In Vitro Anti-Inflammatory and Vasculoprotective Effects of Red Cell Extract from the Black Sea Urchin Arbacia lixula.

Sea urchins have emerged as an important source of bioactive compounds with anti-inflammatory and antioxidant properties relevant to human health. Since inflammation is a crucial pathogenic process in the development and progression of atherosclerosis, we here assessed the potential anti-inflammatory and vasculoprotective effects of coelomic red-cell methanolic extract of the black sea urchin Arbacia lixula in an in vitro model of endothelial cell dysfunction. Human microvascular endothelial cells (HMEC-1) were pretreated with A. lixula red-cell extract (10 and 100 µg/mL) before exposure to the pro-inflammatory cytokine tumor necrosis factor (TNF)-α. The extract was non-toxic after 24 h cell treatment and was characterized by antioxidant power and phenol content. The TNF-α-stimulated expression of adhesion molecules (VCAM-1, ICAM-1) and cytokines/chemokines (MCP-1, CCL-5, IL-6, IL-8, M-CSF) was significantly attenuated by A. lixula red-cell extract. This was functionally accompanied by a reduction in monocyte adhesion and chemotaxis towards activated endothelial cells. At the molecular level, the tested extract significantly counteracted the TNF-α-stimulated activation of the pro-inflammatory transcription factor NF-κB. These results provide evidence of potential anti-atherosclerotic properties of A. lixula red-cell extract, and open avenues in the discovery and development of dietary supplements and/or drugs for the prevention or treatment of cardiovascular diseases.

Effects of elevated temperature on gonadal functions, cellular apoptosis, and oxidative stress in Atlantic sea urchin Arbacia punculata.

Increasing seawater temperature affects growth, reproduction and development in marine organisms. In this study, we examined the effects of elevated temperatures on reproductive functions, heat shock protein 70 (HSP70) and nitrotyrosine protein (NTP, an indicator of reactive nitrogen species) expressions, protein carbonyl (PC, an indicator of oxidative stress) contents, cellular apoptosis, and coelomic fluid (CF) conditions in Atlantic sea urchin. Sea urchins were housed in six aquaria with control (24 °C) and elevated temperatures (28 °C and 32 °C) for a 7-day period. After exposure, sea urchins exhibited decreased percentages of gametes (eggs/sperm), as well as increased HSP70 and NTP expressions in eggs and spermatogenic cells, increased gonadal apoptosis, and decreased CF pH compared to controls. PC contents were also significantly increased in gonadal tissues at higher temperatures. These results suggest that elevated temperature acidifies CF, increases oxidative stress and gonadal apoptosis, and results in impairment of reproductive functions in sea urchins.

Food Modulation Controls Astaxanthin Accumulation in Eggs of the Sea Urchin Arbacia lixula.

The carotenoid astaxanthin has strong antioxidant properties with beneficial effects for various degenerative diseases. This carotenoid is produced by some microalgae species when cultivated in particular conditions, and, interestingly, it is a predominant carotenoid in aquatic animals throughout a broad range of taxa. Recently, astaxanthin was detected in the eggs of the sea urchin Arbacia lixula in relevant concentrations when this organism was maintained in culture. These results have paved the way for deeper research into astaxanthin production by this species, particularly in regards to how astaxanthin production can be modulated by diet. Results showed that the highest content of astaxanthin in eggs was observed in sea urchins fed on a diet enriched with Spirulina platensis. This result was confirmed by the high antioxidant activity recorded in the egg extracts of these animals. Our results suggest that (i) the sea urchin A. lixula is able to synthesize astaxanthin from precursors obtained from food, and (ii) it is possible to modulate the astaxanthin accumulation in sea urchin eggs by modifying the proportions of different food ingredients provided in their diet. This study demonstrates the large potential of sea urchin cultivation for the eco-sustainable production of healthy supplements for nutraceutical applications.

Volume regulation of intestinal cells of echinoderms: Putative role of ion transporters (Na(+)/K(+)-ATPase and NKCC).

Echinoderms are exclusively marine osmoconformer invertebrates. Some species occupy the challenging intertidal region. Upon salinity changes, the extracellular osmotic concentration of these animals also varies, exposing tissues and cells to osmotic challenges. Cells and tissues may then respond with volume regulation mechanisms, which involve transport of ions and water into and/or out of the cells, through ion transporters, such as the Na(+)/K(+)-ATPase and NKCC. The goal of this study was to relate the cell volume regulation capacity of echinoderm intestinal cells Na(+)/K(+)-ATPase and NKCC activities, in three echinoderm species: Holothuria grisea, Arbacia lixula, and Echinometra lucunter. Isolated cells of these species displayed some control of their cell volume upon exposure to anisosmotic media (isolated intestinal cells, calcein fluorescence as indicator of volume change), with a distinct higher capacity shown by H. grisea, which did not swell even upon 50% hyposmotic shock. The holothuroid cells showed indirect evidence (effect of furosemide) of the participation of NKCC in this process, with a secretory function, and of a secondary role by the NKA (effect of ouabain). Other mechanisms are probably responsible for this function in the urchins. Variable expression of these transporters, and others not examined here, may to some extent account for the variability in cell volume regulation capacity in echinoderm cells.

Expression and functional activity of neurotransmitter system components in sea urchins' early development.

Reverse-transcription polymerase chain reaction (RT-PCR) investigation of the expression of the components supposedly taking part in serotonin regulation of the early development of Paracentrotus lividus has shown the presence of transcripts of five receptors, one of which has conservative amino acid residues characteristic of monoaminergic receptors. At the early stages of embryogenesis the expressions of serotonin transporter (SERT) and noradrenaline transporter (NET) were also recognized. The activities of the enzymes of serotonin synthesis and serotonin transporter were shown using immunohistochemistry and incubation with para-chlorophenylalanine (PСРА) and 5-hydroxytryptophan (HTP). Pharmacological experiments have shown a preferential cytostatic activity of ligands characterized as mammalian 5-hydroxytryptamine (5-HT)1-antagonists. On the basis of the sum of the data from molecular biology and embryo physiological experiments, it is suggested that metabotropic serotonin receptors and membrane transporters take part in the regulatory processes of early sea urchin embryogenesis.

Spine and test skeletal matrices of the Mediterranean sea urchin Arbacia lixula--a comparative characterization of their sugar signature.

Calcified structures of sea urchins are biocomposite materials that comprise a minor fraction of organic macromolecules, such as proteins, glycoproteins and polysaccharides. These macromolecules are thought to collectively regulate mineral deposition during the process of calcification. When occluded, they modify the properties of the mineral. In the present study, the organic matrices (both soluble and insoluble in acetic acid) of spines and tests from the Mediterranean black sea urchin Arbacia lixula were extracted and characterized, in order to determine whether they exhibit similar biochemical signatures. Bulk characterizations were performed by mono-dimensional SDS/PAGE, FT-IR spectroscopy, and an in vitro crystallization assay. We concentrated our efforts on characterization of the sugar moieties. To this end, we determined the monosaccharide content of the soluble and insoluble organic matrices of A. lixula spines and tests by HPAE-PAD, together with their respective lectin-binding profiles via enzyme-linked lectin assay. Finally, we performed in situ localization of N-acetyl glucosamine-containing saccharides on spines and tests using gold-conjugated wheatgerm agglutinin. Our data show that the test and spine matrices exhibit different biochemical signatures with regard to their saccharidic fraction, suggesting that future studies should analyse the regulation of mineral deposition by the matrix in these two mineralized structures in detail. This study re-emphasizes the importance of non-protein moieties, i.e. sugars, in calcium carbonate systems, and highlights the need to clearly identify their function in the biomineralization process.

High density and ligand affinity confer ultrasensitive signal detection by a guanylyl cyclase chemoreceptor.

Guanylyl cyclases (GCs), which synthesize the messenger cyclic guanosine 3',5'-monophosphate, control several sensory functions, such as phototransduction, chemosensation, and thermosensation, in many species from worms to mammals. The GC chemoreceptor in sea urchin sperm can decode chemoattractant concentrations with single-molecule sensitivity. The molecular and cellular underpinnings of such ultrasensitivity are not known for any eukaryotic chemoreceptor. In this paper, we show that an exquisitely high density of 3 × 10(5) GC chemoreceptors and subnanomolar ligand affinity provide a high ligand-capture efficacy and render sperm perfect absorbers. The GC activity is terminated within 150 ms by dephosphorylation steps of the receptor, which provides a means for precise control of the GC lifetime and which reduces "molecule noise." Compared with other ultrasensitive sensory systems, the 10-fold signal amplification by the GC receptor is surprisingly low. The hallmarks of this signaling mechanism provide a blueprint for chemical sensing in small compartments, such as olfactory cilia, insect antennae, or even synaptic boutons.

Identification of ABC transporter genes in gonad tissue of two Mediterranean sea urchin species: black, Arbacia lixula L., and rocky, Paracentrotus lividus L.

Multixenobiotic resistance (MXR) represents an important cellular detoxification mechanism in aquatic organisms as it provides them robustness toward natural and man-made contaminants. Several ABC transporters have major roles in the MXR phenotype - P-gp/ABCB1, MRP1-3/ABCC1-3 and BCRP/ABCG2. In this study, we identified the presence of ABC transporters involved in the MXR mechanism of Arbacia lixula and Paracentrotus lividus. AlABCB1/P-gp, AlABCC3/MRP3, AlABCC9/SUR-like and AlABCG-like transcripts were identified in A. lixula; and PlABCC1/P-gp, PlABCC3/MRP3, PlABCC5/MRP5, and PlABCC9/SUR-like transcripts in P. lividus. For each of the new partial sequences, we performed detailed phylogenetic and identity analysis as a first step toward full characterization and understanding of the ecotoxicological role of these ABC transporters.

Direct relationship between osmotic and ionic conforming behavior and tissue water regulatory capacity in echinoids.

Echinoderms are considered marine osmoconforming invertebrates. However, many are intertidal or live next to estuaries, tolerating salinity changes and showing extracellular gradients to dilute seawater. Three species of echinoids - Lytechinus variegatus, which can occur next to estuarine areas, the rocky intertidal Echinometra lucunter, and the mostly subtidal Arbacia lixula - were submitted to a protocol of stepwise (rate of 2-3 psu/h) dilution, down to 15 psu, or concentration, up to 45 psu, of control seawater (35 psu). Coelomic fluid samples were obtained every hour. The seawater dilution experiment lasted 8h, while the seawater concentration experiment lasted 6h. Significant gradients (40-90% above value in 15 psu seawater) for osmolality, sodium, magnesium, and potassium were shown by L. variegatus and E. lucunter. A. lixula showed the smallest gradients, displaying the strongest conforming behavior. The esophagus of the three species was challenged in vitro with 20 and 50% osmotic shocks (hypo- and hyperosmotic). A. lixula, the most "conforming" species, showed the highest capacity to avoid swelling of its tissues upon the -50% hyposmotic shock, and was also the species less affected by salinity changes concerning the observation of spines and ambulacral feet movement in the whole-animal experiments. Thus, the most conforming species (A. lixula) displayed the highest capacity to regulate tissue water/volume, and was also the most euryhaline among the three studied species. In addition, tissues from all three species swelled much more than they shrank under osmotic shocks of same magnitude. This distinct trend to gain water, despite the capacity to hold some gradients upon seawater dilution, helps to explain why echinoderms cannot be fully estuarine, or ever enter fresh water.

Parental dietary effect on embryological development response to toxicants with the sea urchin Arbacia punctulata.

The role of echinoid parental nutrition in early-life stage toxicity is not well understood. Arbacia punctulata were fed either a fresh diet consisting of organic lettuce and carrots or a dry feed. Embryos from parents fed the dry feed exhibited lower sensitivity to copper, whereas the opposite occurred with 1,3,5-trinitrobenzene and sodium dodecyl sulfate (SDS). EC(50) values for the dry and fresh feed treatments, respectively, were 41.0 and 29.9 microg/L for copper, 0.5 and 1.8 mg/L for 1,3,5-trinitrobenzene, and 3.5 and 5.6 mg/L for SDS. The data suggests that nutritional standardization for sea urchins in ecotoxicological laboratories needs to be addressed and further investigated.