Viscerocutaneous form of loxoscelism and erythrocyte glucose-6-phosphate deficiency

In a period of time of five years, all patients who exhibited viscerocutaneous form of loxoscelism were investigated for erythrocyte glucose-6-phosphate deficiency, and in two patients out of seven it was found this deficiency. This finding suggests that this genetical enzyme deficiency could account for the hemolysis after Loxosceles bite, at least in some of the cases.

Glucose-6-phosphate dehydrogenase and glutathione reductase activity in methemoglobin reduction by methylene blue and cyst amine: study on glucose-6-phosphate dehydrogenase-deficient individuals, on normal subjects and on riboflavin-treated subjects

The authors have standardized methods for evaluation of the activity of the glucose-6-phosphate dehydrogenase and of glutathione reductase. The general principle of the first method was based on methemoglobin formation by sodium nitrite followed by stimulation of the glucose-6-phosphate dehydrogenase with methylene blue. Forty six adults (23 males and 23 females) were studied. Subjects were not G6PD deficient and were aged 20 to 30 years. The results showed that methemoglobin reduction by methylene blue was 154.40 and 139.90 mg/min (p<0.05) for males and females, respectively, in whole blood, and 221.10 and 207.85 mg/min (n.s.), respectively, in washed red cells. These data showed that using washed red cells and 0.7g% sodium nitrite concentration produced no differences between sexes and also shortened reading time for the residual amount of methemoglobin to 90 minutes. Glutathione reductase activity was evaluated on the basis of the fact that cystamine (a thiol agent) binds to the SH groups of hemoglobin, forming complexes. These complexes are reversed by the action of glutathione reductase, with methemoglobin reduction occurring simultaneously with this reaction. Thirty two adults (16 males and 16 females) were studied. Subjects were not G6PD deficient and were aged 20 to 30 years. Methemoglobin reduction by cystamine was 81.27 and 91.13 mg/min (p<0.01) for males and females, respectively. These data showed that using washed red cells and 0.1 M cystamine concentration permits a reading of the residual amount of methemoglobin at 180 minutes of incubation. Glutathione reductase activity was evaluated by methemoglobin reduction by cystamine in 14 females before and after treatment with 10 mg riboflavin per day for 8 days. The results were 73.69 and 94.26 jug/min (p<0.01) before and after treatment, showing that riboflavin treatment increase glutathione reductase activity even in normal individuals. Three Black G6PD-deficient individuals (2 males and 1 female) were also studied. The G6PD and glutathione reductase were partially activated, the change being more intense in males. On the basis of race and of the laboratory characteristics observed, it is possible to suggest that the G6PD deficiency of these individuals is of the African type and that the female is heterozygous for this deficiency. Analysis of the results as a whole permitted us to conclude that the methods proposed here were efficient for evaluating the activity of the glucose-6-phosphate dehydrogenase and of glutathione reductase. The latter is dependent on the pentose pathway, which generates NADPH, and on riboflavin, a FAD precursor vitamin.

MEMBRANE FRACTIONS FROM Strongyloides venezuelensis IN THE IMMUNODIAGNOSIS OF HUMAN STRONGYLOIDIASIS

Strongyloides venezuelensis is a parasitic nematode of rodents frequently used to obtain heterologous antigens for the immunological diagnosis of human strongyloidiasis. The aim of this study was to evaluate membrane fractions from S. venezuelensis for human strongyloidiasis immunodiagnosis. Soluble and membrane fractions were obtained in phosphate saline (SS and SM) and Tris-HCl (TS and TM) from filariform larvae of S. venezuelensis. Ninety-two serum samples (n = 92) were obtained from 20 strongyloidiasis patients (Group I), 32 from patients with other parasitic diseases (Group II), and 40 from healthy individuals (Group III), and were analyzed by enzyme-linked immunosorbent assay (ELISA). Soluble fractions (SS and TS) showed 90.0% sensitivity and 88.9% specificity, whereas the membrane fractions (SM and TM) showed 95.0% sensitivity and 94.4% specificity. The present results suggest the possible use of membrane fractions of S. venezuelensis as an alternative antigen for human strongyloidiasis immunodiagnosis.

Isoniazid acetylating phenotype in patients with paracoccidioidomycosis and its relationship with serum sulfadoxin levels, glucose-6-phosphate dehydrogenase and glutathione reductase activities

The authors evaluated the isoniazid acetylating phenotype and measured hematocrit, hemoglobin, glucose-6-phosphate dehydrogenase and glutathione reductase activities plus serum sulfadoxin levels in 39 patients with paracoccidioidomycosis (33 males and 6 females) aged 17 to 58 years. Twenty one (53.84%) of the patients presented a slow acetylatingphenotype and 18(46.16%) a fast acetylating phenotype. Glucose-6-phosphate- dehydrogenase (G6PD) acti vity was decreased in 5(23.80%) slow acetylators and in 4(22.22%) fast acetylators. Glutathione reductase activity was decreased in 14 (66.66%) slow acetylators and in 12 (66.66%) fast acetylators. Serum levels of free and total sulfadoxin Were higher in slow acetylator (p < 0.02). Analysis of the resultspermitted us to conclude that serum sulfadoxin levels are related to the acetylatorphenotype. Furthermore, sulfadoxin levels were always above 50 µg/ml, a value considered therapeutic. Glutathione reductase deficiency observed in 66% of patients may be related to the intestinal malabsorption of nutrients, among them riboflavin, a FAD precursor vitamin, inpatients with paracoceidioidomycosis.

Pyruvate kinase and glucose-6-phosphate dehydrogenase deficies and their association with malaria - population genetics and proteomic studies

A malária é reconhecida como uma das principais forças selectivas a actuar na história recente no genoma humano. Inúmeros polimorfismos genéticos têm sido descritos como protectores contra a gravidade da malária, como o alelo HbS (designado de traço falciforme) e o alelo G6PD A- (associado à deficiência de G6PD). Mais recentemente, também a deficiência de PK foi associada com a protecção contra a malária. Evidências desta associação foram obtidas em estudos com modelos de roedor e estudos in vitro utilizando GV humanos deficientes em PK. Até à data, não foram obtidos dados em populações humanas que revelem esta associação: ainda não foi identificada uma variante de PK com uma prevalência elevada em regiões endémicas de malária e não foram identificadas marcas de selecção na região do gene que codifica para a PK (gene PKLR). Além disso, os mecanismos subjacentes à protecção contra a malária por deficiências enzimáticas dos GV não estão bem esclarecidos. Assim, os objectivos do presente estudo foram: investigar os polimorfismos genéticos humanos com associação com a malária em Cabo Verde; pesquisar marcas de selecção da malária na região do gene PKLR em populações Africanas; determinar a frequência da deficiência em PK e identificar uma eventual variante da enzima que possa estar sob selecção positiva em regiões endémicas de malária; avaliar o efeito das duas deficiências enzimáticas (PK e G6PD) na invasão e maturação do parasita em culturas in vitro de Plasmodium usando GV normais e deficientes; e analisar o perfil proteómico de GV infectados e não infectados, normais e com deficiência (em PK e G6PD), bem como de parasitas isolados de GV tanto deficientes como normais. Em Cabo Verde (área epidémica), não foram identificadas marcas de selecção pela malária, através da análise dos vários polimorfismos. No entanto, quando a análise foi realizada em dois países endémicos (Angola e Moçambique), foram detectadas várias marcas de selecção: a genotipagem de microssatélites (STRs) e polimorfismos de base única (SNPs) localizados na vizinhança do gene PKLR revelou uma diferenciação consideravelmente maior entre as populações Africana e Europeia (Portuguesa), do que a diferenciação determinada aquando da utilização de marcadores genéticos neutros. Além disso, uma região genómica de maior amplitude apresentou um Desequilíbrio de Ligação (LD) significativo no grupo de malária não grave (e não no grupo de malária grave), sugerindo que a malária poderá estar a exercer pressão selectiva sobre a região do genoma humano que envolve o gene PKLR. No estudo que incidiu na determinação da prevalência da deficiência de PK no continente Africano (realizado em Moçambique), esta revelou-se elevada - 4,1% - sendo o valor mais elevado descrito até ao momento a nível mundial para esta enzimopatia. Na pesquisa de mutações que pudessem estar na causa deste fenótipo (baixa actividade de PK), foi identificada uma mutação não sinónima 829G>A (277Glu>Lys), significativamente associada à baixa actividade enzimática. Esta mutação foi também identificada em Angola, São Tomé e Príncipe e Guiné Equatorial, onde a frequência de portadores heterozigóticos foi entre 2,6 e 6,7% (valores que se encontram entre os mais elevados descritos globalmente para mutações associadas à deficiência em PK). Não foi possível concluir acerca da associação entre a deficiência de PK e o grau de severidade da malária e da associação entre o alelo 829A e a mesma, devido ao baixo número de amostras. Os resultados dos ensaios de invasão/maturação do parasita sugeriram que, nos GV com deficiência de PK ou G6PD, a invasão (onde está envolvida a membrana do GV hospedeiro e o complexo apical do parasita) é mais relevante para a eventual protecção contra a malária do que a maturação. Os resultados da análise proteómica revelaram respostas diferentes por parte do parasita nas duas condições de crescimento (GV com deficiência de PK e GV com deficiência de G6PD). Esta resposta parece ser proporcional à gravidade da deficiência enzimática. Nos parasitas que cresceram em GV deficientes em G6PD (provenientes de um indivíduo assintomático), a principal alteração observada (relativamente às condições normais) foi o aumento do número de proteínas de choque térmico e chaperones, mostrando que os parasitas responderam às condições de stress oxidativo, aumentando a expressão de moléculas de protecção. Nos parasitas que cresceram em condições de deficit de PK (GV de indivíduo com crises hemolíticas regulares, dependente de transfusões sanguíneas), houve alteração da expressão de um maior número de proteínas (relativamente ao observado em condições normais), em que a maioria apresentou uma repressão da expressão. Os processos biológicos mais representados nesta resposta do parasita foram a digestão da hemoglobina e a troca de proteínas entre hospedeiro e parasita/remodelação da superfície do GV. Além disso, uma elevada percentagem destas proteínas com expressão alterada está relacionada com as fendas de Maurer, que desempenham um papel importante na patologia da infecção malárica. É colocada a hipótese de que a protecção contra a malária em GV deficientes em PK está relacionada com o processo de remodelação da membrana dos GV pelo parasita, o que pode condicionar a invasão por novos parasitas e a própria virulência da malária. Os resultados da análise do proteoma dos GV contribuirão para confirmar esta hipótese.

Inhibition of SNF1-related protein kinase1 by trehalose 6-phosphate and other metabolites and the interrelation with plant gorwth

All life forms need to monitor carbon and energy availability to survive and this is especially true for plants which must integrate unavoidable environmental conditions with metabolism for cellular homeostasis maintenance. Sugars, in the heart of metabolism, are now recognized as crucial signaling molecules that translate those conditions. One such signal is trehalose 6- phosphate (T6P), a phosphorylated dimer of glucose molecules which levels correlate well with those of sucrose (Suc). Central integrators of stress and energy regulation include the conserved plant Snf1-related kinase1 (SnRK1) which respond to low cellular energy levels by up-regulating energy conserving and catabolic metabolism and down-regulating energy consuming processes. In 2009 T6P was shown to inhibit SnRK1. The in vitro inhibition of SnRK1 by T6P was confirmed in vivo through the observation that genes normally induced by SnRK1 were repressed by T6P and vice-versa, promoting growth processes. These observations provided a model for the regulation of growth by sugar.(...)

Bowel preparation for colonoscopy: comparison of mannitol and sodium phosphate. Results of a prospective randomized study

METHOD: Eighty patients were prospectively randomized for precolonoscopic cleansing either with 750 ml of 10% mannitol (Group M) or 180 ml of a sodium phosphate preparation (Group NaP). Laboratory examinations before and after preparation on all patients included hemoglobin, hematocrit, sodium, potassium, phosphorous, calcium and serum osmolarity. A questionnaire was used to assess undesirable side effects and patient tolerance to the solution. The quality of preparation was assessed by the endoscopist who was unaware of the solution employed. RESULTS: Statistically significant changes were verified in serum sodium, phosphorous, potassium and calcium between the two groups, but no clinical symptoms were observed. There were no significant differences in the frequency of side effects studied. Six of the eight patients in Group NaP who had taken mannitol for a previous colonoscopy claimed better acceptance of the sodium phosphate solution. The endoscopic-blinded trial reported excellent or good bowel preparation in 85% prepared with sodium phosphate versus 82.5% for mannitol (p=0.37). CONCLUSIONS: Quality of preparation and frequency of side effects was similar in the two solutions. The smaller volume of sodium phosphate necessary for preparation seems to be related to its favorable acceptance. Nevertheless, the retention of sodium and phosphate ions contraindicates the use of sodium phosphate in patients with renal failure, cirrhosis, ascites, and heart failure.

Organic-inorganic nanostructured multilayers for calcium phosphate biomineralization

The weak fixation of biomaterials within the bone structure is one of the major reasons of implants failures. Calcium phosphate (CaP) coatings are used in bone tissue engineering to improve implant osseointegration by enhancing cellular adhesion, proliferation and differentiation, leading to a tight and stable junction between implant and host bone. It has also been observed that materials compatible with bone tissue either have a CaP coating or develop such a calcified surface upon implantation. Thus, the development of bioactive coatings becomes essential for further improvement of integration with the surrounding tissue. However, most of current applied CaP coatings methods (e.g. physical vapor deposition), cannot be applied to complex shapes and porous implants, provide poor structural control over the coating and prevent incorporation of bioactive organic compounds (e.g. antibiotics, growth factors) because of the used harsh processing conditions. Layer-by-layer (LbL) is a versatile technology that permits the building-up of multilayered polyelectrolyte films in mild conditions based on the alternate adsorption of cationic and anionic elements that can integrate bioactive compounds. As it is recognized in natureâ s biomineralization process the presence of an organic template to induce mineral deposition, this work investigate a ion based biomimetic method where all the process is based on LbL methodology made of weak natural-origin polyelectrolytes. A nanostructured multilayer component, with 5 or 10 bilayers, was produced initially using chitosan and chondroitin sulphate polyelectrolyte biopolymers, which possess similarities with the extracellular matrix and good biocompatibility. The multilayers are then rinsed with a sequential passing of solutions containing Ca2+ and PO43- ions. The formation of CaP over the polyelectrolyte multilayers was confirmed by QCM-D, SEM and EDX. The outcomes show that 10 polyelectrolyte bilayer condition behaved as a  better site for initiating the formation of CaP as the precipitation occur at earlier stages than in 5 polyelectrolyte bilayers one. This denotes that higher number of bilayers could hold the CaP crystals more efficiently. This work achieved uniform coatings that can be applied to any surface with access to the liquid media in a low-temperature method, which potentiates the manufacture of effective bioactive biomaterials with great potential in orthopedic applications.

State of the art and open questions on cathode preparation based on carbon coated lithium iron phosphate

One important component with particular relevance in battery performance is the cathode, being one of the main responsible elements for cell capacity and cycle life. Carbon coated lithium iron phosphate, C-LiFePO4, active material is one of the most promising cathode materials for the next generation of large scale lithium ion battery applications and strong research efforts are being devoted to it, due to its excellent characteristics, including high capacity, ~170 mAh/g, and safety. This review summarizes the main developments on C-LiFePO4 based cathode film preparation and performance. The effect of the binder, conductive additive, relationship between active material-binder-conductive additive and drying step, in the electrode film fabrication and performance is presented and discussed. Finally, after the presentation of the cell types fabricated with C-LiFePO4 active material and their performance, some conclusions and guidelines for further investigations are outlined.

Estudo do comportamento tribológico de materiais compósitos à base de titânio reforçado com partículas cerâmicas para aplicação em implantes articulares

Dissertação de mestrado integrado em Engenharia Biomédica

Metabolic and hemodynamic effects of saline infusion to maintain volemia on temporary abdominal aortic occlusion

OBJECTIVE: To analyze hemodynamic and metabolic effects of saline solution infusion in the maintenance of blood volume in ischemia-reperfusion syndrome during temporary abdominal aortic occlusion in dogs. METHODS: We studied 20 dogs divided into 2 groups: the ischemia-reperfusion group (IRG, n=10) and the ischemia-reperfusion group with saline solution infusion aiming at maintaining mean pulmonary arterial wedge pressure between 10 and 20 mmHg (IRG-SS, n=10). All animals were anesthetized with sodium thiopental and maintained on spontaneous ventilation. Occlusion of the supraceliac aorta was obtained with inflation of a Fogarty catheter inserted through the femoral artery. After 60 minutes of ischemia, the balloon was deflated, and the animals were observed for another 60 minutes of reperfusion. RESULTS: IRG-SS dogs did not have hemodynamic instability after aortic unclamping, and the mean systemic blood pressure and heart rate were maintained. However, acidosis worsened, which was documented by a greater reduction of arterial pH that occurred especially due to the absence of a respiratory response to metabolic acidosis that was greater with the adoption of this procedure. CONCLUSION: Saline solution infusion to maintain blood volume avoided hemodynamic instability after aortic unclamping. This procedure, however, caused worsening in metabolic acidosis in this experimental model.

Factors affecting phosphate fixation in soils of the state of São Paulo

A laboratory experiment was carried out to study the effects of chemical and physical characteristics of the soil on the phosphate fixing capacity. One hundred samples collected from various localities were at first characterized chemically and their particlesize distribution determined. They were then tested as to their phosphate fixing capacities. The results obtained were statistically analysed by means of both simple linear and multiple correlation. The following conclusions could be drawn: 1. simple linear regression analysis indicated that % C, exchangeable Al+3, CEC, % clay, pH and % sand were the soil characteristics which significantly affected phosphate fixing capacity of São Paulo State soils; 2. multiple linear regression analysis indicated that % C, exchangeable Mg(+2)9 exchangeable- Al+3 and % clay were the soil characteristics which significantly affected the phosphate fixing capacity of São Paulo State soils; 3. the phosphate phonomena fixing as they occur in the soils of the São Paulo State can be best described by the following equation: Y = -2,266 - 3,484 + 3,514 + 5,559 + 1,005 %C Mg+2 Al+3 % clay exchangeable exchangeable 4. phosphate fixation in the soil is affected by the combined effects of both soil chemical and physical characteristics.

The phosphate fixing capacities of two soils from the Piracicaba municipality: latosol and sands

An essay was carried out under laboratory conditions in order to know the phosphate fixing capcity of a Latosol and a Sands from the municipality of Piracicaba, State of São Paulo, Brazil. The results showed that they were able to fix about 1,588 and 274 kg/ha of P2O5> respectively.

Translocation of the yeast dolichol-phosphate-mannose synthase into microsomal membranes.

Dolichol-phosphate-mannose synthase catalyzes the formation of Dolichol-phosphate-mannose from Dolichol-phosphate and GDP-mannose. Analysis of the primary amino acid sequence of the yeast enzyme predicts a luminal orientation of the enzyme in the endoplasmic reticulum. We analysed the translocation of the Dolichol-phosphate-mannose synthase into dog pancreatic microsomal membranes: resistance to proteolytic attack provides evidence of its luminal orientation and asks for a reevaluation of the topology of the reaction.