Membrane-bound alkaline phosphatase from ectopic mineralization and rat bone marrow cell culture

Cells from rat bone marrow exhibit the proliferation-differentiation sequence of osteoblasts, form mineralized extracellular matrix in vitro and release alkaline phosphatase into the medium. Membrane-bound alkaline phosphatase was obtained by method that is easy to reproduce, simpler and fast when compared with the method used to obtain the enzyme from rat osseous plate. The membrane-bound alkaline phosphatase from cultures of rat bone marrow cells has a MWr of about 120 kDa and specific PNPP activity of 1200 U/tng. The ecto-enzyme is anchored to the plasma membrane by the GPI anchor and can be released by PIPLC (selective treatment) or polidocanol (0.2 mg/mL protein and 1% (w/v) detergent). The apparent optimum pH for PNPP hydrolysis by the enzyme was pH 10. This fraction hydrolyzes ATP (240 U/mg), ADP (350 U/ mg), glucose 1-phosphate (1100 U/mg), glucose 6-phosphate (340 Wing), fructose 6-phosphate (460 U/mg), pyrophosphate (330 U/mg) and (3glycerophosphate (600 U/mg). Cooperative effects were observed for the hydrolysis of PPi and beta-glycerophosphate. PNPPase activity was inhibited by 0.1 mM vanadate (46%), 0.1 mM ZnCl2 (68%), 1 mM levamisole (66%), 1 mM arsenate (44%), 10 mM phosphate (21%) and 1 mM theophylline (72%). We report the biochemical characterization of membrane-bound alkaline phosphatase obtained from rat bone marrow cells cultures, using a method that is simple, rapid and easy to reproduce. Its properties are compared with those of rat osseous plate enzyme and revealed that the alkaline phosphatase obtained has some kinetics and structural behaviors with higher levels of enzymatic activity, facilitating the comprehension of the mineralization process and its function. (c) 2006 Elsevier B.V. All rights reserved.

Effect of pelleting and pellet size of a concentrate for intensively reared beef cattle on in vitro fermentation by two different approaches

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Fine root isotropy in Eucalyptus grandis plantations. Towards the prediction of root length densities from root counts on trench walls

The objectives of the study were to assess changes in fine root anisotropy and specific root lengths throughout the development of Eucalyptus grandis ( W. Hill ex Maiden) plantations and to establish a predictive model of root length density (RLD) from root intercept counts on trench walls. Fine root densities (<1 mm in diameter) were studied in 6-, 12-, 22-, 28-, 54-, 68- and 72-month-old E. grandis plantations established on deep Ferralsols in southern Brazil. Fine root intercepts were counted on 3 faces of 90-198 soil cubes (1 dm(3) in volume) in each stand and fine root lengths (L) were measured inside 576 soil cubes, sampled between the depths of 10 cm and 290 cm. The number of fine root intercepts was counted on one vertical face perpendicular to the planting row (N(t)), one vertical face parallel to the planting row (N(l)) and one horizontal face (N(h)), for each soil cube sampled. An overall isotropy of fine roots was shown by paired Student's t-tests between the numbers of fine roots intersecting each face of soil cubes at most stand ages and soil depths. Specific root lengths decreased with stand age in the upper soil layers and tended to increase in deep soil layers at the end of the rotation. A linear regression established between N(t) and L for all the soil cubes sampled accounted for 36% of the variability of L. Such a regression computed for mean Nt and L values at each sampling depth and stand age explained only 55% of the variability, as a result of large differences in the relationship between L and Nt depending on stand productivity. The equation RLD=1.89*LAI*N(t), where LAI was the stand leaf area index (m(2) m(-2)) and Nt was expressed as the number of root intercepts per cm(2), made it possible to predict accurately (R(2)=0.84) and without bias the mean RLDs (cm cm(-3)) per depth in each stand, for the whole data set of 576 soil cubes sampled between 2 years of age and the end of the rotation.

Agregação de ume nitossolo vermelho distroférrico sob sistemas de plantio direto, preparo convencional e mata nativa

This study aimed to evaluate the effect of time since the adoption of the no-till system, in comparison with a native forest area and a conventional tillage area, using the distribution of soil aggregates in a Distroferric Red Nitosol. Treatments were as follows: native forest (NF), conventional tillage (CT), no-till for one year (NT1), no-till for four years (NT4), no-till for five years (NT5), and no-till for 12 years (NT12). Aggregate samples were collected randomly within each treatment at depths of 0-5 and 10-15 cm. After sifting the aggregates in water they were separated into the following aggregate classes > 2 mm; < 2 mm; 2-1 mm, and < 1 mm. The adoption time in the no-till system favored soil aggregation. The mean weighted diameter (MWD) of the soil aggregates and the percentage of aggregates greater than 2 mm increased with adoption time in the no-till system at the 0-5 cm depth. The NF and NT12 treatments had higher MWD values in the 0-5 cm layer. CT had the highest percentage of aggregates smaller than 1 mm.

Isoenzimas na diferenciação de sementes de três espécies do gênero Euterpe

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

Eficácia da utilização de grânulos de argila como veículo para a aplicação aérea de sulfentrazone e isoxaflutole em área de implantação de eucalipto

Realizou-se um experimento em uma área de implantação da cultura do eucalipto no município de São Miguel Arcanjo-SP, com o objetivo de avaliar a eficácia da aplicação aérea de grânulos de argila como veículo dos herbicidas sulfentrazone e isoxaflutole, no controle de plantas daninhas. Foi realizada aplicação aérea dos herbicidas sulfentrazone, nas doses de 500 e 750 g i.a. ha-1, e isoxaflutole, nas doses de 150 e 225 g i.a. ha-1, utilizando-se como veículo grânulos de argila com densidade de 1,05 g cm ³, alta capacidade de absorção (24 mL 100 g-1), alta resistência ao desgaste e tamanho das partículas entre 500 mícrons e 1 mm. Também foram feitas aplicações via líquida dos mesmos herbicidas e doses com um pulverizador convencional, acoplado a um trator. Além desses tratamentos, foi mantida uma parcela testemunha, sem aplicação dos herbicidas. Nas parcelas experimentais foram semeadas as espécies de plantas daninhas Brachiaria decumbens, Ipomoea grandifolia, Merremia cissoides e Panicum maximum, sendo realizadas avaliações visuais de controle aos 75 e 110 dias após a aplicação. em geral, foram observados, nas plantas daninhas avaliadas, resultados de controle semelhantes ou superiores para a aplicação aérea (via grânulos) até 75 DAA e superiores para essa modalidade de aplicação aos 110 DAA, indicando uma extensão no período do efeito do residual dos herbicidas estudados.

Melatonin and its kynurenin-like oxidation products affect the microbicidal activity of neutrophils

Activated phagocytes oxidize the hormone melatonin to N-1-acethyl-N-2-formyl-5-methoxykynuramine (AFMK) in a superoxide anion- and myeloperoxidase-dependent reaction. We examined the effect of melatonin, AFMK and its deformylated-product N-acetyl-5-methoxykynuramine (AMK) on the phagocytosis, the microbicidal activity and the production of hypochlorous acid by neutrophils. Neither neutrophil and bacteria viability nor phagocytosis were affected by melatonin, AFMK or AMK. However these compounds affected the killing of Staphylococcus aureus. After 60 min of incubation, the percentage of viable bacteria inside the neutrophil increased to 76% in the presence of 1 mM of melatonin, 34% in the presence of AFMK and 73% in the presence of AMK. The sole inhibition of HOCl formation, expected in the presence of myeloperoxidase substrates, was not sufficient to explain the inhibition of the killing activity. Melatonin caused an almost complete inhibition of HOCl formation at concentrations of up to 0.05 mM. Although less effective, AMK also inhibited the formation of HOCl However, AFMK had no effect on the production of HOCl These findings corroborate the present view that the killing activity of neutrophils is a complex phenomenon, which involves more than just the production of reactive oxygen species. Furthermore, the action of melatonin and its oxidation products include additional activities beyond their antioxidant property. The impairment of the neutrophils' microbicidal activity caused by melatonin and its oxidation products may have important clinical implications, especially in those cases in which melatonin is pharmacologically administered in patients with infections. (c) 2005 Elsevier SAS. All rights reserved.

A Single Intraoperative Sub-Tenon's Capsule Injection of Triamcinolone and Ciprofloxacin in a Controlled-Release System for Cataract Surgery

PURPOSE. To compare intraoperative injection of triamcinolone and ciprofloxacin in a controlled-release system (DuoCat) with prednisolone and ciprofloxacin eye drops after cataract surgery.METHODS. In this randomized, double-masked, controlled trial, a total of 135 patients undergoing cataract surgery were randomly allocated to two groups: 67 patients treated after surgery with prednisolone 1% and ciprofloxacin 3% eye drops four times daily (week 1), three times daily (week 2), twice daily (week 3), and once daily (week 4) and 0.3% ciprofloxacin drops four times daily (weeks 1 and 2), and 68 patients treated at the end of surgery with a sub-Tenon's injection of 25 mg triamcinolone and 2 mg ciprofloxacin in biodegradable microspheres. The patients were examined on postoperative days 1, 3, 7, 14, and 28. The main outcome measures were postoperative anterior chamber cell and flare, intraocular pressure (IOP), lack of anti-inflammatory response, and presence of infection.RESULTS. No significant differences were observed between the groups in anterior chamber cell (P > 0.14) and flare (P > 0.02) at any postoperative visits. The mean (99% confidence interval) differences in IOP between the prednisolone and triamcinolone groups on days 1, 3, 7, 14, and 28 were -0.4 mm Hg (-2.1 to 1.3), 0.0 mm Hg (-1.4 to 1.3), 0.0 mm Hg (-1.1 to 1.1), -0.2 mm Hg (-1.1 to 0.8), and -0.1 mm Hg (-1.1 to 0.9), respectively. No patient had a postoperative infection.CONCLUSIONS. One injection of DuoCat had a therapeutic response and ocular tolerance that were equivalent to conventional eye drops in controlling inflammation after cataract surgery. (Clinical-Trials. gov number, NCT00431028.) (Invest Ophthalmol Vis Sci. 2009; 50: 3041-3047) DOI: 10.1167/iovs.08-2920

Cocaine induces cell death and activates the transcription nuclear factor kappa-b in pc12 cells

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

Hypothalamic supraoptic but not paraventricular nucleus is involved in cardiovascular responses to carbachol microinjected into the bed nucleus of stria terminalis of unanesthetized rats

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

Oxidation of melatonin by taurine chloramine

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

Experimental Investigation into the Characteristics of In-Pipe Leak Noise in Plastic Water Filled Pipes

This paper presents an experimental investigation of the characteristics of leak noise in plastic water-filled pipes. An experimental set-up was designed to identify the physical mechanisms of leak noise generation. Possible mechanisms include cavitation and turbulence. The experiments show that cavitation is not responsible for leak noise generation and clearly indicate that turbulence is the main mechanism, at least in the experiments conducted. An alternative experimental set-up was also designed to identify the characteristics of leak noise spectra and to investigate how the spectra are affected by the leak size and the leak flow velocity. A number of different hole sizes (leaks) starting from 1 mm diameter, increasing progressively every 0.5 mm until a size of 4 mm diameter were tested for different jet velocities and an empirical model that describes this behaviour is proposed.

Model for determining flow diameter and economic velocity in water elevating systems

The purpose of this paper was to develop a model for calculating the economical flow diameter and velocity, by obtaining the economical diameter, using Swamee's friction factor equation, by minimizing the total annual cost. The application of the model to a regular supply condition showed that the diameter of the actual condition, 250 mm, compared with the diameter calculated by the mode, at the same tariff as that applied to the property ( ground), 284.1 mm, involved the necessity to generate, transmit, and distribute extra electrical energy, due to the higher load loss caused by the original diameter, approximately 30800 kWh/year. This means that in one year, the consumer would spend R$2,804.00 more on pumping cost alone.

Cultura de condrócitos em arcabouço tridimensional: hidrogel de alginato

OBJETIVOS: O presente estudo teve como objetivo cultivar condrócitos retirados da articulação do joelho de coelhos encapsulados em hidrogel de alginato (HA) e caracterizar a produção de matriz extracelular (ECM). MÉTODOS: A cartilagem articular foi removida do joelho de coelhos, com três a seis meses, fragmentada em pedaços de 1mm e submetida à digestão enzimática. Uma concentração de 1x106 céls/mL foram ressuspensas em uma solução de alginato de sódio a 1,5% (w/v), em seguida fez-se o processo de gelatinização em CaCl2 (102 mM), permitindo a formação do HA e cultivo em meio DMEM-F12 durante quatro semanas. A distribuição das células e a ECM foram acessadas através das secções histológicas coradas com e azul de toluidina hematoxilina e eosina (HE). RESULTADOS: Houve um aumento no número e na viabilidade dos condrócitos durante as quatro semanas de cultura. Através das análises histológicas dos HAs corados com azul de toluidina e HE foi possível observar a distribuição definida dos condrócitos no hidrogel, assemelhando-se a grupos isógenos e formação de matriz territorial. CONCLUSÃO: Este estudo demonstrou a eficiência do HA como arcabouço para ser usado na cultura de condrócitos, constituindo uma alternativa no reparo de lesões na cartilagem articular.

Evaluation of Pupillary Diameter in Preschool Children

Few quantitative data exist on pupil diameter, particularly in children. In this article we describe an analysis of pupil diameter and pupil area in normal preschool children during attentive gaze (AG) and spontaneous gaze (SG) using digital imaging. The pupil diameter and pupil area of 200 preschool children aged 4 to 6 years were evaluated. The images were recorded with a digital camera, transferred to a computer, processed by movie software and submitted to the Invert filter of Adobe Photoshop (R) software. Three consecutive measures of pupil diameter and pupil area were taken for both eyes during AG and SG. Arithmetic means of each parameter were calculated and data were expressed in millimeters. Mean pupil diameter was similar for both eyes, presenting values of 4.9 mm during AG and 4.1 mm during SG. Mean pupil area was also similar for both eyes, presenting values of 15.6 mm(2) and 9.1 mm(2) during AG and SG, respectively. There were no differences between boys and girls. However, 6-year-old children showed lower pupil diameter and area under both evaluation conditions.