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1.
Biol Lett ; 18(2): 20210583, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35104429

RESUMO

Puffer and porcupine fishes (families Diodontidae and Tetraodontidae, order Tetradontiformes) are known for their extraordinary ability to triple their body size by swallowing and retaining large amounts of seawater in their accommodating stomachs. This inflation mechanism provides a defence to predation; however, it is associated with the secondary loss of the stomach's digestive function. Ingestion of alkaline seawater during inflation would make acidification inefficient (a potential driver for the loss of gastric digestion), paralleled by the loss of acid-peptic genes. We tested the hypothesis of stomach inflation as a driver for the convergent evolution of stomach loss by investigating the gastric phenotype and genotype of four distantly related stomach inflating gnathostomes: sargassum fish, swellshark, bearded goby and the pygmy leatherjacket. Strikingly, unlike in the puffer/porcupine fishes, we found no evidence for the loss of stomach function in sargassum fish, swellshark and bearded goby. Only the pygmy leatherjacket (Monochanthidae, Tetraodontiformes) lacked the gastric phenotype and genotype. In conclusion, ingestion of seawater for inflation, associated with loss of gastric acid secretion, is restricted to the Tetraodontiformes and is not a selective pressure for gastric loss in other reported gastric inflating fishes.


Assuntos
Perciformes , Estômago , Animais , Digestão , Peixes/genética , Humanos , Água do Mar
2.
Prog Lipid Res ; 86: 101157, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35104467

RESUMO

Marine ecosystems are rich in "omega-3" long-chain (C20-24) polyunsaturated fatty acids (LC-PUFA). Their production has been historically accepted to derive mostly from marine microbes. This long-standing dogma has been challenged recently by the discovery that numerous invertebrates, mostly with an aquatic life-style, have the enzyme machinery necessary for the de novo biosynthesis of polyunsaturated fatty acids (PUFA) and, from them, LC-PUFA. The key breakthrough was the detection in these animals of enzymes called "methyl-end desaturases" enabling PUFA de novo biosynthesis. Moreover, other enzymes with pivotal roles in LC-PUFA biosynthesis, including front-end desaturases and elongation of very long- chain fatty acids proteins, have been characterised in several non-vertebrate animal phyla. This review provides a comprehensive overview of the complement and functions of these gene/protein families in aquatic animals, particularly invertebrates and fish. Therefore, we expand and re-define our previous revision of the LC-PUFA biosynthetic enzymes present in chordates to animals as a whole, discussing how key genomic events have determined the diversity and distribution of desaturase and elongase genes in different taxa. We conclude that both invertebrates and fish display active, but markedly different, LC-PUFA biosynthetic gene networks that result from a complex evolutionary path combined with functional diversification and plasticity.


Assuntos
Ácidos Graxos Dessaturases , Ácidos Graxos Ômega-3 , Animais , Ecossistema , Ácidos Graxos Dessaturases/genética , Elongases de Ácidos Graxos/genética , Ácidos Graxos Insaturados , Peixes/genética
3.
Oper Dent ; 45(4): 435-441, 2020 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-32053463

RESUMO

CLINICAL RELEVANCE: The new formulations of nano-silver fluoride caused less dentin staining than the already available commercial agents 35% silver fluoride and silver diamine fluoride at 30% and 38%. SUMMARY: The objective of this study was to evaluate the dentin staining potential of nano-silver fluoride (NSF 600 and 1500 ppm) compared with the following commercially available cariostatic agents: Advantage Arrest (Elevate Oral Care, West Palm Beach, FL, USA), Riva Star (SDI, Victoria, Australia), and Cariestop (Biodinâmica, Paraná, Brazil). Seventy-five extracted human molars were sectioned at the cementoenamel junction, and the occlusal enamel was removed for exposure of coronary dentin. The samples were divided among the five agents tested (n=15). The dentin staining (ΔE/ΔL) was analyzed with a digital spectrophotometer (VITA Easyshade, VITA Zahnfabrik, Bad Säckingen, Germany) at three different time points (before application, after two weeks, and after four weeks). Photographic images were also performed. The Kruskal-Wallis and Mann-Whitney tests compared the mean ΔE and ΔL values between groups. The NSF 600 and 1500 ppm resulted in the smallest color change (ΔE=1.02 and 1.53) and dentin staining after four weeks (ΔL=-0.76 and -1.2). The new formulations differed significantly from the commercial cariostatic agents (p<0.001). NSF might be an alternative to silver diamine fluoride since it does not compromise esthetics.


Assuntos
Dentina , Fluoretos , Brasil , Humanos , Compostos de Prata , Coloração e Rotulagem
4.
J Hazard Mater ; 358: 508-511, 2018 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-29731175

RESUMO

Biological control of key processes, such as development and reproduction, is largely ascribed to a superfamily of ligand-dependent and independent transcription factors named Nuclear Receptors (NRs). Given their ability to accommodate ligands, NRs are prime targets of man-made compounds that mimic or antagonise the action of endogenous ligands. Accordingly, NRs occupy a prominent role in OECD and EPA guidelines for testing and assessment of Endocrine disrupting chemicals (EDCs). Although NR assays are already a key instrument in the OECD Conceptual Framework for Testing and Assessment of EDCs, the focus is mostly on vertebrate NRs. Here, we address the chief knowledge gaps in the field. More specifically, we (1) verify the growing availability of genomes/transcriptome projects, (2) highlight gaps in the identification and characterization of metazoan NR and in the establishment of (3) life cycle and (4) toxicity testing protocols. An overall bias towards vertebrates and selected invertebrate groups, notably Arthropoda, Annelida and Mollusca, was observed. Hence, if we aim to improve risk assessment of EDCs and emerging pollutants at an ecosystems scale, and understand their mode of action (MOA), we must establish a framework to include a broad phylogenetic sampling of Metazoans.


Assuntos
Disruptores Endócrinos/toxicidade , Poluentes Ambientais/toxicidade , Substâncias Perigosas/toxicidade , Receptores Citoplasmáticos e Nucleares , Testes de Toxicidade/métodos , Animais , Bioensaio , Bases de Dados Factuais , Ligantes , Receptores Citoplasmáticos e Nucleares/genética , Receptores Citoplasmáticos e Nucleares/metabolismo , Medição de Risco , Especificidade da Espécie
5.
J Fish Biol ; 92(4): 1051-1064, 2018 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-29465800

RESUMO

The expression pattern of genes coding for enzymes of the retinoic acid (RA) synthetic and degradation pathways was characterized in adult female zebrafish Danio rerio. Females were conditioned until maturation and post-spawn expression dynamics were determined. A striking upregulation of cyp26b1, but not cyp26a1, was observed following egg deposition, decreasing to initial levels during recovery. A similar, yet lower, fluctuation was observed for aldh1a2 and rdh10a, the enzymes participating in the two-step RA biosynthesis cascade. The present work highlights the dynamics of the adult D. rerio oogenesis and uncovers novel, yet elusive, metabolic contributors. Possible compartmentalized roles for the different gene paralogue isoforms are discussed.


Assuntos
Gônadas/metabolismo , Oogênese , Retinal Desidrogenase/metabolismo , Tretinoína/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Animais , Feminino , Retinal Desidrogenase/genética , Ácido Retinoico 4 Hidroxilase/genética , Ácido Retinoico 4 Hidroxilase/metabolismo , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/genética
6.
J Toxicol Environ Health A ; 80(9): 465-476, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28682217

RESUMO

Nuclear receptors (NR) are involved in the regulation of several metabolic processes and it is well known that these constituents may be modulated by different chemicals classes, including pharmaceuticals that may activate or antagonize NR. In mammals, some pharmaceuticals modulate the transcription of pregnane X receptor, Pxr, peroxisome proliferator activated receptor, Ppars, and aryl hydrocarbon receptor, Ahr, affecting mRNA expression of genes belonging to various regulatory pathways, including lipid metabolism and detoxification mechanisms. The aim of this study was to determine the effects of simvastatin (SIM), an anticholesterolemic drug, on selected NR and AhR mRNA transcription levels during zebrafish early development. Embryos were collected at different development stages (0, 2, 6, 14, 24, 48, and 72 hr post fertilization (hpf)) and mRNA of all target NR was detected at all time points. Embryos (1 and 24 hpf) were exposed to different concentrations of SIM (5 or 50 µg/L) in two differing assays with varying exposure times (2 or 80 hr). The transcription levels of ahr2, raraa, rarab, rarga, pparαa, pparß1, pparγ, pxr, rxraa, rxrab, rxrbb, rxrga, rxrgb, as well as levels of cholesterol (Chol) were measured after exposure. SIM exerted no marked effect on Chol levels, and depending upon exposure duration mRNA levels of NR and AhR either increased or decreased. After 2 hr SIM treatment in 24 hpf embryos, transcription of ppars, pxr, and ahr was up-regulated, while after 80 hr mRNA levels of pxr and ahr were decreased with no marked changes in ppars. Data demonstrate that SIM produced alterations in gene expression of NR which are involved in varying physiological functions and that may disturb regulation of different physiological processes which might impair fish survival and ecosystems regeneration.


Assuntos
Embrião não Mamífero/efeitos dos fármacos , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Receptores Citoplasmáticos e Nucleares/efeitos dos fármacos , Sinvastatina/farmacologia , Peixe-Zebra/genética , Animais
7.
Sci Rep ; 6: 33954, 2016 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-27703170

RESUMO

Carbonic anhydrase plays a key role in CO2 transport, acid-base and ion regulation and metabolic processes in vertebrates. While several carbonic anhydrase isoforms have been identified in numerous vertebrate species, basal lineages such as the cyclostomes have remained largely unexamined. Here we investigate the repertoire of cytoplasmic carbonic anhydrases in the sea lamprey (Petromyzon marinus), that has a complex life history marked by a dramatic metamorphosis from a benthic filter-feeding ammocoete larvae into a parasitic juvenile which migrates from freshwater to seawater. We have identified a novel carbonic anhydrase gene (ca19) beyond the single carbonic anhydrase gene (ca18) that was known previously. Phylogenetic analysis and synteny studies suggest that both carbonic anhydrase genes form one or two independent gene lineages and are most likely duplicates retained uniquely in cyclostomes. Quantitative PCR of ca19 and ca18 and protein expression in gill across metamorphosis show that the ca19 levels are highest in ammocoetes and decrease during metamorphosis while ca18 shows the opposite pattern with the highest levels in post-metamorphic juveniles. We propose that a unique molecular switch occurs during lamprey metamorphosis resulting in distinct gill carbonic anhydrases reflecting the contrasting life modes and habitats of these life-history stages.


Assuntos
Anidrases Carbônicas , Citoplasma , Proteínas de Peixes , Regulação Enzimológica da Expressão Gênica/fisiologia , Petromyzon , Animais , Dióxido de Carbono/metabolismo , Anidrases Carbônicas/biossíntese , Anidrases Carbônicas/genética , Citoplasma/enzimologia , Citoplasma/genética , Proteínas de Peixes/biossíntese , Proteínas de Peixes/genética , Brânquias/enzimologia , Petromyzon/genética , Petromyzon/metabolismo
8.
Gen Comp Endocrinol ; 225: 142-148, 2016 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-26597622

RESUMO

Germ cell commitment and meiosis initiation are among the multitude of physiological roles of retinoic acid (RA) in vertebrates. Acting via receptor-mediated transcription, RA induces the expression of meiotic factors, triggering meiosis. Contrasting with vertebrates, invertebrate RA metabolism is scarcely understood. Still, some physiological processes appear to be conserved. Here we set to evaluate the role of retinoids in the gonad maturation process of the marine gastropod Patella vulgata. We found that retinoid concentration in gonadal tissue, namely RA, varies between breeding and resting specimens, with maxima attained in the latter. Additionally, we isolated and quantified the expression of both the retinoic acid receptor (RAR) and the retinoid X receptor (RXR) in gonads. In view of the stability of retinoid receptor expression, we suggest that the balance of RA levels operates through the enzymatic control of synthetic and catabolic processes. Overall, the reported data are supportive for a developmental role of RA during gonadal maturation in P. vulgata, which should be addressed in other protostome lineages.


Assuntos
Gastrópodes/metabolismo , Gônadas/metabolismo , Receptores do Ácido Retinoico/metabolismo , Receptores X de Retinoides/metabolismo , Retinoides/metabolismo , Animais , Proteínas de Transporte , Expressão Gênica , Reprodução/fisiologia , Tretinoína/metabolismo
9.
Aquat Toxicol ; 164: 155-62, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25974001

RESUMO

Fibrates and other lipid regulator drugs are widespread in the aquatic environment including estuaries and coastal zones, but little is known on their chronic effects on non-target organisms as marine fish. In the present study, turbot juveniles were exposed to the PPARα model agonist WY-14,643 for 21 days by repeated injections at the concentrations of 5mg/kg (lo-WY) and 50mg/kg (hi-WY), and samples taken after 7 and 21 days. Enzyme activity and mRNA expression of palmitoyl-CoA oxidase and catalase in the liver were analyzed as first response, which validated the experiment by demonstrating interactions with the peroxisomal fatty acid oxidation and oxidative stress pathways in the hi-WY treatment. In order to get mechanistic insights, alterations of plasma lipids (free cholesterol, FC; HDL associated cholesterol, C-HDL; triglycerides, TG; non-esterified fatty acids, NEFA) and hepatic mRNA expression of 17 genes involved in fatty acid and lipid metabolism were studied. The exposure to hi-WY reduced the quantity of plasma FC, C-HDL, and NEFA. Microsomal triglyceride transfer protein and apolipoprotein E mRNA expression were higher in hi-WY, and indicated an increased formation of VLDL particles and energy mobilization from liver. It is speculated that energy depletion by PPARα agonists may contribute to a higher susceptibility to environmental stressors.


Assuntos
Proteínas de Peixes/genética , Linguados/fisiologia , Regulação da Expressão Gênica/efeitos dos fármacos , Metabolismo dos Lipídeos , Pirimidinas/toxicidade , Animais , Linguados/genética , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/genética , Lipídeos/sangue , Fígado/efeitos dos fármacos , Fígado/enzimologia , Oxirredução , PPAR alfa/agonistas , PPAR alfa/genética , Poluentes Químicos da Água/toxicidade
10.
Mar Genomics ; 10: 17-25, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23517768

RESUMO

Thorough evaluation of normalization approaches is a fundamental aspect in real-time quantitative RT-PCR experiments to avoid artificial introduced intergroup variations. In our study, we tested three normalization strategies in an experimental data set derived from a toxicological exposure of Scophthalmus maximus to the peroxisome proliferator-activated receptor alpha (PPARα) agonist WY-14643. Juvenile turbots were exposed by repeated injections to 5 mg or 50 mg WY-14643/kg, and liver samples were taken at day 1, 7 and 21. Specifically, the mRNA expression of peroxiredoxin 5 (prdx5) was normalized to the cDNA content, to the mRNA expression of single reference genes (b-actin, b-act; elongation factor 1 α, ef1a; glyceraldehyde-3-phosphate dehydrogenase, gapdh; ribosomal protein L8, rpl8; tata-box binding protein, tbp; tubulin beta 2C chain, tubb2c; ubiquitin-conjugating enzyme E2L 3, ub2l3) or to a combination of multiple reference genes using geNorm, BestKeeper or NormFinder algorithms. Four single reference genes (ef1a, rpl8, tubb2c, tbp) did not show any significant differences between the treatment groups over time, while significant intergroup variations were observed for cDNA content, gapdh, b-act and ub2l3. The normalization of prdx5 to the valid (not altered) single reference genes led to significant up-regulated (prdx5/rpl8), not-regulated (prdx5/ef1a; prdx5/tbp) or down-regulated (prdx5/tubb2c) mRNA expression pattern. The multiple reference gene approaches resulted in different rankings and combinations of the most stable expressed reference genes (geNorm: ef1a>rpl8>b-act; BestKeeper: ub2l3>gapdh>ef1a; NormFinder: b-act>ef1a). However, the normalization with the three multiple reference gene procedures demonstrated consistent expression pattern with a significant up-regulation of prdx5 in response to the higher concentration after 21 days. Concluding, even if not yet established as "gold" standard for expression profiling in environmental toxicology or physiology using freshwater or marine fish models, the multiple reference gene approach is recommended, since it eliminates any biased results, which represented the major flaw of single reference genes.


Assuntos
Linguados/genética , Regulação da Expressão Gênica/fisiologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Animais , DNA Complementar/genética , DNA Complementar/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo
11.
Comp Biochem Physiol B Biochem Mol Biol ; 164(4): 247-58, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23380517

RESUMO

Dietary long chain polyunsaturated fatty acids (FA) have been recognized of crucial importance in early development of vertebrates, contributing to the impressive morphological and physiological changes both as building blocks and to energy production. The importance of lipids along development depends on ontogenetic, phylogenetic and environmental parameters. The expression patterns of FA metabolism genes have not been characterized in developing fish embryos nor compared to lipid class profiles. Full lipid metabolism only occurred after hatching, as revealed by alterations in lipid profiles and FA gene expression. Nonetheless, transcriptional changes of some FA genes were already present in embryos at notochord formation. Many genes displayed an expression profile opposed to the decrease of lipids along the development, while others responded solely to starvation. Transcription of most genes involved in FA metabolism had a strong correlation to PPARs' mRNA levels (α1, α2, ß, γ). The comparison of mRNA expression of the genes with the lipid profiles produced new insights into the FA metabolism and regulation during the development of turbot larvae, providing the basis for future studies including comparative approaches with other vertebrate species.


Assuntos
Linguados/metabolismo , Metabolismo dos Lipídeos/genética , Lipídeos/química , Óvulo/metabolismo , Receptores Ativados por Proliferador de Peroxissomo/metabolismo , Animais , Ácidos Graxos/genética , Ácidos Graxos/metabolismo , Linguados/genética , Larva/genética , Larva/metabolismo , Lipídeos/genética , Transcriptoma
12.
Fish Physiol Biochem ; 39(4): 1043-55, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23266856

RESUMO

Peroxisome proliferator-activated receptors (PPARs) are involved in the regulation of lipid and carbohydrate metabolism and can be activated either by natural ligands as fatty acids or by synthetic ligands including several environmental chemicals. In this study, two PPARα isoforms (α1 and α2) were analyzed in turbot (Scophthalmus maximus) for a different tissue distribution. PPARα1 was ubiquitously expressed, while the PPARα2 was predominantly expressed in the heart. Following this result, turbot juveniles were exposed by injection to a synthetic selective PPARα agonist, WY-14643, for 14 days. Suppression subtractive hybridization (SSH) was performed with pools of heart samples of control and exposed fish to get insights into PPARα-regulated genes in the heart of juvenile turbot. Four genes were positively identified in the forward-subtracted and 12 genes in the reverse-subtracted cDNA SSH library, corresponding to the down-regulated and up-regulated genes in response to the WY-14643 treatment, respectively. The confirmation of these results in individual samples of juvenile turbot exposed to WY-14643 revealed a statistically significant mRNA induction of two cardiac muscle proteins (myosin light chain 2 and tropomyosin 4), which were shown to be involved in heart contraction and heartbeat regulation in other teleost species. Herewith, we showed for the first time that PPARα2 is predominantly expressed in the heart and that a PPARα agonist can induce the mRNA expression of cardiac muscle proteins in teleosts.


Assuntos
Miosinas Cardíacas/metabolismo , Linguados/metabolismo , Miocárdio/metabolismo , Cadeias Leves de Miosina/metabolismo , PPAR alfa/genética , Pirimidinas , Tropomiosina/metabolismo , Animais , Linguados/genética , Regulação da Expressão Gênica/efeitos dos fármacos , PPAR alfa/agonistas , PPAR alfa/biossíntese , Isoformas de Proteínas/metabolismo , Pirimidinas/farmacologia , Reação em Cadeia da Polimerase em Tempo Real , Transcrição Gênica
13.
Lett Appl Microbiol ; 50(3): 246-51, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20025649

RESUMO

AIMS: To investigate the effect of lactic acid (LA), copper (II), and monolaurin as natural antimicrobials against Cronobacter in infant formula. METHODS AND RESULTS: The effect of LA (0.1, 0.2 and 0.3% v/v), copper (II) (10, 50 and 100 microg ml(-1)) and monolaurin (1000, 2000, and 3000 microg ml(-1)) suspended into tween-80 or dissolved in ethanol against Cronobacter in infant formula was investigated. Reconstituted infant formula and powdered infant formula were inoculated with five strains of Cronobacter spp. at the levels of c. 1 x 10(6) CFU ml(-1) and 1 x 10(3) CFU g(-1), respectively. LA at 0.2% v/v had a bacteriostatic effect on Cronobacter growth, whereas 0.3% v/v LA resulted in c. 3 log(10) reduction. Copper (II) at the levels of 50 microg ml(-1) and 100 microg ml(-1) elicited c. 1 and 2 log(10) reductions, respectively. The combination of 0.2% LA and 50 microg ml(-1) copper (II) resulted in a complete elimination of the organism. Monolaurin exhibited a slight inhibitory activity against Cronobacter (c. 1.5 log(10) difference) compared to the control when ethanol was used to deliver monolaurin. CONCLUSIONS: A complete elimination of Cronobacter was obtained when a combination of sublethal concentrations of LA (0.2%) and copper (II) (50 microg ml(-1)) was used. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of the synergistic interactive combination of LA and copper (II) could be beneficial to control Cronobacter in the infant formula industry.


Assuntos
Antibacterianos/farmacologia , Sulfato de Cobre/farmacologia , Cronobacter sakazakii/crescimento & desenvolvimento , Microbiologia de Alimentos , Fórmulas Infantis , Ácido Láctico/farmacologia , Lauratos/farmacologia , Monoglicerídeos/farmacologia , Contagem de Colônia Microbiana , Cronobacter sakazakii/efeitos dos fármacos , Enterobacteriaceae/efeitos dos fármacos , Enterobacteriaceae/crescimento & desenvolvimento , Humanos , Lactente , Testes de Sensibilidade Microbiana
14.
Evol Dev ; 5(5): 459-65, 2003.
Artigo em Inglês | MEDLINE | ID: mdl-12950625

RESUMO

Homeobox genes encode DNA-binding proteins, many of which are implicated in the control of embryonic development. Evolutionarily, most homeobox genes fall into two related clades: the ANTP and the PRD classes. Some genes in ANTP class, notably Hox, ParaHox, and NK genes, have an intriguing arrangement into physical clusters. To investigate the evolutionary history of these gene clusters, we examined homeobox gene chromosomal locations in the cephalochordate amphioxus, Branchiostoma floridae. We deduce that 22 amphioxus ANTP class homeobox genes localize in just three chromosomes. One contains the Hox cluster plus AmphiEn, AmphiMnx, and AmphiDll. The ParaHox cluster resides in another chromosome, whereas a third chromosome contains the NK type homeobox genes, including AmphiMsx and AmphiTlx. By comparative analysis we infer that clustering of ANTP class homeobox genes evolved just once, during a series of extensive cis-duplication events of genes early in animal evolution. A trans-duplication event occurred later to yield the Hox and ParaHox gene clusters on different chromosomes. The results obtained have implications for understanding the origin of homeobox gene clustering, the diversification of the ANTP class of homeobox genes, and the evolution of animal genomes.


Assuntos
Cordados não Vertebrados/genética , Mapeamento Cromossômico , Evolução Molecular , Genes Homeobox/genética , Modelos Genéticos , Animais , Sondas de DNA , Hibridização in Situ Fluorescente
15.
J Mol Recognit ; 15(4): 209-20, 2002.
Artigo em Inglês | MEDLINE | ID: mdl-12382239

RESUMO

A continuum solvent model based on the generalized Born (GB) or finite-difference Poisson-Boltzmann (FDPB) approaches has been employed to compare the binding of 4'-6-diamidine-2-phenyl indole (DAPI) to the minor groove of various DNA sequences. Qualitative agreement between the results of GB and FDPB approaches as well as between calculated and experimentally observed trends regarding the sequence specificity of DAPI binding to B-DNA was obtained. Calculated binding energies were decomposed into various contributions to solvation and DNA-ligand interaction. DNA conformational adaptation was found to make a favorable contribution to the calculated total interaction energy but did not change the DAPI binding affinity ranking of different DNA sequences. The calculations indicate that closed complex formation is mainly driven by nonpolar contributions and was found to be disfavored electrostatically due to a desolvation penalty that outbalances the attractive Coulomb interaction. The calculated penalty was larger for DAPI binding to GC-rich sequences compared with AT-rich target sequences and generally larger for the FDPB vs the GB continuum model. A radial interaction profile for DAPI at different distances from the DNA minor groove revealed an electrostatic energy minimum a few Angstroms farther away from the closed binding geometry. The calculated electrostatic interaction up to this distance is attractive and it may stabilize a nonspecific binding arrangement.


Assuntos
DNA/metabolismo , Indóis/metabolismo , Substâncias Intercalantes/metabolismo , Nucleotídeos de Adenina/química , Sequência de Bases , Sítios de Ligação , DNA/análise , DNA/química , Conformação de Ácido Nucleico , Solventes
16.
J Med Chem ; 39(4): 929-36, 1996 Feb 16.
Artigo em Inglês | MEDLINE | ID: mdl-8632416

RESUMO

For further studies on the structural and conformational requirements of positions 2,3, and 7 in the bradykinin sequence, we replaced the proline residues by the more hydrophobic and conformationally restricted N-methyl-L- and D-phenylalanine (NMF). The biological activities of the new analogs were evaluated on rat uterus, guinea pig ileum, and guinea pig lung strip. Receptor binding of the analogs was studied in membranes from rat uterus and guinea pig ileum. Influence of bradykinin analogs on the release of cytokines from mouse spleen cell cultures was also measured. Bradykinin analogs were synthesized by the solid phase method, using Boc strategy on PAM or Merrifield resins. The best results in the formation of the N-methylamide bond were obtained with the coupling reagent PyBrop. In position 7 the substitution of D-Phe by D-NMF, retaining the configuration of the amino acid, converts bradykinin antagonists into agonists. The bradykinin analogs with D-NMF at position 7 gave the highest known tissue selectivity for rat uterus among agonists. [L-NMF(2)]bradykinin has moderate agonist activity on rat uterus but antagonist activity on guinea pig lung strip. It represents a new antagonist for B(2) receptors without any replacement at position 7. The same analog completely inhibits bradykinin-evoked cytokine expression by mononuclear cells.


Assuntos
Bradicinina/análogos & derivados , Bradicinina/síntese química , Fenilalanina/análogos & derivados , Prolina , Sequência de Aminoácidos , Animais , Bradicinina/agonistas , Bradicinina/antagonistas & inibidores , Membrana Celular/metabolismo , Feminino , Cobaias , Íleo/efeitos dos fármacos , Íleo/fisiologia , Técnicas In Vitro , Indicadores e Reagentes , Masculino , Dados de Sequência Molecular , Músculo Liso/efeitos dos fármacos , Músculo Liso/fisiologia , Conformação Proteica , Ratos , Receptores da Bradicinina/efeitos dos fármacos , Receptores da Bradicinina/metabolismo , Estereoisomerismo , Relação Estrutura-Atividade , Útero/efeitos dos fármacos , Útero/fisiologia
17.
Int J Pept Protein Res ; 46(2): 155-65, 1995 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-8567170

RESUMO

Continuing our studies of the bioactive conformation of bradykinin (BK) antagonists, we synthesized a first series of analogues with side-chain cyclization in the N-terminal sequence. Through this conformational constraint it should be possible to gain insight into their three-dimensional structure. The cycles were proposed on the basis of existing ideas and hypotheses about the receptor bound conformation of BK and its antagonists. The reported peptides contain D-Phe at position 7 or D-Tic-Oic (D-Tetrahydroisoquinoline-3 -carboxyl-octahydroindole-2-carboxylic acid) at positions 7 and 8, respectively, and a disulfide or lactam bridge between positions 0 and 6. Syntheses, including cyclization reactions, were carried out on PAM resin. The biological activity of the lead compound [DPhe7]-BK, the linear precursors and the cyclic peptides, as estimated on isolated rat uterus, guinea pig ileum and lung strips, are in the same range. The conformational properties of the new cyclic analogues were studied through energy minimization on a model compound. The results of the calculations support the existence of low-energy structures containing a beta-turn. Therefore, such a turn in the N-terminal segment of the molecule can be proposed as an important structural feature of the bioactive conformation of BK antagonists.


Assuntos
Bradicinina/antagonistas & inibidores , Peptídeos Cíclicos/síntese química , Precursores de Proteínas/síntese química , Útero/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Bradicinina/análogos & derivados , Cromatografia Líquida de Alta Pressão , Cromatografia em Camada Fina , Simulação por Computador , Dissulfetos/química , Dissulfetos/metabolismo , Eletroforese em Gel de Poliacrilamida , Feminino , Lactamas/química , Lactamas/metabolismo , Dados de Sequência Molecular , Peptídeos Cíclicos/química , Peptídeos Cíclicos/farmacologia , Conformação Proteica , Precursores de Proteínas/química , Precursores de Proteínas/farmacologia , Ratos , Ratos Wistar , Espectrometria de Massas de Bombardeamento Rápido de Átomos , Relação Estrutura-Atividade , Contração Uterina/efeitos dos fármacos
18.
Artigo em Inglês | MEDLINE | ID: mdl-9346870

RESUMO

With the aim of synthesizing cyclic antagonists of the nonapeptide hormone bradykinin with minimal side chain modification, we performed backbone to backbone and backbone to side chain cyclization. To probe and compare different strategies for this new kind of cyclization, the branched peptide bonds were formed by both reductive alkylation on the solid phase and by using preformed building units. Lactam bridges between the modified amide groups were formed by the use of the phenylalanine derivatives N(CH2COOH)Phe and N(CH2CH2NH2)Phe. The best results in the formation of the N-alkylamide bond were obtained with the coupling reagent PyBrop. The coupling rate was monitored by estimation of the N-terminal Fmoc-group. The cyclization was performed on the solid support. Unexpected difficulties resulted from the instability of the N-alkylamide bond under strong acidic conditions, as used for deprotection and for removal from the resin. We synthesized peptides with backbone to backbone cyclization between positions 2 and 5, as well as backbone to side chain cyclizations between positions 0 and 5, and between 2 and 6. The relatively high biological activities of some of the cyclic analogues support the supposed receptor-bound conformation of bradykinin antagonists with a beta-turn in the N-terminal sequence.


Assuntos
Bradicinina/antagonistas & inibidores , Antagonistas de Hormônios/química , Antagonistas de Hormônios/síntese química , Animais , Desenho de Fármacos , Humanos , Peptídeos/síntese química , Peptídeos/química , Relação Estrutura-Atividade
19.
Rev. bras. anestesiol ; 36(4): 285-8, jul.-ago. 1986. tab
Artigo em Português | LILACS | ID: lil-38159

RESUMO

Foi realizado um estudo prospectivo com bloqueio do plexo braquial pela interescalênica em 60 crianças, entre 2 e 15 anos, submetidas a procedimentos cirúrgicos ou manipulaçöes do membro superior, em regime de urgência e/ou ambulatorial. Como guia para cálculo do volume da soluçäo anestésica (lidocaína), foi utilizada a tabela proposta por Winnie em 1970, sendo usada uma concentraçäo entre 0,5 e 2%. Os resultados obtidos demonstram facilidade na execuçäo da técnica quando associada a uma leve sedaçäo e/ou medicaçäo pré-anestésica. Foi obtido bloqueio completo em 70% dos casos, havendo falhas parciais em 30% deles. A maior incidência de falha ocorreu na regiäo do nervo ulnar. Em 20% dos casos, foi feita complementaçäo com bloqueio isolado de nervo ou sedaçäo, e em 10% o local da intervençäo näo se estendia pela área näo bloqueada. Näo foi registrado nenhuma complicaçäo durante o estudo


Assuntos
Pré-Escolar , Criança , Adolescente , Humanos , Masculino , Feminino , Procedimentos Cirúrgicos Ambulatórios , Plexo Braquial , Emergências , Lidocaína , Bloqueio Nervoso
20.
Arq Neuropsiquiatr ; 33(2): 151-5, 1975 Jun.
Artigo em Português | MEDLINE | ID: mdl-1080045

RESUMO

A case of hereditary multiple exostoses successfully operated is reported. The patient, a 15 year-old white brazilian boy, was admitted with tetraplegia and Babinski's sign. Early diagnosis followed by prompt surgery may prevent permanent spinal cord damage.


Assuntos
Exostose Múltipla Hereditária/complicações , Compressão da Medula Espinal/etiologia , Adolescente , Vértebras Cervicais/cirurgia , Exostose Múltipla Hereditária/genética , Exostose Múltipla Hereditária/cirurgia , Humanos , Laminectomia , Masculino , Compressão da Medula Espinal/cirurgia
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