Characterization of the Channel Constriction Allowing the Access of the Substrate to the Active Site of Yeast Oxidosqualene Cyclase

In oxidosqualene cyclases (OSCs), an enzyme which has been extensively studied as a target for hypocholesterolemic or antifungal drugs, a lipophilic channel connects the surface of the protein with the active site cavity. Active site and channel are separated by a narrow constriction operating as a mobile gate for the substrate passage. In Saccharomyces cerevisiae OSC, two aminoacidic residues of the channel/constriction apparatus, Ala525 and Glu526, were previously showed as critical for maintaining the enzyme functionality. In this work sixteen novel mutants, each bearing a substitution at or around the channel constrictions, were tested for their enzymatic activity. Modelling studies showed that the most functionality-lowering substitutions deeply alter the H-bond network involving the channel/constriction apparatus. A rotation of Tyr239 is proposed as part of the mechanism permitting the access of the substrate to the active site. The inhibition of OSC by squalene was used as a tool for understanding whether the residues under study are involved in a pre-catalytic selection and docking of the substrate oxidosqualene

Assessment of in‐situ gelling microemulsion systems upon temperature and dilution condition for corneal delivery of bevacizumab

Bevacizumab (BVZ), a recombinant humanized monoclonal antibody, has recently been proposed as a topical application in the treatment of anterior segment neovascularization; however, as there are some disadvantages in the administration of common eye-drops, ophthalmic topical drug delivery systems are under study to improve the precorneal residence time, reducing the frequency of administration. In this work, oil-in-water and water-in-oil BVZ-loaded microemulsions are developed, able to increase their viscosity, either by the formation of a liquid-crystalline structure upon aqueous dilution, thanks to the presence of Epikuron® 200 and polysorbate 80, or by body-temperature-induced jellification for the presence of Pluronic® F127 aqueous solution as an external phase. In oil-in-water microemulsion, hydrophobic ion pairs of BVZ were also prepared, and their incorporation was determined by release studies. Microemulsions were characterized for rheological behavior, corneal opacity, in vitro corneal permeation, and adhesion properties. The studied microemulsions were able to incorporate BVZ (from 1.25 to 1.6 mg/mL), which maintained dose-dependent activity on retinal pigment epithelial ARPE-19 cell lines. BVZ loaded in microemulsions permeated the excised cornea easier (0.76–1.56% BVZ diffused, 4–20% BVZ accumulated) than BVZ commercial solution (0.4% BVZ diffused, 5% accumulated) and only a mild irritation effect on the excised cornea was observed. The good adhesion properties as well the increased viscosity after application, under conditions that mimic the corneal environment (from 1 × 103 to more than 100 × 103 mPa·s), might prolong precorneal residence time, proving these systems could be excellent topical BVZ release systems

Ferramentas computacionais de apoio às boas práticas de manejo de tilápia.

A piscicultura brasileira cresce cerca de 30% ao ano, por tratar-se de fonte de proteína de alta qualidade e pela lucratividade. De origem africana, a tilapicultura está presente hoje em grande parte do território nacional, mas ainda sofrendo as pressões para a maior qualidade do produto e adoção de práticas menos impactantes ao ambiente. Entre alguns desses impactos citam-se as poluição e interferências em níveis de biodiversidade e de qualidade da água. Assim, o produtor carece de informação e ferramentas que favoreçam a rápida transferência de práticas já identificadas pela pesquisa, que aumentem seu conhecimento e agilidade na incorporação destas às atividades da propriedade. O projeto "Manejo e Gestão Ambiental da Aqüicultura" da Embrapa Meio Ambiente, componente do projeto em rede Aquabrasil, vem identificando Boas Práticas de Manejo (BPM) de tilápia passíveis de uso imediato pelo produtor a partir de sua rápida observação da propriedade. Nele, o sistema informatizado Aquisys está em desenvolvimento para vialibizar o acesso dinâmico, via web, às avaliações e indicadores de produção, manejo e qualidade de água em atenção às BPM. Este trabalho apresenta duas de suas ferramentas computacionais desenvolvidas para tilápia: a) um sistema especialista de rápida avaliação de BPM na propriedade (em linguagem CLIPS); e b) um programa para determinar a %ração e %proteína bruta a ser oferecida, considerando os pesos dos peixes e temperatura local (em linguagens PHP e HTML)

Phenolic profile and biological activity of table grapes (Vitis vinifera L.)

Table grapes are largely produced and consumed in the Mediterranean area. Furthermore, in the last years, the commercial interest in table grapes and other non-alcoholic grape products has notably increased worldwide. In addition to the nutritional aspects, polyphenol compounds in fresh grapes could exert positive effects on human health. The aim of this study was the characterization of the phenolic pattern of sixteen grape varieties and the evaluation of the associated antioxidant capacity and anti-inflammatory activity at gastric level. The methods used were: 1) Folin-Cocalteau\u2019s assay for the quantification of total polyphenol content; 2) High-Performance Liquid Chromatography (HPLC) coupled with Diode-Array Detector (DAD) to perform the quantitative analysis of grapes phenolic fraction 3) DPPH (1,1,-diphenil-2-picrylhydrazyl) spectrophotometric assay for the assessment of radical scavenging activity; 4) assessment of IL-8 release from human gastric epithelial cells to evaluate the anti-inflammatory activity of grape extracts. Some grapes, including seedless varieties, showed that the phenolic pattern was highly correlated with the biological activities, and, in particular, with peel and seeds portion. These results suggest that selected grape varieties could represent, also for consumers who do not drink wine, a source of healthy compounds potentially able to counteract oxidative stress and gastric inflammation

Human Cryptic Host Defence Peptide GVF27 Exhibits Anti-Infective Properties against Biofilm Forming Members of the Burkholderia cepacia Complex

Therapeutic solutions to counter Burkholderia cepacia complex (Bcc) bacteria are challenging due to their intrinsically high level of antibiotic resistance. Bcc organisms display a variety of potential virulence factors, have a distinct lipopolysaccharide naturally implicated in antimicrobial resistance. and are able to form biofilms, which may further protect them from both host defence peptides (HDPs) and antibiotics. Here, we report the promising anti-biofilm and immunomodulatory activities of human HDP GVF27 on two of the most clinically relevant Bcc members, Burkholderia multivorans and Burkholderia cenocepacia. The effects of synthetic and labelled GVF27 were tested on B. cenocepacia and B. multivorans biofilms, at three different stages of formation, by confocal laser scanning microscopy (CLSM). Assays on bacterial cultures and on human monocytes challenged with B. cenocepacia LPS were also performed. GVF27 exerts, at different stages of formation, anti-biofilm effects towards both Bcc strains, a significant propensity to function in combination with ciprofloxacin, a relevant affinity for LPSs isolated from B. cenocepacia as well as a good propensity to mitigate the release of pro-inflammatory cytokines in human cells pre-treated with the same endotoxin. Overall, all these findings contribute to the elucidation of the main features that a good therapeutic agent directed against these extremely leathery biofilm-forming bacteria should possess

Relationship Between Vancomycin Trough Concentrations and Nephrotoxicity: A Prospective Multicenter Trial

Several single-center studies have suggested that higher doses of vancomycin, aimed at producing trough concentrations of \u3e15 mg/liter, are associated with increased risk of nephrotoxicity. We prospectively assessed the relative incidence of nephrotoxicity in relation to trough concentration in patients with documented methicillin-resistant Staphylococcus aureus (MRSA) infections at seven hospitals throughout South Carolina. Adult patients receiving vancomycin for at least 72 h with at least one vancomycin trough concentration determined under steady-state conditions were prospectively studied. The relationship between vancomycin trough concentrations of \u3e15 mg/ml and the occurrence of nephrotoxicity was assessed using univariate and multivariate analyses, controlling for age, gender, race, dose, length of therapy, use of other nephrotoxins (including contrast media), intensive care unit (ICU) residence, episodes of hypotension, and comorbidities. Nephrotoxicity was defined as an increase in serum creatinine of 0.5 mg/dl or a ≥ 50% increase from the baseline for two consecutive measurements. MICs of vancomycin for the MRSA isolates were also determined. A total of 288 patients were studied between February 2008 and June 2010, with approximately one-half having initial trough concentrations of ≥ 15 mg/ml. Nephrotoxicity was observed for 42 patients (29.6%) with trough concentrations \u3e15 mg/ml and for 13 (8.9%) with trough concentrations of ≤ 15 mg/ml. Multivariate analysis revealed vancomycin trough concentrations of \u3e15 mg/ml and race (black) as risk factors for nephrotoxicity in this population. Vancomycin trough concentrations of \u3e15 mg/ml appear to be associated with a 3-fold increased risk of nephrotoxicity

The use of chest ultrasonography in suspected cases of COVID-19 in the emergency department

Aim: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus-specific reverse transcriptase-polymerase chain reaction (RT-PCR) represents the diagnostic gold standard. We explored the value of chest ultrasonography to predict positivity to SARS-CoV-2 on RT-PCR in suspected COVID-19 cases. Patients & methods: Consecutive patients with suspect COVID-19 were included if they had fever and/or history of cough and/or dyspnea. Lung ultrasound score (LUSS) was computed according to published methods. Results: A total of 76 patients were included. A 3-variable model based on aspartate transaminase (AST) > upper limit of normal, LUSS >12 and body temperature >37.5°C yielded an overall accuracy of 91%. Conclusion: A simple LUSS-based model may represent a powerful tool for initial assessment in suspected cases of COVID-19. The gold standard for diagnosis of COVID-19 is RT-PCR. During a pandemic emergency, it may be useful to identify suspect symptomatic patients who may safely be observed without undergoing testing for COVID-19. In this work, a simple model based on the findings of lung ultrasound, AST levels and fever showed an overall accuracy of 91% to predict the results of RT-PCR

Relationship Between Biogenic Amines and Free Amino Acid Contents of Winesand Musts from Alentejo (Portugal)

The concentration of biogenic amines and free amino acids was studied in 102 Portuguese wines and 18 musts from Alentejo demarcated (D.O.C.) regions. Most wines were commercial, except for 38 monovarietals obtained by micro vinification. Musts from the varieties used to produce the latter wines were also studied. Both biogenic amines and free amino acids were analyzed by HPLC using fluorescence detection for their o-phthalaldehyde/fluorenylmethyl chloroformate (OPA/FMOC) derivatives. The most significant amines (average 10.8 mg/L for histamine+tyramine in red, and 7.4 mg/L for white wines) were found to be present at low levels and, although no important relationship between each individual biogenic amine could be obtained, the total amine content depends significantly on the assimilable amino acid content in wine