Sarcosine oxidase composite screen-printed electrode for sarcosine determination in biological samples

XIX Meeting of the Portuguese Electrochemical Society - XVI Iberic Meeting of ElectrochemistryProstate Cancer (PCa) is the most common form of cancer in men in Europe with a 61.4 % incidence among all cancer cases and a 12.1 % mortality [1] and, therefore, its early detection is fundamental for increasing the survival rate. Currently, diagnosis and management of patients with PCa is only based on the determination of the biomarker Prostate Specific Antigen (PSA). However, the method used for PCa detection has poor sensitivity and specificity, leading to false negative and false positive test results and many patients are sent to unnecessary biopsy procedures [2]. Therefore, there is a need to seek for new biomarkers and more effective screening. In this work, a biosensor device was developed for the quantification of sarcosine via electrochemical detection of H2O2 (at 0.6 V) generated from the catalyzed oxidation of sarcosine. The detection was carried out after the modification of carbon screen printed electrodes (SPEs) by immobilization of sarcosine oxidase (SOX) on the electrode surface. The strategies used herein included the activation of the carbon films by an electrochemical step and the formation of an NHS/EDAC layer to bond the enzyme to the electrode, the use of metallic or semiconductor nanoparticles layer previously or during the enzyme immobilization. In order to improve the sensor stability and selectivity a polymeric layer with extra enzyme content was further added. The proposed methodology for the detection of sarcosine allowed obtaining a limit of detection (LOD) of 1.6x10-5 mM, using a linear concentration range between 1x10-5 and 1x10-4 mM. The biosensor was successfully applied to the analysis of sarcosine in urine samples.

Geographic variation in cork oak and its implications for expected impacts of climate change

Cork oak (Quercus suber L.) is a protected tree species in Portugal, being also the source of raw material for the cork industry, a major player in Portuguese economy (representing 3% of GDP in 2010). The future climatic scenarios for Portugal point to an increase in average summer temperatures from 0.3 to 0.7ºC between 2016 and 2035, and up to 4.6ºC until 2100. In addition, precipitation estimates suggest a reduction of annual rainfall from 20 to 40%, especially in southern Portugal. Water stress will, therefore, be a leading constraint to primary production. The combined effects of drought and high temperatures will lead to decreases in carbon assimilation and increases in tree mortality, and consequently current reforestation efforts will need to account for these expected adverse outcomes through the sustainable use of suitable genetic material. There are several reasons that can be highlighted to emphasize the need for an efficient management of cork oak genetic resources in Portugal, namely: i) to avoid cork import, and thus to increase cork production to meet the industry demands; ii) to overcome a generally poor area of natural regeneration, which does not help to ensure an in situ conservation of genetic resources; iii) to deploy adapted genetic material for afforestation/reforestation; and iv) to develop a gene resources conservation program, as cork oak is a vital component of agro-silvopastoral systems in the Mediterranean region. Given the broad native range of the species, involving significant environmental and geographic gradients, a high level of genetic variation can be expected. It is possible that disruptive selection has caused a large differentiation in adaptive traits among populations, namely in the ability to tolerate different environmental stress events (e.g. drought and frost) and to cope with pests and diseases. Between 1998 and 2011, we have collected data involving survival, growth, phenology and water-use efficiency traits from five common-garden provenance trials (including family structure in two of the trials), that were established in Portugal under a concerted action launched by the EUFORGEN’s network. These multi-site field experiments are based on up to 35 tested provenances covering the entire natural distribution of cork oak, and results obtained from the genetic evaluation of the trials have indicated significant differences among populations for all the measured traits at all observed ages. Four of the tested provenances (Alpujarras – Haza de Lino, Puglia – Lucci - S. Teresa, Landes - Soustons, Rif Occidental – Ain Rami) were then chosen according to their contrasting field performance for growth, phenology and water-use efficiency (WUE), and were further evaluated under controlled-environment conditions where drought stress was induced. In this context, the main drivers of drought adaptation appeared to be early stomatal closure and root investment, which also showed significant differences among the selected provenances. The responses to drought over time also varied among these studied populations, and seemed to be related to their differences in growth rhythm. The Ain Rami population seemed to be most prone population to endure drought conditions. Facing a water deficit scenario this population, with highest growth, showed a higher investment on roots compared to the Haza de Lino population, that even under optimal hydration status, had lower biomass values, more reduced transpiration area (smallest size, with lowest Specific Leaf Area), leading to a lower water consumption. This population showed a delay in onset of stress when compared to other populations, only revealed no stomatal limitations with high stress levels. Furthermore, Ain Rami showed higher WUE under drought conditions both in the field trials and under controlled conditions, but average WUE in wet conditions. The results from the field and controlled-environment experiments were consistent in that geographic origin had an important influence on the performance of fitness surrogates and functional traits, and thus providing a strong indication that seed origin must be considered in cork oak reforestation programs.info:eu-repo/semantics/publishedVersio

It Takes Two to Tango, Part II : Synthesis of A-Ring Functionalised Quinones Containing Two Redox-Active Centres with Antitumour Activities

In 2021, our research group published the prominent anticancer activity achieved through the successful combination of two redox centres (ortho-quinone/para-quinone or quinone/seleniumcontaining triazole) through a copper-catalyzed azide-alkyne cycloaddition (CuAAC) reaction. The combination of two naphthoquinoidal substrates towards a synergetic product was indicated, but not fully explored. Herein, we report the synthesis of 15 new quinone-based derivatives prepared from click chemistry reactions and their subsequent evaluation against nine cancer cell lines and the murine fibroblast line L929. Our strategy was based on the modification of the A-ring of paranaphthoquinones and subsequent conjugation with different ortho-quinoidal moieties. As anticipated, our study identified several compounds with IC50 values below 0.5 µM in tumour cell lines. Some of the compounds described here also exhibited an excellent selectivity index and low cytotoxicity on L929, the control cell line. The antitumour evaluation of the compounds separately and in their conjugated form proved that the activity is strongly enhanced in the derivatives containing two redox centres. Thus, our study confirms the efficiency of using A-ring functionalized para-quinones coupled with ortho-quinones to obtain a diverse range of two redox centre compounds with potential applications against cancer cell lines. Here as well, it literally takes two for an efficient tango

Ranked Adjusted Rand: integrating distance and partition information in a measure of clustering agreement

BACKGROUND: Biological information is commonly used to cluster or classify entities of interest such as genes, conditions, species or samples. However, different sources of data can be used to classify the same set of entities and methods allowing the comparison of the performance of two data sources or the determination of how well a given classification agrees with another are frequently needed, especially in the absence of a universally accepted "gold standard" classification. RESULTS: Here, we describe a novel measure – the Ranked Adjusted Rand (RAR) index. RAR differs from existing methods by evaluating the extent of agreement between any two groupings, taking into account the intercluster distances. This characteristic is relevant to evaluate cases of pairs of entities grouped in the same cluster by one method and separated by another. The latter method may assign them to close neighbour clusters or, on the contrary, to clusters that are far apart from each other. RAR is applicable even when intercluster distance information is absent for both or one of the groupings. In the first case, RAR is equal to its predecessor, Adjusted Rand (HA) index. Artificially designed clusterings were used to demonstrate situations in which only RAR was able to detect differences in the grouping patterns. A study with larger simulated clusterings ensured that in realistic conditions, RAR is effectively integrating distance and partition information. The new method was applied to biological examples to compare 1) two microbial typing methods, 2) two gene regulatory network distances and 3) microarray gene expression data with pathway information. In the first application, one of the methods does not provide intercluster distances while the other originated a hierarchical clustering. RAR proved to be more sensitive than HA in the choice of a threshold for defining clusters in the hierarchical method that maximizes agreement between the results of both methods. CONCLUSION: RAR has its major advantage in combining cluster distance and partition information, while the previously available methods used only the latter. RAR should be used in the research problems were HA was previously used, because in the absence of inter cluster distance effects it is an equally effective measure, and in the presence of distance effects it is a more complete one

Comparative effectiveness and predictors of response to tumour necrosis factor inhibitor therapies in rheumatoid arthritis

Funding Information: positions on two Pfizer sponsored trials and has directed an educational course supported by Bristol Myers Squibb. He serves as an epidemiology consultant to CORRONA. J.A.P.S. has received honoraria as a speaker or consultant and benefited from research support from several pharmaceutical companies involved in the production of biologic agents (Abbott, Amgen, MSD, Pfizer and Roche), always at sums less than E10 000. All other authors have declared no conflicts of interest. Funding Information: Funding: This work was supported by a grant from Harvard-Portugal Program HMSP-ICS/SAU-ICT/0002/ 2010.Objectives: Adalimumab, etanercept and infliximab are effective TNF inhibitors (TNFis) in the treatment of RA, but no randomized clinical trials have compared the three agents. Prior observational data are not consistent. We compared their effectiveness over 1 year in a prospective cohort.Methods: Analyses were performed on subjects' first episode of TNFi use in the Rheumatic Diseases Portuguese Register, Reuma.pt. The primary outcome was the proportion of patients with European League Against Rheumatism good response sustained at two consecutive observations separated by 3 months during the first year of TNFi use. Comparisons were performed using conventional adjusted logistic regression, as well as matching subjects across the three agents using a propensity score. In addition, baseline predictors of treatment response to TNFi were identified.Results: The study cohort included 617 RA patients, 250 starting etanercept, 206 infliximab and 161 adalimumab. Good response was achieved by 59.6% for adalimumab, 59.2% for etanercept and 51.9% for infliximab (P = 0.21). The modelled probability of good response did not significantly differ across agents (etanercept vs adalimumab OR = 0.97, 95% CI 0.55, 1.71; etanercept vs infliximab OR = 1.25, 95% CI 0.74, 2.12; infliximab vs adalimumab OR = 0.80, 95% CI 0.47, 1.36). Matched propensity score analyses also showed no significant treatment response differences. Greater educational attainment was a predictor of better response, while smoking, presence of ACPA, glucocorticoid use and worse physician assessment of disease activity at baseline each predicted a reduced likelihood of treatment response.Conclusion: Over 1 year, we found no difference in effectiveness between adalimumab, etanercept and infliximab.publishersversionpublishe

Nanostructured 3D Constructs Based on Chitosan and Chondroitin Sulphate Multilayers for Cartilage Tissue Engineering

Nanostructured three-dimensional constructs combining layer-by-layer technology (LbL) and template leaching were processed and evaluated as possible support structures for cartilage tissue engineering. Multilayered constructs were formed by depositing the polyelectrolytes chitosan (CHT) and chondroitin sulphate (CS) on either bidimensional glass surfaces or 3D packet of paraffin spheres. 2D CHT/CS multi-layered constructs proved to support the attachment and proliferation of bovine chondrocytes (BCH). The technology was transposed to 3D level and CHT/CS multi-layered hierarchical scaffolds were retrieved after paraffin leaching. The obtained nanostructured 3D constructs had a high porosity and water uptake capacity of about 300%. Dynamical mechanical analysis (DMA) showed the viscoelastic nature of the scaffolds. Cellular tests were performed with the culture of BCH and multipotent bone marrow derived stromal cells (hMSCs) up to 21 days in chondrogenic differentiation media. Together with scanning electronic microscopy analysis, viability tests and DNA quantification, our results clearly showed that cells attached, proliferated and were metabolically active over the entire scaffold. Cartilaginous extracellular matrix (ECM) formation was further assessed and results showed that GAG secretion occurred indicating the maintenance of the chondrogenic phenotype and the chondrogenic differentiation of hMSCs

TRAF1/C5 but Not PTPRC Variants Are Potential Predictors of Rheumatoid Arthritis Response to Anti-Tumor Necrosis Factor Therapy

Background. The aim of our work was to replicate, in a Southern European population, the association reported in Northern populations between PTPRC locus and response to anti-tumor necrosis factor (anti-TNF) treatment in rheumatoid arthritis (RA). We also looked at associations between five RA risk alleles and treatment response. Methods. We evaluated associations between anti-TNF treatment responses assessed by DAS28 change and by EULAR response at six months in 383 Portuguese patients. Univariate and multivariate linear and logistic regression analyses were performed. In a second step to confirm our findings, we pooled our population with 265 Spanish patients. Results. No association was found between PTPRC rs10919563 allele and anti-TNF treatment response, neither in Portuguese modeling for several clinical variables nor in the overall population combining Portuguese and Spanish patients. The minor allele for RA susceptibility, rs3761847 SNP in TRAF1/C5 region, was associated with a poor response in linear and logistic univariate and multivariate regression analyses. No association was observed with the other allellic variants. Results were confirmed in the pooled analysis. Conclusion. This study did not replicate the association between PTPRC and the response to anti-TNF treatment in our Southern European population. We found that TRAF1/C5 risk RA variants potentially influence anti-TNF treatment response.This work was supported by a grant from Harvard-Portugal Program HMSP-ICS/SAU-ICT/0002/2010. Daniel H. Solomon received support for this work from the NIH (K24-AR-055989). Elizabeth W. Karlson received support for this work from NIH (K24-AR-AR0524). Reuma.pt received unrestricted grants from Abbott, Bristol-Myers Squibb, Merck Sharp and Dohme, Pfizer, Roche, and UCB Pharma

Synthesis and antiproliferative activity of novel limonene derivatives with a substituted thiourea moiety

A series of R-(+)-limonene derivatives bearing a substituted thiourea moiety (3-13) and five S-methyl analogs (14-18) were synthesized and evaluated for their in vitro antiproliferative activity against human cancer cell lines. Compounds bearing aromatic substituents (3-6) exhibit cytotastic activity in the full panel of cell lines tested, with GI50 values in the range of 2.5 to 24 µmol L-1. Compounds 3, 10, 12 and 16 were the most active with GI50 values in the range of 0.41 to 3.0 µmol L-1, against different cell lines.No presente trabalho descrevemos a síntese e a avaliação da atividade antiproliferativa, frente a linhagens de células tumorais humanas, de derivados do R-(+)-limoneno (3-18) contendo uma unidade tiouréia substituída. Os derivados com substituintes arílicos (3-6) exibiram atividade citostática frente a todas linhagens testadas, com inibição de 50% do crescimento celular (GI50) em concentrações na faixa de 2,5 a 24 µmol L-1. Os compostos 3, 10, 12 e 16 foram os mais ativos, com GI50 na faixa de 0,41 a 3,0 mmol L-1, frente a diferentes linhagens celulares.954960Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES

Correction: Serologically Defined Variations in Malaria Endemicity in Pará State, Brazil.

[This corrects the article DOI: 10.1371/journal.pone.0113357.]