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1.
Molecules ; 26(11)2021 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-34205146

RESUMO

The objective of the present study was to investigate the proximate composition, antiradical properties and hepatoprotective activity of three species of shellfish, Corbicula japonica, Spisula sachalinensis, and Anadara broughtonii, from the coastal areas of Far East Russia. Biologically active peptides such as taurine (3.74 g/100 g protein) and ornithine (2.12 g/100 g protein) have been found in the tissues of A. broughtonii. C. japonica contains a high amount of ornithine (5.57 g/100 g protein) and taurine (0.85 g/100 g protein). The maximum DPPH and ABTS radical scavenging activity (36.0 µg ascorbic acid/g protein and 0.68 µmol/Trolox equiv/g protein, respectively) was determined for the tissue of C. japonica. The protein and peptide molecular weight distribution of the shellfish tissue water extracts was investigated using HPLC. It was found that the amount of low molecular weight proteins and peptides were significantly and positively correlated with radical scavenging activity (Pearson's correlation coefficient = 0.96), while the amount of high molecular weight proteins negatively correlated with radical scavenging activity (Pearson's correlation coefficient = -0.86). Hepatoprotective activity, measured by the survival rate of HepG2 hepatocytes after cotreatment with t-BHP, was detected for C. japonica. The highest protection (95.3 ± 2.4%) was achieved by the cold water extract of C. japonica at the concentration of 200 mg/mL. Moreover, oral administration of hot water extract of C. japonica to rats before the treatment with CCl4 exhibited a markedly protective effect by lowering serum levels of ALT and AST, inhibiting the changes in biochemical parameters of functional state of rat liver, including MDA, SOD, GSH and GST.


Assuntos
Antioxidantes/farmacologia , Arcidae/química , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Corbicula/química , Hepatócitos/citologia , Frutos do Mar/análise , Spisula/química , terc-Butil Hidroperóxido/efeitos adversos , Administração Oral , Animais , Antioxidantes/química , Tetracloreto de Carbono/efeitos adversos , Sobrevivência Celular , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Humanos , Masculino , Peso Molecular , Ornitina/isolamento & purificação , Ratos , Federação Russa , Frutos do Mar/classificação , Taurina/isolamento & purificação
2.
Mar Pollut Bull ; 161(Pt B): 111740, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33128982

RESUMO

In this study, we assessed the Atlantic surfclam (Spisula solidissima) energy budget under different ocean acidification conditions (OA). During 12 weeks, 126 individuals were maintained at three different ρCO2 concentrations. Every two weeks, individuals were sampled for physiological measurements and scope for growth (SFG). In the high ρCO2 treatment, clearance rate decreased and excretion rate increased relative to the low ρCO2 treatment, resulting in reduced SFG. Moreover, oxygen:nitrogen (O:N) excretion ratio dropped, suggesting that a switch in metabolic strategy occurred. The medium ρCO2 treatment had no significant effects upon SFG; however, metabolic loss increased, suggesting a rise in energy expenditure. In addition, a significant increase in food selection efficiency was observed in the medium treatment, which could be a compensatory reaction to the metabolic over-costs. Results showed that surfclams are particularly sensitive to OA; however, the different compensatory mechanisms observed indicate that they are capable of some temporary resilience.


Assuntos
Spisula , Animais , Homeostase , Humanos , Concentração de Íons de Hidrogênio , Oceanos e Mares , Água do Mar
3.
PLoS One ; 13(6): e0199212, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29902260

RESUMO

The Atlantic surfclam (Spisula solidissima) is a commercially important species in North American waters, undergoing biological and ecological shifts. These are attributed, in part, to environmental modifications in its habitat and driven by climate change. Investigation of shell growth patterns, trace elements, and isotopic compositions require an examination of growth lines and increments preserved in biogenic carbonates. However, growth pattern analysis of S. solidissima is challenging due to multiple disturbance lines caused by environmental stress, erosion in umbonal shell regions, and constraints related to sample size and preparation techniques. The present study proposes an alternative method for describing chronology. First, we analyzed growth patterns using growth lines within the shell and hinge. To validate the assumption of annual periodicity of growth line formation, we analyzed the oxygen isotope composition of the outer shell layer of two specimens (46°54'20"N; 56°18'58"W). Maximum δ18Oshell values occurred at the exact same location as internal growth lines in both specimens, confirming that they are formed annually and that growth ceases during winter. Next, we used growth increment width data to build a standardized growth index (SGI) time-series (25-year chronology) for each of the three parts of the shell. Highly significant correlations were found between the three SGI chronologies (p < 0.001; 0.55 < τ < 0.68) of all specimens. Thus, ligament growth lines provide a new method of determining ontogenetic age and growth rate in S. solidissima. In a biogeographic approach, the shell growth performance of S. solidissima in Saint-Pierre and Miquelon was compared to those in other populations along its distribution range in order to place this population in a temporal and regional context.


Assuntos
Exoesqueleto/crescimento & desenvolvimento , Ligamentos/crescimento & desenvolvimento , Oceanos e Mares , Spisula/crescimento & desenvolvimento , Animais , América do Norte , Isótopos de Oxigênio/análise , Spisula/química
4.
Artigo em Inglês | MEDLINE | ID: mdl-29689308

RESUMO

Atlantic surfclam populations have significantly declined in state and federal waters from the south shore of Long Island, New York to the Delmarva Peninsula since the early 2000s. Previous studies have demonstrated that surfclams in this geographic range show signs of physiological stress, suggested to be a result of increasing ocean temperatures. In this study, we examined the effect of 2 temperature regimes (19 °C and 23 °C) on surfclam physiology. These temperatures were chosen because they represent maximal (23 °C) and minimal (19 °C) temperatures prevailing in New York clamming areas during summer. Results demonstrated enhanced energy metabolism and significant reductions in filtration rate, scope for growth, and immune functions in clams exposed to the warmer temperature treatment. Although net energy gains remained positive in both treatments under our experimental conditions, the findings suggest that temperature stress is involved in the recent observations of surfclams in poor condition. The impact of elevated temperatures on phytoplankton quantity/quality and other environmental variables in combination with the direct impact on surfclam filtration and metabolic rates could lead to a negative energy balance. While some uncertainties remain about population-scale impacts of overall warming trends, we fear that future increases in temperature may lead to the collapse of the Atlantic surfclam between New York and Virginia, especially within inshore regions.


Assuntos
Spisula/fisiologia , Estresse Fisiológico , Temperatura , Animais , Metabolismo Energético , Estações do Ano , Spisula/crescimento & desenvolvimento , Spisula/imunologia , Spisula/metabolismo
5.
Sci Rep ; 7(1): 6930, 2017 07 31.
Artigo em Inglês | MEDLINE | ID: mdl-28761142

RESUMO

Tandemly repeated DNAs usually constitute significant portions of eukaryotic genomes. In bivalves, however, repetitive DNAs are habitually not widespread. In our search for abundant repetitive DNAs in trough shells, we discovered a novel satellite DNA, SSUsat, which constitutes at least 1.3% of the genome of Spisula subtruncata. As foreseen by the satellite DNA library hypothesis, we confirmed that this satellite DNA is also present in two other Mactridae species, showing a highly conserved nucleotide sequence together with a dramatic diminution in the number of repeats. Predominantly located at the G + C-rich intercalary heterochromatin of S. subtruncata, SSUsat displays several DNA methylation peculiarities. The level of methylation of SSUsat is high (3.38%) in comparison with bivalve standards and triplicates the mean of the S. subtruncata genome (1.13%). Methylation affects not only the cytosines in CpG dinucleotides but also those in CHH and CHG trinucleotides, a feature common in plants but scarce and without any clear known relevance in animals. SSUsat segments enriched in methylated cytosines partly overlap those showing higher sequence conservation. The presence of a chromosome pair showing an accumulation of markedly under-methylated SSUsat monomers additionally indicates that the methylation processes that shape repetitive genome compartments are quite complex.


Assuntos
Metilação de DNA , DNA Satélite/genética , Spisula/genética , Animais , Composição de Bases , Mapeamento Cromossômico , Heterocromatina/genética , Análise de Sequência de DNA
6.
J Gerontol A Biol Sci Med Sci ; 70(4): 434-43, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24780585

RESUMO

The susceptibility of membrane lipids to peroxidation (peroxidation index [PI]) increases with the double bond content of fatty acids and is inversely correlated to longevity in mammals, birds, and bivalve molluscs. In molluscs, membrane polyunsaturated fatty acids content can be affected by temperature, nutrition, and the individual's age. In this study, we evaluated how these three parameters may alter correlations between PI and longevity. We determined the fatty acid and dimethyl acetal compositions of phospholipids from gill mitochondrial and nonmitochondrial preparations from the short-lived Spisula solidissima (maximum longevity = 37 years) and the long-lived Arctica islandica (maximum longevity = 507 years) exposed to diet abundance and temperature (season) treatments. We also evaluated the effect of individual age on PI in S. solidissima (from 6 to 23 years). The temperature increase from winter to summer (2 to 12°C) coincided with decreases in values of PI, proportions of eicosapentaenoic acid, and dimethyl acetals. Higher microalgae supplementation increased polyunsaturated fatty acids and PI and decreased dimethyl acetals; age did not affect the PI in S. solidissima. Our finding that the PI of A. islandica remained significantly lower than that of S. solidissima in corresponding fractions throughout treatments suggests that longevity-related differences in PI are resilient to environmental conditions.


Assuntos
Envelhecimento/metabolismo , Dieta , Peroxidação de Lipídeos , Longevidade , Mercenaria , Mitocôndrias/metabolismo , Estações do Ano , Spisula , Temperatura , Animais , Ácido Eicosapentaenoico/metabolismo , Ácidos Graxos Insaturados/metabolismo , Brânquias/metabolismo , Isobutiratos/metabolismo , Lipídeos de Membrana/metabolismo , Mercenaria/metabolismo , Spisula/metabolismo
7.
Cryo Letters ; 35(6): 537-43, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25583015

RESUMO

BACKGROUND: The signaling of insulin-like growth factor-I (IGF-I) is involved in the development, growth, reproduction and aging of vertebrates. However, few studies have investigated the involvement of IGF-I during states of extreme shock, such as those induced by potently toxic cryoprotective agents (CPAs) or low temperature conditions, in bivalves. OBJECTIVE: We investigated the toxicity of CPAs and the potential relationship between larval viability and the IGF-I receptor (IGF-IR) after treatment with CPAs or freezing in surf clam (Spisula sachalinensis) larvae. MATERIALS AND METHODS: The umbo larvae and different concentrations of CPAs (dimethyl sulfoxide, DMSO; ethylene glycol, EG) were used to investigate the toxicity of CPAs and the vitrification of surf clam larvae. The relationship between larval viability and the IGF-I receptor (IGF-IR) after treatment with CPAs or freezing was investigated using immunoblot analysis. RESULTS: An increase in concentration greater than 4M DMSO was fatal in larvae; however, 5M EG combined with a mixture of CPAs had no harmful effects. Moreover, live larvae immersed in a 5M EG solution remained intact and maintained their normal shape and organs. However, even though the larvae survived the CPA toxicity test, none of the vitrified larvae survived. After immersion into CPAs and vitrification, 97-kDa IGF-IR ß-subunits could be detected in all larvae; but tyrosine phosphorylation of the intracellular ß-subunits was detected only in the control and live groups. CONCLUSION: IGF-IR was activated in the umbo larvae but not in dead surf clam larvae treated with CPA and frozen. Activation of IGF-IR has relevance to the umbo larval stage in live surf clams treated with CPAs.


Assuntos
Crioprotetores/toxicidade , Dimetil Sulfóxido/toxicidade , Etilenoglicol/toxicidade , Receptor IGF Tipo 1/metabolismo , Spisula/efeitos dos fármacos , Spisula/enzimologia , Animais , Criopreservação , Larva/efeitos dos fármacos , Larva/enzimologia , Larva/fisiologia , Fosforilação , Spisula/fisiologia , Vitrificação
8.
Aging Cell ; 12(4): 584-92, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23566066

RESUMO

The observation of an inverse relationship between lifespan and mitochondrial H2O2 production rate would represent strong evidence for the disputed oxidative stress theory of aging. Studies on this subject using invertebrates are surprisingly lacking, despite their significance in both taxonomic richness and biomass. Bivalve mollusks represent an interesting taxonomic group to challenge this relationship. They are exposed to environmental constraints such as microbial H2S, anoxia/reoxygenation, and temperature variations known to elicit oxidative stress. Their mitochondrial electron transport system is also connected to an alternative oxidase that might improve their ability to modulate reactive oxygen species (ROS) yield. Here, we compared H2O2 production rates in isolated mantle mitochondria between the longest-living metazoan--the bivalve Arctica islandica--and two taxonomically related species of comparable size. In an attempt to test mechanisms previously proposed to account for a reduction of ROS production in long-lived species, we compared oxygen consumption of isolated mitochondria and enzymatic activity of different complexes of the electron transport system in the two species with the greatest difference in longevity. We found that A. islandica mitochondria produced significantly less H2O2 than those of the two short-lived species in nearly all conditions of mitochondrial respiration tested, including forward, reverse, and convergent electron flow. Alternative oxidase activity does not seem to explain these differences. However, our data suggest that reduced complex I and III activity can contribute to the lower ROS production of A. islandica mitochondria, in accordance with previous studies. We further propose that a lower complex II activity could also be involved.


Assuntos
Peróxido de Hidrogênio/metabolismo , Longevidade/fisiologia , Mitocôndrias/metabolismo , Mya/metabolismo , Spisula/metabolismo , Animais , Respiração Celular , Transporte de Elétrons , Ativação Enzimática , Proteínas Mitocondriais/metabolismo , Mya/fisiologia , Oxirredutases/metabolismo , Consumo de Oxigênio , Proteínas de Plantas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Spisula/fisiologia , Fatores de Tempo
9.
Toxicon ; 65: 41-58, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23369833

RESUMO

The periodic occurrence of Paralytic Shellfish Poisoning (PSP) toxins in UK surf clams and the recent move away from biological assays for PSP testing resulted in the need to determine method performance characteristics for the replacement analytical method in this species. With the requirement for laboratory reference materials to aid this validation together with known issues relating to toxin transformation in live clams and homogenised tissue, there was the need to assess the toxin transformation characteristics of PSP toxins in surf clam tissue. Initial work examined the rates of toxin transformation in UK surf clam tissue incubated with toxin standards, showing rapid transformation of N-sulfocarbamoyl toxins with slower transformation of carbamate toxins. Full transformational pathways were determined using a combination of three different analytical methods and confirmed the major expected transformations involving decarbamoylation, with some evidence for additional reaction pathways. Results obtained from the analysis of surf clam and oyster tissues incubated with varying concentrations of toxic Alexandrium algae highlighted expected transformation reactions, although significant differences were observed in the extent of the transformations amongst the range of toxins studied, with less efficient transformation of N-hydroxylated toxins as compared with other carbamate and N-sulfocarbamoyl toxins. Analysis of PSP-toxic incurred oyster, scallop and mussel tissues incubated with variable proportions of surf clam tissue showed large differences in the extent of the transformations. Total conversion of N-sulfocarbamoyl toxins was confirmed at low relative proportions of surf clam tissue in all three species, whereas transformation of carbamate toxins was found to occur only in the presence of higher proportions of surf clam tissue in oysters and mussels in comparison with scallops. Results enabled the production of three laboratory reference materials prepared following incubation of incurred homogenates with optimum proportions of surf clam tissue, resulting in materials containing a large number of PSP toxins. Stability experiments provided good preliminary evidence for the stability of these targeted materials under storage conditions. The work therefore provides both additional information relating to the transformational activity in UK surf clams and highlights a good potential method for the targeted production of reference materials which include a wider range of toxins than normally present in naturally incurred shellfish.


Assuntos
Inspeção de Alimentos/métodos , Saxitoxina/análise , Spisula , Animais , Monitoramento Ambiental/métodos , Contaminação de Alimentos/análise , Saxitoxina/química , Intoxicação por Frutos do Mar/prevenção & controle
10.
RNA Biol ; 10(3): 391-6, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23324608

RESUMO

The nucleolinus is a nuclear subcompartment long ago posited to play a role in cell division. In a recent study using surf clam oocytes, cytoplasmic foci containing a nucleolinar protein were shown to later recruit γ-tubulin, identifying them as centrosomal precursors. (1) We now demonstrate the presence of structural RNAs from the nucleolinus in these procentrosomes. They include the well-known but poorly understood rRNA-transcribed spacer regions. In situ hybridization revealed a specific and dynamic association of these structural RNAs with the cell division apparatus that extends through the early stages of meiosis. In addition to their bearing on the debate over the nature of centrosome- and spindle-associated RNAs, the observations also suggest that rRNA spacer regions are not simply waste products to be discarded immediately, but may be functional byproducts that play a role in formation of the cell division apparatus.


Assuntos
Estruturas do Núcleo Celular/metabolismo , Centrossomo/fisiologia , RNA Ribossômico/genética , Spisula/genética , Tubulina (Proteína)/metabolismo , Animais , Estruturas do Núcleo Celular/genética , Citoplasma/metabolismo , DNA Espaçador Ribossômico/genética , Feminino , Meiose , Oócitos/fisiologia , RNA Ribossômico/metabolismo , Fuso Acromático/fisiologia , Spisula/metabolismo
11.
Artigo em Inglês | MEDLINE | ID: mdl-22356233

RESUMO

In 2006, a group of scientists studying centrosomes of Spisula solidissima mollusc oocytes under the leadership of Alliegro (Alliegro, M.C.; Alliegro, M.A.; Palazzo, R.E. Centrosome-associated RNA in surf clam oocytes. Proc. Natl. Acad. Sci. USA 2006, 103(24), 9034-9038) reliably demonstrated the existence of specific RNA in centrosome, called centrosomal RNA (cnRNA). In their first article, five different RNAs (cnRNAs 11, 102, 113, 170, and 184) were described. During the process of full sequencing of the first transcript (cnRNA 11), it was discovered that the transcript contained a conserved structure-a reverse transcriptase domain located together with the most important centrosomal protein, γ-tubulin. In an article published in 2005, we made assumptions about several possible mechanisms for determining the most important functions of centrosomal structures and referred to one of them as a "RNA-dependent mechanism." This idea about participation of hypothetic centrosomal small interference RNA and/or microRNA in the process was made one year prior to the discovery of cnRNA by Alliegro's group. The discovery of specific RNA in a centrosome is indirect evidence of a centrosomal hypothesis of cellular ageing and differentiation. The presence of a reverse transcriptase domain in this type of RNA, together with its uniqueness and specificity, makes the centrosome a place of information storage and reproduction.


Assuntos
Diferenciação Celular/fisiologia , Senescência Celular/fisiologia , Centrossomo/química , Modelos Biológicos , RNA/genética , Spisula/genética , Animais , Diferenciação Celular/genética , Senescência Celular/genética , DNA Polimerase Dirigida por RNA/genética , Spisula/enzimologia
12.
J Biomed Opt ; 17(1): 015001, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-22352645

RESUMO

This study combines microfluidics with optical microablation in a microscopy system that allows for high-throughput manipulation of oocytes, automated media exchange, and long-term oocyte observation. The microfluidic component of the system transports oocytes from an inlet port into multiple flow channels. Within each channel, oocytes are confined against a microfluidic barrier using a steady fluid flow provided by an external computer-controlled syringe pump. This allows for easy media replacement without disturbing the oocyte location. The microfluidic and optical-laser microbeam ablation capabilities of the system were validated using surf clam (Spisula solidissima) oocytes that were immobilized in order to permit ablation of the 5 µm diameter nucleolinus within the oocyte nucleolus. Oocytes were the followed and assayed for polar body ejection.


Assuntos
Técnicas Analíticas Microfluídicas/métodos , Microcirurgia/instrumentação , Microcirurgia/métodos , Oócitos/citologia , Animais , Núcleo Celular , Desenho de Equipamento , Ensaios de Triagem em Larga Escala/instrumentação , Ensaios de Triagem em Larga Escala/métodos , Terapia a Laser , Óptica e Fotônica/instrumentação , Reprodutibilidade dos Testes , Spisula/citologia
13.
J Biol Chem ; 287(9): 6702-13, 2012 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-22219192

RESUMO

The nucleolinus is a little-known cellular structure, discovered over 150 years ago (Agassiz, L. (1857) Contributions to the Natural History of the United States of America, First Monograph, Part IIL, Little, Brown and Co., Boston) and thought by some investigators in the late 19th to mid-20th century to function in the formation of the centrosomes or spindle. A role for the nucleolinus in formation of the cell division apparatus has recently been confirmed in oocytes of the surf clam, Spisula solidissima (Alliegro, M. A., Henry, J. J., and Alliegro, M. C. (2010) Proc. Natl. Acad. Sci. U.S.A. 107, 13718-13723). However, we know so little about the composition and dynamics of this compartment, it is difficult to construct mechanistic hypotheses or even to be sure that prior reports were describing analogous structures in the cells of mammals, amphibians, plants, and other organisms where it was observed. Surf clam oocytes are an attractive model to approach this problem because the nucleolinus is easily visible by light microscopy, making it accessible by laser microsurgery as well as isolation by common cell fractionation techniques. In this report, we analyze the macromolecular composition of isolated Spisula nucleolini and examine the relationship of this structure to the nucleolus and cell division apparatus. Analysis of nucleolinar RNA and protein revealed a set of molecules that overlaps with but is nevertheless distinct from the nucleolus. The proteins identified were primarily ones involved in nucleic acid metabolism and cell cycle regulation. Monoclonal antibodies generated against isolated nucleolini revealed centrosomal forerunners in the oocyte cytoplasm. Finally, induction of damage to the nucleolinus by laser microsurgery altered the trafficking of α- and γ-tubulin after fertilization. These observations strongly support a role for the nucleolinus in cell division and represent our first clues regarding mechanism.


Assuntos
Nucléolo Celular/fisiologia , Nucléolo Celular/ultraestrutura , Meiose/fisiologia , Oócitos/citologia , Animais , Divisão Celular/fisiologia , Centríolos/fisiologia , Centrossomo/fisiologia , Microscopia Confocal , Proteínas Nucleares/isolamento & purificação , Proteínas Nucleares/metabolismo , Oócitos/ultraestrutura , RNA Nuclear/genética , RNA Nuclear/isolamento & purificação , Fuso Acromático/fisiologia , Spisula , Tubulina (Proteína)/metabolismo
14.
Biomed Khim ; 56(3): 351-9, 2010.
Artigo em Russo | MEDLINE | ID: mdl-20695214

RESUMO

A possibility of adhesion inhibition of Corynebacterium diphtheriae to human buccal epithelium by glycoside hydrolases of marine hydrobiontes was investigated using alpha-galactosidase from marine bacterium Pseudoalteromonas sp. KMM 701, total enzyme preparation and beta-1,3-glucanase from marine fungi Chaetomium, total enzyme preparation and beta-1,3-glucanase from marine mollusk Littorina kurila, and total enzyme preparation from crystalline style of marine mollusk Spisula sachalinensis were used. The enzymes were added to test-tubes containing buccal epithelial cells and/or the toxigenic bacterial strain C. diphtheriae No 1129, v. gravis. All the investigated enzymes were able to abort C. diphtheriae adherence, to human buccal epithelocytes. Inhibition of adhesion was more pronounced in the case of treatment of epithelocytes with highly purified enzymes of marine hydrobiontes in comparison with total enzyme preparations. The significant inhibition of C. diphtheriae adhesion was observed when the enzymes were added to the epithelocytes with the attached microorganisms. The results obtained show that glycoside hydrolases of marine hydrobiontes degrade any carbohydrates expressed on cell surface of bacterium or human buccal epithelocytes, impair unique lectin-carbohydrate interaction and prevent the adhesion.


Assuntos
Aderência Bacteriana , Corynebacterium diphtheriae/fisiologia , Glicosídeo Hidrolases/farmacologia , Mucosa Bucal/efeitos dos fármacos , Animais , Chaetomium/enzimologia , Crustáceos/enzimologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/microbiologia , Glucana 1,3-beta-Glucosidase/isolamento & purificação , Glucana 1,3-beta-Glucosidase/farmacologia , Glicosídeo Hidrolases/isolamento & purificação , Humanos , Técnicas In Vitro , Mucosa Bucal/microbiologia , Pseudoalteromonas/enzimologia , Spisula/enzimologia , alfa-Galactosidase/isolamento & purificação , alfa-Galactosidase/farmacologia
15.
Proc Natl Acad Sci U S A ; 107(31): 13718-23, 2010 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-20643950

RESUMO

The nucleolinus is an RNA-rich compartment, closely apposed to or embedded within the nucleolus. Discovered over 150 y ago, fewer than two dozen articles have been published on the nucleolinus, probably because complex histochemical stains are required for its visualization in the great majority of cells. The nucleolinus has been reported in invertebrate oocytes, mammalian and amphibian epithelial cells, neurons, and several transformed cell lines. A prominent nucleolinus, clearly visible with transmitted light microscopes at 10x magnification, is present in each oocyte of the surf clam, Spisula solidissima. We observed a consistent relationship between the nucleolinus and the developing meiotic apparatus following Spisula oocyte activation. Through sonication and sucrose gradient fractionation of purified oocyte nuclei, we isolated nucleolini, extracted their RNA, and prepared an in situ riboprobe (NLi-1), which is associated specifically with the nucleolinus, confirming its unique composition. Other in situ observations revealed a NLi-1 and nucleolinar association with the developing spindle and centrosomes. Laser microsurgery that targeted the nucleolinus resulted in failed meiotic cell division in parthenogenetically activated oocytes and failed mitosis in fertilized oocytes. Although the nucleolinus may be a forgotten organelle, its demonstrated role in spindle formation suggests it deserves renewed attention.


Assuntos
Nucléolo Celular , Centrossomo , Fuso Acromático , Spisula/citologia , Animais , Divisão Celular , Dados de Sequência Molecular , Oócitos/citologia , RNA/isolamento & purificação
16.
Mar Environ Res ; 68(1): 1-11, 2009 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-19342090

RESUMO

The coast of The Netherlands is protected by nourishing sand. Generally, two different techniques are used, beach nourishment and shoreface nourishment. The latter technique supplies sand at a water depth of about 5-8m in the surf zone, and has been used on a regular basis since 1997 with increasing volumes since 2001. Observations on the bivalve mollusc Spisula subtruncata that was abundant before 1997 and a key food species for wintering seaduck show a decline since 2001. This coincided with a decrease in the abundance of the Common Scoter Melanitta nigra, the most numerous wintering seaduck off the Dutch coast. These observations raised concern about shoreface nourishments. This study analyses the timing and locations of shoreface nourishments in combination with S. subtruncata abundance and spatial distribution. Against the expectation, no causal relationship was found between the decline of S. subtruncata and shoreface nourishments. Other causes, such as climate change, fisheries, unsuccessful settlement or predation of spatfall are more likely behind the decline of Spisula along the Dutch coast.


Assuntos
Patos , Dióxido de Silício/efeitos adversos , Spisula , Animais , Biomassa , Demografia , Mortalidade , Países Baixos , Oceanos e Mares , Crescimento Demográfico
17.
Mol Cancer Ther ; 8(3): 592-601, 2009 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-19258425

RESUMO

HMN-176 is a potential new cancer therapeutic known to retard the proliferation of tumor cell lines. Here, we show that this compound inhibits meiotic spindle assembly in surf clam oocytes and delays satisfaction of the spindle assembly checkpoint in human somatic cells by inducing the formation of short and/or multipolar spindles. HMN-176 does not affect centrosome assembly, nuclear envelope breakdown, or other aspects of meiotic or mitotic progression, nor does it affect the kinetics of Spisula or mammalian microtubule (MT) assembly in vitro. Notably, HMN-176 inhibits the formation of centrosome-nucleated MTs (i.e., asters) in Spisula oocytes and oocyte extracts, as well as from isolated Spisula or mammalian centrosomes in vitro. Together, these results reveal that HMN-176 is a first-in-class anticentrosome drug that inhibits proliferation, at least in part, by disrupting centrosome-mediated MT assembly during mitosis.


Assuntos
Compostos de Benzilideno/farmacologia , Microtúbulos/efeitos dos fármacos , Piridinas/farmacologia , Fuso Acromático/efeitos dos fármacos , Animais , Bovinos , Centrossomo/efeitos dos fármacos , Centrossomo/metabolismo , Avaliação Pré-Clínica de Medicamentos , Células HeLa , Humanos , Microtúbulos/metabolismo , Oócitos/efeitos dos fármacos , Oócitos/metabolismo , Multimerização Proteica/efeitos dos fármacos , Fuso Acromático/metabolismo , Spisula , Células Tumorais Cultivadas
18.
Proc Natl Acad Sci U S A ; 105(19): 6993-7, 2008 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-18458332

RESUMO

The evolutionary origin of centriole/kinetosomes, centrosomes, and other microtubule organizing centers (MTOCs), whether by direct filiation or symbiogenesis, has been controversial for >50 years. Centrioles, like mitochondria and chloroplasts, duplicate independently of the nucleus and constitute a heritable system independent of chromosomal DNA. Nucleic acids endogenous to the MTOC would support evolutionary origin by symbiogenesis. To date, most reports of centrosome-associated nucleic acids have used generalized reagents such as RNases and nucleic acid dyes. Here, from a library of RNAs extracted from isolated surf clam (Spisula solidissima) centrosomes, we describe a group of centrosome-associated transcripts representing a structurally unique intron-poor collection of nuclear genes skewed toward nucleic acid metabolism. Thus, we resolve the debate over the existence of centrosome-associated RNA (cnRNA). A subset of cnRNAs contain functional domains that are highly conserved across distant taxa, such as nucleotide polymerase motifs. In situ localization of cnRNA65, a molecule with an RNA polymerase domain, showed it is present in the intact oocyte nucleus (germinal vesicle). Its expression, therefore, precedes the appearance of gamma-tubulin-containing centrosomes. At this stage, the in situ signal resembles the nucleolinus, a poorly understood organelle proposed to play a role in spindle formation. After oocyte activation and germinal vesicle breakdown, cnRNA65 persists as a cytoplasmic patch within which gamma-tubulin-stained centrosomes can be seen. These observations provoke the question of whether cnRNAs and the nucleolinus serve as cytological progenitors of the centrosome and may support a symbiogenetic model for its evolution.


Assuntos
Núcleo Celular/genética , Centrossomo/metabolismo , Íntrons/genética , Oócitos/metabolismo , RNA/metabolismo , Spisula/citologia , Spisula/genética , Animais , DNA/metabolismo , Regulação da Expressão Gênica , Genoma , Oócitos/citologia , Reação em Cadeia da Polimerase , RNA/química , Transporte de RNA
19.
Cell Motil Cytoskeleton ; 64(10): 739-55, 2007 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-17630664

RESUMO

Nonmuscle myosin II (Myo2) has been shown to associate with membranes of the trans-Golgi network and to be involved in Golgi to ER retrograde protein transport. Here, we provide evidence that Myo2 not only associates with membranes but functions to transport vesicles on actin filaments (AFs). We used extracts from unactivated clam oocytes for these studies. AFs assembled spontaneously in these extracts and myosin-dependent vesicle transport was observed upon activation. In addition, actin bundles formed and moved relative to each other at an average speed of 0.30 microm/s. Motion analysis revealed that vesicles moved on the spontaneously assembled AFs at speeds greater than 1 microm/s. The motor on these vesicles was identified as a member of the nonmuscle Myo2 family based on sequence determination by Edman chemistry. Vesicles in these extracts were purified by sucrose gradient centrifugation and movement was reconstituted in vitro using skeletal muscle actin coated coverslips. When peripheral membrane proteins of vesicles including Myo2 were removed by salt stripping or when extracts were treated with an antibody specific to clam oocyte nonmuscle Myo2, vesicle movement was inhibited. Blebbistatin, a Myo2 specific inhibitor, also blocked vesicle movement. Myo2 light chain kinase activity was found to be essential for vesicle movement and sliding of actin bundles. Together, our data provide direct evidence that nonmuscle Myo2 is involved in actin-dependent vesicle transport in clam oocytes.


Assuntos
Actinas/metabolismo , Miosina Tipo II/metabolismo , Vesículas Secretórias/metabolismo , Spisula/metabolismo , Sequência de Aminoácidos , Animais , Transporte Biológico , Citoesqueleto/metabolismo , Compostos Heterocíclicos de 4 ou mais Anéis/efeitos adversos , Dados de Sequência Molecular , Miosina Tipo II/antagonistas & inibidores , Miosina Tipo II/química , Oócitos/química , Oócitos/metabolismo , Homologia de Sequência de Aminoácidos
20.
Genesis ; 45(5): 282-91, 2007 May.
Artigo em Inglês | MEDLINE | ID: mdl-17458867

RESUMO

The cell cycle is driven by the activity of cyclin/cdk complexes. In somatic cells, cyclin E/cdk2 oscillates throughout the cell cycle and has been shown to promote S-phase entry and initiation of DNA replication. In contrast, cyclin E/cdk2 activity remains constant throughout the early embryonic development of the sea urchin and localizes to the sperm nucleus following fertilization. We now show that cyclin E localization to the sperm nucleus following fertilization is not unique to the sea urchin, but also occurs in the surf clam, and inhibition of cyclin E/cdk2 activity by roscovitine inhibits the morphological changes indicative of male pronuclear maturation in sea urchin zygotes. Finally, we show that inhibition of cyclin E/cdk2 activity does not block DNA replication in the early cleavage cycles of the sea urchin. We conclude that cyclin E/cdk2 activity is required for male pronuclear maturation, but not for initiation of DNA replication in early sea urchin development.


Assuntos
Ciclina E/metabolismo , Quinase 2 Dependente de Ciclina/metabolismo , Maturação do Esperma/fisiologia , Strongylocentrotus purpuratus/embriologia , Strongylocentrotus purpuratus/metabolismo , Animais , Ciclo Celular/efeitos dos fármacos , Quinase 2 Dependente de Ciclina/antagonistas & inibidores , Replicação do DNA , Feminino , Masculino , Inibidores de Proteínas Quinases/farmacologia , Purinas/farmacologia , Roscovitina , Especificidade da Espécie , Cabeça do Espermatozoide/metabolismo , Spisula/embriologia , Spisula/metabolismo , Strongylocentrotus purpuratus/efeitos dos fármacos , Zigoto/metabolismo
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