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1.
Rev. biol. trop ; 70(1)dic. 2022.
Artigo em Espanhol | LILACS-Express | LILACS, SaludCR | ID: biblio-1387705

RESUMO

Resumen Introducción: Durante millones de años, los organismos marinos han venido desarrollando estrategias para adaptarse a los cambios ambientales y con esto, sintetizando una gran variedad de metabolitos secundarios con actividades biológicas. Objetivo: Evaluar las actividades antimicrobiana y antioxidante e identificar los ácidos grasos del extracto metanólico de la esponja marina Tetilla rodriguesi recolectada en la bahía de Cispatá. Métodos: Los especímenes de T. rodriguesi se sometieron a extracción para obtener el extracto metanólico, del cual se obtuvieron los ácidos grasos que fueron esterificados e identificados haciendo uso de técnicas cromatográficas. Con el extracto metanólico se realizó la actividad antioxidante frente a los radicales ABTS+• y DPPH• y la actividad antimicrobiana por el método de microdilución frente a cepas de referencia de Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Enterococcus faecalis y Candida albicans; además de aislados clínicos de Candida albicans (obtenida en sangre) y Candida krusei (obtenida en catéter). Resultados: Los resultados arrojaron la identificación de 19 ácidos grasos de diferente naturaleza. En el análisis de la actividad antioxidante se pudo encontrar, que la inhibición de los radicales evaluados fue moderada (296.98 ppm para ABTS+• y 3 523.62 ppm para DPPH•). La evaluación de la actividad antimicrobiana mostró, que el extracto metanólico de T. rodriguesi logró reducir en gran medida el crecimiento de todos los microorganismos evaluados. Conclusión: A pesar de que los poríferos tienden a poseer ácidos grasos de gran tamaño, en este trabajó no se encontró la presencia de ácidos grasos de cadena carbonada mayor a 20 miembros. Los resultados de la actividad antioxidante, se asemejan en gran medida al encontrado en otras especies del mismo phylum. En esta investigación, se pudo establecer que el extracto metanólico de T. rodriguesi logró disminuir en gran medida el crecimiento de todas las cepas bacterianas y fúngicas utilizadas.


Abstract Introduction: For millions of years, organisms that inhabit the marine environment have been developing strategies to adapt to environmental changes and with this, synthesizing a great variety of secondary metabolites with biological activities. Objective: Evaluate the antimicrobial and antioxidant activities and identify the fatty acids of the methanolic extract of the marine sponge Tetilla rodriguesi collected in Cispatá bay, Colombian Caribbean. Methods: T. rodriguesi specimens were subjected to extraction to obtain the methanolic extract, of which the fatty acids were obtained, esterified and identified it using chromatographic techniques. With the methanolic extract, the antioxidant activity was carried out against the radicals ABTS+• and DPPH•, and the antimicrobial activity by the microdilution method against reference strains of Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Klebsiella pneumoniae, Enterococcus faecalis and Candida albicans; in addition to clinical isolates of Candida albicans (obtained in blood) and Candida krusei (obtained in catheter). Result: The results yielded the identification of 19 fatty acids of different nature. In the analysis of the antioxidant activity could be found that inhibition of radical evaluated was moderate (296.98 ppm for ABTS+• and 3 523.62 ppm for DPPH•). The evaluation of antimicrobial activity showed that the methanol extract of T. rodriguesi managed to greatly reduce the growth of all microorganisms tested. Conclusions: Despite the fact that porifers tend to have large fatty acids, in this study the presence of fatty acids with a carbon chain greater than 20 members was not found. The result of antioxidant activity is largely resembled that found in other species of the same phylum. In this research, it was established that the methanolic extract of T. rodriguesi managed to greatly reduce the growth of all the bacterial and fungal strains used.


Assuntos
Animais , Crambe (Esponja) , Ácidos Graxos , Ecossistema , Invertebrados , Biologia Marinha
2.
Mar Drugs ; 17(10)2019 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-31600933

RESUMO

Regulated cell death (RCD) results from the activation of one or more signal transduction modules both in physiological or pathological conditions. It is now established that RCD is involved in numerous human diseases, including cancer. As regulated cell death processes can be modulated by pharmacological tools, the research reported here aims to characterize new marine compounds acting as RCD modulators. Protein kinases (PKs) are key signaling actors in various RCDs notably through the control of either mitosis (e.g., the PKs Aurora A and B) or necroptosis (e.g., RIPK1 and RIPK3). From the primary screening of 27 various extracts of marine organisms collected in the Mediterranean Sea, an extract and subsequently a purified high molecular weight compound dubbed P3, were isolated from the marine sponge Crambe tailliezi and characterized as a selective inhibitor of PKs Aurora A and B. Furthermore, P3 was shown to induce apoptosis and to decrease proliferation and mitotic index of human osteosarcoma U-2 OS cells.


Assuntos
Produtos Biológicos/farmacologia , Crambe (Esponja)/química , Crambe (Esponja)/metabolismo , Citotoxinas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Células HEK293 , Células Hep G2 , Humanos , Células MCF-7 , Mar Mediterrâneo , Peso Molecular , Inibidores de Proteínas Quinases/farmacologia , Proteínas Quinases/metabolismo , Transdução de Sinais/efeitos dos fármacos
3.
FEMS Microbiol Lett ; 366(24)2019 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-31960895

RESUMO

Marine fungi are part of the huge and understudied biodiversity hosted in the sea. To broaden the knowledge on fungi inhabiting the Mediterranean Sea and their role in sponge holobiont, three sponges namely Aplysina cavernicola, Crambe crambe and Phorbas tenacior were collected in Villefranche sur Mer, (France) at about 25 m depth. The fungal communities associated with the sponges were isolated using different techniques to increase the numbers of fungi isolated. All fungi were identified to species level giving rise to 19, 13 and 3 species for P. tenacior, A. cavernicola and C. crambe, respectively. Of note, 35.7% and 50.0% of the species detected were either reported for the first time in the marine environment or in association with sponges. The mini-satellite analysis confirmed the uniqueness of the mycobiota of each sponge, leading to think that the sponge, with its metabolome, may shape the microbial community.


Assuntos
Crambe (Esponja)/microbiologia , Microbiota , Animais , Biodiversidade , Fungos/isolamento & purificação , Mar Mediterrâneo , Filogenia , Poríferos/microbiologia , Água do Mar/microbiologia
4.
Mar Drugs ; 16(5)2018 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-29734790

RESUMO

Isolation of marine compounds from living invertebrates represents a major challenge for sustainable and environmentally friendly exploitation of marine bio-resources. To develop innovative technology to trap invertebrate compounds in the open sea, the proof of concept of a system combining external continuous circulation of water with XAD-amberlite solid-phase extraction was validated in an aquarium. In this work, we reported the elicitation of guanidine alkaloid production of Crambe crambe in the presence of Anemonia sulcata, both collected from the Mediterranean Sea. Besides the previously reported crambescidin 359 (1), and crambescidin acid (2), three new compounds were isolated; one carboxylated analog of 1 named crambescidin 401 (3), and two analogs of crambescin B, crambescin B 281 (4) and crambescin B 253 (5). Based on these results, a technology named Somartex® for “Self Operating MARine Trapping Extractor” was patented and built to transfer the concept from closed aquarium systems to open marine ecosystems.


Assuntos
Organismos Aquáticos/química , Invertebrados/química , Alcaloides/química , Animais , Biotecnologia/métodos , Crambe (Esponja)/química , Ecossistema , Guanidina/química , Mar Mediterrâneo , Pirimidinas/química , Compostos de Espiro/química
5.
Nutrients ; 10(1)2018 Jan 02.
Artigo em Inglês | MEDLINE | ID: mdl-29301302

RESUMO

Pyrroloquinoline and guanidine-derived alkaloids present distinct groups of marine secondary metabolites with structural diversity that displayed potentialities in biological research. A considerable number of these molecular architectures had been recorded from marine sponges belonging to different marine genera, including Batzella, Crambe, Monanchora, Clathria, Ptilocaulis and New Caledonian starfishes Fromia monilis and Celerina heffernani. In this review, we aim to comprehensively cover the chemodiversity and the bioactivities landmarks centered around the chemical constituents exclusively isolated from these three marine genera including Batzella, Crambe and Monanchora over the period 1981-2017, paying a special attention to the polycyclic guanidinic compounds and their proposed biomimetic landmarks. It is concluded that these marine sponge genera represent a rich source of novel compounds with potential applications for cancer and other therapeutic areas.


Assuntos
Alcaloides/farmacologia , Antineoplásicos/farmacologia , Crambe (Esponja)/metabolismo , Guanidinas/farmacologia , Neoplasias/tratamento farmacológico , Poríferos/metabolismo , Pirróis/farmacologia , Quinolinas/farmacologia , Alcaloides/química , Alcaloides/isolamento & purificação , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Crambe (Esponja)/classificação , Guanidinas/química , Guanidinas/isolamento & purificação , Humanos , Mimetismo Molecular , Estrutura Molecular , Neoplasias/metabolismo , Neoplasias/patologia , Poríferos/classificação , Pirróis/química , Pirróis/isolamento & purificação , Quinolinas/química , Quinolinas/isolamento & purificação , Relação Estrutura-Atividade
6.
FEMS Microbiol Lett ; 364(11)2017 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-28520957

RESUMO

Marine sponges have been shown to harbor diverse microbial symbiont communities that play key roles in host functioning, yet little is known about how anthropogenic disturbances impact sponge-microbe interactions. The Mediterranean sponge Crambe crambe is known to accumulate heavy metals in polluted harbors. In this study, we investigated whether the microbiome of C. crambe differed between sponges inhabiting a polluted harbor in Blanes (Spain) and a nearby (<1 km) natural environment. Triplicate sponge and ambient seawater samples were collected from each site and the microbial composition of each sample was determined by 16S rRNA gene sequence analysis (Illumina Hi-Seq platform). No significant differences in the diversity or structure of microbial communities in C. crambe were detected between habitats, while a significant difference in community structure was observed in ambient seawater inside and outside of the polluted harbor. The microbiome of C. crambe was clearly differentiated from free-living seawater microbes and dominated by Proteobacteria, specifically a single betaproteobacterium that accounted for >86% of all sequence reads. These results indicate that sponge microbiomes exhibit greater stability and pollution tolerance than their free-living microbial counterparts, potentially mitigating the effects of pollutants on coastal marine communities.


Assuntos
Betaproteobacteria/isolamento & purificação , Crambe (Esponja)/microbiologia , Microbiota/genética , Filogenia , Animais , Técnicas de Tipagem Bacteriana , Betaproteobacteria/classificação , DNA Bacteriano/genética , Proteobactérias/classificação , Proteobactérias/isolamento & purificação , RNA Ribossômico 16S/genética , Água do Mar/microbiologia , Análise de Sequência de DNA , Espanha , Microbiologia da Água
7.
Sci Rep ; 6: 29474, 2016 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-27381941

RESUMO

Ecological interactions in the marine environment are now recognized to be partly held by chemical cues produced by marine organisms. In particular, sponges are sessile animals thought to rely on the bioactive substances they synthesize to ensure their development and defense. However, the mechanisms leading the sponges to use their specialized metabolites as chemical cues remain unknown. Here we report the constant release of bioactive polycyclic guanidinic alkaloids by the Mediterranean sponge Crambe crambe into the dissolved and the particulate phases using a targeted metabolomics study. These compounds were proven to be stored into already described specialized (spherulous) sponge cells and dispersed into the water column after release through the sponge exhaling channels (oscula), leading to a chemical shield surrounding the sponge. Low concentrations of these compounds were demonstrated to have teratogenic effects on embryos of a common sea squirt (ascidian). This mechanism of action called spherulization may therefore contribute to the ecological success of encrusting sponges that need to extend their substrate cover to expand.


Assuntos
Alcaloides/química , Crambe (Esponja)/fisiologia , Animais , Transporte Biológico , Cromatografia Líquida de Alta Pressão , Crambe (Esponja)/química , Citometria de Fluxo , Metaboloma , Microscopia Eletrônica de Varredura , Microscopia de Fluorescência , Água do Mar , Teratógenos/química , Urocordados/efeitos dos fármacos
8.
Mar Drugs ; 13(8): 4633-53, 2015 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-26225985

RESUMO

The Mediterranean marine sponge Crambe crambe is the source of two families of guanidine alkaloids known as crambescins and crambescidins. Some of the biological effects of crambescidins have been previously reported while crambescins have undergone little study. Taking this into account, we performed comparative transcriptome analysis to examine the effect of crambescin-C1 (CC1) on human tumor hepatocarcinoma cells HepG2 followed by validation experiments to confirm its predicted biological activities. We report herein that, while crambescin-A1 has a minor effect on these cells, CC1 protects them against oxidative injury by means of metallothionein induction even at low concentrations. Additionally, at high doses, CC1 arrests the HepG2 cell cycle in G0/G1 and thus inhibits tumor cell proliferation. The findings presented here provide the first detailed approach regarding the different effects of crambescins on tumor cells and provide a basis for future studies on other possible cellular mechanisms related to these bioactivities.


Assuntos
Fatores Biológicos/farmacologia , Citoproteção/efeitos dos fármacos , Metalotioneína/metabolismo , Pirimidinas/farmacologia , Alcaloides/farmacologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Crambe (Esponja)/metabolismo , Fase G1/efeitos dos fármacos , Células Hep G2 , Humanos , Fase de Repouso do Ciclo Celular/efeitos dos fármacos , Transcriptoma/efeitos dos fármacos
9.
Sci Total Environ ; 475: 71-82, 2014 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-24419288

RESUMO

Crambe crambe is a Mediterranean marine sponge known to produce original natural substances belonging to two families of guanidine alkaloids, namely crambescins and crambescidins, which exhibit cytotoxic and antiviral activities. These compounds are therefore considered as potential anticancer drugs. The present study focuses on the environmental assessment of a novel in vivo process for the production of pure crambescin and crambescidin using sponge specimens cultured in aquarium. The assessment was performed following the ISO 14040 standard and extended from the production of the different mass and energy flows to the system to the growth of the sponge in indoor aquarium and further periodic extraction and purification of the bioactive compounds. According to the results, the two stages that have a remarkable contribution to all impact categories are the purification of the bioactive molecules followed by the maintenance of the sponge culture in the aquarium. Among the involved activities, the production of the chemicals (particularly methanol) together with the electricity requirements (especially due to the aquarium lighting) are responsible for up to 90% of the impact in most of the assessed categories. However, the contributions of other stages to the environmental burdens, such as the collection of sponges, considerably depend on the assumptions made during the inventory stage. The simulation of alternative scenarios has led to propose improvement alternatives that may allow significant reductions ranging from 20% to 70%, mainly thanks to the reduction of electricity requirements as well as the partial reuse of methanol.


Assuntos
Alcaloides/análise , Produtos Biológicos/análise , Conservação dos Recursos Naturais/métodos , Crambe (Esponja)/crescimento & desenvolvimento , Pesqueiros/métodos , Guanidinas/análise , Animais , Crambe (Esponja)/química
10.
Sci Rep ; 3: 2780, 2013 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-24071658

RESUMO

Sponges harbour microbial communities that contribute to the genetic and metabolic potential of their host. Among metabolites produced by sponge-associated microbial communities, halogenated compounds are of special interest because of their biotechnological potential. In this study, we have examined the diversity of the cultivable fraction of the marine demosponge Crambe crambe microbiota. Application of complementary cultivation methods yielded 107 bacterial isolates, some of which may be sponge-specific based on their phylogenetic analysis. Among these, Psychrobacter sp. was found to contain a putative halogenase gene. In addition to the culture-dependent approach for discovering halogenase genes, a cDNA library was constructed to determine the diversity of halogenase genes expressed in situ by the C. crambe microbiota. To this end, seventeen putative tryptophan halogenase cDNA sequences were identified, most of which were only remotely related to known halogenase genes, indicating the potential for novel bioactive compounds being produced by the C. crambe microbiota.


Assuntos
Bactérias/genética , Crambe (Esponja)/microbiologia , Microbiota , Animais , Bactérias/classificação , Bactérias/enzimologia , Variação Genética , Dados de Sequência Molecular , Oxirredutases/genética , Filogenia , RNA Bacteriano , RNA Ribossômico 16S
11.
Sci Rep ; 3: 2583, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-24002533

RESUMO

Crambe crambe is a marine sponge that produces high concentrations of the pharmacologically significant pentacyclic guanidine alkaloids (PGAs), Crambescines and Crambescidines. Although bio-mimetic chemical synthesis of PGAs suggests involvement of microorganisms in their biosynthesis, there are conflicting reports on whether bacteria are associated with this sponge or not. Using 16S rRNA gene pyrosequencing we show that the associated bacterial community of C. crambe is dominated by a single bacterial species affiliated to the Betaproteobacteria. Microscopy analysis of sponge tissue sections using a specific probe and in situ hybridization confirmed its dominance in the sponge mesohyl and a single microbial morphology was observed by transmission electron microscopy. If confirmed the presence of a simple bacteria community in C. crambe makes this association a very pertinent model to study sponge-bacteria interactions and should allow further research into the possible implication of bacteria in PGA biosynthesis.


Assuntos
Betaproteobacteria/fisiologia , Crambe (Esponja)/metabolismo , Crambe (Esponja)/microbiologia , Microbiota , Compostos de Espiro/metabolismo , Animais , Betaproteobacteria/classificação , Betaproteobacteria/genética , Biodiversidade , Crambe (Esponja)/ultraestrutura , Filogenia , RNA Ribossômico 16S
12.
Int J Cancer ; 130(7): 1671-81, 2012 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-21544815

RESUMO

Cancer stem cells (CSCs) are suggested as reason for resistance of tumors toward conventional tumor therapy including pancreatic and advanced prostate cancer. New therapeutic agents are urgently needed for targeting of CSCs. Marine sponges harbor novel and undefined compounds with antineoplastic activity but their potential to eliminate CSC characteristics is not examined so far. We collected 10 marine sponges and one freshwater sponge by diving at the seaside and prepared crude methanolic extracts. The effect to established pancreatic and prostate CSC lines was evaluated by analysis of apoptosis, cell cycle, side population, colony and spheroid formation, migratory potential in vitro and tumorigenicity in vivo. While each sponge extract at a 1:10 dilution efficiently diminished viability, Crambe crambe marine sponge extract (CR) still strongly reduced viability of tumor cells at a dilution of 1:1,000 but was less toxic to normal fibroblasts and endothelial cells. CR inhibited self-renewal capacity, apoptosis resistance, and proliferation even in gemcitabine-selected pancreatic cancer cells with acquired therapy resistance and enhanced CSC characteristics. CR pretreatment of tumor cells diminished tumorigenicity of gemcitabine-resistant tumor cells in mice and totally abolished tumor take upon combination with gemcitabine. Our data suggest that CR contains substances, which render standard cancer therapy more effective by targeting of CSC characteristics. Isolation of bioactive metabolites from CR and evaluation in mice are required for development of new CSC-specific chemotherapeutic drugs from a marine sponge.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Crambe (Esponja)/química , Células-Tronco Neoplásicas/efeitos dos fármacos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias da Próstata/tratamento farmacológico , Animais , Apoptose/efeitos dos fármacos , Caspases/metabolismo , Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Transformação Celular Neoplásica/efeitos dos fármacos , Células Cultivadas , Desoxicitidina/administração & dosagem , Desoxicitidina/análogos & derivados , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Humanos , Masculino , Camundongos , Camundongos Nus , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia
13.
Aquat Toxicol ; 81(2): 137-43, 2007 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-17196674

RESUMO

Metals and polycyclic aromatic hydrocarbons (PAHs) contaminate sediments and waters of coastal areas threatening early stages of invertebrate development. Effects on these stages may largely determine the decline and even disappearance of invertebrate populations in polluted environments. Our study aimed to determine the possible influence of metals (Cu and Cd) and PAHs on larval settlement and consecutive survival of two widespread sponges of the Mediterranean: Crambe crambe and Scopalina lophyropoda. Larvae of both species were exposed to Cu and Cd for a short period during 1 week, and settlement and following (6 months) survival of juvenile were monitored. Short exposures to copper and cadmium at the concentrations used did not affect C. crambe settlement compared with SW control, and no effect on consecutive survival of juveniles was observed. In contrast, short pulses of copper and cadmium at the concentrations used enhanced Scopalina lophyropoda settlement and did not affect the consecutive survival of juveniles with respect to SW controls. Furthermore, experiments designed to assess the effects of short exposures to PAHs and the combined effect of contamination by Cu(2+) and PAHs on larval settlement, were conduced during 10 days on C. crambe larvae. Hydrocarbons, differently than copper and cadmium, inhibited the settlement of sponge larvae to a certain extent. The synergetic negative effect of copper and hydrocarbons on C. crambe settlers may cause a decline of populations in areas with both sources of contamination. The present study provides the only available data on toxicity of copper, cadmium and hydrocarbon toxicants on sponge larval settlement.


Assuntos
Cádmio/toxicidade , Cobre/toxicidade , Hidrocarbonetos Policíclicos Aromáticos/toxicidade , Poríferos/efeitos dos fármacos , Poluentes Químicos da Água/toxicidade , Animais , Crambe (Esponja)/efeitos dos fármacos , Larva/efeitos dos fármacos , Mar Mediterrâneo , Análise de Sobrevida , Fatores de Tempo
14.
Appl Microbiol Biotechnol ; 73(3): 525-32, 2006 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-16862426

RESUMO

Marine sponges are sources of high-value bioactives. Engineering aspects of in vitro culture of sponges from cuttings (explants) are poorly understood. This work develops a diffusion-controlled growth model for sponge explants. The model assumes that the explant growth is controlled by diffusive transport of at least some nutrients from the surrounding medium into the explant that generally has a poorly developed aquiferous system for internal irrigation during early stages of growth. Growth is assumed to obey Monod-type kinetics. The model is shown to satisfactorily explain the measured growth behavior of the marine sponge Crambe crambe in two different growth media. In addition, the model is generally consistent with published data for growth of explants of the sponges Disidea avara and Hemimycale columella. The model predicted that nutrient concentration profiles for nutrients, such as dissolved oxygen within the explant, are consistent with data published by independent researchers. In view of the proposed model's ability to explain available data for growth of several species of sponge explants, diffusive transport does play a controlling role in explant growth at least until a fully developed aquiferous system has become established. According to the model and experimental observations, the instantaneous growth rate depends on the size of the explant and all those factors that influence the diffusion of critical nutrients within the explant. Growth follows a hyperbolic profile that is consistent with the Monod kinetics.


Assuntos
Reatores Biológicos , Crambe (Esponja)/crescimento & desenvolvimento , Animais , Crambe (Esponja)/metabolismo , Meios de Cultura/química , Difusão , Cinética , Modelos Biológicos
15.
Biotechnol Prog ; 22(3): 781-90, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16739962

RESUMO

Marine sponges are potential sources of many unique metabolites, including cytotoxic and anticancer compounds. Natural sponge populations are insufficient or inaccessible for producing commercial quantities of metabolites of interest. It is commonly accepted that tissue (fragments, explants, and primmorphs) and in vitro cell cultivation show great potential. However, there is little knowledge of the nutritional requirements of marine sponges to carry out efficient and sustained in vitro culture and progress has been slow. In marine invertebrate fila many unsuccessful attempts have been made with in vitro cultures using typical commercial animal cell media based on sources of dissolved organic carbon (DOC) (e.g., DMEM, RPMI, M199, L-15, etc.). One of the reasons for this failure is the use of hardly identifiable growth promoters, based on terrestrial animal sera. An alternative is the use of extracts from marine animals, since they may contain nutrients necessary for growth. In this work we have cultivated in vitro explants of the encrusting marine sponge Crambe crambe. It is one of the most abundant sponges on the Mediterranean coastline and also possesses an array of potentially active metabolites (crambines and crambescidins). Initially a new approach was developed in order to show consumption of DOC by explants. Thus, different initial DOC concentrations (300, 400, 700 and 1200 mg DOC L(-1)) were assayed. Consumption was evident in all four assays and was more marked in the first 6 h. The DOC assimilation data were adjusted to an empirical model widely used for uptake kinetics of organic dissolved compounds in marine invertebrates. Second, a protocol was established to cultivate explants in vitro. Different medium formulations based on RPMI 1640 commercial medium enriched with amino acids and inorganic salts to emulate seawater salinity were assayed. The enrichment of this medium with an Octopus aqueous extract in the proportions of 10% and 20% (v/v) resulted in an evident sustained long-term growth of C. crambe explants. This growth enhancement produced high metabolic activity in the explants, as is confirmed by the high ammonium and lactate content in the medium a few days after its renewal and by the consumption of glucose. The lactate accumulation increased with the size and age of explants. Prior to these experiments, we successfully developed a robust new alternative method, based on digital image treatment, for accurate determination of the explant apparent volume as growth measure.


Assuntos
Crambe (Esponja)/efeitos dos fármacos , Crambe (Esponja)/crescimento & desenvolvimento , Meios de Cultura/farmacologia , Animais , Técnicas de Cultura de Células/métodos , Células Cultivadas , Crambe (Esponja)/metabolismo , Relação Dose-Resposta a Droga , Cinética , Mar Mediterrâneo , Fatores de Tempo
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