Identification of Novel Interaction between ADAM17 (a Disintegrin and Metalloprotease 17) and Thioredoxin-1

ADAM17, which is also known as TNF alpha-converting enzyme, is the major sheddase for the EGF receptor ligands and is considered to be one of the main proteases responsible for the ectodomain shedding of surface proteins. How a membrane-anchored proteinase with an extracellular catalytic domain can be activated by inside-out regulation is not completely understood. We characterized thioredoxin-1 (Trx-1) as a partner of the ADAM17 cytoplasmic domain that could be involved in the regulation of ADAM17 activity. We induced the overexpression of the ADAM17 cytoplasmic domain in HEK293 cells, and ligands able to bind this domain were identified by MS after protein immunoprecipitation. Trx-1 was also validated as a ligand of the ADAM17 cytoplasmic domain and full-length ADAM17 recombinant proteins by immunoblotting, immunolocalization, and solid phase binding assay. In addition, using nuclear magnetic resonance, it was shown in vitro that the titration of the ADAM17 cytoplasmic domain promotes changes in the conformation of Trx-1. The MS analysis of the cross-linked complexes showed cross-linking between the two proteins by lysine residues. To further evaluate the functional role of Trx-1, we used a heparin-binding EGF shedding cell model and observed that the overexpression of Trx-1 in HEK293 cells could decrease the activity of ADAM17, activated by either phorbol 12-myristate 13-acetate or EGF. This study identifies Trx-1 as a novel interaction partner of the ADAM17 cytoplasmic domain and suggests that Trx-1 is a potential candidate that could be involved in ADAM17 activity regulation.

The EXTRIP (EXtracorporeal TReatments In Poisoning) workgroup: Guideline methodology

Extracorporeal treatments (ECTRs), such as hemodialysis and hemoperfusion, are used in poisoning despite a lack of controlled human trials demonstrating efficacy. To provide uniform recommendations, the EXTRIP group was formed as an international collaboration among recognized experts from nephrology, clinical toxicology, critical care, or pharmacology and supported by over 30 professional societies. For every poison, the clinical benefit of ECTR is weighed against associated complications, alternative therapies, and costs. Rigorous methodology, using the AGREE instrument, was developed and ratified. Methods rely on evidence appraisal and, in the absence of robust studies, on a thorough and transparent process of consensus statements. Twenty-four poisons were chosen according to their frequency, available evidence, and relevance. A systematic literature search was performed in order to retrieve all original publications regardless of language. Data were extracted on a standardized instrument. Quality of the evidence was assessed by GRADE as: High = A, Moderate = B, Low = C, Very Low = D. For every poison, dialyzability was assessed and clinical effect of ECTR summarized. All pertinent documents were submitted to the workgroup with a list of statements for vote (general statement, indications, timing, ECTR choice). A modified Delphi method with two voting rounds was used, between which deliberation was required. Each statement was voted on a Likert scale (1-9) to establish the strength of recommendation. This approach will permit the production of the first important practice guidelines on this topic.

Crotamine Pharmacology Revisited: Novel Insights Based on the Inhibition of K-V Channels

Crotamine, a 5-kDa peptide, possesses a unique biological versatility. Not only has its cell-penetrating activity become of clinical interest but, moreover, its potential selective antitumor activity is of great pharmacological importance. In the past, several studies have attempted to elucidate the exact molecular target responsible for the crotamine-induced skeletal muscle spasm. The aim of this study was to investigate whether crotamine affects voltage-gated potassium (K-V) channels in an effort to explain its in vivo effects. Crotamine was studied on ion channel function using the two-electrode voltage clamp technique on 16 cloned ion channels (12 K-V channels and 4 Na-V channels), expressed in Xenopus laevis oocytes. Crotamine selectively inhibits K-V 1.1, K-V 1.2, and K-V 1.3 channels with an IC50 of similar to 300 nM, and the key amino acids responsible for this molecular interaction are suggested. Our results demonstrate for the first time that the symptoms, which are observed in the typical crotamine syndrome, may result from the inhibition of K-V channels. The ability of crotamine to inhibit the potassium current through K-V channels unravels it as the first snake peptide with the unique multifunctionality of cell-penetrating and antitumoral activity combined with K-V channel-inhibiting properties. This new property of crotamine might explain some experimental observations and opens new perspectives on pharmacological uses.

Estimating total body fat using a skinfold prediction equation in Brazilian children

Background: The double burden of obesity and underweight is increasing in developing countries and simple methods for the assessment of fat mass in children are needed. Aim: To develop and validate a new anthropometric predication equation for assessment of fat mass in children. Subjects and methods: Body composition was assessed in 145 children aged 9.8 +/- 1.3 (SD) years from Sao Paulo, Brazil using dual energy X-ray absorptiometry (DEXA) and skinfold measurements. The study sample was divided into development and validation sub-sets to develop a new prediction equation for FM (PE). Results: Using multiple linear regression analyses, the best equation for predicting FM (R-2 - 0.77) included body weight, triceps skinfold, height, gender and age as independent variables. When cross-validated, the new PE was valid in this sample (R-2 = 0.80), while previously published equations were not. Conclusion: The PE was more valid for Brazilian children that existing equations, but further studies are needed to assess the validity of this PE in other populations.

Permanent Genetic Resources added to Molecular Ecology Resources Database 1 February 2012-31 March 2012

This article documents the addition of 171 microsatellite marker loci and 27 pairs of single nucleotide polymorphism (SNP) sequencing primers to the Molecular Ecology Resources Database. Loci were developed for the following species: Bombus pauloensis, Cephalorhynchus heavisidii, Cercospora sojina, Harpyhaliaetus coronatus, Hordeum vulgare, Lachnolaimus maximus, Oceanodroma monteiroi, Puccinia striiformis f. sp. tritici, Rhea americana, Salmo salar, Salmo trutta, Schistocephalus solidus, Sousa plumbea and Tursiops aduncus. These loci were cross-tested on the following species: Aquila heliaca, Bulweria bulwerii, Buteo buteo, Buteo swainsoni, Falco rusticolus, Haliaeetus albicilla, Halobaena caerulea, Hieraaetus fasciatus, Oceanodroma castro, Puccinia graminis f. sp. Tritici, Puccinia triticina, Rhea pennata and Schistocephalus pungitii. This article also documents the addition of 27 sequencing primer pairs for Puffinus baroli and Bulweria bulwerii and cross-testing of these loci in Oceanodroma castro, Pelagodroma marina, Pelecanoides georgicus, Pelecanoides urinatrix, Thalassarche chrysostoma and Thalassarche melanophrys.

Fatores prognósticos associados ao tratamento cirúrgico da mielorradiculopatia espondilótica cervical

OBJETIVO: Identificar os fatores clínicos dos indivíduos, fatores sociais, ambientais e dos exames de imagem que se correlacionam ao resultado final de melhora neurológica em pacientes submetidos ao tratamento cirúrgico da mielopatia espondilótica cervical. MÉTODOS: A avaliação clínica foi quantificada pela escala deficitária da JOA. Analisamos 200 casos de mielorradiculopatia cervical, operados no HC-FMUSP, no período de janeiro de 1993 a janeiro de 2007. A média de segmento foi de 06 anos e 08 meses. A análise radiológica foi baseada nos critérios de instabilidade de White e scala de Kellgren. RESULTADOS: Em 80% houve melhora, 14% estabilização e em 6% piora do quadro neurológico. A piora neurológica não foi associada com nenhum fator clínico, ambiental ou de imagem. A melhora neurológica foi diretamente proporcional a menor idade na cirurgia, ausência de co-morbidade, sinal de Hoffman, atrofia muscular, hipersinal medular na RNM, menor período de evolução pré-operatório, melhor status neurológico pré-operatório e inversamente proporcional ao diâmetro AP do canal medular e multiplicidade de compressões. Identificou-se associação com o tabagismo. Mais de 70 anos, evolução superior a 24 meses, atrofia muscular, pontuação JOA igual ou inferior a sete pontos e diâmetro AP do canal inferior ou igual a seis mm não foram associado à melhora.