Umbelliferone induces changes in the structure and pharmacological activities of Bn IV, a phospholipase A(2) isoform isolated from Bothrops neuwiedi

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Effect of umbelliferone (7-hydroxycoumarin, 7-HOC) on the enzymatic, edematogenic and necrotic activities of secretory phospholipase A2 (sPLA2) isolated from Crotalus durissus collilineatus venom

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Renal and cardiovascular effects of Bothrops marajoensis venom and phospholipase A(2)

Bothrops marajoensis is found in the savannah of Marajo Island in the State of Par S and regions of Amapa State, Brazil. The aim of the work was to study the renal and cardiovascular effects of the B. marajoensis venom and phospholipase A(2) (PLA(2)). The venom was fractionated by Protein Pack 5PW. N-terminal amino acid sequencing of sPLA(2) showed amino acid identity with other lysine K49sPLA(2)s of snake venom. B. marajoensis venom (30 mu g/mL) decreased the perfusion pressure, renal vascular resistance, urinary flow, glomerular filtration rate and sodium tubular transport. PLA(2) did not change the renal parameters. The perfusion pressure of the mesenteric bed did not change after infusion of venom. In isolated heart, the venom decreased the force of contraction and increased PP but did not change coronary flow. In the arterial pressure, the venom and PLA(2) decreased mean arterial pressure and cardiac frequency. The presence of atrial flutter and late hyperpolarisation reversed, indicating QRS complex arrhythmia and dysfunction in atrial conduction. In conclusion, B. marajoensis venom and PLA(2) induce hypotension and bradycardia while simultaneously blocking electrical conduction in the heart. Moreover, the decrease in glomerular filtration rate, urinary flow and electrolyte transport demonstrates physiological changes to the renal system. (C) 2009 Elsevier Ltd. All rights reserved.

Harpalycin 2 inhibits the enzymatic and platelet aggregation activities of PrTX-III, a D49 phospholipase A(2) from Bothrops pirajai venom

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Crystal structure of Bn IV in complex with myristic acid: A Lys49 myotoxic phospholipase A(2) from Bothrops neuwiedi venom

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Purification, characterization, crystallization and theoretical molecular modeling of gyroxin fraction from Crotalus durissus terrificus venom (Laurenti, 1768)

The venom of Crotalus durissus terrificus snakes presents various substances, including a serine protease with thrombin-like activity, called gyroxin, that clots plasmatic fibrinogen and promote the fibrin formation. The aim of this study was to purify and structurally characterize the gyroxin enzyme from Crotalus durissus terrificus venom. For isolation and purification, the following methods were employed: gel filtration on Sephadex G75 column and affinity chromatography on benzamidine Sepharose 6B; 12% SDS-PAGE under reducing conditions; N-terminal sequence analysis; cDNA cloning and expression through RT-PCR and crystallization tests. Theoretical molecular modeling was performed using bioinformatics tools based on comparative analysis of other serine proteases deposited in the NCBI (National Center for Biotechnology Information) database. Protein N-terminal sequencing produced a single chain with a molecular mass of similar to 30 kDa while its full-length cDNA had 714 bp which encoded a mature protein containing 238 amino acids. Crystals were obtained from the solutions 2 and 5 of the Crystal Screen Kit (R), two and one respectively, that reveal the protein constitution of the sample. For multiple sequence alignments of gyroxin-like B2.1 with six other serine proteases obtained from snake venoms (SVSPs), the preservation of cysteine residues and their main structural elements (alpha-helices, beta-barrel and loops) was indicated. The localization of the catalytic triad in His57, Asp102 and Ser198 as well as S1 and S2 specific activity sites in Thr193 and Gli215 amino acids was pointed. The area of recognition and cleavage of fibrinogen in SVSPs for modeling gyroxin B2.1 sequence was located at Arg60, Arg72, Gln75, Arg81, Arg82, Lis85, Glu86 and Lis87 residues. Theoretical modeling of gyroxin fraction generated a classical structure consisting of two alpha-helices, two beta-barrel structures, five disulfide bridges and loops in positions 37, 60, 70, 99, 148, 174 and 218. These results provided information about the functional structure of gyroxin allowing its application in the design of new drugs.

Occurrence of Cryptosporidium (Apicomplexa, Cryptosporidiidae) in Crotalus durissus terrificus (Serpentes, Viperidae) in Brazil

The objective of the present study was to investigate the prevalence of Cryptosporidium (Apicomplexa, Cryptosporidiidae) in the snake Crotalus durissus terrificus (Serpentes, Viperidae). Fifty animals were evaluated for the presence of oocysts of Cryptosporidium sp. at the time of arrival and 30 and 60 days later. Intestinal washings with saline solution (1% body weight), fecal samples, and organ scrapings were collected during the study. Oocysts were concentrated by an ether-phosphate-buffered saline sedimentation technique and then separated by a density gradient centrifugation technique. Smears were made with the sediment and submitted to modified acid-fast and auramine-rhodamine staining. Cryptosporidium-positive smears were used as controls for the experimental findings. The overall prevalence of Cryptosporidium sp. oocysts was 14%. Among the positive snakes, oocysts were detected only in the intestinal washing in two specimens, only in the feces in four specimens, and in both materials at least once in one specimen. The positive snakes were predominantly from Santa Maria da Serra city State of São Paulo (57.1%). We also observed that all of the examinations that presented positive results were obtained at least 27 days after the capture of the animals.

The occurrence of Crepidobothrium sp. (Cestoda, Proteocephalidae) in Bothrops moojeni (Hoge) (Serpentes, Viperidae)

The occurrence of Crepidobothrium sp.(Cestoda, Proteocephalidae) in the intestine of Bothrops moojeni (Hoge,1965)(Serpentes, Viperidae) is reported. The host snake was rescued from the fauna in Porto Primavera dam, Mato Grosso do Sul State, Brazil. The snake died in captivity on July 13,1999. At necropsy, 28 tapeworms were found in the snake intestine. The analysis of specimens morphology allowed the conclusion that they belong to the Crepidobothrium (Monticelli, 1900) genus. It was not possible to determine the Crepidobothrium species due to the lack of the gravid proglottids. This is the first report of B. moojeni as a host of cestodes.

Antibacterial and antiparasitic effects of Bothrops marajoensis venom and its fractions: Phospholipase A(2) and L-amino acid oxidase

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Cutaneous Basophilia in the Resistance of Goats to Amblyomma cajennense Nymphs after Repeated Infestations

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Guinea Pigs Develop Cutaneous Basophilia after Repeated Infestations by Nymphs of the Tick Amblyomma triste

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

The adrenergic regulation of the cardiovascular system in the South American rattlesnake, Crotalus durissus

The present study investigates adrenergic regulation of the systemic and pulmonary circulations of the anaesthetised South American rattlesnake, Crotalus durissus. Haemodynamic measurements were made following bolus injections of adrenaline and adrenergic antagonists administered through a systemic arterial catheter. Adrenaline caused a marked systemic vasoconstriction that was abolished by phentolamine, indicating this response was mediated through alpha-adrenergic receptors. Injection of phentolamine gave rise to a pronounced vasodilatation (systemic conductance (G(sys)) more than doubled), while injection of propranolol caused a systemic vasoconstriction, pointing to a potent alpha-adrenergic, and a weaker beta-adrenergic tone in the systemic vasculature of Crotalus. Overall, the pulmonary vasculature was far less responsive to adrenergic stimulation than the systemic circulation. Adrenaline caused a small but non-significant pulmonary vasodilatation and there was tendency of reducing this dilatation after either phentolamine or propranolol. Injection of phentolamine increased pulmonary conductance (G(pul)), while injection of propranolol produced a small pulmonary constriction, indicating that alpha-adrenergic and beta-adrenergic receptors contribute to a basal regulation of the pulmonary vasculature. Our results suggest adrenergic regulation of the systemic vasculature, rather than the pultrionary, may be an important factor in the development of intracardiac shunts. (c) 2007 Elsevier B.V. All rights reserved.

Comportamento de ovinos em capim-tanzânia sob lotação rotativa com quatro níveis de suplementação concentrada

Avaliou-se o comportamento de ovinos em pastagem de capim-tanzânia irrigado sob suplementação (0,0; 0,6; 1,2 e 1,8% PV) com concentrado. O delineamento utilizado foi o inteiramente casualizado com seis repetições (ovinos). A porcentagem do tempo total sob o sombrite, pastejando, ruminando, em outras atividades, em ócio, o número de ingestões de suplemento/sal, de defecações, de micções e de ingestão de água e a taxa de bocado foram estimados dividindo-se o dia em oito períodos de 3 horas (das 5 às 8 h; 8 às 11 h; 11 às 14 h; 14 às 17 h; 17 às 20 h; 20 às 23 h; 23 às 2 h; 2 às 5 h). O tempo de pastejo foi maior entre os animais que não receberam suplementação e decresceu progressivamente, voltando a elevar somente com 1,8% de suplementação. O tempo de ruminação foi maior nos animais com suplementação de 0,6% PV, principalmente das 14 às 17 h e das 17 às 20 h, enquanto o tempo de ócio foi maior no nível de 1,2% PV. A ingestão de água elevou-se com o aumento da suplementação até 1,2% PV e concentrou-se entre 11 e 14 h. O número de micções e de defecações foi maior com suplementação no nível de 1,8% PV. A elevação progressiva na taxa de bocado até o nível de suplementação de 1,2% PV no período das 11 às 20 h com posterior redução sugere efeito aditivo do suplemento sobre o pasto até esse nível, no qual se inicia efeito substitutivo. O tempo sob o sombrite concentrou-se no período das 8 às 14 h e reduziu com a suplementação até 1,2% PV. A suplementação afeta o comportamento ingestivo de ovinos em pastejo, uma vez que, no nível de 1,2% PV, houve otimização da capacidade ingestiva dos animais. Níveis superiores de suplementação comprometem o comportamento ingestivo, por isso, são necessárias alterações na formulação do suplemento quando utilizados níveis mais elevados.

Relação entre hábitos bucais, alterações oronasofaringianas e mal-oclusões em pré-escolares de Vitória, Espírito Santo, Brasil

O objetivo deste estudo foi estimar a prevalência das mal-oclusões e variáveis a elas associadas, como hábitos deletérios (HD) e as alterações oronasofaringianas (AO), respiração bucal, deglutição atípica e fonação atípica, em crianças com idade de três anos, no Município de Vitória, Espírito Santo, Brasil. A amostra constituiu-se de 291 crianças de ambos os sexos, matriculadas nos Centros de Educação Infantil, selecionadas por meio de amostragem probabilística por conglomerados. A análise de regressão logística indicou maior risco relativo (RR) de crianças com sobressaliência alterada, mordida aberta e mordida cruzada, em apresentar: respiração bucal (RR = 1,89; IC: 1,56-2,03), (RR = 2,46; IC: 2,00-3,02), (RR = 1,45; IC: 1,23-1,72); deglutição atípica (RR = 2,57; IC: 1,87-3,52), (RR = 3,49; IC: 2,53-4,81), (RR = 1,86; IC: 1,46-2,39) e fonação atípica (RR = 2,25; IC: 1,66-3,05), (RR = 3,18; IC: 2,38-4,25), (RR = 1,71; IC: 1,32-2,22), respectivamente. Foi mostrado haver associação entre sucção de dedo e de chupeta com sobressaliência alterada (p < 0,001) e sucção de chupeta e mordida aberta (p < 0,001). Esses resultados indicam que a prevalência das mal-oclusões está associada aos HD e às AO.

Extração semiautomática de contornos de telhado de edifícios com base em snakes e programação dinâmica

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)