χ-conopeptide MrIA partially overlaps desipramine and cocaine binding sites on the human norepinephrine transporter

The interactions of chi-conopeptide MrIA with the human norepinephrine transporter (hNET) were investigated by determining the effects of hNET point mutations on the inhibitory potency of MrIA. The mutants were produced by site-directed mutagenesis and expressed in COS-7 cells. The potency of MrIA was greater for inhibition of uptake by hNET of [H-3] norepinephrine (K-i 1.89 muM) than [H-3] dopamine (K-i 4.33 muM), and the human dopamine transporter and serotonin transporter were not inhibited by MrIA ( to 7 muM). Of 18 mutations where hNET amino acid residues were exchanged with those of the human dopamine transporter, MrIA had increased potency for inhibition of [H-3] norepinephrine uptake for three mutations ( in predicted extracellular loops 3 and 4 and transmembrane domain (TMD) 8) and decreased potency for one mutation (in TMD6 and intracellular loop (IL) 3). Of the 12 additional mutations in TMDs 2, 4, 5, and 11 and IL1, three mutations (in TMD2 and IL1) had reduced MrIA inhibitory potency. All of the other mutations tested had no influence on MrIA potency. A comparison of the results with previous data for desipramine and cocaine inhibition of norepinephrine uptake by the mutant hNETs reveals that MrIA binding to hNET occurs at a site that is distinct from but overlaps with the binding sites for tricyclic antidepressants and cocaine.

Surface stickiness of drops of carbohydrate and organic acid solutions during convective drying: Experiments and modeling

Drying kinetics of low molecular weight sugars such as fructose, glucose, sucrose and organic acid such as citric acid and high molecular weight carbohydrate such as maltodextrin (DE 6) were determined experimentally using single drop drying experiments as well as predicted numerically by solving the mass and heat transfer equations. The predicted moisture and temperature histories agreed with the experimental ones within 6% average relative (absolute) error and average difference of +/- 1degreesC, respectively. The stickiness histories of these drops were determined experimentally and predicted numerically based on the glass transition temperature (T-g) of surface layer. The model predicted the experimental observations with good accuracy. A nonsticky regime for these materials during spray drying is proposed by simulating a drop, initially 120 mum in diameter, in a spray drying environment.

Identification of slow and fast-acting toxins in a highly ciguatoxic barracuda (Sphyraena barracuda) by HPLC/MS and radiolabelled ligand binding

A barracuda implicated in ciguatera fish poisoning in Guadeloupe was estimated to have an overall flesh toxicity of 15 MUg/g using mouse bioassay. A lipid soluble extract was separated into two toxic fractions, FrA and FrB, on a LH20 Sephadex column eluted with dichloromethane/methanol (1:1). When intraperitoneal injected into mice, FrA provoked symptoms characteristic of slow-acting ciguatoxins, whereas FrB produced symptoms indicative of fast-acting toxins (FAT). High performance liquid chromatography/mass spectrometry/radio-ligand binding (HPLC/MS/RLB) analysis confirmed the two fractions were distinct, because only a weak overlap of some compounds was observed. HPLC/MS/RLB analysis revealed C-CTX-1 as the potent toxin present in FrA, and two coeluting active compounds at m/z 809.43 and 857.42 in FrB, all displaying the characteristic pattern of ion formation for hydroxy-polyethers. Other C-CTX congeners and putative hydroxy-polyether-like compounds were detected in both fractions, however, the RLB found them inactive. C-CTX-1 accounted for >90% of total toxicity in this barracuda and was confirmed to be a competitive inhibitor of brevetoxin binding to voltage-sensitive sodium channels (VSSCs) with a potency two-times lower than P-CTX-1. However, FAT active on VSSCs and

The peroxisome proliferator-activated receptor beta/delta agonist, GW501516, regulates the expression of genes involved in lipid catabolism and energy uncoupling in skeletal muscle cells

Lipid homeostasis is controlled by the peroxisome proliferator-activated receptors (PPARalpha, -beta/delta, and -gamma) that function as fatty acid-dependent DNA-binding proteins that regulate lipid metabolism. In vitro and in vivo genetic and pharmacological studies have demonstrated PPARalpha regulates lipid catabolism. In contrast, PPARgamma regulates the conflicting process of lipid storage. However, relatively little is known about PPARbeta/delta in the context of target tissues, target genes, lipid homeostasis, and functional overlap with PPARalpha and -gamma. PPARbeta/delta, a very low-density lipoprotein sensor, is abundantly expressed in skeletal muscle, a major mass peripheral tissue that accounts for approximately 40% of total body weight. Skeletal muscle is a metabolically active tissue, and a primary site of glucose metabolism, fatty acid oxidation, and cholesterol efflux. Consequently, it has a significant role in insulin sensitivity, the blood-lipid profile, and lipid homeostasis. Surprisingly, the role of PPARbeta/delta in skeletal muscle has not been investigated. We utilize selective PPARalpha, -beta/delta, -gamma, and liver X receptor agonists in skeletal muscle cells to understand the functional role of PPARbeta/delta, and the complementary and/or contrasting roles of PPARs in this major mass peripheral tissue. Activation of PPARbeta/delta by GW501516 in skeletal muscle cells induces the expression of genes involved in preferential lipid utilization, beta-oxidation, cholesterol efflux, and energy uncoupling. Furthermore, we show that treatment of muscle cells with GW501516 increases apolipoprotein-A1 specific efflux of intracellular cholesterol, thus identifying this tissue as an important target of PPARbeta/delta agonists. Interestingly, fenofibrate induces genes involved in fructose uptake, and glycogen formation. In contrast, rosiglitazone-mediated activation of PPARgamma induces gene expression associated with glucose uptake, fatty acid synthesis, and lipid storage. Furthermore, we show that the PPAR-dependent reporter in the muscle carnitine palmitoyltransferase-1 promoter is directly regulated by PPARbeta/delta, and not PPARalpha in skeletal muscle cells in a PPARgamma coactivator-1-dependent manner. This study demonstrates that PPARs have distinct roles in skeletal muscle cells with respect to the regulation of lipid, carbohydrate, and energy homeostasis. Moreover, we surmise that PPARgamma/delta agonists would increase fatty acid catabolism, cholesterol efflux, and energy expenditure in muscle, and speculate selective activators of PPARbeta/delta may have therapeutic utility in the treatment of hyperlipidemia, atherosclerosis, and obesity.

Self-optimization module for scheduling using case-based reasoning

Metaheuristics performance is highly dependent of the respective parameters which need to be tuned. Parameter tuning may allow a larger flexibility and robustness but requires a careful initialization. The process of defining which parameters setting should be used is not obvious. The values for parameters depend mainly on the problem, the instance to be solved, the search time available to spend in solving the problem, and the required quality of solution. This paper presents a learning module proposal for an autonomous parameterization of Metaheuristics, integrated on a Multi-Agent System for the resolution of Dynamic Scheduling problems. The proposed learning module is inspired on Autonomic Computing Self-Optimization concept, defining that systems must continuously and proactively improve their performance. For the learning implementation it is used Case-based Reasoning, which uses previous similar data to solve new cases. In the use of Case-based Reasoning it is assumed that similar cases have similar solutions. After a literature review on topics used, both AutoDynAgents system and Self-Optimization module are described. Finally, a computational study is presented where the proposed module is evaluated, obtained results are compared with previous ones, some conclusions are reached, and some future work is referred. It is expected that this proposal can be a great contribution for the self-parameterization of Metaheuristics and for the resolution of scheduling problems on dynamic environments.

Contextual intelligent load management with ANN adaptive learning module

With the current increase of energy resources prices and environmental concerns intelligent load management systems are gaining more and more importance. This paper concerns a SCADA House Intelligent Management (SHIM) system that includes an optimization module using deterministic and genetic algorithm approaches. SHIM undertakes contextual load management based on the characterization of each situation. SHIM considers available generation resources, load demand, supplier/market electricity price, and consumers’ constraints and preferences. The paper focus on the recently developed learning module which is based on artificial neural networks (ANN). The learning module allows the adjustment of users’ profiles along SHIM lifetime. A case study considering a system with fourteen discrete and four variable loads managed by a SHIM system during five consecutive similar weekends is presented.

Development and validation of the Portuguese version of the WHOQOL-OLD module

OBJECTIVE: The increasing proportion of older adults in the general population and the specific characteristics of this age group show the need for the development of specific instruments to measure quality of life in older adults. The study aimed at describing the development and validation of the Portuguese version of the World Health Organization Quality of Life for Older Persons (WHOQOL-OLD) module. METHODS: The WHOQOL-OLD instrument was administered in a sample of 424 older adults in the city of Porto Alegre, Southern Brazil, in 2005. The questionnaire comprises 24 items divided into six facets: sensory abilities; autonomy; past, present and future activities; social participation; death and dying; and intimacy. Besides the WHOQOL-OLD module, the WHOQOL-BREF, BDI and BHS instruments were also applied. The instrument's internal consistency was assessed using Cronbach's alpha coefficient. RESULTS: The instrument showed adequate internal consistency (Cronbach's coefficients ranging from 0.71 to 0.88), discriminant validity (p<0.01), concurrent validity (correlation coefficients ranging from -0.61 to -0.50) and test-retest reliability (correlation coefficients ranging from 0.58 to 0.82). Findings concerning criterion validity need further studies. CONCLUSIONS: The WHOQOL-Old module is a useful alternative with good psychometric performance in the investigation of quality of life in older adults.

Brazilian WHOQOL-OLD Module version: a Rasch analysis of a new instrument

OBJECTIVE: To evaluate the Brazilian version of WHOQOL-OLD Module and to test potential changes to the instrument to increase its psychometric adequacy. METHODS: A total of 424 older adults living in a city in Southern Brazil completed the WHOQOL-OLD instrument, in 2005. Rasch analysis was used to explore the psychometric performance of the scale, as implemented by the RUMM2020 software. Item-trait interaction, threshold disorders, presence of differential item functioning and item fit, were analyzed. RESULTS: Two ("death and dying" and "sensory abilities") out of six domains showed inadequate item-trait interactions. Rescoring the response scale and deleting the most misperforming items led to scale improvement. The evaluation of domains and items individually showed that the "intimacy" domain does perform well in contrast to the findings using the classical approach. In addition, the "sensory abilities" domain does not derive an interval measure in its current format. CONCLUSIONS: Unidimensionality and local independence were seen in all domains. Changes in the response scale and deletion of problematic items improved the scale's performance.

Novel potentiometric sensors of molecular imprinted polymers for specific binding of chlormequat

Molecularly imprinted polymers (MIP) were used as potentiometric sensors for the selective recognition and determination of chlormequat (CMQ). They were produced after radical polymerization of 4-vinyl pyridine (4-VP) or methacrylic acid (MAA) monomers in the presence of a cross-linker. CMQwas used as template. Similar nonimprinted (NI) polymers (NIP) were produced by removing the template from reaction media. The effect of kind and amount of MIP or NIP sensors on the potentiometric behavior was investigated. Main analytical features were evaluated in steady and flow modes of operation. The sensor MIP/4-VP exhibited the best performance, presenting fast near-Nernstian response for CMQover the concentration range 6.2×10-6 – 1.0×10-2 mol L-1 with detection limits of 4.1×10-6 mol L-1. The sensor was independent from the pH of test solutions in the range 5 – 10. Potentiometric selectivity coefficients of the proposed sensors were evaluated over several inorganic and organic cations. Results pointed out a good selectivity to CMQ. The sensor was applied to the potentiometric determination of CMQin commercial phytopharmaceuticals and spiked water samples. Recoveries ranged 96 to 108.5%.

Cholesterol 24S-Hydroxylase overexpression inhibits the liver X receptor (LXR) pathway by activating small guanosine triphosphate-binding proteins (sGTPases) in neuronal cells

The neuronal-specific cholesterol 24S-hydroxylase (CYP46A1) is important for brain cholesterol elimination. Cyp46a1 null mice exhibit severe deficiencies in learning and hippocampal long-term potentiation, suggested to be caused by a decrease in isoprenoid intermediates of the mevalonate pathway. Conversely, transgenic mice overexpressing CYP46A1 show an improved cognitive function. These results raised the question of whether CYP46A1 expression can modulate the activity of proteins that are crucial for neuronal function, namely of isoprenylated small guanosine triphosphate-binding proteins (sGTPases). Our results show that CYP46A1 overexpression in SH-SY5Y neuroblastoma cells and in primary cultures of rat cortical neurons leads to an increase in 3-hydroxy-3-methyl-glutaryl-CoA reductase activity and to an overall increase in membrane levels of RhoA, Rac1, Cdc42 and Rab8. This increase is accompanied by a specific increase in RhoA activation. Interestingly, treatment with lovastatin or a geranylgeranyltransferase-I inhibitor abolished the CYP46A1 effect. The CYP46A1-mediated increase in sGTPases membrane abundance was confirmed in vivo, in membrane fractions obtained from transgenic mice overexpressing this enzyme. Moreover, CYP46A1 overexpression leads to a decrease in the liver X receptor (LXR) transcriptional activity and in the mRNA levels of ATP-binding cassette transporter 1, sub-family A, member 1 and apolipoprotein E. This effect was abolished by inhibition of prenylation or by co-transfection of a RhoA dominant-negative mutant. Our results suggest a novel regulatory axis in neurons; under conditions of membrane cholesterol reduction by increased CYP46A1 expression, neurons increase isoprenoid synthesis and sGTPase prenylation. This leads to a reduction in LXR activity, and consequently to a decrease in the expression of LXR target genes.

Preservation of catechin antioxidant properties loaded in carbohydrate nanoparticles

Epigallocatechin gallate (EGCG), an antioxidant with several pharmacological and biological activities, was encapsulated in carbohydrate particles to preserve its antioxidant properties and improve its bioavailability. Gum arabic–maltodextrin particles loaded with EGCG (EGCG/P) were successfully produced by homogenization and spray-drying, with an EGCG loading efficiency of 96 ± 3%. Spray-dried particles are spherical or corrugated and polydisperse with diameters less than 20 m. The particles in aqueous suspension revealed two main populations, with mean average diameters of 40 nm and 400 nm. Attenuated total reflection-infrared spectroscopy (ATR-IR) confirmed that EGCG was incorporated in the carbohydrate matrix by intermolecular interactions, maintaining its chemical integrity. Atomic force microscopy imaging proved the particle spherical shape and size. The present study demonstrates that the carbohydrate matrix is able to preserve EGCG antioxidant properties, as proof of concept to be used as polymeric drug carrier.

Overexpression of tumour-associated carbohydrate antigen sialyl-Tn in advanced bladder tumours

Little is known on the expression of the tumour-associated carbohydrate antigen sialyl-Tn (STn), in bladder cancer. We report here that 75% of the high-grade bladder tumours, presenting elevated proliferation rates and high risk of recurrence/progression expressed STn. However, it was mainly found in non-proliferative areas of the tumour, namely in cells invading the basal and muscle layers. STn was also found in tumour-adjacent mucosa, which suggests its dependence on a field effect of the tumour. Furthermore, it was not expressed by the normal urothelium, demonstrating the cancer-specific nature of this antigen. STn expression correlated with that of sialyltransferase ST6GalNAc.I, its major biosynthetic enzyme. The stable expression of ST6GalNAc.I in the bladder cancer cell line MCR induced STn expression and a concomitant increase of cell motility and invasive capability. Altogether, these results indicate for the first time a link between STn expression and malignancy in bladder cancer. Hence, therapies targeting STn may constitute new treatment approaches for these tumours.

Análise psicométrica inicial da versão brasileira do DISABKIDS Atopic Dermatitis Module

OBJETIVO: Analisar as propriedades psicométricas iniciais da versão brasileira de instrumento de avaliação da qualidade de vida relacionada à saúde de crianças e adolescentes com dermatite atópica. MÉTODOS: Estudo transversal realizado com amostra de 52 crianças e adolescentes, com idades entre oito e 18 anos, diagnosticados com dermatite atópica, e seus responsáveis, recrutados em serviço de dermatologia de hospital universitário na cidade de São Paulo, SP, em 2009. Foram avaliadas a validade de construto, a confiabilidade de consistência interna e a correlação entre as respostas de crianças e adolescentes e seus responsáveis da versão brasileira do Atopic Dermatitis Module (DISABKIDS-ADM). RESULTADOS: A confiabilidade de consistência interna foi satisfatória, com coeficiente alfa de Cronbach aceitável para as dimensões constantes no instrumento (0,7024/0,8124 e 0,7239/0,8604). A análise multitraço-multimétodo para validade convergente mostrou valores maiores que 0,30 para todos os itens. Quanto à validade discriminante, a análise revelou resultados satisfatórios. A concordância entre as versões self e proxy foi avaliada pelo coeficiente de correlação intra-classe, com valores de 0,8173 para impacto e 0,7629 para estigma. CONCLUSÕES: Diante dos resultados encontrados, considera-se que o instrumento DISABKIDS-ADM pode ser utilizado por pesquisadores brasileiros depois de finalizado seu processo de validação, por seus resultados iniciais apontarem propriedades psicométricas satisfatórias, que permitem considerá-lo um instrumento válido e confiável.

Ligand-Induced structural changes in TEM‑1 probed by molecular dynamics and relative binding free energy calculations

The TEM family of enzymes has had a crucial impact on the pharmaceutical industry due to their important role in antibiotic resistance. Even with the latest technologies in structural biology and genomics, no 3D structure of a TEM- 1/antibiotic complex is known previous to acylation. Therefore, the comprehension of their capability in acylate antibiotics is based on the protein macromolecular structure uncomplexed. In this work, molecular docking, molecular dynamic simulations, and relative free energy calculations were applied in order to get a comprehensive and thorough analysis of TEM-1/ampicillin and TEM-1/amoxicillin complexes. We described the complexes and analyzed the effect of ligand binding on the overall structure. We clearly demonstrate that the key residues involved in the stability of the ligand (hot-spots) vary with the nature of the ligand. Structural effects such as (i) the distances between interfacial residues (Ser70−Oγ and Lys73−Nζ, Lys73−Nζ and Ser130−Oγ, and Ser70−Oγ−Ser130−Oγ), (ii) side chain rotamer variation (Tyr105 and Glu240), and (iii) the presence of conserved waters can be also influenced by ligand binding. This study supports the hypothesis that TEM-1 suffers structural modifications upon ligand binding.

Embedding instruments & modules into an IEEE1451-FPGA-Based Weblab infrastructure

Adopting standard-based weblab infrastructures can be an added value for spreading their influence and acceptance in education. This paper suggests a solution based on the IEEE1451.0 Std. and FPGA technology for creating reconfigurable weblab infrastructures using Instruments and Modules (I&Ms) described through standard Hardware Description Language (HDL) files. It describes a methodology for creating and binding I&Ms into an IEEE1451-module embedded in a FPGA-based board able to be remotely controlled/accessed using IEEE1451-HTTP commands. At the end, an example of a step-motor controller module bond to that IEEE1451-module is described.