsula, West Sardinia, Italy) that includes a Marine Reserve was divided into five sectors. These display combinations of the environmental constraints influencing sea urchin population dynamics, namely type of habitat (calcareous rock, granite, basalt, patchy and continuous meadows of Posidonia oceanica), average bottom current speed and predatory fish abundance. Size-frequency distribution of sea urchins under commercial size (<5 cm diameter size) assessed during the period from 2004 to 2007, before the population collapse in 2010, were compared for sectors and types of habitat. Specific correlations between recruits (0-1 cm diameter size) and bottom current speeds and between middle-sized sea urchins (2-5 cm diameter size) and predatory fish abundance were assessed. Parameters representing habitat spatial configuration (patch density, perimeter-to-area ratio, mean patch size, largest patch index, interspersion/juxtaposition index) were calculated and their influence on sea urchin density assessed. The density of sea urchins under commercial size was significantly higher in calcareous rock and was positively and significantly influenced by the density and average size of the rocky habitat patches. Recruits were significantly abundant in rocky habitats, while they were almost absent in P. oceanica meadows. The density of middle-sized sea urchins was more abundant in calcareous rock than in basalt, granite or P. oceanica. High densities of recruits resulted significantly correlated to low values of average bottom current speed, while a negative trend between the abundance of middle-sized sea urchins and predatory fish was found. Our results point out the need to account for the environmental constraints influencing local sea urchin density in fisheries management.

Dataset Information

Sea urchin Paracentrotus lividus immune cells in culture: formulation of the appropriate harvesting and culture media and maintenance conditions.

Publications

Sea urchin <i>Paracentrotus lividus</i> immune cells in culture: formulation of the appropriate harvesting and culture media and maintenance conditions.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets