Mode of Peroxisome Proliferator-Activated Receptor gamma Activation by Luteolin

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

Thrombomodulin-independent activation of protein C and specificity of hemostatically active snake venom serine proteinases - Crystal structures of native and inhibited Agkistrodon contortrix contortrix protein C activator

Protein C activation initiated by the thrombin-thrombomodulin complex forms the major physiological anticoagulant pathway. Agkistrodon contortrix contortrix protein C activator, a glycosylated single-chain serine proteinase, activates protein C without relying on thrombomodulin. The crystal structures of native and inhibited Agkistrodon contortrix contortrix protein C activator determined at 1.65 and 1.54 angstrom resolutions, respectively, indicate the pivotal roles played by the positively charged belt and the strategic positioning of the three carbohydrate moieties surrounding the catalytic site in protein C recognition, binding, and activation. Structural changes in the benzamidine-inhibited enzyme suggest a probable function in allosteric regulation for the anion-binding site located in the C-terminal extension, which is fully conserved in snake venom serine proteinases, that preferentially binds Cl1- instead of SO42-.

Activation of Ca2+-independent membrane-damaging activity in Lys49-phospholipase A(2) promoted by amphiphilic molecules

Association of class-II phospholipase A(2) (PLA(2)) with aggregated phospholipid substrate results in elevated levels of the Ca2+-dependent hydrolytic activity. The Asp49 residue participates in coordination of the Ca2+ ion cofactor, however, in Lys49-PLA(2) homologues (Lys49-PLA(2)S), substitution of the Asp49 by Lys results in loss of Ca2+ binding and lack of detectable phospholipid hydrolysis. Nevertheless, Lys49-PLA2S cause Ca2+-independent damage of liposome membranes. Bothropstoxin-I is a homodimeric Lys49-PLA(2) from the venom of Bothrops jararacussu, and in fluorescent marker release and dynamic light scattering experiments with DPPC liposomes we demonstrate activation of the Ca2+-independent membrane damaging activity by similar to4 molecules of sodium dodecyl sulphate (SDS) per protein monomer. Activation is accomparlied by significant changes in the intrinsic tryptophan fluorescence emission (ITFE) and near UV circular dichroism (UVCD) spectra of the protein. Subsequent binding of 7-10 SDS molecules results in further alterations in the ITFE and far UVCD spectra. Reduction in the rate of N-bromosuccinimide modification of Trp77 at the dimer interface suggests that initial binding of SDS to this region accompanies the activation of the membrane damaging activity. 1-anilinonaphthalene-8-sulphonic acid binding studies indicate that subsequent SDS binding to the active site is concomitant with the second structural transition. These results provide insights in the structural basis of amphiphile/protein coupling in class-II PLA(2)s. (C) 2004 Published by Elsevier B.V.

INTRAPULPAL TEMPERATURE VARIATION DURING BLEACHING WITH VARIOUS ACTIVATION MECHANISMS

Objectives: The aim of this study was to evaluate the intrapulpal temperature variation after bleaching treatment with 35% hydrogen peroxide using different sources of activation. Material and Methods: Twenty-four human teeth were sectioned in the mesiodistal direction providing 48 specimens, and were divided into 4 groups (n=12): (G1) Control - Bleaching gel without light activation, (G2) Bleaching gel + halogen light, (G3) Bleaching gel + LED, (G4) Bleaching gel + Nd: YAG Laser. The temperatures were recorded using a digital thermometer at 4 time points: before bleaching gel application, 1 min after bleaching gel application, during activation of the bleaching gel, and after the bleaching agent turned from a dark-red into a clear gel. Data were analyzed statistically by the Dunnet's test, ANOVA and Tukey's test (alpha=0.05). Results: The mean intrapulpal temperature values (degrees C) in the groups were: G1: 0.617 +/- 0.41; G2: 1.800 +/- 0.68; G3: 0.975 +/- 0.51; and G4: 4.325 +/- 1.09. The mean maximum temperature variation (MTV) values were: 1.5 degrees C (G1), 2.9 degrees C (G2), 1.7 degrees C (G3) and 6.9 degrees C (G4). When comparing the experimental groups to the control group, G3 was not statistically different from G1 (p>0.05), but G2 and G4 presented significantly higher (p<0.05) intrapulpal temperatures and MTV. The three experimental groups differed significantly (p<0.05) from each other. Conclusions: The Nd: YAG laser was the activation method that presented the highest values of intrapulpal temperature variation when compared with LED and halogen light. The group activated by LED light presented the lowest values of temperature variation, which were similar to that of the control group.

In vitro evaluation of dental bleaching effectiveness using hybrid lights activation

OBJETIVO: Avaliar se fontes de luz aumentam a eficácia do peróxido de hidrogênio na técnica de clareamento profissional. METODOLOGIA: Foram empregados 60 dentes incisivos bovinos, com dimensões coronárias e radiculares padronizadas a partir do limite amelo-cementário, sendo descartada a porção lingual. Os corpos-de-prova (cp) foram limpos em ultra-som por 20 min e a dentina condicionada com H3PO4 a 38% por 15 s, sendo os (cp) imersos em solução de café solúvel a 25% por duas semanas. A dentina foi impermeabilizada com esmalte e os (cp) divididos em 5 grupos, sendo a cor inicial mensurada através do espectofotômetro-EasyShade (VITA). Todos os (cp) receberam três aplicações por 10 min do gel clareador Opalescence Xtra-Boost (Ultradent) conforme segue: Grupo 1 - controle, não recebeu fotoativação, Grupo 2 - ativado com luz halôgena, Grupo 3 - ativado com LED azul/LASER, Grupo 4 - ativado com LED verde/LASER e Grupo 5 - ativado com LED vermelho. Após o clareamento foi mensurada a variação de cor E, a*, b*e L* e as referentes à escala de cor Vita Clássico. Os dados foram submetidos à análise de variância, teste de Tukey e de Dunn (α=5%). RESULTADOS: A diferença geral da cor foi reduzida quando se empregou LED Azul e Luz Halógena, sendo que o desempenho do peróxido de hidrogênio a 38% foi intensificado dependendo da fonte de luz utilizada. A avaliação quantitativa de cor, obtida por espectrofotômetro e pela escala de cor Vita Clássico, foram coincidentes. CONCLUSÃO: O tipo de fonte de luz empregada interfere na eficácia do agente clareador.

Influence of chemical activation of a 35% hydrogen peroxide bleaching gel on its penetration and efficacy-In vitro study

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

The Influence of Chemical Activation on Tooth Bleaching Using 10% Carbamide Peroxide

The aim of this study was to assess the influence of manganese gluconate, a chemical activator of bleaching agents, at a concentration of 0.01% on the efficiency of a 10% carbamide peroxide-based bleaching agent. Forty bovine incisors were immersed in a 25% instant coffee solution for seven days and randomly divided into two groups. Group 1 was the control group and consisted of 10% carbamide peroxide-based bleaching gel only. Group 2 consisted of 10% carbamide peroxide-based bleaching gel and 0.01% manganese gluconate. Three readings of color were taken using the Vita Easy-shade spectrophotometer: the initial reading, a reading at seven days, and a reading at 14 days. Total color variation was calculated by Delta E*Lab. Data were submitted to the statistical t-test (5%), which showed that after seven days group 2 had a significant increase in the degree of tooth bleaching compared with group 1. The mean values (+/-SD) were 16.33 (+/-3.95) for group 1 and 19.29 (+/-4.97) for group 2. However, the results for group 1 and group 2 were similar after 14 days. Adding 0.01% manganese gluconate to 10% carbamide peroxide bleaching gel increased the degree of tooth bleaching after a seven-day treatment and did not influence the resulting shade after 14 days.

In Vitro Assessment of Chemical Activation Efficiency During In-office Dental Bleaching

Purpose: This study compared five types of chemical catalyzing agents added to 35% hydrogen peroxide gel, with regard to their capacity of intensifying in-office dental bleaching results.Methods: One-hundred and twenty bovine incisors were used, of which the crowns and roots were cut in the incisor-apical direction, to acquire the dimensions of a human central incisor. The specimens were sectioned in the mesiodistal direction by means of two longitudinal cuts, the lingual halves being discarded. The vestibular halves received prophylaxis with a bicarbonate jet, ultrasound cleaning and acid etching on the dentinal portion. Next, the specimens were stored in receptacles containing a 25% instant coffee solution for two weeks. After the darkening period, initial measurement of the shade obtained was taken with the Easy Shade appliance, which allowed it to be quantified by the CIELab* method. The samples were divided into six groups, corresponding to the chemical activator used: a) none (CON); b) ferric chloride (CF); c) ferrous sulphate (SF); d) manganese gluconate (GM); e) manganese chloride (CM); f) mulberry root extract (RA). Each group received three 10-minute applications of the gels containing the respective activating agents. Next, a new shade measurement was made.Results: The Analysis of Variance and Tukey tests (alpha=5%) showed statistically significant differences for the shade perception values (p=0.002). Groups GM, CM and RA showed significantly higher means than the control group.Conclusion: The presence of some chemical activators is capable of resulting in a significant increase in tooth shade variation.

Activation energy for the reverse eutectoid reaction in hypo-eutectoid Cu-Al alloys

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

Influence of Ag additions on the activation energy for the reverse eutectoid reaction in Cu-Al alloys

The eutectoid transformation may be defined as a solid-state diffusion-controlled decomposition process of a high-temperature phase into a two-phase lamellar aggregate behind a migrating boundary on cooling below the eutectoid temperature. In substitutional solid solutions, the eutectoid reaction involves diffusion of the solute atoms either through the matrix or along the boundaries or ledges. The effect of Ag on the non-isothermal kinetics of the reverse eutectoid reaction in the Cu-9 mass%Al, Cu-10 mass%Al, and Cu-11 mass%Al alloys were studied using differential scanning calorimetry (DSC), X-ray diffraction (XRD), and scanning electron microscopy (SEM). The activation energy for this reaction was obtained using the Kissinger and Ozawa methods. The results indicated that Ag additions to Cu-Al alloys interfere on the reverse eutectoid reaction, increasing the activation energy values for the Cu-9 mass%Al and Cu-10 mass%Al alloys and decreasing these values for the Cu-11 mass%Al alloy for additions up to 6 mass%Ag. The changes in the activation energy were attributed to changes in the reaction solute and in Ag solubility due to the increase in Al content.

Macrophage activation by Paepalanthus spp. extracts

Macrófagos são conhecidos por exercerem uma importante função de mecanismo de defesa quando estas células são estimuladas com produtos naturais e produtos bacterianos (dentre outros). Uma variedade de citocinas e compostos químicos são liberados para induzir sistema de defesa fundamental. Entre outros peróxido de hidrogênio (H2O2) tem sido identificado como moléculas tendo multifunções. Entretanto está bem estabelecido que (H2O2) está envolvido em inúmeros processos fisiológicos, como por exemplo, neurotransmissão, relaxamento da musculatura lisa ou regulação imune. Para a determinação de peróxido de hidrogênio (H2O2) em macrófago peritoneal em camundongos, determinou-se a ação imunomodulatória de extratos (etanólico e etanólico 70%) obtidos de quatro espécies do gênero Paepalanthus (Eriocaulaceae) na concentração de 10 mg/mL. Os estratos etanólicos 70 % de capítulos de P. Hilairei, P. robustus, P. vellozioides e P. speciocus apresentaram maior liberação de (H2O2) do que os outros extratos etanólicos.

Influence of activation on the multipoint immobilization of penicillin G acylase on macroporous silica

Penicillin G acylase is the second most important enzyme used by industry in an immobilized form. Penicillin hydrolysis is its main application. This reaction is used to produce 6-aminopenicillanic acid (6-APA), an intermediate in the synthesis of semisynthetic antibiotics. This work aims to compare catalytic properties of different penicillin G acylase (PGA) derivatives obtained by multipoint immobilization of the enzyme on macroporous silica. Enzyme amino groups react with different aldehyde groups produced in the support using either glutaraldehyde or glyoxyl activation. In the former method, silica reacts with g-aminopropyltriethoxysilane (g-APTS) and glutaraldehyde; in the latter, a reaction with glycidoxypropyltrimethoxysilane (GPTMS) is followed by acid hydrolysis and oxidation using sodium periodate. This work determines the influence of degree of activation, using glutaraldehyde, on immobilization parameters. PGA was immobilized on these two different supports. Maximum enzyme load, immobilized enzyme activity (derivative activity), rate of immobilization and thermal stability were checked for both cases. For glutaraldehyde activation, the results showed that 0.5% of the g-APTS is sufficient for all the hydroxyl groups in the silica to react. They also showed that degree of activation only affects immobilization yield and reaction velocity and that reduction of the glutaraldehyde derivatives with sodium borohydride does not affect their thermal stability. In comparing the derivatives obtained using glyoxyl and glutaraldehyde activation, it was observed that the glyoxyl derivatives presented better immobilization parameters, with a maximum enzyme load of 264 IU/g silica and a half-life of 20 minutes at 60 °C.

THE ROLE OF ANGIOTENSIN AT1 RECEPTORS IN THE DIURETIC, NATRIURETIC, KALIURETIC AND BLOOD-PRESSURE RESPONSES INDUCED BY ANGIOTENSIN-II ACTIVATION OF THE MEDIAN PREOPTIC NUCLEUS IN CONSCIOUS RATS

We determined the effects of two classical angiotensin II (ANG II) antagonists, [Sar(1), Ala(8)]-ANG II and [Sar(1), Thr(8)]-ANG II, and losartan (a nonpeptide and selective antagonist for the AT 1 angiotensin receptors) on diuresis, natriuresis, kaliuresis and arterial blood pressure induced by ANG II administration into the median preoptic nucleus (MnPO) of male Holtzman rats weighing 250-300 g. Urine was collected in rats submitted to a water load (5% body weight) by gastric gavage, followed by a second water load (5% body weight) 1 h later. The volume of the drug solutions injected was 0.5 mu l over 10-15 s. Pre-treatment with [Sar(1), Ala(8)]-ANG II (12 rats) and [Sar(1), Thr(8)]-ANG II (9 rats), at the dose of 60 ng reduced (13.7 +/- 1.0 vs 11.0 +/- 1.0 and 10.7 +/- 1.2, respectively), whereas losartan (14 rats) at the dose of 160 ng totally blocked (13.7 +/- 1.0 vs 7.6 +/- 1.5) the urine excretion induced by injection of 12 ng of ANG II (14 rats). [Sar(1), Ala(8)]-ANG II impaired Na+ excretion (193 +/- 16 vs 120 +/- 19): whereas [Sar(1), Thr(8)]-ANG II and losartan blocked Na+ excretion (193 +/- 16 vs 77 +/- 15 and 100 +/- 12, respectively) induced by ANG II. Similar effects induced by ANG II on K+ excretion were observed with [Sar(1), Ala(8)]-ANG II, [Sar(1), Thr(8)]-ANG II, and losartan pretreatment (133 +/- 18 vs 108 +/- 11, 80 +/- 12, and 82 +/- 15, respectively). The same doses as above of [Sar(1), Ala(8)]-ANG II (8 rats), [Sar(1), Thr(8)]-ANG II (8 rats). and losartan (9 rats) blocked the increase in the arterial blood pressure induced by 12 ng of ANG II (12 rats) (32 +/- 4 ru 4 +/- 2, 3.5 +/- 1, and 2 +/- 1: respectively. The results indicate that the AT1 receptor subtype participates in the increases of diuresis, natriuresis. kaliuresis and arterial blood pressure induced by the administration of ANG II into the MnPO.