A facile blow spinning technique for green cellulose acetate/polystyrene composite separator for flexible energy storage devices - Amjid Rafique; Inês Sequeira; Ana Sofia Bento; Mariana Moniz; João Carmo; Eduardo Oliveira; Oliveira, J.P.; Marques, Ana; Ferreira, Isabel; Baptista, Ana (XXI Congresso da Sociedade Portuguesa de Materiais and XII International Symposium on Materials, 2023)

Green cellulose-based polymer electrolyte suitable for e-Textiles - João Carmo; Mariana Moniz; Amji Rafique; Ferreira, Isabel; Baptista, Ana (XXI Congresso da Sociedade Portuguesa de Materiais and XII International Symposium on Materials, 2023)

PEDOT:PSS Electrospray Functionalization of Carbon Yarns for Integration in Flexible Fibre-Shaped Supercapacitors - Amjid Rafique; João Carmo; Marques, Ana; Ferreira, Isabel; Baptista, Ana (XXI Congresso da Sociedade Portuguesa de Materiais and XII International Symposium on Materials, 2023)

Arsenite oxidase in complex with antimonite and arsenite oxyanions: Insights into the catalytic mechanism - Engrola, Filipa; Correia, Márcia A. S.; Cameron Watson; Romão, Carlos C.; Veiros, Luis F.; Romão, Maria João; Santos-Silva, Teresa; Joanne M. Santini (Journal of Biological ChemistryJournal of Biological Chemistry, 2023)
Arsenic contamination of groundwater is among one of the biggest health threats affecting millions of people in the world. There is an urgent need for efficient arsenic biosensors where the use of arsenic metabolizing enzymes can be explored. In this work, we have solved four crystal structures of arsenite oxidase (Aio) in complex with arsenic and antimony oxyanions and the structures determined correspond to intermediate states of the enzymatic mechanism. These structural data were complemented with density-functional theory calculations providing a unique view of the molybdenum active site at different time points that, together with mutagenesis data, enabled to clarify the enzymatic mechanism and the molecular determinants for the oxidation of As(III) to the less toxic As(V) species.Arsenic contamination of groundwater is among one of the biggest health threats affecting millions of people in the world. There is an urgent need for efficient arsenic biosensors where the use of arsenic metabolizing enzymes can be explored. In this work, we have solved four crystal structures of arsenite oxidase (Aio) in complex with arsenic and antimony oxyanions and the structures determined correspond to intermediate states of the enzymatic mechanism. These structural data were complemented with density-functional theory calculations providing a unique view of the molybdenum active site at different time points that, together with mutagenesis data, enabled to clarify the enzymatic mechanism and the molecular determinants for the oxidation of As(III) to the less toxic As(V) species.
doi: 10.1016/j.jbc.2023.105036

Elucidating the concentration-dependent effects of thiocyanate binding to carbonic anhydrase - Silva, José Malanho; Cerofolini, Linda; Ana Luísa Carvalho; Ravera, Enrico; Fragai, Marco; Parigi, Giacomo; Macedo, Anjos L.; Geraldes, Carlos F. G. C.; Luchinat, Claudio (2023)
Many proteins naturally carry metal centers, with a large share of them being in the active sites of several enzymes. Paramagnetic effects are a powerful source of structural information and, therefore, if the native metal is paramagnetic, or it can be functionally substituted with a paramagnetic one, paramagnetic effects can be used to study the metal sites, as well as the overall structure of the protein. One notable example is cobalt(II) substitution for zinc(II) in carbonic anhydrase. In this manuscript we investigate the effects of sodium thiocyanate on the chemical environment of the metal ion of the human carbonic anhydrase II. The electron paramagnetic resonance (EPR) titration of the cobalt(II) protein with thiocyanate shows that the EPR spectrum changes from A-type to C-type on passing from 1:1 to 1:1000-fold ligand excess. This indicates the occurrence of a change in the electronic structure, which may reflect a sizable change in the metal coordination environment in turn caused by a modification of the frozen solvent glass. However, paramagnetic nuclear magnetic resonance (NMR) data indicate that the metal coordination cage remains unperturbed even in 1:1000-fold ligand excess. This result proves that the C-type EPR spectrum observed at large ligand concentration should be ascribed to the low temperature at which EPR measurements are performed, which impacts on the structure of the protein when it is destabilized by a high concentration of a chaotropic agent.
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Keywords: EPR; Human carbonic anhydrase II; NMR; Paramagnetism; Sodium thiocyanate; Structural biology

Integrated approaches for the separation and purification of recombinant HPV16 E6 protein from Escherichia coli crude extracts - Gomes, D.; Correia, M. A. S.; Romão, M.J.; Passarinha, L.A.; Sousa, A. (2023)
Human papillomavirus (HPV) is a sexually transmissible virus responsible for 5% of global human cancers and associated with 99% of cervical cancer cases. The oncogenic potential of high-risk HPVs is mainly related to the E6 and E7 oncoproteins, which are responsible, at least in part, for inactivating the p53 and pRb tumor suppressor proteins. Due to the critical role of the E6 protein in malignant tumorigenesis, it is widely recognized as a therapeutic target for anti-HPV drug development. Nevertheless, it is required to obtain large amounts of protein with high purity to perform biointeraction studies with the potential inhibitor drugs. In this work, recombinant dual-tagged E6 protein (His6-MBP-E6) was expressed from Escherichia coli (E. coli) cultures and successfully extracted by sonication/ice cycles. Affinity chromatography using MBPtrap columns allowed 85 ± 5% protein recovery with the elimination of major host heterologous proteins in a single fraction. Subsequently, a polishing step was studied by applying anionic exchange (QSepharose), size exclusion (Superdex), or immobilized-metal affinity chromatography (HisTrap). The combination of affinity chromatography with size exclusion or two affinity chromatography techniques allowed us to obtain 82 ± 2% and 94 ± 3%, of highly pure His6-MBP-E6, respectively. Also, the secondary structure of His6-MBP-E6 is preserved in both purification strategies, as appraised by circular dichroism and western-blot studies. Thermal shift assay confirmed the CD results and suggested potential additives for protein stabilization. Altogether, the reproducible strategies established for the purification of His6-MBP-E6 protein could be successfully applied to later perform biointeraction studies and structural characterization of protein–ligand complexes.
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Keywords: Affinity purification; Chromatography; E6 protein; HPV; Protein solubility; Protein stability

The structure of a Bacteroides thetaiotamicron carbohydrate-binding module provides new insight into the recognition of complex pectic polysaccharides by the human microbiome - Trovão, Filipa; Correia, Viviana G.; Lourenço, Frederico M.; Ribeiro, Diana O.; Ana Luísa Carvalho; Palma, Angelina S.; Pinheiro, Benedita A. (2023)
TheBacteroides thetaiotaomicronhas developed a consortium of enzymes capable of overcoming steric constraints and degrading, in a sequential manner, the complex rhamnogalacturonan II (RG-II) polysaccharide. BT0996 protein acts in the initial stages of the RGII depolymerisation, where its two catalytic modules remove the terminal monosaccharides from RG-II side chains A and B. BT0996 is modular and has three putative carbohydrate-binding modules (CBMs) for which the roles in the RG-II degradation are unknown. Here, we present the characterisation of themoduleat the C-terminal domain, which we designated BT0996C. The high-resolution structure obtained by X-ray crystallography reveals that the protein displays a typical β-sandwich fold with structural similarity to CBMs assigned to families 6 and 35. The distinctive features are: 1) the presence of several charged residues at the BT0996-C surface creating a large, broad positive lysine-rich patch that encompasses the putative binding site; and 2) the absence of the highly conserved binding-site signatures observed in CBMs from families 6 and 35, such as region A tryptophan and region C asparagine. These findings hint at a binding mode of BT0996-C not yet observed in its homologues. In line with this, carbohydrate microarrays and microscale thermophoresis show the ability of BT0996-C to bind α1-4-linked polygalacturonic acid, and that electrostatic interactions are essential for the recognition of the anionic polysaccharide. The results support the hypothesis that BT0996-C may have evolved to potentiate the action of BT0996 catalytic modules on the complex structure of RG-II by binding to the polygalacturonic acid backbone sequence.
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Keywords: Carbohydrate Binding Module; Carbohydrates; Human Gut Microbiota; Rhamnogalacturonan II

Structure-function studies can improve binding affinity of cohesin-dockerin interactions for multi-protein assemblies - Duarte, Marlene; Alves, Victor D.; Correia, Márcia; Caseiro, Catarina; Ferreira, Luís M. A.; Romão, Maria João; Ana Luísa Carvalho; Najmudin, Shabir; Bayer, Edward A.; Fontes, Carlos M. G. A.; Bule, Pedro (2023)
The cellulosome is an elaborate multi-enzyme structure secreted by many anaerobic microorganisms for the efficient degradation of lignocellulosic substrates. It is composed of multiple catalytic and non-catalytic components that are assembled through high-affinity protein-protein interactions between the enzyme-borne dockerin (Doc) modules and the repeated cohesin (Coh) modules present in primary scaffoldins. In some cellulosomes, primary scaffoldins can interact with adaptor and cell-anchoring scaffoldins to create structures of increasing complexity. The cellulosomal system of the ruminal bacterium, Ruminococcus flavefaciens, is one of the most intricate described to date. An unprecedent number of different Doc specificities results in an elaborate architecture, assembled exclusively through single-binding-mode type-III Coh-Doc interactions. However, a set of type-III Docs exhibits certain features associated with the classic dual-binding mode Coh-Doc interaction. Here, the structure of the adaptor scaffoldin-borne ScaH Doc in complex with the Coh from anchoring scaffoldin ScaE is described. This complex, unlike previously described type-III interactions in R. flavefaciens, was found to interact in a dual-binding mode. The key residues determining Coh recognition were also identified. This information was used to perform structure-informed protein engineering to change the electrostatic profile of the binding surface and to improve the affinity between the two modules. The results show that the nature of the residues in the ligand-binding surface plays a major role in Coh recognition and that Coh-Doc affinity can be manipulated through rational design, a key feature for the creation of designer cellulosomes or other affinity-based technologies using tailored Coh-Doc interactions.
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Keywords: Biomass degradation; Carbohydrates; cellulosome; cohesin; dockerin; protein complex

Cellulose-based supercapacitors - Baptista, Ana; Amjid Rafique; Mariana Moniz; Inês Sequeira; João Carmo; Ferreira, Isabel (1st Iberian Symposium on Functional Organic Polymers, 2023)

Artificial Generative Intelligence and Work – Portugal - Moniz, António B.; Candeias, Marta; Nuno Boavida (EPTA, 2023)

Arsenite oxidase in complex with antimonite and arsenite oxyanions: Insights into the catalytic mechanism - Engrola, Filipa; Correia, Márcia A. S.; Cameron Watson; Romão, Carlos C.; Veiros, Luis F.; Romão, Maria João; Santos-Silva, Teresa; Joanne M. Santini (Journal of Biological Chemistry, 2023)

Cell division protein FtsK coordinates bacterial chromosome segregation and daughter cell separation - H. Veiga; Jousselin, A.; Schaeper, S.; Marques, L. B.; P. Reed; B.M. Saraiva; Wilton, J.; S. R. Filipe; M. G. Pinho (EMBO J, 2023)

Chemical stability of new nafion membranes doped with bisphosphonic acids under Fenton oxidative conditions - F. C. Teixeira; A. P. S. Teixeira; C.M. Rangel (Int. J. Hydrogen Energy, 2023)

Design of Magnetic Fe3O4/CeO2 “Core/Shell”-Like Nanocomposites with Pronounced Antiamyloidogenic and Antioxidant Bioactivity - Shlapa, Yu.; Siposova, K.; Veltruska, K.; Maraloiu, V.-A.; Rajnak, M.; Garcarova, I.; Timko, M.; Musatov, A.; Belous A. (ACS Appl. Mater. Interfaces, 2023)

Development of photosensitive fibers for photocurrent therapy - Júlio Rafael Cunha (FCT Nova, 2023)

Electrospray Deposition of PEDOT:PSS on Carbon Yarn Electrodes for Solid-State Flexible Supercapacitors - Mariana Moniz; Amjid Rafique; Marques, Ana; Ferreira, Isabel; Baptista, Ana; João Carmo; Oliveira, J.P. (ACS Applied Materials & Interfaces 2023, 2023)

Exploring electrospinning for sustainable triboelectric applications - António Maria Martins (FCT Nova, 2023)

A facile blow spinning technique for green cellulose acetate/polystyrene composite separator for flexible energy storage devices - Amjid Rafique; Inês Sequeira; Ana Sofia Bento; Mariana Moniz; João Carmo; Eduardo Oliveira; Oliveira, J.P.; Marques, Ana; Ferreira, Isabel (Chemical Engineering Journal, 2023)

Fibre-shaped photovoltaic devices for electronic textiles - Catarina Reis (FCT Nova, 2023)

Functionalisation of Electrospun Cellulose Acetate Membranes with PEDOT and PPy for Electronic Controlled Drug Release - Beatriz Lago; M Brito; C.M.M. Almeida; Ferreira, Isabel; Baptista, Ana (Nanomaterials 2023, 2023)

Light-responsive on-demand drug-delivery systems for dermal applications - Mariana Mendes (FCT Nova, 2023)

Lithium recovery from brines by lithium membrane flow capacitive deionization (Li-MFCDI) – A proof of concept - H.M. Saif; Crespo, J. G.; S. Pawlowski (Journal of Membrane Science Letters, 2023)

Mimicking Natural-Colored Photonic Structures with Cellulose-Based Materials - Quelhas, A.R.; Trindade, A.C. (Crystals, 2023)

Nature-Inspired Cellulose-Based Active Materials: From 2D to 4D - Marta I. Magalhães; Ana P.C. Almeida (Applied Biosciences, 2023)

Optoelectronic stimulation of drug release from membranes for dermal applications - Carolina Nascimento (FCT Nova, 2023)

Photovoltaic fibers for wearable energy harvesting applications - Francisco Baptista (FCT Nova, 2023)

Recent Advances and Challenges Towards Application of Fibers and Textiles in Integrated Photovoltaic Energy Storage Devices - Amjid Rafique; Ferreira, Isabel; G.Abbas; Baptista, Ana (Nano-Micro Letters, 2023)

Surface-modified cerium dioxide nanoparticles with improved anti-amyloid and preserved nanozymatic activity - Garcarova, I.; Valusova, E.; Shlapa, Yu.; Belous, A.; Musatov, A.; Siposova, K. (Colloids and Surfaces B: Biointerfaces, 2023)

A Biochemical Deconstruction-Based Strategy to Assist the Characterization of Bacterial Electric Conductive Filaments - Silva, , Marta A.; Fernandes, Ana P.; Turner, David L.; Salgueiro, , Carlos A. (International Journal of Molecular Sciences, 2023)
Periplasmic nanowires and electric conductive filaments made of the polymeric assembly of c-type cytochromes from Geobacter sulfurreducens bacterium are crucial for electron storage and/or extracellular electron transfer. The elucidation of the redox properties of each heme is fundamental to the understanding of the electron transfer mechanisms in these systems, which first requires the specific assignment of the heme NMR signals. The high number of hemes and the molecular weight of the nanowires dramatically decrease the spectral resolution and make this assignment extremely complex or unattainable. The nanowire cytochrome GSU1996 ( 42 kDa) is composed of four domains (A to D) each containing three c-type heme groups. In this work, the individual domains (A to D), bi-domains (AB, CD) and full-length nanowire were separately produced at natural abundance. Sufficient protein expression was obtained for domains C ( 11 kDa/three hemes) and D ( 10 kDa/three hemes), as well as for bi-domain CD ( 21 kDa/six hemes). Using 2D-NMR experiments, the assignment of the heme proton NMR signals for domains C and D was obtained and then used to guide the assignment of the corresponding signals in the hexaheme bi-domain CD. This new biochemical deconstruction-based procedure, using nanowire GSU1996 as a model, establishes a new strategy to functionally characterize large multiheme cytochromes.
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Characterization of the inner membrane cytochrome ImcH from Geobacter reveals its importance for extracellular electron transfer and energy conservation - Pimenta, Andreia I.; Paquete, Catarina M.; Leonor Morgado; Edwards, Marcus J.; Clarke, Thomas A.; Salgueiro, , Carlos A.; Pereira, Inês A. C.; Duarte, Américo G. (Protein Science, 2023)
Abstract Electroactive bacteria combine the oxidation of carbon substrates with an extracellular electron transfer (EET) process that discharges electrons to an electron acceptor outside the cell. This process involves electron transfer through consecutive redox proteins that efficiently connect the inner membrane to the cell exterior. In this study, we isolated and characterized the quinone-interacting membrane cytochrome c ImcH from Geobacter sulfurreducens, which is involved in the EET process to high redox potential acceptors. Spectroscopic and electrochemical studies show that ImcH hemes have low midpoint redox potentials, ranging from −150 to −358 mV, and connect the oxidation of the quinol-pool to EET, transferring electrons to the highly abundant periplasmic cytochrome PpcA with higher affinity than to its homologues. Despite the larger number of hemes and transmembrane helices, the ImcH structural model has similarities with the NapC/NirT/NrfH superfamily, namely the presence of a quinone-binding site on the P-side of the membrane. In addition, the first heme, likely involved on the quinol oxidation, has apparently an unusual His/Gln coordination. Our work suggests that ImcH is electroneutral and transfers electrons and protons to the same side of the membrane, contributing to the maintenance of a proton motive force and playing a central role in recycling the menaquinone pool.
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Keywords: electroactive bacteria; energy conservation; extracellular electron transfer; Geobacter

The Chlamydia trachomatis IncM Protein Interferes with Host Cell Cytokinesis, Centrosome Positioning, and Golgi Distribution and Contributes to the Stability of the Pathogen-Containing Vacuole - Maria Pequito Luís; Inês Serrano Pereira; Joana N. Bugalhão; Catarina N. Simões; Mota, Cristiano; Romão, Maria João; Mota, Luís Jaime (Infection and Immunity, 2023)
Chlamydia trachomatis is an obligate intracellular bacterial pathogen that causes ocular and urogenital infections in humans. The ability of C. trachomatis to grow intracellularly in a pathogen-containing vacuole (known as an inclusion) depends on chlamydial effector proteins transported into the host cell by a type III secretion system. Chlamydia trachomatis is an obligate intracellular bacterial pathogen that causes ocular and urogenital infections in humans. The ability of C. trachomatis to grow intracellularly in a pathogen-containing vacuole (known as an inclusion) depends on chlamydial effector proteins transported into the host cell by a type III secretion system. Among these effectors, several inclusion membrane proteins (Incs) insert in the vacuolar membrane. Here, we show that human cell lines infected by a C. trachomatis strain deficient for Inc CT288/CTL0540 (renamed IncM) displayed less multinucleation than when infected by IncM-producing strains (wild type or complemented). This indicated that IncM is involved in the ability of Chlamydia to inhibit host cell cytokinesis. The capacity of IncM to induce multinucleation in infected cells was shown to be conserved among its chlamydial homologues and appeared to require its two larger regions predicted to be exposed to the host cell cytosol. C. trachomatis-infected cells also displayed IncM-dependent defects in centrosome positioning, Golgi distribution around the inclusion, and morphology and stability of the inclusion. The altered morphology of inclusions containing IncM-deficient C. trachomatis was further affected by depolymerization of host cell microtubules. This was not observed after depolymerization of microfilaments, and inclusions containing wild-type C. trachomatis did not alter their morphology upon depolymerization of microtubules. Overall, these findings suggest that IncM may exert its effector function by acting directly or indirectly on host cell microtubules.
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Combining the amplification refractory mutation system and high-resolution melting analysis for KRAS mutation detection in clinical samples - Oliveira, Beatriz B.; Costa, Beatriz; Morão, Bárbara; Faias, Sandra; Bruno Veigas; Pereira, Lucília Pebre; Albuquerque, Cristina; Maio, Rui; Marília Cravo; Fernandes, Alexandra R; Baptista, Pedro Viana (Analytical and Bioanalytical Chemistry, 2023)
The success of personalized medicine depends on the discovery of biomarkers that allow oncologists to identify patients that will benefit from a particular targeted drug. Molecular tests are mostly performed using tumor samples, which may not be representative of the tumor’s temporal and spatial heterogeneity. Liquid biopsies, and particularly the analysis of circulating tumor DNA, are emerging as an interesting means for diagnosis, prognosis, and predictive biomarker discovery. In this study, the amplification refractory mutation system (ARMS) coupled with high-resolution melting analysis (HRMA) was developed for detecting two of the most relevant KRAS mutations in codon 12. After optimization with commercial cancer cell lines, KRAS mutation screening was validated in tumor and plasma samples collected from patients with pancreatic ductal adenocarcinoma (PDAC), and the results were compared to those obtained by Sanger sequencing (SS) and droplet digital polymerase chain reaction (ddPCR). The developed ARMS-HRMA methodology stands out for its simplicity and reduced time to result when compared to both SS and ddPCR but showing high sensitivity and specificity for the detection of mutations in tumor and plasma samples. In fact, ARMS-HRMA scored 3 more mutations compared to SS (tumor samples T6, T7, and T12) and one more compared to ddPCR (tumor sample T7) in DNA extracted from tumors. For ctDNA from plasma samples, insufficient genetic material prevented the screening of all samples. Still, ARMS-HRMA allowed for scoring more mutations in comparison to SS and 1 more mutation in comparison to ddPCR (plasma sample P7). We propose that ARMS-HRMA might be used as a sensitive, specific, and simple method for the screening of low-level mutations in liquid biopsies, suitable for improving diagnosis and prognosis schemes. Graphical Abstract: [Figure not available: see fulltext.]
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/LA%2FP%2F0140%2F2020/PT# info:eu-repo/grantAgreement/FCT/OE/2020.07660.BD/PT# Publisher Copyright: © 2023, The Author(s).
Keywords: ARMS-HRMA; circulating tumor DNA; Mutation detection; pancreatic cancer

Doxorubicin-sensitive and -resistant colorectal cancer spheroid models: assessing tumor microenvironment features for therapeutic modulation - Valente, Ruben; Cordeiro, Sandra; Luz, André; Rodrigues, Catarina Roma; Baptista, Pedro V; Fernandes, Alexandra R (Frontiers in Cell and Developmental Biology, 2023)
Introduction: The research on tumor microenvironment (TME) has recently been gaining attention due to its important role in tumor growth, progression, and response to therapy. Because of this, the development of three-dimensional cancer models that mimic the interactions in the TME and the tumor structure and complexity is of great relevance to cancer research and drug development. Methods: This study aimed to characterize colorectal cancer spheroids overtime and assess how the susceptibility or resistance to doxorubicin (Dox) or the inclusion of fibroblasts in heterotypic spheroids influence and modulate their secretory activity, namely the release of extracellular vesicles (EVs), and the response to Dox-mediated chemotherapy. Different characteristics were assessed over time, namely spheroid growth, viability, presence of hypoxia, expression of hypoxia and inflammation-associated genes and proteins. Due to the importance of EVs in biomarker discovery with impact on early diagnostics, prognostics and response to treatment, proteomic profiling of the EVs released by the different 3D spheroid models was also assessed. Response to treatment was also monitored by assessing Dox internalization and its effects on the different 3D spheroid structures and on the cell viability. Results and Discussion: The results show that distinct features are affected by both Dox resistance and the presence of fibroblasts. Fibroblasts can stabilize spheroid models, through the modulation of their growth, viability, hypoxia and inflammation levels, as well as the expressions of its associated transcripts/proteins, and promotes alterations in the protein profile exhibit by EVs. Summarily, fibroblasts can increase cell-cell and cell-extracellular matrix interactions, making the heterotypic spheroids a great model to study TME and understand TME role in chemotherapies resistance. Dox resistance induction is shown to influence the internalization of Dox, especially in homotypic spheroids, and it is also shown to influence cell viability and consequently the chemoresistance of those spheroids when exposed to Dox. Taken together these results highlight the importance of finding and characterizing different 3D models resembling more closely the in vivo interactions of tumors with their microenvironment as well as modulating drug resistance.
Funding Information: info:eu-repo/grantAgreement/FCT/Concurso para Financiamento de Projetos de Investigação Científica e Desenvolvimento Tecnológico em Todos os Domínios Científicos - 2020/PTDC%2FQUI-QOR%2F1304%2F2020/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/2022.04315.PTDC/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/LA%2FP%2F0140%2F2020/PT# info:eu-repo/grantAgreement/FCT//SFRH%2FBPD%2F124612%2F2016/PT# info:eu-repo/grantAgreement/FCT/OE/2022.12161.BD/PT# info:eu-repo/grantAgreement/FCT//2022.09845.BD/PT# info:eu-repo/grantAgreement/FCT/OE/2021.08629.BD/PT# The author(s) declare financial support was received for the research, authorship, and/or publication of this article. This work was financed by national funds from FCT–Fundação para a Ciência e a Tecnologia, I.P./MCTES through the projects PTDC/QUIQIN/0146/2020. Publisher Copyright: Copyright © 2023 Valente, Cordeiro, Luz, Melo, Rodrigues, Baptista and Fernandes.
Keywords: chemoresistance; colorectal cancer; Doxorubicin; extracellular vesicles; heterotypic spheroid models; Tumor microenvironment

Exploring oxidative stress pathways in Geobacter sulfurreducens: the redox network between MacA peroxidase and triheme periplasmic cytochromes - Portela, Pilar C.; Leonor Morgado; Silva, , Marta A.; Denkhaus, Lukas; Einsle, Oliver; Salgueiro, , Carlos A. (Frontiers in Microbiology, 2023)
The recent reclassification of the strict anaerobe Geobacter sulfurreducens bacterium as aerotolerant brought attention for oxidative stress protection pathways. Although the electron transfer pathways for oxygen detoxification are not well established, evidence was obtained for the formation of a redox complex between the periplasmic triheme cytochrome PpcA and the diheme cytochrome peroxidase MacA. In the latter, the reduction of the high-potential heme triggers a conformational change that displaces the axial histidine of the low-potential heme with peroxidase activity. More recently, a possible involvement of the triheme periplasmic cytochrome family (PpcA-E) in the protection from oxidative stress in G. sulfurreducens was suggested. To evaluate this hypothesis, we investigated the electron transfer reaction and the biomolecular interaction between each PpcA-E cytochrome and MacA. Using a newly developed method that relies on the different NMR spectral signatures of the heme proteins, we directly monitored the electron transfer reaction from reduced PpcA-E cytochromes to oxidized MacA. The results obtained showed a complete electron transfer from the cytochromes to the high-potential heme of MacA. This highlights PpcA-E cytochromes’ efficient role in providing the necessary reducing power to mitigate oxidative stress situations, hence contributing to a better knowledge of oxidative stress protection pathways in G. sulfurreducens.
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Hemes on a string: insights on the functional mechanisms of PgcA from Geobacter sulfurreducens - Fernandes, Tomás M.; Silva, , Marta A.; Leonor Morgado; Salgueiro, , Carlos A. (Journal of Biological Chemistry, 2023)
Microbial extracellular reduction of insoluble compounds requires soluble electron shuttles that diffuse in the extracellular environment, freely diffusing cytochromes or direct contact with cellular conductive appendages that release or harvest electrons to assure a continuous balance between cellular requirements and environmental conditions. In this work, we produced and characterized the three cytochrome domains of PgcA, an extracellular triheme cytochrome that contributes to Fe(III) and Mn(IV) oxides reduction in Geobacter sulfurreducens. The three domains are structurally homologous, but their heme groups show variable axial coordination and reduction potential values. Electron transfer experiments monitored by NMR and visible spectroscopy show the variable extent to which the domains promiscuously exchange electrons, while reducing different electron acceptors. The results suggest that PgcA is part of a new class of cytochromes - microbial heme-tethered redox strings - that use low-complexity protein stretches to bind metals and promote intra- and intermolecular electron transfer events through its cytochrome domains.
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Keywords: Electron transfer; Metalloprotein; Multiheme cytochromes; Nuclear Magnetic Resonance (NMR); Protein domain; Respiratory chain

Hierarchical self-assembly of a reflectin-derived peptide - Dias, Ana Margarida Gonçalves Carvalho; Moreira, Inês Pimentel; Iana Lychko; Lopes Soares, Cátia; Nurrito, Arianna; Moura Barbosa, Arménio Jorge; Lutz-Bueno, Viviane; Mezzenga, Raffaele; Ana Luísa Carvalho; Pina, Ana Sofia; Roque, Ana Cecília Afonso (Frontiers in Chemistry, 2023)
Reflectins are a family of intrinsically disordered proteins involved in cephalopod camouflage, making them an interesting source for bioinspired optical materials. Understanding reflectin assembly into higher-order structures by standard biophysical methods enables the rational design of new materials, but it is difficult due to their low solubility. To address this challenge, we aim to understand the molecular self-assembly mechanism of reflectin’s basic unit—the protopeptide sequence YMDMSGYQ—as a means to understand reflectin’s assembly phenomena. Protopeptide self-assembly was triggered by different environmental cues, yielding supramolecular hydrogels, and characterized by experimental and theoretical methods. Protopeptide films were also prepared to assess optical properties. Our results support the hypothesis for the protopeptide aggregation model at an atomistic level, led by hydrophilic and hydrophobic interactions mediated by tyrosine residues. Protopeptide-derived films were optically active, presenting diffuse reflectance in the visible region of the light spectrum. Hence, these results contribute to a better understanding of the protopeptide structural assembly, crucial for the design of peptide- and reflectin-based functional materials.
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Hybrid Digital-Droplet Microfluidic Chip for Applications in Droplet Digital Nucleic Acid Amplification: Design, Fabrication and Characterization - Coelho, Beatriz J.; Neto, Joana P.; Sieira, Bárbara; Moura, André T.; Elvira Fortunato; Rodrigo Martins; Baptista, Pedro V; Rui Igreja; Hugo Águas (Sensors, 2023)
Microfluidic-based platforms have become a hallmark for chemical and biological assays, empowering micro- and nano-reaction vessels. The fusion of microfluidic technologies (digital microfluidics, continuous-flow microfluidics, and droplet microfluidics, just to name a few) presents great potential for overcoming the inherent limitations of each approach, while also elevating their respective strengths. This work exploits the combination of digital microfluidics (DMF) and droplet microfluidics (DrMF) on a single substrate, where DMF enables droplet mixing and further acts as a controlled liquid supplier for a high-throughput nano-liter droplet generator. Droplet generation is performed at a flow-focusing region, operating on dual pressure: negative pressure applied to the aqueous phase and positive pressure applied to the oil phase. We evaluate the droplets produced with our hybrid DMF–DrMF devices in terms of droplet volume, speed, and production frequency and further compare them with standalone DrMF devices. Both types of devices enable customizable droplet production (various volumes and circulation speeds), yet hybrid DMF–DrMF devices yield more controlled droplet production while achieving throughputs that are similar to standalone DrMF devices. These hybrid devices enable the production of up to four droplets per second, which reach a maximum circulation speed close to 1540 µm/s and volumes as low as 0.5 nL.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/LA%2FP%2F0037%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F50025%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F50025%2F2020/PT# info:eu-repo/grantAgreement/FCT/Concurso para Financiamento de Projetos de Investigação Científica e Desenvolvimento Tecnológico em Todos os Domínios Científicos - 2017/PTDC%2FBTM-SAL%2F31201%2F2017/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/LA%2FP%2F0140%2F2020/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F132904%2F2017/PT# info:eu-repo/grantAgreement/FCT/OE/COVID%2FBD%2F152453%2F2022/PT# Publisher Copyright: © 2023 by the authors.
Keywords: digital microfluidics; droplet microfluidics; flow focusing; negative pressure

Structural Characterization of Neisseria gonorrhoeae Bacterial Peroxidase—Insights into the Catalytic Cycle of Bacterial Peroxidases - Nóbrega, Cláudia S.; Ana Luísa Carvalho; Romão, Maria João; Pauleta, Sofia R. (International Journal of Molecular Sciences, 2023)
Neisseria gonorrhoeae is an obligate human pathogenic bacterium responsible for gonorrhea, a sexually transmitted disease. The bacterial peroxidase, an enzyme present in the periplasm of this bacterium, detoxifies the cells against hydrogen peroxide and constitutes one of the primary defenses against exogenous and endogenous oxidative stress in this organism. The 38 kDa heterologously produced bacterial peroxidase was crystallized in the mixed-valence state, the active state, at pH 6.0, and the crystals were soaked with azide, producing the first azide-inhibited structure of this family of enzymes. The enzyme binds exogenous ligands such as cyanide and azide, which also inhibit the catalytic activity by coordinating the P heme iron, the active site, and competing with its substrate, hydrogen peroxide. The inhibition constants were estimated to be 0.4 ± 0.1 µM and 41 ± 5 mM for cyanide and azide, respectively. Imidazole also binds and inhibits the enzyme in a more complex mechanism by binding to P and E hemes, which changes the reduction potential of the latest heme. Based on the structures now reported, the catalytic cycle of bacterial peroxidases is revisited. The inhibition studies and the crystal structure of the inhibited enzyme comprise the first platform to search and develop inhibitors that target this enzyme as a possible new strategy against N. gonorrhoeae.
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Tracking W-Formate Dehydrogenase Structural Changes During Catalysis and Enzyme Reoxidation - Vilela-Alves, Guilherme; Manuel, Rita Rebelo; Ana Rita Oliveira; Pereira, Inês Cardoso; Romão, Maria João; Mota, Cristiano (International Journal of Molecular Sciences, 2023)
Metal-dependent formate dehydrogenases (Fdh) catalyze the reversible conversion of CO2 to formate, with unrivalled efficiency and selectivity. However, the key catalytic aspects of these enzymes remain unknown, preventing us from fully benefiting from their capabilities in terms of biotechnological applications. Here, we report a time-resolved characterization by X-ray crystallography of the Desulfovibrio vulgaris Hildenborough SeCys/W-Fdh during formate oxidation. The results allowed us to model five different intermediate structures and to chronologically map the changes occurring during enzyme reduction. Formate molecules were assigned for the first time to populate the catalytic pocket of a Fdh. Finally, the redox reversibility of DvFdhAB in crystals was confirmed by reduction and reoxidation structural studies.
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DMSO/IL solvent systems for cellulose dissolution: Binary or ternary mixtures? - Paiva, Tiago G.; Zanatta, Marcileia; Cabrita, Eurico J.; Bernardes, Carlos E. S.; Corvo, Marta C. (Journal of Molecular Liquids, 2022)
{The mechanism of cellulose dissolution in ionic liquid (IL)/dimethyl sulfoxide (DMSO) solvent systems has attracted much attention due to the possible replacement of synthetic materials. However, the solvent behaviour is not completely understood. This work has found an explanation for the solvent behaviour in cellulose dissolution, considering the almost unavoidable presence of the water. Ternary {[}C(4)mim] Cl/DMSO/H2O mixtures were studied with Nuclear Magnetic Resonance experiments and molecular dynamics simulations to explore IL/molecular solvents interactions and disclose the water interactions in these complex media. Titration of binary and ternary solvent systems with water and DMSO disclosed a relation between water's proton chemical shift and the molar fraction of the mixture components, creating an unprecedent theory to predict the cellulose solvation ability. A ``working range{''} for IL/DMSO/H2O ratio was observed, tested in cellulose dissolution, and rationalized using cellobiose interaction. Within this solvent ratio, the interactions between components are maximized, being switched on, while out of the range, the interactions are no longer detected. (C) 2021 Elsevier B.V. All rights reserved.}
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Effect of Iodide-Based Organic Salts and Ionic Liquid Additives in Dye-Sensitized Solar Cell Performance - Sarrato, Joao; Pinto, Ana Lucia; Cruz, Hugo; Jordao, Noemi; Malta, Gabriela; Branco, Paula S.; Carlos Lima, J.; Branco, Luis C. (NANOMATERIALS, 2022)
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Antiproliferative Activity of Buddleja saligna (Willd.) against Melanoma and In Vivo Modulation of Angiogenesis - Twilley, Danielle; Thipe, Velaphi C.; Kishore, Navneet; Bloebaum, Pierce; Catarina Roma-Rodrigues; Baptista, Pedro V; Fernandes, Alexandra R; Selepe, Mamoalosi A.; Langhansova, Lenka; Katti, Kattesh; Namrita Lall (Pharmaceuticals, 2022)
Melanoma cells secrete pro-angiogenic factors, which stimulates growth, proliferation and metastasis, and therefore are key therapeutic targets. Buddleja saligna (BS), and an isolated triterpenoid mixture (DT-BS-01) showed a fifty percent inhibitory concentration (IC50) of 33.80 ± 1.02 and 5.45 ± 0.19 µg/mL, respectively, against melanoma cells (UCT-MEL-1) with selectivity index (SI) values of 1.64 and 5.06 compared to keratinocytes (HaCat). Cyclooxygenase-2 (COX-2) inhibition was observed with IC50 values of 35.06 ± 2.96 (BS) and 26.40 ± 4.19 µg/mL (DT-BS-01). BS (30 µg/mL) significantly inhibited interleukin (IL)-6 (83.26 ± 17.60%) and IL-8 (100 ± 0.2%) production, whereas DT-BS-01 (5 µg/mL) showed 51.07 ± 2.83 (IL-6) and 0 ± 6.7% (IL-8) inhibition. Significant vascular endothelial growth factor (VEGF) inhibition, by 15.84 ± 4.54 and 12.21 ± 3.48%, respectively, was observed. In the ex ovo chick embryo yolk sac membrane assay (YSM), BS (15 µg/egg) significantly reduced new blood vessel formation, with 53.34 ± 11.64% newly formed vessels. Silver and palladium BS nanoparticles displayed noteworthy SI values. This is the first report on the significant anti-angiogenic activity of BS and DT-BS-01 and should be considered for preclinical trials as there are currently no US Food and Drug Administration (FDA) approved drugs to inhibit angiogenesis in melanoma.
Funding Information: info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# This research was funded by the University of Pretoria, the National Research Foundation-DAAD (SFD13080220333), the Department of Science and Innovation (DST/CON 0059/2019), the Innovation Hub, L’Oréal-UNESCO. Publisher Copyright: © 2022 by the authors.
Keywords: angiogenesis; Antiproliferative activity; Buddleja saligna; ex ovo YSM; Melanoma

In Vitro and In Vivo Biological Activity of Ruthenium 1,10-Phenanthroline-5,6-dione Arene Complexes - Lenis-Rojas, Oscar A; Catarina Roma-Rodrigues; Carvalho, Beatriz; Cabezas-Sainz, Pablo; Fernández Vila, Sabela; Sánchez, Laura; Baptista, Pedro V; Fernandes, Alexandra R; Royo, Beatriz (International Journal of Molecular Sciences, 2022)
Ruthenium(II) arene complexes exhibit promising chemotherapeutic properties. In this study, the effect of the counter anion in Ru(II) complexes was evaluated by analyzing the biological effect of two Ru(II) p-cymene derivatives with the 1,10-phenanthroline-5,6-dione ligand of general-formula [(η6-arene)Ru(L)Cl][X] X = CF3SO3 (JHOR10) and PF6 (JHOR11). The biological activity of JHOR10 and JHOR11 was examined in the ovarian carcinoma cell line A2780, colorectal carcinoma cell line HCT116, doxorubicin-resistant HCT116 (HCT116-Dox) and in normal human dermal fibroblasts. Both complexes JHOR10 and JHOR11 displayed an antiproliferative effect on A2780 and HCT116 cell lines, and low cytotoxicity in fibroblasts. Interestingly, JHOR11 also showed antiproliferative activity in the HCT116-Dox cancer cell line, while JHOR10 was inactive. Studies in A2780 cells showed that JHOR11 induced the production of reactive oxygen species (ROS) that trigger autophagy and cellular senescence, but no apoptosis induction. Further analysis showed that JHOR11 presented no tumorigenicity, with no effect in the cellular mobility, as evaluated by thye wound scratch assay, and no anti- or pro-angiogenic effect, as evaluated by the ex-ovo chorioallantoic membrane (CAM) assay. Importantly, JHOR11 presented no toxicity in chicken and zebrafish embryos and reduced in vivo the proliferation of HCT116 injected into zebrafish embryos. These results show that these are suitable complexes for clinical applications with improved tumor cell cytotoxicity and low toxicity, and that counter-anion alteration might be a viable clinical strategy for improving chemotherapy outcomes in multidrug-resistant (MDR) tumors.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04612%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04612%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# Funding Information: We are grateful to Fundação para a Ciência e a Tecnologia, I.P., through LS4FUTURE Associated Laboratory (LA/P/0087/2020). The NMR spectrometers at CERMAX are integrated in the national NMR Network and partially supported through project 022162. Oscar A. Lenis-Rojas acknowledge national funds through FCT, POPH-Programa Operacional Potencial Humano, and FSE (European Social Fund) for the CEEC 2017 Initiative. Additionally, this work is financed by national funds from FCT—Fundação para a Ciência e a Tecnologia, I.P., in the scope of the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy—i4HB. Publisher Copyright: © 2022 by the authors.
Keywords: anticancer activity; autophagy; ROS; Ru-dione complexes; Zebrafish

Learning to see VOCs with Liquid Crystal Droplets - Susana I C J Palma; Frazao, Jose; Alves, Rita; Henrique M.A. Costa; Claudia Alves; Gamboa, Hugo; Silveira, Margarida; Roque, Ana C A (2022 IEEE International Symposium on Olfaction and Electronic Nose (ISOEN), 2022)
In hybrid gels with immobilized liquid crystal (LC) droplets, fast and unique optical texture variations are generated when distinct volatile organic compounds (VOCs) interact with the LC and disturb its molecular order. The optical texture variations can be observed under a polarized optical microscope or transduced into a signal representing the variations of light transmitted through the LC. We show how hybrid gels can accurately identify 11 distinct VOCs by using deep learning to analyze optical texture variations of individual droplets (0.93 average F1-score) and by using machine learning to analyze 1D optical signals from multiple droplets in hybrid gels (0.88 average F1-score)
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VOC Sensing in Humid and Dry Environments - Carina Esteves; Susana Palma; Costa, Henrique; Claudia Alves; Santos, Goncalo; Ramou, Efthymia; Roque, Ana Cecilia (2022 IEEE International Symposium on Olfaction and Electronic Nose (ISOEN), 2022)
We report the development of gas-sensing multicomponent hybrid materials to be used under humidified and dried environments without the need of sample preconditioning or heavy signal processing. The easy tunability and the unique characteristics presented by the multicomponent hybrid materials suggests their use in nearterm applications in electronic nose systems able to operate in dry or humidified environments.
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Digital Microfluidics-Powered Real-Time Monitoring of Isothermal DNA Amplification of Cancer Biomarker - Coelho, Beatriz Jorge; Bruno Veigas; Luís Bettencourt; Hugo Águas; Elvira Fortunato; Rodrigo Martins; Baptista, Pedro V; Rui Igreja (Biosensors, 2022)
We introduce a digital microfluidics (DMF) platform specifically designed to perform a loop-mediated isothermal amplification (LAMP) of DNA and applied it to a real-time amplification to monitor a cancer biomarker, c-Myc (associated to 40% of all human tumors), using fluorescence microscopy. We demonstrate the full manipulation of the sample and reagents on the DMF platform, resulting in the successful amplification of 90 pg of the target DNA (0.5 ng/µL) in less than one hour. Furthermore, we test the efficiency of an innovative mixing strategy in DMF by employing two mixing methodologies onto the DMF droplets—low frequency AC (alternating current) actuation as well as back-and-forth droplet motion—which allows for improved fluorescence readouts. Fluo-rophore bleaching effects are minimized through on-chip sample partitioning by DMF processes and sequential droplet irradiation. Finally, LAMP reactions require only 2 µL volume droplets, which represents a 10-fold volume reduction in comparison to benchtop LAMP.
Funding Information: info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F50025%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F50025%2F2020/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FBTM-SAL%2F31201%2F2017/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F132904%2F2017/PT# LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy?i4HB. Additional funding was received from FCT under the scope of the project reference LA/P/0037/2020, Beatriz J. Coelho and grant COVID/BD/152453/2022. Publisher Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland.
Keywords: cancer biomarker; digital microfluidics; fluorescence detection; loop-mediated isothermal amplification; real-time nucleic acid amplification monitoring

Endogenous Fluorescent Proteins in the Mucus of an Intertidal Polychaeta: Clues for Biotechnology - Rodrigo, Ana P.; Lopes, Ana Catarina; Pereira, Ricardo; Anjo, Sandra I.; Manadas, Bruno; Grosso, Ana R.; Baptista, Pedro V; Fernandes, Alexandra R; Costa, Pedro M. (Marine Drugs, 2022)
The vast ocean holds many unexplored organisms with unique adaptive features that enable them to thrive in their environment. The secretion of fluorescent proteins is one of them, with reports on the presence of such compounds in marine annelids being scarce. The intertidal Eulalia sp. is an example. The worm secretes copious amounts of mucus, that when purified and concentrated extracts, yield strong fluorescence under UV light. Emission has two main maxima, at 400 nm and at 500 nm, with the latter responsible for the blue–greenish fluorescence. Combining proteomics and transcriptomics techniques, we identified ubiquitin, peroxiredoxin, and 14-3-3 protein as key elements in the mucus. Fluorescence was found to be mainly modulated by redox status and pH, being consistently upheld in extracts prepared in Tris-HCl buffer with reducing agent at pH 7 and excited at 330 nm. One of the proteins associated with the fluorescent signal was localized in secretory cells in the pharynx. The results indicate that the secretion of fluorescent proteinaceous complexes can be an important defense against UV for this dweller. Additionally, the internalization of fluorescent complexes by ovarian cancer cells and modulation of fluorescence of redox status bears important considerations for biotechnological application of mucus components as markers.
Funding Information: info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FBTA-BTA%2F28650%2F2017/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F109462%2F2015/PT# info:eu-repo/grantAgreement/FCT/CEEC IND 2017/CEECIND%2F02699%2F2017%2FCP1462%2FCT0018/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04539%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04539%2F2020/PT# project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy-i4HB. The National Mass Spectrometry Network (RNEM) under the contract POCI-01-0145-FEDER-402-022125 (ref.: ROTEIRO/0028/2013). Publisher Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland.
Keywords: Fluorescence; marine Annelida; Mass Spectrometry; mucosecretions; poisonous secretions; proteome; redox status; transcriptomics; ubiquitin conjugates; UV light

Molecular Beacon for Detection miRNA-21 as a Biomarker of Lung Cancer - Alexandre, Daniela; Teixeira, Bernardo; Rico, André; Valente, Salete; Craveiro, Ana; Baptista, Pedro V; Cruz, Carla (International Journal of Molecular Sciences, 2022)
Lung cancer (LC) is the leading cause of cancer-related death worldwide. Although the diagnosis and treatment of non-small cell lung cancer (NSCLC), which accounts for approximately 80% of LC cases, have greatly improved in the past decade, there is still an urgent need to find more sensitive and specific screening methods. Recently, new molecular biomarkers are emerging as potential non-invasive diagnostic agents to screen NSCLC, including multiple microRNAs (miRNAs) that show an unusual expression profile. Moreover, peripheral blood mononuclear cells’ (PBMCs) miRNA profile could be linked with NSCLC and used for diagnosis. We developed a molecular beacon (MB)-based miRNA detection strategy for NSCLC. Following PBMCs isolation and screening of the expression profile of a panel of miRNA by RT-qPCR, we designed a MB targeting of up-regulated miR-21-5p. This MB 21-5p was characterized by FRET-melting, CD, NMR and native PAGE, allowing the optimization of an in-situ approach involving miR-21-5p detection in PBMCs via MB. Data show the developed MB approach potential for miR-21-5p detection in PBMCs from clinical samples towards NSCLC.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F00709%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F00709%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/POR_CENTRO/2021.07695.BD/PT# UID/Multi/04349/ 2019 POCI-01-0145-FEDER-022122 ROTEIRO/0031/2013-PINFRA/22161/2016 LA/P/0140/2020
Keywords: Diagnosis; Lung cancer; microRNAs; molecular beacon; Peripheral blood mononuclear cells

Experimental Investigation on the Behaviour of Hybrid HPFRC Flat Slabs - Brisid Isufi; Carla Marchão; Rui Marreiros; António Pinho Ramos (fib Congress 2022, Oslo, 2022)

Synergy between silk fibroin and ionic liquids for active gas-sensing materials - Inês P. Moreira; Carina Esteves; Susana I C J Palma; Ramou, Efthymia; Ana L.M. Carvalho; Roque, Ana C A (Materials Today Bio, 2022)
Silk fibroin is a biobased material with excellent biocompatibility and mechanical properties, but its use in bioelectronics is hampered by the difficult dissolution and low intrinsic conductivity. Some ionic liquids are known to dissolve fibroin but removed after fibroin processing. However, ionic liquids and fibroin can cooperatively give rise to functional materials, and there are untapped opportunities in this combination. The dissolution of fibroin, followed by gelation, in designer ionic liquids from the imidazolium chloride family with varied alkyl chain lengths (2–10 carbons) is shown here. The alkyl chain length of the anion has a large impact on fibroin secondary structure which adopts unconventional arrangements, yielding robust gels with distinct hierarchical organization. Furthermore, and due to their remarkable air-stability and ionic conductivity, fibroin ionogels are exploited as active electrical gas sensors in an electronic nose revealing the unravelled possibilities of fibroin in soft and flexible electronics.
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Quercetin Liposomal Nanoformulation for Ischemia and Reperfusion Injury Treatment - Margarida Ferreira‐silva; Catarina Faria‐silva; Carvalheiro, Manuela C.; Simões, Sandra; Marinho, H. Susana; Marcelino, Paulo; Campos, Maria Celeste; Metselaar, Josbert M.; Fernandes, Eduarda; Baptista, Pedro V; Fernandes, Alexandra R; Corvo, M. Luísa (Pharmaceutics, 2022)
Ischemia and reperfusion injury (IRI) is a common complication caused by inflammation and oxidative stress resulting from liver surgery. Current therapeutic strategies do not present the desirable efficacy, and severe side effects can occur. To overcome these drawbacks, new therapeutic alternatives are necessary. Drug delivery nanosystems have been explored due to their capacity to improve the therapeutic index of conventional drugs. Within nanocarriers, liposomes are one of the most successful, with several formulations currently in the market. As improved therapeutic outcomes have been demonstrated by using liposomes as drug carriers, this nanosystem was used to deliver quercetin, a flavonoid with anti-inflammatory and antioxidant properties, in hepatic IRI treatment. In the present work, a stable quercetin liposomal formulation was developed and characterized. Additionally, an in vitro model of ischemia and reperfusion was developed with a hypoxia chamber, where the anti-inflammatory potential of liposomal quercetin was evaluated, revealing the downregulation of pro-inflammatory markers. The anti-inflammatory effect of quercetin liposomes was also assessed in vivo in a rat model of hepatic IRI, in which a decrease in inflammation markers and enhanced recovery were observed. These results demonstrate that quercetin liposomes may provide a significant tool for addressing the current bottlenecks in hepatic IRI treatment.
PD/BD/135264/2017 UID/DTP/04138/2020 UIDP/04138/2020 UIDP/04378/2020 UIDB/04378/2020 LA/P/0140/2020 UIDB/50006/2020 UIDB/00100/2020
Keywords: Anti-inflammatory therapy; Drug delivery nanosystems; Hepatic ischemia and reperfusion injury; Inflammation; Liposomes; Liver; Quercetin

Supercapacitors for a wearable All-FIBRE device - Marques, Ana Claudia (3nd Condensed Matter Physics National Conference, 2022)

Classification of Volatile Compounds with Morphological Analysis of e-nose Response - Alves, Rita; João Rodrigues; Ramou, Efthymia; Susana Palma; Roque, Ana; Gamboa, Hugo (Proceedings of the 15th International Joint Conference on Biomedical Engineering Systems and Technologies - BIOSIGNALS, 2022)
Electronic noses (e-noses) mimic human olfaction, by identifying Volatile Organic Compounds (VOCs). This work presents a novel approach that successfully classifies 11 known VOCs using the signals generated by sensing gels in an in-house developed e-nose. The proposed signals’ analysis methodology is based on the generated signals’ morphology for each VOC since different sensing gels produce signals with different shapes when exposed to the same VOC. For this study, two different gel formulations were considered, and an average f1-score of 84% and 71% was obtained, respectively. Moreover, a standard method in time series classification was used to compare the performances. Even though this comparison reveals that the morphological approach is not as good as the 1-nearest neighbour with euclidean distance, it shows the possibility of using descriptive sentences with text mining techniques to perform VOC classification.
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Incorporation of VOC-Selective Peptides in Gas Sensing Materials - Oliveira, Ana; Ramou, Efthymia; Teixeira, Gonçalo; Susana Palma; Roque, Ana (Proceedings of the 15th International Joint Conference on Biomedical Engineering Systems and Technologies, 2022)
Enhancing the selectivity of gas sensing materials towards specific volatile organic compounds (VOCs) is challenging due to the chemical simplicity of VOCs as well as the difficulty in interfacing VOC selective biological elements with electronic components used in the transduction process. We aimed to tune the selectivity of gas sensing materials through the incorporation of VOC-selective peptides into gel-like gas sensing materials. Specifically, a peptide (P1) known to discriminate single carbon deviations among benzene and derivatives, along with two modified versions (P2 and P3), were integrated with gel compositions containing gelatin, ionic liquid and without or with a liquid crystal component (ionogels and hybrid gels respectively). These formulations change their electrical or optical properties upon VOC exposure, and were tested as sensors in an in-house developed e-nose. Their ability to distinct and identify VOCs was evaluated via a supervised machine learning classifier. Enhanced discrimination of benzene and hexane was detected for the P1-based hybrid gel. Additionally, complementarity of the electrical and optical sensors was observed considering that a combination of both their accuracy predictions yielded the best classification results for the tested VOCs. This indicates that a combinatorial array in a dual-mode e-nose could provide optimal performance and enhanced selectivity.
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Light Triggered Enhancement of Antibiotic Efficacy in Biofilm Elimination Mediated by Gold-Silver Alloy Nanoparticles - Alves-Barroco, Cinthia; Lorenzo Rivas-García; Fernandes, Alexandra R; Baptista, Pedro Viana (Frontiers in Microbiology, 2022)
Bacterial biofilm is a tri-dimensional complex community of cells at different metabolic stages involved in a matrix of self-produced extracellular polymeric substances. Biofilm formation is part of a defense mechanism that allows the bacteria to survive in hostile environments, such as increasing resistance or tolerance to antimicrobial agents, causing persistent infections hard to treat and impair disease eradication. One such example is bovine mastitis associated with Streptococcus dysgalactiae subsp. dysgalactiae (SDSD), whose worldwide health and economic impact is on the surge. As such, non-conventional nanobased approaches have been proposed as an alternative to tackle biofilm formation and to which pathogenic bacteria fail to adapt. Among these, metallic nanoparticles have gained significant attention, particularly gold and silver nanoparticles, due to their ease of synthesis and impact against microorganism growth. This study provides a proof-of-concept investigation into the use of gold-silver alloy nanoparticles (AuAgNPs) toward eradication of bacterial biofilms. Upon visible light irradiation of AuAgNPs there was considerable disturbance of the biofilms' matrix. The hindering of structural integrity of the biofilm matrix resulted in an increased permeability for entry of antibiotics, which then cause the eradication of biofilm and inhibit subsequent biofilm formation. Additionally, our results that AuAgNPs inhibited the formation of SDSD biofilms via distinct stress pathways that lead to the downregulation of two genes critical for biofilm production, namely, brpA-like encoding biofilm regulatory protein and fbpA fibronectin-binding protein A. This study provides useful information to assist the development of nanoparticle-based strategies for the active treatment of biofilm-related infections triggered by photoirradiation in the visible.
Funding Information: info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F118350%2F2016/PT# This work was financed by national funds from FCT-Fundação para a Ciência e a Tecnologia, I.P., in the scope of the project UIDP/04378/2020 and UIDB/04378/2020 of the Research Unit on Applied Molecular Biosciences - UCIBIO and the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy-i4HB. FCT–MEC have also acknowledged to CA-B, and Inn-Indigo 00002/2015 RA Detect to LR-G. Funding Information: info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F118350%2F2016/PT# This work was financed by national funds from FCT-Funda??o para a Ci?ncia e a Tecnologia, I.P., in the scope of the project UIDP/04378/2020 and UIDB/04378/2020 of the Research Unit on Applied Molecular Biosciences - UCIBIO and the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy-i4HB. FCT?MEC have also acknowledged for SFRH/BD/118350/2016 to CA-B, and Inn-Indigo 00002/2015 RA Detect to LR-G. Publisher Copyright: Copyright © 2022 Alves-Barroco, Rivas-García, Fernandes and Baptista.
Keywords: biofilm inhibition; biofilms; eradication mature biofilms; gold-silver alloy nanoparticles; Streptococcus

Manganese(I) tricarbonyl complexes as potential anticancer agents - Lenis-Rojas, Oscar A; Carvalho, Beatriz; Cabral, Rui; Silva, Margarida; Friães, Sofia; Catarina Roma-Rodrigues; Meireles, Marta S. H.; Gomes, Clara S B; Fernández, Jhonathan A. A.; Vila, Sabela F.; Rubiolo, Juan A.; Laura Sanchez; et al. (JBIC Journal of Biological Inorganic Chemistry, 2022)
The antiproliferative activity of [Mn(CO)3(N^N)Br] (N^N = phendione 1, bipy 3) and of the two newly synthesized Mn complexes [Mn(CO)3(acridine)(phendione)]OTf (2) and [Mn(CO)3(di-triazole)Br] (4) has been evaluated by MTS against three tumor cell lines A2780 (ovarian carcinoma), HCT116 (colorectal carcinoma), HCT116doxR (colorectal carcinoma resistant to doxorubicin), and in human dermal fibroblasts. The antiproliferative assay showed a dose-dependent effect higher in complex 1 and 2 with a selectivity toward ovarian carcinoma cell line 21 times higher than in human fibroblasts. Exposure of A2780 cells to IC50 concentrations of complex 1 and 2 led to an increase of reactive oxygen species that led to the activation of cell death mechanisms, namely via intrinsic apoptosis for 2 and autophagy and extrinsic apoptosis for 1. Both complexes do not target DNA or interfere with cell cycle progression but are able to potentiate cell migration and neovascularization (for 2) an indicative that their application might be directed for initial tumor stages to avoid tumor invasion and metastization and opening a new avenue for complex 2 application in regenerative medicine. Interestingly, both complexes do not show toxicity in both in vivo models (CAM and zebrafish). Graphical abstract: [Figure not available: see fulltext.]
Funding Information: info:eu-repo/grantAgreement/FCT/3599-PPCDT/125363/PT# This work was funded by national funds through FCT–Fundação para a Ciência e a Tecnologia, I.P., Project MOSTMICRO-ITQB with refs UIDB/04612/2020 and UIDP/04612/2020. The NMR spectrometers at CERMAX are integrated in the National NMR, Network (PTNMR) and are partially supported by Infrastructure Project No. 022161 (co-financed by FEDER through COMPETE 2020, POCI and PORL and FCT through PIDDAC). This work is financed by national funds from FCT—Fundação para a Ciência e a Tecnologia, I.P., in the scope of the project UIDP/04378/2020 and UIDB/04378/2020 of the Research Unit on Applied Molecular Biosciences—UCIBIO and the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy—i4HB. Oscar A. Lenis-Rojas acknowledges Project LISBOA-01-0145-FEDER-007660 (Microbiologia Molecular, Estrutural e Celular) funded by FEDER funds through COMPETE2020-Programa Operacional Competitividade e Internacionalização (POCI) and by national funds through FCT, POPH-Programa Operacional Potencial Humano, and FSE (European Social Fund) for the CEEC 2017 Initiative. Jhonathan A.A Fernández acknowledges Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES) and the program CAPES/PRINT Proc. 88887.470075/2019-00 for the financial support. Sabela F Vila, Juan A. Rubiolo, and Laura Sanchez acknowledge ACUIGEN (GI-1251) from the Universidad de Santiago de Compostela and Xunta de Galicia proyect ED431C 2018/28 for the financial support. Clara S. B. Gomes acknowledges the Associate Laboratory for Green Chemistry–LAQV and the Applied Molecular Biosciences Unit–UCIBIO. LAQV is financed by national funds from Fundação para a Ciência e a Tecnologia (UIDB/50006/2020, UIDP/50006/2020). The authors acknowledge M. Baleia and I. Goncalves for pDNA, gDNA, and preliminary BAX/BCL-2 data, respectively.
Keywords: A2780 tumor cells; Antiproliferative activity; Cancer; In vivo models; Mn(I) complexes

Boron clusters (ferrabisdicarbollides) shaping the future as radiosensitizers for multimodal (chemo/radio/PBFR) therapy of glioblastoma - Miquel Nuez-Martínez; María Queralt-Martín; Muñoz-Juan, Amanda; Aguilella, Vicente M.; Laromaine, Anna; Teixidor, Francesc; Viñas, Clara; Pinto, Catarina G.; Pinheiro, Teresa; Guerreiro, Joana F.; Filipa Mendes; Catarina Roma-Rodrigues; et al. (Journal of Materials Chemistry B, 2022)
Glioblastoma multiforme (GBM) is the most common and fatal primary brain tumor, and is highly resistant to conventional radiotherapy and chemotherapy. Therefore, the development of multidrug resistance and tumor recurrence are frequent. Given the poor survival with the current treatments, new therapeutic strategies are urgently needed. Radiotherapy (RT) is a common cancer treatment modality for GBM. However, there is still a need to improve RT efficiency, while reducing the severe side effects. Radiosensitizers can enhance the killing effect on tumor cells with less side effects on healthy tissues. Herein, we present our pioneering study on the highly stable and amphiphilic metallacarboranes, ferrabis(dicarbollides) ([o-FESAN]− and [8,8′-I2-o-FESAN]−), as potential radiosensitizers for GBM radiotherapy. We propose radiation methodologies that utilize secondary radiation emissions from iodine and iron, using ferrabis(dicarbollides) as iodine/iron donors, aiming to achieve a greater therapeutic effect than that of a conventional radiotherapy. As a proof-of-concept, we show that using 2D and 3D models of U87 cells, the cellular viability and survival were reduced using this treatment approach. We also tested for the first time the proton boron fusion reaction (PBFR) with ferrabis(dicarbollides), taking advantage of their high boron (11B) content. The results from the cellular damage response obtained suggest that proton boron fusion radiation therapy, when combined with boron-rich compounds, is a promising modality to fight against resistant tumors. Although these results are encouraging, more developments are needed to further explore ferrabis(dicarbollides) as radiosensitizers towards a positive impact on the therapeutic strategies for GBM.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UID%2FMulti%2F04349%2F2019/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/UTAP-EXPL%2FFMT%2F0020%2F2021/PT# info:eu-repo/grantAgreement/FCT/9471 - RIDTI/PTDC%2FBTM-TEC%2F29256%2F2017/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04565%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# Funding Information: The authors received support from the Spanish Ministerio de Economía y Competitividad (PID2019-106832RB-100), the Generalitat de Catalunya (2017SGR1720), FCT - Fundação para a Ciência e a Tecnologia, in the scope of the projects LISBOA-01-0247-FEDER-045904, and the project LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy - i4HB. The Metrology Laboratory of Ionizing Radiation team of Centro Tecnológico e Nuclear, Instituto Superior Técnico (CTN/IST) is acknowledged for their support in the irradiation setups. Miquel Nuez-Martínez is enrolled in the PhD program of the UAB. MQM and VMA acknowledge financial support by the Spanish Government MCIN/AEI/10.13039/501100011033 (project 2019-108434GB-I00 to VMA and project IJC2018-035283-I to MQM), and Universitat Jaume I (project UJI-B2018-53 to V. M. A. and project UJI-A2020-21 to MQM). SV thanks Croatian Science Foundation (project IP-2018-01-3168). Catarina I.G. Pinto is enrolled in the PhD scholarship 689 DFA/BD/07119/2020. Publisher Copyright: © 2022 The Royal Society of Chemistry.

Ecstasy metabolites and monoamine neurotransmitters upshift the Na+/K+ ATPase activity in mouse brain synaptosomes - Barbosa, Daniel Jose; Capela, Joao Paulo; Ferreira, Luisa Maria; Branco, Paula Serio; Fernandes, Eduarda; Bastos, Maria de Lourdes; Carvalho, Felix (ARCHIVES OF TOXICOLOGY, 2022)
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Protein Adsorption Performance of a Novel Functionalized Cellulose-Based Polymer - Gago, Diana; Corvo, Marta C.; Chagas, Ricardo; Ferreira, Luisa M.; Coelhoso, Isabel (POLYMERS, 2022)
Dicarboxymethyl cellulose (DCMC) was synthesized and tested for protein adsorption. The prepared polymer was characterized by inductively coupled plasma atomic emission spectrometry (ICP-AES), attenuated total reflection Fourier-transform infrared spectroscopy (ATR-FTIR) and solid state nuclear magnetic resonance (ssNMR) to confirm the functionalization of cellulose. This work shows that protein adsorption onto DCMC is charge dependent. The polymer adsorbs positively charged proteins, cytochrome C and lysozyme, with adsorption capacities of 851 and 571 mg g(-1), respectively. In both experiments, the adsorption process follows the Langmuir adsorption isotherm. The adsorption kinetics by DCMC is well described by the pseudo second-order model, and adsorption equilibrium was reached within 90 min. Moreover, DCMC was successfully reused for five consecutive adsorption-desorption cycles, without compromising the removal efficiency (98-99%).
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Short Lecture 4 {"}Evaluation of antiproliferative and anti-angiogenic activity of an ethanolic extract of Helichrysum odoratissimum (L.) Sweet against skin cancer{"} - Twilley, D; Meyer, D; Langhansova, L; Mcgaw, L. J.; Madikizela, B; Roma-Rodrigues, C.; Baptista, {P., V}; Fernandes, A. R.; Lall, N (Planta Medica, 2022)
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Tackling Humidity with Designer Ionic Liquid-Based Gas Sensing Soft Materials - Carina Esteves; Susana I C J Palma; Henrique M.A. Costa; Cláudia Alves; Gonçalo M.C. Santos; Ramou, Efthymia; Ana Luísa Carvalho; Vitor Alves; Roque, Ana C A (Advanced Materials, 2022)
Relative humidity is simultaneously a sensing target and a contaminant in gas and volatile organic compound (VOC) sensing systems, where strategies to control humidity interference are required. An unmet challenge is the creation of gas-sensitive materials where the response to humidity is controlled by the material itself. Here, humidity effects are controlled through the design of gelatin formulations in ionic liquids without and with liquid crystals as electrical and optical sensors, respectively. In this design, the anions [DCA]− and [Cl]− of room temperature ionic liquids from the 1-butyl-3-methylimidazolium family tailor the response to humidity and, subsequently, sensing of VOCs in dry and humid conditions. Due to the combined effect of the materials formulations and sensing mechanisms, changing the anion from [DCA]− to the much more hygroscopic [Cl]−, leads to stronger electrical responses and much weaker optical responses to humidity. Thus, either humidity sensors or humidity-tolerant VOC sensors that do not require sample preconditioning or signal processing to correct humidity impact are obtained. With the wide spread of 3D- and 4D-printing and intelligent devices, the monitoring and tuning of humidity in sustainable biobased materials offers excellent opportunities in e-nose sensing arrays and wearable devices compatible with operation at room conditions.
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Keywords: anion-tunability; gas sensing; gelatin; humidity; ionogels; liquid crystals; methylimidazolium ionic liquids

Automatic Generation of Contracts for Concurrent Java Programs - Pereira, H. G.; Sousa, D. G.; Bradbury, J. S.; Lourenço, J. M. (Atas do INForum 2022, 2022)

Empowering a Relational Database with LSD: Lazy State Determination - Thales, P.; Vale, T. M.; Dias, R.J.; Lourenço, J. M. (Atas do INForum 2022, 2022)

OSCAR - A Java Noise Injection Framework - Luna, F.; Bradbury, J. S.; Lourenço, J. M. (Atas do INForum 2022, 2022)
Best paper of the CPDLA track and runner-up for the Best Paper Award.

A Study of Latency-Aware Data-Placement in Heterogeneous (PMEM) Memory Systems - Antão, J.; Barreto, J.; Lourenço, J. M. (Atas do INForum 2022, 2022)

Cellulose-based energy storage devices - Baptista, Ana (International Summer School “BEST Summer Course on Technology”, 2022)

The W/SeCys-FdhAB formate dehydrogenase from Desulfovibrio vulgaris Hildenborough - Oliveira, Ana R.; Mota, Cristiano; Romão, Maria J.; Pereira, Inês A. C. (Encyclopedia of Inorganic and Bioinorganic Chemistry, 2022)
Abstract The W/SeCys-FdhAB formate dehydrogenase from Desulfovibrio vulgaris Hildenborough is a dimeric periplasmic enzyme that catalyzes the reversible oxidation of formate and reduction of CO2. It belongs to the group of metal-dependent FDHs, with a tungsten at the active site coordinated by two pyranopterin guanine dinucleotides, a selenocysteine, and one labile sulfur atom. FdhAB has a remarkably high activity and unusual tolerance to oxygen, making it an ideal model system to study biological CO2 reduction.
Major Reference Works
Keywords: CO2 reduction; formate dehydrogenase; pyranopterin; Selenocysteine; tungsten

Solution Combustion Synthesis of Hafnium-Doped Indium Oxide Thin Films for Transparent Conductors - Firmino, Rita; Carlos, Emanuel; Pinto, Joana Vaz; Deuermeier, Jonas; Rodrigo Martins; Elvira Fortunato; Barquinha, Pedro; Rita Branquinho (Nanomaterials, 2022)
Indium oxide (In2O3)-based transparent conducting oxides (TCOs) have been widely used and studied for a variety of applications, such as optoelectronic devices. However, some of the more promising dopants (zirconium, hafnium, and tantalum) for this oxide have not received much attention, as studies have mainly focused on tin and zinc, and even fewer have been explored by solution processes. This work focuses on developing solution-combustion-processed hafnium (Hf)-doped In2O3 thin films and evaluating different annealing parameters on TCO’s properties using a low environmental impact solvent. Optimized TCOs were achieved for 0.5 M% Hf-doped In2O3 when produced at 400 °C, showing high transparency in the visible range of the spectrum, a bulk resistivity of 5.73 × 10−2 Ω.cm, a mobility of 6.65 cm2/V.s, and a carrier concentration of 1.72 × 1019 cm−3. Then, these results were improved by using rapid thermal annealing (RTA) for 10 min at 600 °C, reaching a bulk resistivity of 3.95 × 10 −3 Ω.cm, a mobility of 21 cm2/V.s, and a carrier concentration of 7.98 × 1019 cm−3, in air. The present work brings solution-based TCOs a step closer to low-cost optoelectronic applications.
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Emergent solution based IGZO memristor towards neuromorphic applications - Martins, Raquel Azevedo; Carlos, Emanuel; Deuermeier, Jonas; Pereira, Maria Elias; Rodrigo Martins; Elvira Fortunato; Kiazadeh, Asal (Journal of Materials Chemistry C, 2022)
Solution-based memristors are emergent devices, due to their potential in electrical performance for neuromorphic computing combined with simple and cheap fabrication processes.
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Molecular Mechanisms of Microbial Extracellular Electron Transfer: The Importance of Multiheme Cytochromes - Paquete, Catarina M.; Leonor Morgado; Salgueiro, , Carlos A.; Ricardo O. Louro (FBL, 2022)
Extracellular electron transfer is a key metabolic process of many organismsthat enables them to exchange electrons with extracellular electrondonors/acceptors. The discovery of organisms with these abilities and theunderstanding of their electron transfer processes has become a priority for thescientific and industrial community, given the growing interest on the use ofthese organisms in sustainable biotechnological processes. For example, inbioelectrochemical systems electrochemical active organisms can exchangeelectrons with an electrode, allowing the production of energy and added-valuecompounds, among other processes. In these systems, electrochemical activeorganisms exchange electrons with an electrode through direct or indirectmechanisms, using, in most cases, multiheme cytochromes. In numerouselectroactive organisms, these proteins form a conductive pathway that allowselectrons produced from cellular metabolism to be transferred across the cellsurface for the reduction of an electrode, or vice-versa. Here, the mechanisms bywhich the most promising electroactive bacteria perform extracellular electrontransfer will be reviewed, emphasizing the proteins involved in these pathways.The ability of some of the organisms to perform bidirectional electron transferand the pathways used will also be highlighted.
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2-Hydroxychalcone-?-Cyclodextrin Conjugate with pH-Modulated Photoresponsive Binding Properties - Paulino, Micael; Perez-Juste, Ignacio; Magdalena Cid, Maria; Da Silva, Jose P.; Pereira, M Manuela A; Basilio, Nuno (JOURNAL OF ORGANIC CHEMISTRY, 2022)
Stimuli-responsive supramolecular receptors are important building blocks for the construction of self-assembled functional materials. We report the design and synthesis of a pH and light-responsive 2-hydroxychalcone-beta-cyclodextrin conjugate (1-Ct) and its characterization by spectroscopic and computational methods. 1-Ct follows the typical reaction network of trans-chalcone-flavylium photoswitches. Upon light irradiation, 1-Ct can be photochemically converted into the cis-chalcone/hemiketal forms (1-Cc/1-B) under neutral pH conditions or to the flavylium cation (1-AH+) at acidic pH values. This stimuli-responsive beta-cyclodextrin host, 1Ct, was found to form stronger intramolecular self-inclusion complexes (Kintra = 14) than 1-AH+ (Kintra = 3) and weaker than 1-Cc/1-B (overall Kintra = 179), allowing control over their stability and binding properties by combinations of pH and light stimuli.
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Binding of VIVO2+, VIVOL, VIVOL2 and VVO2L Moieties to Proteins: X-ray/Theoretical Characterization and Biological Implications - Santos, Marino F. A.; Sciortino, Giuseppe; Isabel Correia; Fernandes, Andreia C. P.; Santos-Silva, Teresa; Pisanu, Federico; Garribba, Eugenio; João Costa Pessoa (Chemistry – A European JournalChemistry – A European Journal, 2022)
Abstract Vanadium compounds have frequently been proposed as therapeutics, but their application has been hampered by the lack of information on the different V-containing species that may form and how these interact with blood and cell proteins, and with enzymes. Herein, we report several resolved crystal structures of lysozyme with bound VIVO2+ and VIVOL2+, where L=2,2?-bipyridine or 1,10-phenanthroline (phen), and of trypsin with VIVO(picolinato)2 and VVO2(phen)+ moieties. Computational studies complete the refinement and shed light on the relevant role of hydrophobic interactions, hydrogen bonds, and microsolvation in stabilizating the structure. Noteworthy is that the trypsin?VVO2(phen) and trypsin?VIVO(OH)(phen) adducts correspond to similar energies, thus suggesting a possible interconversion under physiological/biological conditions. The obtained data support the relevance of hydrolysis of VIV and VV complexes in the several types of binding established with proteins and the formation of different adducts that might contribute to their pharmacological action, and significantly widen our knowledge of vanadium?protein interactions.
https://doi.org/10.1002/chem.202200105
Keywords: metalloproteins; Molecular modeling; structure elucidation; Vanadium; X-ray diffraction

Spectroscopic and Structural Characterization of Reduced Desulfovibrio vulgaris Hildenborough W-FdhAB Reveals Stable Metal Coordination during Catalysis - Ana Rita Oliveira; Mota, Cristiano; Klymanska, Kateryna; Biaso, Frédéric; Romão, Maria João; Guigliarelli, Bruno; Pereira, Inês Cardoso (ACS Chemical BiologyACS Chemical Biology, 2022)
Metal-dependent formate dehydrogenases are important enzymes due to their activity of CO2 reduction to formate. The tungsten-containing FdhAB formate dehydrogenase from Desulfovibrio vulgaris Hildenborough is a good example displaying high activity, simple composition, and a notable structural and catalytic robustness. Here, we report the first spectroscopic redox characterization of FdhAB metal centers by EPR. Titration with dithionite or formate leads to reduction of three [4Fe–4S]1+ clusters, and full reduction requires Ti(III)–citrate. The redox potentials of the four [4Fe–4S]1+ centers range between −250 and −530 mV. Two distinct WV signals were detected, WDV and WFV, which differ in only the g2-value. This difference can be explained by small variations in the twist angle of the two pyranopterins, as determined through DFT calculations of model compounds. The redox potential of WVI/V was determined to be −370 mV when reduced by dithionite and −340 mV when reduced by formate. The crystal structure of dithionite-reduced FdhAB was determined at high resolution (1.5 Å), revealing the same structural alterations as reported for the formate-reduced structure. These results corroborate a stable six-ligand W coordination in the catalytic intermediate WV state of FdhAB.Metal-dependent formate dehydrogenases are important enzymes due to their activity of CO2 reduction to formate. The tungsten-containing FdhAB formate dehydrogenase from Desulfovibrio vulgaris Hildenborough is a good example displaying high activity, simple composition, and a notable structural and catalytic robustness. Here, we report the first spectroscopic redox characterization of FdhAB metal centers by EPR. Titration with dithionite or formate leads to reduction of three [4Fe–4S]1+ clusters, and full reduction requires Ti(III)–citrate. The redox potentials of the four [4Fe–4S]1+ centers range between −250 and −530 mV. Two distinct WV signals were detected, WDV and WFV, which differ in only the g2-value. This difference can be explained by small variations in the twist angle of the two pyranopterins, as determined through DFT calculations of model compounds. The redox potential of WVI/V was determined to be −370 mV when reduced by dithionite and −340 mV when reduced by formate. The crystal structure of dithionite-reduced FdhAB was determined at high resolution (1.5 Å), revealing the same structural alterations as reported for the formate-reduced structure. These results corroborate a stable six-ligand W coordination in the catalytic intermediate WV state of FdhAB.
doi: 10.1021/acschembio.2c00336

Synthesis and characterisation of ionic liquid crystals based on substituted pyridinium cations - Santos, Andreia F. M.; Cruz, Carlos; Godinho, Maria H; Madalena Dionísio; Figueirinhas, João L.; Branco, Luis C. (Liquid CrystalsLiquid Crystals, 2022)
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doi: 10.1080/02678292.2022.2070783

Utilização de BEDRF em Ligações Laje-Pilar sujeitas a Ações Horizontais Cíclicas - António Ramos; Brisid Isufi; Rui Marreiros; Carla Marchão (6ªs Jornadas Portuguesas de Engenharia de Estruturas, 2022)

Utilização Racional de BEDRF em Ligações Laje-Pilar - António Ramos; Brisid Isufi; Carla Marchão; Rui Marreiros (6ªs Jornadas Portuguesas de Engenharia de Estruturas, 2022)

Elucidation of complex respiratory chains: a straightforward strategy to monitor electron transfer between cytochromes - Leonor Morgado; Salgueiro, , Carlos A. (Metallomics, 2022)
{Cytochromes are electron transfer proteins essential in various biological systems, playing crucial roles in the respiratory chains of bacteria. These proteins are particularly abundant in electrogenic microorganisms and are responsible for the efficient delivery of electrons to the cells’ exterior. The capability of sending electron outside the cells open new avenues to be explored for emerging biotechnological applications in bioremediation, microbial electrosynthesis and bioenergy fields. To develop these applications, it is critical to identify the different redox partners and elucidate the stepwise electron transfer along the respiratory paths. However, investigating direct electron transfer events between proteins with identical features in nearly all spectroscopic techniques is extremely challenging. NMR spectroscopy offers the possibility to overcome this difficulty by analysing the alterations of the spectral signatures of each protein caused by electron exchange events. The uncrowded NMR spectral regions containing the heme resonances of the cytochromes display unique and distinct signatures in the reduced and oxidized states, which can be explored to monitor electron transfer within the redox complex. In this study, we present a strategy for a fast and straightforward monitorization of electron transfer between c-type cytochromes, using as model a triheme periplasmic cytochrome (PpcA) and a membrane associated monoheme cytochrome (OmcF) from the electrogenic bacterium Geobacter sulfurreducens. The comparison between the 1D 1H NMR spectra obtained for samples containing the two cytochromes and for samples containing the individual proteins clearly demonstrated a unidirectional electron transfer within the redox complex. This strategy provides a simple and straightforward means to elucidate complex biologic respiratory electron transfer chains.}
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Activated carbon materials with a rich surface chemistry prepared from L-cysteine amino acid - Reljic, S.; Cuadrado-Collados, C.; Oliveira Jardim, E.; Farrando-Perez, J.; Martinez-Escandell, M.; Silvestre-Albero, J (Fluid Phase Equilibria, 2022)

Automatic Generation of Contracts for Concurrent Java Progra - Hugo Pereira (FCT-NOVA, 2022)

Automation trends in Portugal: implications in productivity and employment - Candeias, Marta; Nuno Boavida; António Brandão Moniz (GEE Papers, 2022)

Automation trends in Portugal: implications in productivity and employment - Candeias, Marta; Moniz, António B.; Nuno Boavida (GEE Paper, 2022)

Behavior of flat slabs with partial use of high-performance fiber reinforced concrete under monotonic vertical loading - Brisid Isufi; João Pedro Relvas; Carla Marchão; António Ramos (Engineering Structures, 2022)
Reinforced concrete flat slabs are used worldwide in multi-story buildings. In these slabs, the design is often governed by punching shear and serviceability. The mitigation of these issues during design usually leads to increased raw material consumption and costs. Previous studies have shown that using Fiber Reinforced Concrete (FRC) or High-Strength Concrete (HSC) only at the vicinity of the column, while casting the rest of the slab with Normal Strength Concrete (NSC), can lead to an improved behavior under gravity loads in terms of both serviceability and ultimate capacity. Motivated by these results and the scarcity of previous tests, the present paper experimentally investigates the applicability of High-Performance Fiber Reinforced Concrete (HPFRC) as an alternative material that can be seen as an improvement over FRC and HSC, allowing a combination of ductility and strength. In addition, the HPFRC used in this paper is self-compacting, thus reducing the labor costs associated with concrete vibration. Five 150 mm thick flat slabs were tested under monotonically increasing punching load. The experimental variables were the flexural reinforcement ratio and the extent of the HPFRC zone. One of the specimens was cast only with NSC and served as a reference slab. Results show that the solution was effective for both flexural reinforcement ratios considered. Cracking load, maximum load, as well as the displacement capacity were increased significantly, even for a small extent of HPFRC (1.5 times the effective depth from the face of the column). Regarding the ultimate load capacity, it was observed an increase of 44% to 58% for the specimens with lower reinforcement ratio (0.64%) and between 15%–21% for the specimens with higher reinforcement ratio (0.96%). The results indicate that the use of HPFRC is a promising solution regarding both serviceability and ultimate limit state design of reinforced concrete flat slabs under gravity loading, with obvious advantages in material savings and labor costs.

Carbon-based monoliths with improved thermal and mechanical properties for methane storage - Reljic, S.; Cuadrado-Collados, C.; Farrando-Perez, J.; Jardim, E.O.; Martinez-Escandell, M.; Silvestre-Albero, J (Fuel, 2022)

Cellulose acetate fibres loaded with daptomycin for metal implant coatings - Faria, Jaime; Bruno Dionísio; Irís Soares; Baptista, Ana Catarina; Marques, Ana; Lídia Gonçalves; Ana Bettencourt; Carlos Baleizão; Ferreira, Isabel (Carbohydrate Polymers, 2022)

Cellulose Nanocrystal Aqueous Colloidal Suspensions: Evidence of Density Inversion at the Isotropic-liquid Crystal Phase Transition - da Rosa, Rafaela R.; Silva, Pedro E. S.; Saraiva, Diogo V.; Kumar, Anant; de Sousa, António P. Mendes; Sebastião, Pedro; Fernandes, Susete N.; Godinho, Maria Helena (Advanced Materials, 2022)
Abstract The colloidal suspensions of aqueous cellulose nanocrystals (CNCs) are known to form liquid crystalline systems above certain critical concentrations. From an isotropic phase; tactoid formation; growth; and sedimentation have been determined as the genesis of a high-density cholesteric phase; which after drying; originates solid iridescent films. Herein; we report the coexistence of a liquid crystal upper phase and an isotropic bottom phase in CNC aqueous suspensions at isotropic-nematic phase separation for the first time. Furthermore; isotropic spindle-like domains are observed in the low-density liquid crystalline phase; and high-density liquid crystalline phases are also prepared. The CNCs isolated from the low- and high-density liquid crystalline phases are found to have similar average lengths; diameters; and surface charges. The existence of a liquid crystalline low-density phase is explained by the presence of air dissolved in the water present within the CNCs. The air dissolves out when the water solidifies into ice and remains within the CNCs. The self-adjustment of the cellulose chain conformation enables the entrapment of air within the CNCs and CNC buoyancy in aqueous suspensions. This article is protected by copyright. All rights reserved
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Keywords: aqueous suspensions; cellulose nanocrystals (CNCs); liquid crystals; low-density liquid crystalline phase

Cerium dioxide nanoparticles synthesized via precipitation at constant pH: Synthesis, physical-chemical and antioxidant properties - Shlapa, Yu.; Solopan, S.; Sarnatskaya, V.; Siposova, K.; Garcarova, I.; Veltruska, K.; Timashkov, I.; Lykhova, O.; Kolesnik, D.; Musatov, A.; Nikolaev, V.; Belous, A. (Colloids and Surfaces B: Biointerface, 2022)

Changes in productivity and labour relations: artificial intelligence in the automotive sector in Portugal - António Brandão Moniz; Candeias, Marta; Nuno F.F.G. Boavida (Int. J. Automotive Technology and Management, 2022)

Changes in productivity and labour relations: artificial intelligence in the automotive sector in Portugal - Moniz, A. B.; Candeias, M.; Boavida, N. (Int. J. Automotive Technology and Management, 2022)

Characterization and modeling of resistive switching phenomena in IGZO devices - https://aip.scitation.org/doi/full/10.1063/5.0098145 (AIP Advances, 2022)

Characterization of a novel cytochrome involved in Geobacter sulfurreducens’ electron harvesting pathways - Liliana R. Teixeira; Fernandes, Tomás M.; Silva, , Marta A.; Leonor Morgado; Salgueiro, , Carlos A. (Chemistry – A European Journal, 2022)
Electron harvesting bacteria are key targets to develop microbial electrosynthesis technologies, which are valid alternatives for the production of value-added compounds without utilization of fossil fuels. Geobacter sulfurreducens, that is capable of donating and accepting electrons from electrodes, is one of the most promising electroactive bacteria. Its electron transfer mechanisms to electrodes have been progressively elucidated, however the electron harvesting pathways are still poorly understood. Previous studies showed that the periplasmic cytochromes PccH and GSU2515 are overexpressed in current-consuming G. sulfurreducens biofilms. PccH was characterized, though no putative partners have been identified. In this work, GSU2515 was characterized by complementary biophysical techniques and in silico simulations using the AlphaFold neural network. GSU2515 is a low-spin monoheme cytochrome with a disordered N-terminal region and an α-helical C-terminal domain harboring the heme group. The cytochrome undergoes a redox-linked heme axial ligand switch, with Met91 and His94 as distal axial ligand in the reduced and oxidized state, respectively. The reduction potential of the cytochrome is negative and is modulated by the pH in the physiological range: -78 mV at pH 6 and -113 mV at pH 7. Such pH-dependence coupled to the redox-linked switch of the axial ligand allows the cytochrome to drive a proton-coupled electron transfer step that is crucial to confer directionality to the respiratory chain. Biomolecular interactions and electron transfer experiments indicated that GSU2515 and PccH form a redox complex. Overall, the data obtained highlights for the first time how periplasmic proteins bridge the electron transfer between the outer and inner membrane in the electron harvesting pathways of G. sulfurreducens.
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Keywords: Biomolecular interactions; c-Type cytochrome; Electron transfer; Microbial electrosynthesis; Nuclear magnetic resonance

Digital Transformation in the Automotive Sector in Portugal: Data Analysis of Industrial R&D Projects - Candeias, Marta; Moniz, António B.; Nuno Boavida (Inclusive Futures for Eu- rope: Addressing the Digitalisation Challenges, Inclusive Futures for Europe, 2022)

Discovery and characterization of a novel Dyp-type peroxidase from a marine actinobacterium isolated from Trondheim fjord, Norway - Cordas, C. M.; G.S. Nguyen; G.V. Valério; M. Jønsson; K. Sóllner; I. Aune; A. Wentzel; Moura, J. J. G. (J Inorg Biochem, 2022)

Drug Delivery from PCL/Chitosan Multilayer Coatings for Metallic Implants - Íris Soares; Faria, Jaime; Marques, Ana; Isabel A C Ribeiro; Carlos Baleizão; Ana Bettencourt; Ferreira, Isabel; Baptista, Ana Catarina (ACS omega, 2022)

Drug Delivery from PCL/Chitosan Multilayer Coatings for Metallic Implants - Íris Soares; Faria, Jaime; Marques, Ana; Isabel Ribeiro; Carlos Baleizão; Ana Bettencourt; Ferreira, Isabel; Baptista, Ana (ACS Omega, 2022)

Drug delivery from PCL/Chitosan multilayer coatings for metallic implants, - I. Soares,; Faria, J.; AC. Marques; I. Ribeiro; C. Baleizão; A. Bettencourt; Ferreira, I.; Baptista, A. C. (ACS Omega, 2022)

Dual-Functional Antioxidant and Anti-amyloid Cerium Oxide Nanoparticles Fabricated by Controlled Synthesis in Water-Alcohol Solutions - Siposova, K.; Huntosova, V.; Garcarova, I.; Shlapa, Yu.; Timashkov, I.; Belous, A.; Musatov, A. (Biomedicines, 2022)

Effect of ferrofluid magnetization on transformer temperature rise - Rajnak, M.; Franko, M.; Paulovicova, K.; Karpets, M.; Parekh, K.; Upadhyay, R.; Kurimsky, J.; Dolnik, B.; Cimbala, R.; Havran, P.; Timko, M.; Kopcansky, P. (Journal of Physics D: Applied Physics, 2022)

Electric field-induced assembly of magnetic nanoparticles from dielectric ferrofluids on planar interface - Karpets, M.; Rajnak, M.; Petrenko, V.I.; Gapon, I.; Avdeev, M.; Bulavin, L.; Timko, M.; Kopcansky, P. (Journal of Molecular Liquids, 2022)

Electrode Kinetics of Ion Jelly and Ion Sol-Gel Redox Materials on Screen-Printed Electrodes - R.N.H. Carvalho; Cordas, C. M.; Fonseca, L. P. (Appl Sci, 2022)

Emergent solution based IGZO memristor towards neuromorphic applications - https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9241358/ (Materials Chemistry C, 2022)

Empowering a Relational Database with LSD: Lazy State Determination - Thales Parreira (FCT-NOVA, 2022)

Encapsulation of the septal cell wall protects Streptococcus pneumoniae from its major peptidoglycan hydrolase and host defenses - Figueiredo, J; M. X. Henriques; M. J. Catalão; Pinheiro, S.; A. R. Narciso; Mesquita, F.; B.M. Saraiva; M. Carido; D. Cabanes; M. G. Pinho; S. R. Filipe (PLoS Pathogens, 2022)

Evaluation of Sweat Sampling Procedures for Human Stress Biomarkers Detection - Nunes, M. J.; Moura, J. J. G.; J. P. Noronha; Branco, L. C.; Samhan-Arias, A.; Sousa, J.P.; C. Rouco; Cordas, C. (Analytica, 2022)

Experimental investigation on punching shear behaviour of RC-(R)UHPFRC composite flat slabs without transverse reinforcement - Aurelio Sine; Mario Pimentel; Sandra Nunes (Engineering Structures, 2022)

Fabric-based supercapacitors toward wearable applications - Eduardo Oliveira (FCT Nova, 2022)

Flexible Active Crossbar Arrays Using Amorphous Oxide Semiconductor Technology toward Artificial Neural Networks Hardware - https://doi.org/10.1002/aelm.202200642 (Advanced Electronic Materials, 2022)

From cellulose-based waste to advanced materials - Sara Portugal Gonçalves (FCT Nova, 2022)

Functionalization of fibers and textiles suitable for electronic textiles - Inês Brandão Sequeira (FCT Nova, 2022)

How fullerenes inhibit the amyloid fibril formation of hen lysozyme - One-Sun Lee, O.-S.; Petrenko, V.I.; Siposova, K.; Musatov, A.; Park, H.; S. Lanceros-Méndez (Journal of Industrial and Engineering Chemistry, 2022)

Hybrid use of HPFRC in Slab-Column Connections under Cyclic Laterial Loading - António Pinho Ramos; Brisid Isufi; Rui Marreiros; Carla Marchão (fib Congress 2022, Oslo, 2022)

Imidazolium and picoline-based electrolytes for electrochemical reduction of CO2 at high pressure - S. Messias; V. Paz; Cruz, H.; C.M. Rangel; Branco, L. C.; A. S. Reis Machado (Energy Advances, 2022)

The impact of alternating and rotating regimes on the heating characteristics of magnetic bacterial cellulose structure - Molcan, M.; Safarik, I.; Prochazkova, J.; Kopcansky, P.; Timko, M.; Skumiel, A. (Journal of Magnetism and Magnetic Materials, 2022)

Improving Key-Value Database Scalability with LSD: Lazy State Determination - David Carpinteiro (FCT-NOVA, 2022)

The Influence of a Rotating Magnetic Field on the Thermal Effect in Magnetic Fluid - Skumiel, A.; Kocansky, P.; Timko, M.; Molcan, M.; Paulovicova, K.; Wojciechowski, R. (International Journal of Thermal Sciences, 2022)

The intriguing dose-dependent effect of selected amphiphilic compounds on insulin amyloid aggregation: Focus on a cholesterol-based detergent - Siposova, K.; Petrenko, V.I.; Garcarova, I.; Sedlakova, D.; Almasy, L.; Kyzyma, O.A.; Kriechbaum, M.; Musatov, A. (Chobimalt, 2022)

Iron-Sulfur centers: Functions of an ancient metal site - Pauleta, S. R.; Carepo, M.; Grazina, R.; Moura, I.; Moura, J. J. G. (Comprehensive Inorganic Chemistry III From Biology to Nanotechnology, vol. 2, 2022)

Nanoscale Events on Cyanobiphenyl-Based Self-Assembled Droplets Triggered by Gas Analytes - Ramou, E.; S. I. C. J. Palma; Roque, A. C. A. (ACS Applied Materials and Interfaces, 2022)

Native Protein Template Assisted Synthesis of Non-Native Metal-Sulfur Clusters - Maiti B.K.; Moura J.J.G. (BioChem, 2022)

Observation of Grain Boundary Passivation and Charge Distribution in Perovskite Films Improved with Anti-solvent Treatment - Shrabani Panigrahi; Calmeiro, Tomás; Manuel Joao Mendes; Hugo Águas; Elvira Fortunato; Rodrigo Martins (Journal of Physical Chemistry C, 2022)

OSCAR — A Noise Injection Framework for Testing Concurrent Software - Filipe de Luna (FCT-NOVA, 2022)

Photonic-Structured Perovskite Solar Cells: Detailed Optoelectronic Analysis - Haque, Sirazul; Alexandre, Miguel; Clemens Baretzky; Daniele Rossi; Francesca De Rossi; António T. Vicente; Francesca Brunetti; Hugo Águas; Rute A. S. Ferreira; Elvira Fortunato; Matthias Auf Der Maur; Uli Wurfel; et al. (ACS Photonics, 2022)

Rational Use of FPFRC in Slab-Connections Under Reversed Horizontal Cyclic Loading - António Ramos; Brisid Isufi; Rui Marreiros; Mária Bolesova; Katarina Gajdsova (Engineering Structures, 2022)
Slab – column connections that are subjected to combined gravity and horizontal loading during an earthquake are prone to premature failure due to punching shear. Traditional solutions to avoid punching failure and to increase the displacement capacity of this type of connection include using stirrups and double-headed studs as shear reinforcement. The use of High-Performance Fiber Reinforced Concrete (HPFRC) in a small region of the slab around the column as a substitute for traditional solutions is investigated in this paper, because this material has the potential to reduce labor and material costs. To fulfill this objective, four slab specimens with a thickness of 150 mm were tested under combined gravity and reversed horizontal drifts. The results are discussed in detail. The experimental variables considered were the top flexural reinforcement ratio, the size of the HPFRC zone and the intensity of the gravity load. Previously published tests that serve as reference specimens are used to compare the results. The behavior of the specimens with HPFRC was substantially improved compared to the reference specimens in terms of drift capacity: from only 1.0% drift to above 5.5%, even though a very small quantity of HPFRC was used, extended up to only 1.5 times the effective depth of the slab from the face of the column. Specimens with HPFRC also behaved better when compared to specimens with High-Strength Concrete (HSC). Side effects of using HPFRC in the slab in the vicinity of the column include an increase of the unbalanced moment transfer capacity and lateral stiffness, as well as a reduction of the deflections of the slab.

Reduction Kinetic of Water Soluble Metal Salts by Geobacter sulfurreducens: Fe2+/Hemes Stabilize and Regulate Electron Flux Rates - Karamash, Maksym; Stumpe, Michael; Dengjel, Jörn; Salgueiro, , Carlos A.; Giese, Bernd; Fromm, Katharina M (Frontiers in Microbiology, 2022)
Geobacter sulfurreducens is a widely applied microorganism for the reduction of toxic metal salts, as an electron source for bioelectrochemical devices, and as a reagent for the synthesis of nanoparticles. In order to understand the influence of metal salts, and of electron transporting, multiheme c-cytochromes on the electron flux during respiration of G. sulfurreducens, the reduction kinetic of Fe3+, Co3+, V5+, Cr6+, and Mn7+ containing complexes were measured. Starting from the resting phase, each G. sulfurreducens cell produced an electron flux of 3.7 × 105 electrons per second during the respiration process. Reduction rates were within ± 30% the same for the 6 different metal salts, and reaction kinetics were of zero order. Decrease of c-cytochrome concentrations by downregulation and mutation demonstrated that c-cytochromes stabilized respiration rates by variation of their redox states. Increasing Fe2+/heme levels increased electron flux rates, and induced respiration flexibility. The kinetic effects parallel electrochemical results of G. sulfurreducens biofilms on electrodes, and might help to optimize bioelectrochemical devices.
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Screen‐Printed Electrodes Testing for Detection of Potential Stress Biomarkers in Sweat - Nunes M.J.; Valério G.N.; Samhan‐Arias A.; Moura J.J.G.; Rouco C.; Sousa, J.P.; Cordas C.M. (Electrocatalysis, 2022)

Slab – column connection punching and ductility improvement methods for seismic response of buildings with flat slabs - Brisid Isufi; André Almeida; Rui Marreiros; António Ramos; Válter Lúcio (Structural Concrete, 2022)

Structure and Biological Activity of Particles Produced from Highly Activated Carbon Adsorbent - Sarnatskaya, V.; Shlapa, Yu.; Lykhova, A.; Brieieva, O.; Prokopenko, I.; Sidorenko, A.; Solopan, S.; Kolesnik, D.; Belous, A.; Nikolaev, V. (Heliyon, 2022)

Sulfide and transition metals - A partnership for life - B.K. Maiti; Maia, L. B.; Moura, J. J. G. (J Inorg Biochem, 2022)

Tailoring the synaptic properties of a-IGZO memristors for artificial deep neural networks - https://aip.scitation.org/doi/full/10.1063/5.0073056 (APL material, 2022)

Utilização Racional de Betões de Alta Resistência Reforçados com Fibras de Aço em Lajes Fungiformes - João Pedro Relvas (NOVA School of Science and Technology, 2022)

“Manufacturing Life” in Real Work Processes? New Manufacturing Environments with Micro and Nanorobotics - António Brandão Moniz; Bettina-Johanna Krings (NanoEthics, 2022)

Advances in Membrane-Bound Catechol-O-Methyltransferase Stability Achieved Using a New Ionic Liquid-Based Storage Formulation - Gonçalves, Ana M.; Sousa, Ângela; Pedro, Augusto Q.; Romão, Maria J.; João A. Queiroz; Gallardo, Eugénia; Passarinha, Luís A. (International Journal of Molecular Sciences, 2022)
Membrane-bound catechol-O-methyltransferase (MBCOMT), present in the brain and involved in the main pathway of the catechol neurotransmitter deactivation, is linked to several types of human dementia, which are relevant pharmacological targets for new potent and nontoxic inhibitors that have been developed, particularly for Parkinson’s disease treatment. However, the inexistence of an MBCOMT 3D-structure presents a blockage in new drugs’ design and clinical studies due to its instability. The enzyme has a clear tendency to lose its biological activity in a short period of time. To avoid the enzyme sequestering into a non-native state during the downstream processing, a multi-component buffer plays a major role, with the addition of additives such as cysteine, glycerol, and trehalose showing promising results towards minimizing hMBCOMT damage and enhancing its stability. In addition, ionic liquids, due to their virtually unlimited choices for cation/anion paring, are potential protein stabilizers for the process and storage buffers. Screening experiments were designed to evaluate the effect of distinct cation/anion ILs interaction in hMBCOMT enzymatic activity. The ionic liquids: choline glutamate [Ch][Glu], choline dihydrogen phosphate ([Ch][DHP]), choline chloride ([Ch]Cl), 1- dodecyl-3-methylimidazolium chloride ([C12mim]Cl), and 1-butyl-3-methylimidazolium chloride ([C4mim]Cl) were supplemented to hMBCOMT lysates in a concentration from 5 to 500 mM. A major potential stabilizing effect was obtained using [Ch][DHP] (10 and 50 mM). From the DoE 146% of hMBCOMT activity recovery was obtained with [Ch][DHP] optimal conditions (7.5 mM) at −80 °C during 32.4 h. These results are of crucial importance for further drug development once the enzyme can be stabilized for longer periods of time.
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Analysis of the In Vitro Toxicity of Nanocelluloses in Human Lung Cells as Compared to Multi-Walled Carbon Nanotubes - Pinto, Fátima; Lourenço, Ana Filipa; Pedrosa, Jorge F. S.; Lídia Gonçalves; Ventura, Célia; Vital, Nádia; Ana Bettencourt; Fernandes, Susete N.; da Rosa, Rafaela R.; Godinho, Maria Helena; Louro, Henriqueta; Ferreira, Paulo J. T.; et al. (Nanomaterials, 2022)
Cellulose micro/nanomaterials (CMNM), comprising cellulose microfibrils (CMF), nanofibrils (CNF), and nanocrystals (CNC), are being recognized as promising bio-nanomaterials due to their natural and renewable source, attractive properties, and potential for applications with industrial and economical value. Thus, it is crucial to investigate their potential toxicity before starting their production at a larger scale. The present study aimed at evaluating the cell internalization and in vitro cytotoxicity and genotoxicity of CMNM as compared to two multi-walled carbon nanotubes (MWCNT), NM-401 and NM-402, in A549 cells. The exposure to all studied NM, with the exception of CNC, resulted in evident cellular uptake, as analyzed by transmission electron microscopy. However, none of the CMNM induced cytotoxic effects, in contrast to the cytotoxicity observed for the MWCNT. Furthermore, no genotoxicity was observed for CNF, CNC, and NM-402 (cytokinesis-block micronucleus assay), while CMF and NM-401 were able to significantly raise micronucleus frequency. Only NM-402 was able to induce ROS formation, although it did not induce micronuclei. Thus, it is unlikely that the observed CMF and NM-401 genotoxicity is mediated by oxidative DNA damage. More studies targeting other genotoxicity endpoints and cellular and molecular events are underway to allow for a more comprehensive safety assessment of these nanocelluloses.
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Editorial: Rational drug design of metal complexes for cancer therapy - Morais, Tânia S.; Fernandes, Alexandra R; Baptista, Pedro V; Gambino, Dinorah (Frontiers in Chemistry, 2022)
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info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F00100%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/LA%2FP%2F0056%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/LA%2FP%2F0140%2F2020/PT# info:eu-repo/grantAgreement/FCT/Concurso para Financiamento de Projetos de Investigação Científica e Desenvolvimento Tecnológico em Todos os Domínios Científicos - 2020/PTDC%2FQUI-QIN%2F0146%2F2020/PT# info:eu-repo/grantAgreement/FCT/CEEC IND 2017/CEECIND%2F00630%2F2017%2FCP1387%2FCT0012/PT#
Keywords: cancer chemotherapy; Drug delivery; inorganic/organometallic; mechanism of action; Medicinal chemistry; metal-based drug design; Nanomedicine

Electron Flow From the Inner Membrane Towards the Cell Exterior in Geobacter sulfurreducens: Biochemical Characterization of Cytochrome CbcL - Antunes, Jorge M. A.; Silva, , Marta A.; Salgueiro, , Carlos A.; Leonor Morgado (Frontiers in Microbiology, 2022)
Exoelectrogenic microorganisms are in the spotlight due to their unique respiratory mechanisms and potential applications in distinct biotechnological fields, including bioremediation, bioenergy production and microbial electrosynthesis. These applications rely on the capability of these microorganisms to perform extracellular electron transfer, a mechanism that allows the bacteria to transfer electrons to the cell’s exterior by establishing functional interfaces between different multiheme cytochromes at the inner membrane, periplasmic space, and outer membrane. The multiheme cytochrome CbcL from Geobacter sulfurreducens is associated to the inner membrane and plays an essential role in the transfer of electrons to final electron acceptors with a low redox potential, as Fe(III) oxides and electrodes poised at −100 mV. CbcL has a transmembranar di-heme b-type cytochrome domain with six helices, linked to a periplasmic cytochrome domain with nine c-type heme groups. The complementary usage of ultraviolet-visible, circular dichroism and nuclear magnetic resonance permitted the structural and functional characterization of CbcL’s periplasmic domain. The protein was found to have a high percentage of disordered regions and its nine hemes are low-spin and all coordinated by two histidine residues. The apparent midpoint reduction potential of the CbcL periplasmic domain was determined, suggesting a thermodynamically favorable transfer of electrons to the putative redox partner in the periplasm − the triheme cytochrome PpcA. The establishment of a redox complex between the two proteins was confirmed by probing the electron transfer reaction and the molecular interactions between CbcL and PpcA. The results obtained show for the first time how electrons are injected into the periplasm of Geobacter sulfurreducens for subsequent transfer to the cell’s exterior.
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From iron to bacterial electroconductive filaments: Exploring cytochrome diversity using Geobacter bacteria - Salgueiro, , Carlos A.; Leonor Morgado; Silva, , Marta A.; Ferreira, Marisa R.; Fernandes, Tomás M.; Portela, Pilar C. (Coordination Chemistry Reviews, 2022)
Iron is the most versatile of all biochemically active metals, with variability encompassing its electronic configuration, number of unpaired electrons, type of ligands and iron-complexes stability. The versatility of iron properties is transposed to the proteins it can be associated to, especially relevant in the case of heme proteins. In this Review, the structural and functional properties of heme proteins are revisited, with particular focus on c-type cytochromes. The genome of Geobacter bacteria encodes for an unusually high number of assorted c-type cytochromes and, for this reason, they are used in this Review as a showcase of the cytochrome diversity. In the last decades, a vast portfolio of cytochromes has been revealed in these bacteria, with most of them defining new classes, ranging from monoheme to the recently identified polymeric assembly of multiheme cytochromes that forms micrometer-long electrically conductive filaments. These discoveries were on pace with the development of modern NMR equipment and advances in protein isotopic labeling methods, which are also revisited in this Review. Finally, following the description of the current state of the art of Geobacter cytochromes, examples on how the available structural and functional information was explored to structurally map protein–protein and protein–ligand interacting regions in redox complexes, and hence elucidate Geobacter’s respiratory pathways, are presented.
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Keywords: Bacterial electroconductive nanowires; Cytochrome; Heme group; Iron atom; Structure–function relationship

Genotoxicity of Three Micro/Nanocelluloses with Different Physicochemical Characteristics in MG-63 and V79 Cells - Ventura, Célia; Marques, Catarina; Cadete, João; Vilar, Madalena; Pedrosa, Jorge F. S.; Pinto, Fátima; Fernandes, Susete Nogueira; da Rosa, Rafaela Raupp; Godinho, Maria Helena; Ferreira, Paulo J. T.; Louro, Henriqueta; Silva, Maria João (Journal of Xenobiotics, 2022)
(1) Background: Nanocellulose is an innovative engineered nanomaterial with an enormous potential for use in a wide array of industrial and biomedical applications and with fast growing economic value. The expanding production of nanocellulose is leading to an increased human exposure, raising concerns about their potential health effects. This study was aimed at assessing the potential toxic and genotoxic effects of different nanocelluloses in two mammalian cell lines; (2) Methods: Two micro/nanocelluloses, produced with a TEMPO oxidation pre-treatment (CNFs) and an enzymatic pre-treatment (CMFs), and cellulose nanocrystals (CNCs) were tested in osteoblastic-like human cells (MG-63) and Chinese hamster lung fibroblasts (V79) using the MTT and clonogenic assays to analyse cytotoxicity, and the micronucleus assay to test genotoxicity; (3) Results: cytotoxicity was observed by the clonogenic assay in V79 cells, particularly for CNCs, but not by the MTT assay; CNF induced micronuclei in both cell lines and nucleoplasmic bridges in MG-63 cells; CMF and CNC induced micronuclei and nucleoplasmic bridges in MG-63 cells, but not in V79 cells; (4) Conclusions: All nanocelluloses revealed cytotoxicity and genotoxicity, although at different concentrations, that may be related to their physicochemical differences and availability for cell uptake, and to differences in the DNA damage response of the cell model.
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Molecular geometries of the heme axial ligands from the triheme cytochrome PpcF from Geobacter metallireducens reveal a conserved heme core architecture - Ferreira, Marisa R.; Fernandes, Tomás M.; Turner, David L.; Salgueiro, , Carlos A. (Archives of Biochemistry and Biophysics, 2022)
Electroactive Geobacter bacteria can perform extracellular electron transfer and present a wide metabolic versatility. These bacteria reduce organic, toxic and radioactive compounds, and produce electric current while interacting with electrodes, making them interesting targets for numerous biotechnological applications. Their global electrochemical responses rely on an efficient interface between the inside and the cell's exterior, which is driven by the highly abundant periplasmic triheme PpcA-family cytochromes. The functional features of these cytochromes have been studied in G. sulfurreducens and G. metallireducens, and although they share a high degree of structural homology and sequence identity, their properties are quite distinct. In this work, the heme axial ligand geometries and the magnetic properties of PpcF from G. metallireducens were determined. The data obtained constitute important constraints for the determination of its solution structure in the oxidized state and indicate that the (i) heme core architecture; (ii) axial ligands geometries and (iii) magnetic properties of the cytochrome are conserved compared to the other members of the PpcA-families. Furthermore, the results also indicate that the heme arrangement is crucial to maintain an intrinsic regulation of the protein's redox properties and hence its electron transfer efficiency and functionality.
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Keywords: Multiheme cytochromes; Nuclear magnetic resonance; Paramagnetic redox centers

Nitrate Removal by Donnan Dialysis and Anion-Exchange Membrane Bioreactor Using Upcycled End-of-Life Reverse Osmosis Membranes - Lejarazu-Larrañaga, Amaia; Ortiz, Juan M.; Molina, Serena; Sylwin Pawlowski; Claudia F. Galinha; Otero, Vanessa; García-Calvo, Eloy; Velizarov, Svetlozar; João G. Crespo (Membranes, 2022)
This work explores the application of Reverse Osmosis (RO) upcycled membranes, as Anion Exchange Membranes (AEMs) in Donnan Dialysis (DD) and related processes, such as the Ion Exchange Membrane Bioreactor (IEMB), for the removal of nitrate from contaminated water, to meet drinking water standards. Such upcycled membranes might be manufactured at a lower price than commercial AEMs, while their utilization reinforces the commitment to a circular economy transition. In an effort to gain a better understanding of such AEMs, confocal µ-Raman spectroscopy was employed, to assess the distribution of the ion-exchange sites through the thickness of the prepared membranes, and 2D fluorescence spectroscopy, to evaluate alterations in the membranes caused by fouling and chemical cleaning The best performing membrane reached a 56% average nitrate removal within 24 h in the DD and IEMB systems, with the latter furthermore allowing for simultaneous elimination of the pollutant by biological denitrification, thus avoiding its discharge into the environment. Overall, this work validates the technical feasibility of using RO upcycled AEMs in DD and IEMB processes for nitrate removal. This membrane recycling concept might also find applications for the removal and/or recovery of other target negatively charged species.
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A solvent‐free strategy to prepare amorphous salts of folic acid with enhanced solubility and cell permeability - Martins, Inês C. B.; Forte, Andreia; Diogo, Hermínio P.; Raposo, Luís R.; Baptista, Pedro V; Fernandes, Alexandra R; Branco, Luís C.; Duarte, M. Teresa (Chemistry–Methods, 2022)
Eight new amorphous organic salts of folic acid (FA) were prepared by mechanochemistry. FA can prevent cardiovascular and neurological diseases. Mechanochemistry overcomes serious FA solubility issues avoiding the use of toxic solvents. Due to low FA solubility, therapeutic effects in supplements and drugs are not achieved. Current strategies to improve FA solubility include its derivatization by using complex synthetic procedures. Herein, a simple and green procedure, avoiding structural modifications, was designed using mechanochemistry. Biocompatible amine-derivative coformers were strategically combined with FA to obtain salts with good physicochemical properties. New 1 : 1 and 1 : 2 amorphous FA salts offer 10 to 10,000 times better aqueous solubility and 10 to 100 times better octanol-water partition coefficient values (Koctanol/water) than FA alone. Koctanol/water is considered as a surrogate of cell permeability. No toxic effects in normal human primary dermal fibroblasts were detected for the prepared FA salts. Our findings suggest that 1 : 2 FA salts of choline hydroxide and derivatives could be good candidates for future pharmaceutical/nutraceutical applications.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UID%2FQUI%2F00100%2F2013/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/PEst-OE%2FQUI%2FUI0100%2F2013/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F93140%2F2013/PT#

Synergy between silk fibroin and ionic liquids for active gas-sensing materials - Inês P. Moreira; Carina Esteves; Susana I C J Palma; Ramou, Efthymia; Ana L.M. Carvalho; Roque, Ana C A (Materials Today Bio, 2022)
Silk fibroin is a biobased material with excellent biocompatibility and mechanical properties, but its use in bioelectronics is hampered by the difficult dissolution and low intrinsic conductivity. Some ionic liquids are known to dissolve fibroin but removed after fibroin processing. However, ionic liquids and fibroin can cooperatively give rise to functional materials, and there are untapped opportunities in this combination. The dissolution of fibroin, followed by gelation, in designer ionic liquids from the imidazolium chloride family with varied alkyl chain lengths (2–10 carbons) is shown here. The alkyl chain length of the anion has a large impact on fibroin secondary structure which adopts unconventional arrangements, yielding robust gels with distinct hierarchical organization. Furthermore, and due to their remarkable air-stability and ionic conductivity, fibroin ionogels are exploited as active electrical gas sensors in an electronic nose revealing the unravelled possibilities of fibroin in soft and flexible electronics.
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Keywords: Bioelectronics; gas sensing; Ionic conductivity; ionic liquids; Physical ionogels; Silk fibroin

Tackling Humidity with Designer Ionic Liquid-Based Gas Sensing Soft Materials - Carina Esteves; Susana I C J Palma; Henrique M.A. Costa; Cláudia Alves; Gonçalo M.C. Santos; Ramou, Efthymia; Ana Luísa Carvalho; Vitor Alves; Roque, Ana C A (Advanced Materials, 2022)
Abstract Relative humidity is simultaneously a sensing target and a contaminant in gas and volatile organic compound (VOC) sensing systems, where strategies to control humidity interference are required. An unmet challenge is the creation of gas-sensitive materials where the response to humidity is controlled by the material itself. Here, humidity effects are controlled through the design of gelatin formulations in ionic liquids without and with liquid crystals as electrical and optical sensors, respectively. In this design, the anions [DCA]− and [Cl]− of room temperature ionic liquids from the 1-butyl-3-methylimidazolium family tailor the response to humidity and, subsequently, sensing of VOCs in dry and humid conditions. Due to the combined effect of the materials formulations and sensing mechanisms, changing the anion from [DCA]− to the much more hygroscopic [Cl]−, leads to stronger electrical responses and much weaker optical responses to humidity. Thus, either humidity sensors or humidity-tolerant VOC sensors that do not require sample preconditioning or signal processing to correct humidity impact are obtained. With the wide spread of 3D- and 4D-printing and intelligent devices, the monitoring and tuning of humidity in sustainable biobased materials offers excellent opportunities in e-nose sensing arrays and wearable devices compatible with operation at room conditions.
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Keywords: anion-tunability; gas sensing; gelatin; humidity; ionogels; liquid crystals; methylimidazolium ionic liquids

Advanced porous materials from poly(ionic liquid)s: Challenges, applications and opportunities - Barrulas, Raquel V; Zanatta, Marcileia; Casimiro, Teresa; Corvo, Marta C. (CHEMICAL ENGINEERING JOURNAL, 2021)
{Over the past few years porous materials have become a topic of intense research. Porous poly(ionic liquid)s combine the porous architecture with intrinsic ionic liquids properties. In all research areas, the quest for new and improved materials has targeted functional materials with enhanced specificity and efficiency towards the final application. The application of porous materials ranges from sensing, protein separation, solid-phase extraction, catalysis, to CO2 capture and reuse. Recently, the design, synthesis, and porosity control of poly (ionic liquid)s have been attempted through strategies that include classic polymerization techniques as well as molecular imprinting and aerogels production. This review aims at providing the recent advances on porous poly (ionic liquid)s, giving a critical perspective about the works in which key requirements for porosity induction are discussed. Several applications that rely on molecular interactions between the porous material and target compounds are presented, focusing mainly on CO2 capture and reuse, along with some challenges that the scientific community in this field need to be aware of.}

Ibuprofen incorporated into unmodified and modified mesoporous silica: From matrix synthesis to drug release - Inocencio, Sara; Cordeiro, Teresa; Matos, Ines; Danede, Florence; Sotomayor, Joao C.; Fonseca, Isabel M.; Correia, Natalia T.; Corvo, Marta C.; Dionisio, Madalena (MICROPOROUS AND MESOPOROUS MATERIALS, 2021)
{Aiming to rationalize the release profile of an incorporated pharmaceutical drug in terms of its mobility, driven by guest-host interactions, the poorly water-soluble ibuprofen drug was loaded in a mesoporous inorganic silica matrix with unmodified (MCM-41) and modified surface (MCM-41sil) by post-synthesis silylation, both having pore sizes similar to 3 nm. The single calorimetric detection of a broad glass transition step for both ibuprofen com-posites indicates full drug amorphization, confirmed by the only appearance of an amorphous halo in the powder XRD patterns. Moreover, a gradient profile is disclosed by the heat flux derivative plot in the glass transition, in coherence with the thermogravimetric profile that shows a multi-step decomposition trace for confined ibuprofen in these matrixes. While identical guest dynamics, as probed by dielectric relaxation spectroscopy, were found in both dehydrated composites, a significant molecular population with faster relaxation exists in the hydrated state for the drug inside the unmodified matrix. This was rationalized as the concurrence of true confinement effects, which manifest under nanometer dimensions, and greater water affinity of the unmodified matrix, forcing the drug molecules to be placed mostly in the pore core. Finite size effects are also felt in both dehydrated composites, however guest-host interactions give origin to a dominant population with slowed down mobility that governs the overall guest dynamics. In spite of an inferior number of active sites for drug adsorption in the silylated matrix, a faster ibuprofen delivery in phosphate buffer (pH = 6.8) was observed when the drug is released from unmodified MCM-41 in the hydrated state. Therefore, our results suggest that a relevant role is played by water molecules, which impair a strong guest adsorption in the host surface more efficiently than the limited surface modification, influence the higher ratio of a faster population in the pore core and facilitate the diffusion of the aqueous releasing media inside pores.}
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Tuning the H-1 NMR Paramagnetic Relaxation Enhancement and Local Order of {[}Aliquat](+)-Based Systems Mixed with DMSO - Cordeiro, Rui; Beira, Maria J.; Cruz, Carlos; Figueirinhas, Joao L.; Corvo, Marta C.; Almeida, Pedro L; Rosatella, Andreia A.; Afonso, Carlos A. M.; Daniel, Carla I.; Sebastiao, Pedro J. (International Journal of Molecular Sciences, 2021)
{Understanding the behavior of a chemical compound at a molecular level is fundamental, not only to explain its macroscopic properties, but also to enable the control and optimization of these properties. The present work aims to characterize a set of systems based on the ionic liquids {[}Aliquat]{[}Cl] and {[}Aliquat]{[}FeCl4] and on mixtures of these with different concentrations of DMSO by means of H-1 NMR relaxometry, diffusometry and X-ray diffractometry. Without DMSO, the compounds reveal locally ordered domains, which are large enough to induce order fluctuation as a significant relaxation pathway, and present paramagnetic relaxation enhancement for the {[}Aliquat]{[}Cl] and {[}Aliquat]{[}FeCl4] mixture. The addition of DMSO provides a way of tuning both the local order of these systems and the relaxation enhancement produced by the tetrachloroferrate anion. Very small DMSO volume concentrations (at least up to 1%) lead to enhanced paramagnetic relaxation without compromising the locally ordered domains. Larger DMSO concentrations gradually destroy these domains and reduce the effect of paramagnetic relaxation, while solvating the ions present in the mixtures. The paramagnetic relaxation was explained as a correlated combination of inner and outer-sphere mechanisms, in line with the size and structure differences between cation and anion. This study presents a robust method of characterizing paramagnetic ionic systems and obtaining a consistent analysis for a large set of samples having different co-solvent concentrations.}
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Combined cancer therapeutics—Tackling the complexity of the tumor microenvironment - Catarina Roma-Rodrigues; Raposo, Luís R.; Valente, Rúben; Fernandes, Alexandra R; Baptista, Pedro V (Wiley Interdisciplinary Reviews: Nanomedicine and Nanobiotechnology, 2021)
Cancer treatment has yet to find a “silver bullet” capable of selectively and effectively kill tumor cells without damaging healthy cells. Nanomedicine is a promising field that can combine several moieties in one system to produce a multifaceted nanoplatform. The tumor microenvironment (TME) is considered responsible for the ineffectiveness of cancer therapeutics and the difficulty in the translation from the bench to bed side of novel nanomedicines. A promising approach is the use of combinatorial therapies targeting the TME with the use of stimuli-responsive nanomaterials which would increase tumor targeting. Contemporary combined strategies for TME-targeting nanoformulations are based on the application of external stimuli therapies, such as photothermy, hyperthermia or ultrasounds, in combination with stimuli-responsive nanoparticles containing a core, usually composed by metal oxides or graphene, and a biocompatible stimuli-responsive coating layer that could also contain tumor targeting moieties and a chemotherapeutic agent to enhance the therapeutic efficacy. The obstacles that nanotherapeutics must overcome in the TME to accomplish an effective therapeutic cargo delivery and the proposed strategies for improved nanotherapeutics will be reviewed. This article is categorized under: Therapeutic Approaches and Drug Discovery > Nanomedicine for Oncologic Disease Nanotechnology Approaches to Biology > Nanoscale Systems in Biology Therapeutic Approaches and Drug Discovery > Emerging Technologies.
Fundacao para a Ciencia e a Tecnologia, Grant/Award Numbers: SFRH/BPD/124612/2016, UIDB/04378/2020
Keywords: Cancer; cancer therapeutics; Nanomedicine; nanoparticles; Tumor microenvironment

The Effect of Dicarboxymethyl Cellulose on the Prevention of Protein Haze Formation on White Wine - Gago, Diana; Chagas, Ricardo; Ferreira, Luisa M. (BEVERAGES, 2021)
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New directions for inline inspection of automobile laser welds using non-destructive testing - Machado, Miguel Araújo; Rosado, Luís Filipe Soldado Granadeiro; Mendes, Nuno Alberto Marques; Miranda, Rosa Maria Mendes; dos Santos, Telmo Jorge Gomes (The International Journal of Advanced Manufacturing Technology, 2021)
An innovative pilot installation and eddy current testing (ECT) inspection system for laser-brazed joints is presented. The proposed system detects both surface and sub-surface welding defects operating autonomously and integrated with a robotized arm. Customized eddy current probes were designed and experimentally validated detecting pore defects with 0.13 mm diameter and sub-surface defects buried 1 mm deep. The integration of the system and the manufacturing process towards an Industry 4.0 quality control paradigm is also discussed.
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Modelling CO2 absorption in aqueous solutions of cholinium lysinate ionic liquid - Martins, C. F.; Neves, L. A.; Chagas, R.; Ferreira, LM; C. A. M. Afonso; Coelhoso, I. M.; Crespo, J. G.; Mota, J. PB. (CHEMICAL ENGINEERING JOURNAL, 2021)
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DCMC as a Promising Alternative to Bentonite in White Wine Stabilization. Impact on Protein Stability and Wine Aromatic Fraction - Saracino, Francesco; Brinco, Joao; Gago, Diana; Gomes da Silva, Marco; Ferreira, Ricardo Boavida; Ricardo-da-Silva, Jorge; Chagas, Ricardo; Ferreira, Luisa Maria (MOLECULES, 2021)
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Inflammatory Factors, Genetic Variants and Predisposition for Preterm Birth - Couceiro, Joana; Matos, Irina; Mendes, José João; Baptista, Pedro V; Fernandes, Alexandra R; Quintas, Alexandre (Clinical Genetics, 2021)
Preterm birth is a major clinical and public health challenge, with a prevalence of 11% worldwide. It is the leading cause of death in children younger than five years old and represents 70% of neonatal deaths and 75% of neonatal morbidity. Despite the clinical and public health significance, this condition's aetiology is still unclear, and most of the cases are spontaneous. There are several known preterm birth risk factors, including inflammatory diseases and the genetic background, although the underlying molecular mechanisms are far from understood. The present review highlights the research advances on the association between inflammatory-related genes and the increased risk for preterm delivery. The most associated genetic variants are the TNFα rs1800629, the IL1α rs17561, and the IL1RN rs2234663. Moreover, many of the genes discussed in this review are also implicated in pathologies involving inflammatory or autoimmune systems, such as periodontal disease, bowel inflammatory disease, and autoimmune rheumatic diseases. This review presents evidence suggesting a common genetic background to preterm birth, autoimmune and inflammatory diseases susceptibility. This article is protected by copyright. All rights reserved.
Egas Moniz Cooperativa de Ensino Superior, CRL, Grant/Award Number: CEI/11/2018; Fundacao para a Ciencia e a Tecnologia, Grant/Award Numbers: UIDB/04378/2020, UIDP/04378/2020

Vanadium(IV) complexes with methyl-substituted 8-hydroxyquinolines: Catalytic potential in the oxidation of hydrocarbons and alcohols with peroxides and biological activity - Joanna Palion-Gazda; Luz, André; Raposo, Luis R.; Choroba, Katarzyna; Nycz, Jacek E.; Alina Bieńko; Agnieszka Lewińska; Erfurt, Karol; Baptista, Pedro V; Machura, Barbara; Fernandes, Alexandra R; Shul’pina, Lidia S.; et al. (Molecules, 2021)
Methyl-substituted 8-hydroxyquinolines (Hquin) were successfully used to synthetize five-coordinated oxovanadium(IV) complexes: [VO(2,6-(Me)2-quin)2 ] (1), [VO(2,5-(Me)2-quin)2 ] (2) and [VO(2-Me-quin)2 ] (3). Complexes 1–3 demonstrated high catalytic activity in the oxidation of hydrocarbons with H2 O2 in acetonitrile at 50◦ C, in the presence of 2-pyrazinecarboxylic acid (PCA) as a cocatalyst. The maximum yield of cyclohexane oxidation products attained was 48%, which is high in the case of the oxidation of saturated hydrocarbons. The reaction leads to the formation of a mixture of cyclohexyl hydroperoxide, cyclohexanol and cyclohexanone. When triphenylphosphine is added, cyclohexyl hydroperoxide is completely converted to cyclohexanol. Consideration of the regioand bond-selectivity in the oxidation of n-heptane and methylcyclohexane, respectively, indicates that the oxidation proceeds with the participation of free hydroxyl radicals. The complexes show moderate activity in the oxidation of alcohols. Complexes 1 and 2 reduce the viability of colorectal (HCT116) and ovarian (A2780) carcinoma cell lines and of normal dermal fibroblasts without showing a specific selectivity for cancer cell lines. Complex 3 on the other hand, shows a higher cytotoxicity in a colorectal carcinoma cell line (HCT116), a lower cytotoxicity towards normal dermal fibroblasts and no effect in an ovarian carcinoma cell line (order of magnitude HCT116 > fibroblasts > A2780).
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# Nos. 19-03-00142 RFMEFI61917X0007 State Task 0082-2014-0007 (CITIS): AAAA-A17-117040610283-3 LA/P/0140/2020
Keywords: 8-hydroxyquinoline; biological activity; Catalytic properties; Cytotoxicity studies; Vanadium(IV) complexes

Removing CO2 from Xenon anaesthesia circuits using an amino-acid ionic liquid solution in a membrane contactor - Martins, C. F.; Neves, L. A.; Chagas, R.; Ferreira, LM; Coelhoso, I. M.; Crespo, J. G. (SEPARATION AND PURIFICATION TECHNOLOGY, 2021)
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Double active transient thermography - Miguel A. Machado; Silva, Maria I.; Martins, Ana P.; Carvalho, Marta S.; Telmo G. Santos (NDT & E International, 2021)
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Genetic biomarkers in chronic myeloid leukemia: What have we learned so far? - Abdulmawjood, Bilal; Costa, Beatriz; Catarina Roma-Rodrigues; Baptista, Pedro V; Fernandes, Alexandra R (International Journal of Molecular Sciences, 2021)
Chronic Myeloid Leukemia (CML) is a rare malignant proliferative disease of the hematopoietic system, whose molecular hallmark is the Philadelphia chromosome (Ph). The Ph chromosome originates an aberrant fusion gene with abnormal kinase activity, leading to the buildup of reactive oxygen species and genetic instability of relevance in disease progression. Several genetic abnormalities have been correlated with CML in the blast phase, including chromosomal aberrations and common altered genes. Some of these genes are involved in the regulation of cell apoptosis and proliferation, such as the epidermal growth factor receptor (EGFR), tumor protein p53 (TP53), or Schmidt-Ruppin A-2 proto-oncogene (SRC); cell adhesion, e.g., catenin beta 1(CTNNB1); or genes associated to TGF-β, such as SKI like proto-oncogene (SKIL), transforming growth factor beta 1 (TGFB1) or transforming growth factor beta 2 (TGFB2); and TNF-α pathways, such as Tumor necrosis factor (TNFA) or Nuclear factor kappa B subunit 1 (NFKB1). The involvement of miRNAs in CML is also gaining momentum, where dysregulation of some critical miRNAs, such as miRNA-451 and miRNA-21, which have been associated to the molecular modulation of pathogenesis, progression of disease states, and response to therapeutics. In this review, the most relevant genomic alterations found in CML will be addressed.
UIDP/04378/2020 UIDB/04378/2020 LA/P/0140/2020
Keywords: chronic myeloid leukemia; Genetic biomarkers; Genomic instability; MiRNAs; Philadelphia chromosome

High-speed inspection of delamination defects in unidirectional CFRP by non-contact eddy current testing - Miguel A. Machado; Kim-Niklas Antin; Rosado, Luís S.; Pedro Vilaça; Telmo G. Santos (Composites Part B: Engineering, 2021)
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Multisensor Inspection of Laser-Brazed Joints in the Automotive Industry - Miguel A. Machado; Rosado, Luís S.; Mendes, Nuno M.; Miranda, Rosa M.; Telmo G. Santos (Sensors, 2021)
Automobile laser brazing remains a complex process whose results are affected by several process variables that may result in nonacceptable welds. A multisensory customized inspection system is proposed, with two distinct non-destructive techniques: the potential drop method and eddy current testing. New probes were designed, simulated, produced, and experimentally validated in automobile's laser-brazed weld beads with artificially introduced defects. The numerical simulations allowed the development of a new four-point probe configuration in a non-conventional orthogonal shape demonstrating a superior performance in both simulation and experimental validation. The dedicated inspection system allowed the detection of porosities, cracks, and lack of bonding defects, demonstrating the redundancy and complementarity these two techniques provide.
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New non-toxic n-alkyl cholinium-based ionic liquids as excipients to improve the solubility of poorly water-soluble drugs - Jesus, Ana R.; Raposo, Luís R.; Soromenho, Mário R.C.; Agostinho, Daniela A. S.; Esperança, José M. S. S.; Baptista, Pedro V; Fernandes, Alexandra R; Reis, Patrícia M. (Symmetry, 2021)
In this work, we prepared new biocompatible N-alkyl cholinium-based ionic liquids to be used as cosolvents to improve the solubility of poorly water-soluble drugs, namely, sodium diclo-fenac and paracetamol. In this set of ionic liquids, we intend to understand the effect of increasing the asymmetry of the ionic liquid cation/anion by growing the length of one of the alkyl chains attached to the nitrogen center/sulfonate center on the dissolution capacity of the ionic liquid. The addition of these new ionic liquids to water increased the dissolution capacity of the drugs up to four-times that in water, and improved the pharmacodynamic properties of these drugs, especially the case of sodium diclofenac. The intermolecular interactions between the drugs and ionic liquids were investigated by NMR. Two-dimensional1H/1H nuclear overhauser effect spectroscopy (NO-ESY) revealed an interaction between sodium diclofenac and the alaninate anion from the [C2Ch]2[SucAla]. In the case of paracetamol and [C4Ch][C2SO3], it was possible to observe two inter-molecular interactions between the hydroxyl group of paracetamol and two protons from the cation [C4Ch]+. Interestingly, the ionic liquid bearing a succinyl-DL-alaninate anion, [SucAla]2−, and a N-ethyl cholinium cation, [C2Ch]+, which presented the highest ability to dissolve sodium diclofenac, showed no cytotoxicity up to 500 mM. Therefore, this ionic liquid is a potential candidate for drug delivery applications.
info:eu-repo/grantAgreement/FCT/Investigador FCT/IF%2F00621%2F2015%2FCP1284%2FCT0002/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F50006%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F50006%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FCVT-EPI%2F6685%2F2014/PT# LA/P/0140/2020 ROTEIRO/0031/2013—PINFRA/22161/2016
Keywords: Bioavailability; cytotoxicity; Drug delivery; ionic liquids; Oral drugs

New 3-Ethynylaryl Coumarin-Based Dyes for DSSC Applications: Synthesis, Spectroscopic Properties, and Theoretical Calculations - Sarrato, Joao; Pinto, Ana Lucia; Malta, Gabriela; Rock, Eva H.; Pina, Joao; Lima, Joao Carlos; Jorge Parola, A.; Branco, Paula S. (MOLECULES, 2021)
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Copper(ii) complexes with tridentate halogen-substituted Schiff base ligands: synthesis, crystal structures and investigating the effect of halogenation, leaving groups and ligand flexibility on antiproliferative activities - Kordestani, Nazanin; Amiri Rudbari, Hadi; Fernandes, Alexandra R; Raposo, Luís R.; Luz, André; Baptista, Pedro V; Bruno, Giuseppe; Scopelliti, Rosario; Fateminia, Zohreh; Micale, Nicola; Nikolay Tumanov; Johan Wouters; et al. (Dalton Transactions, 2021)
To investigate the effect of different halogen substituents and leaving groups and the flexibility of ligands on the anticancer activity of copper complexes, sixteen copper(ii) complexes with eight different tridentate Schiff-base ligands containing pyridine and 3,5-halogen-substituted phenol moieties were synthesized and characterized by spectroscopic methods. Four of these complexes were also characterized by X-ray crystallography. The cytotoxicity of the complexes was determined in three different tumor cell lines (i.e.the A2780 ovarian, HCT116 colorectal and MCF7 breast cancer cell line) and in a normal primary fibroblast cell line. Complexes were demonstrated to induce a higher loss of cell viability in the ovarian carcinoma cell line (A2780) with respect to the other two tumor cell lines, and therefore the biological mechanisms underlying this loss of viability were further investigated. Complexes with ligandL1(containing a 2-pycolylamine-type motif) were more cytotoxic than complexes withL2(containing a 2-(2-pyridyl)ethylamine-type motif). The loss of cell viability in A2780 tumor cells was observed in the orderCu(Cl2-L1)NO3>Cu(Cl2-L1)Cl>Cu(Br2-L1)Cl>Cu(BrCl-L1)Cl. All complexes were able to induce reactive oxygen species (ROS) that could be related to the loss of cell viability. ComplexesCu(BrCl-L1)ClandCu(Cl2-L1)NO3were able to promote A2780 cell apoptosis and autophagy and for complexCu(BrCl-L1)Clthe increase in apoptosis was due to the intrinsic pathway.Cu(Cl2-L1)ClandCu(Br2-L1)Clcomplexes lead to cellular detachment allowing to correlate with the results of loss of cell viability. Despite the ability of theCu(BrCl-L1)Clcomplex to induce programmed cell death in A2780 cells, its therapeutic window turned out to be low making theCu(Cl2-L1)NO3complex the most promising candidate for additional biological applications.
The authors are grateful to the Research Council of the University of Isfahan for financial support of this work. This work was supported by the Applied Molecular Biosciences Unit-UCIBIO which is financed by national funds from FCT/MCTES (UIDB/04378/2020).

In Vitro and in Vivo Effect of Palladacycles: Targeting A2780 Ovarian Carcinoma Cells and Modulation of Angiogenesis - Reigosa-Chamorro, Francisco; Raposo, Luís R.; Paula Munín-Cruz; Pereira, M. Teresa; Catarina Roma-Rodrigues; Baptista, Pedro V; Fernandes, Alexandra R; Vila, José M. (Inorganic Chemistry, 2021)
Palladacycles are versatile organometallic compounds that show potential for therapeutic use. Here are described the synthesis and characterization of mono- and dinuclear palladacycles bearing diphosphines. Their biological effect was investigated in A2780, an ovarian-derived cancer line, and in normal dermal fibroblasts. The compounds displayed selective cytotoxicity toward the A2780 cell line. Compound 3 decreased the cell viability through cell cycle retention in G0/G1, triggered apoptosis through the intrinsic pathway, and induced autophagy in A2780 cells. Compound 9 also induced cell cycle retention, apoptosis, and cellular detachment. Notably, compound 9 induced the production of intracellular reactive oxygen species (ROS). Our work demonstrated that compound 3 enters A2780 cells via active transport, which requires energy, while compound 9 enters A2780 cells mostly passively. The potential effect of palladacycles in angiogenesis was investigated for the first time in an in vivo chorioallantoic membrane model, showing that while compound 3 displayed an antiangiogenic effect crucial to fighting cancer progression, compound 9 promoted angiogenesis. These results show that palladacycles may be used in different clinical applications where pro- or antiangiogenic effects may be desirable.
This work was financed by national funds from FCT through the Applied Molecular Biosciences Unit, UCIBIO (Grants UIDP/04378/2020 and UIDB/04378/2020), and fellowships SFRH/BPD/124612/2016 (to C.R.-R.) and PTDC/CVTEPI/6685/2014 (to L.R.R.). This work was also made possible thanks to financial support received from the Xunta de Galicia (Galicia, Spain) under the Grupos de Referencia Competitiva Programme (Project GRC2019/14). F.R.-C. thanks the Spanish Ministry of Education (Grant FPU15/07145).

Rosa x hybrida extracts with dual actions: Antiproliferative effects against tumour cells and inhibitor of Alzheimer disease - Lorenzo Rivas-García; Quiles, José L.; Catarina Roma-Rodrigues; Raposo, Luis R.; Navarro-Hortal, María D.; Romero-Márquez, Jose M.; Esteban-Muñoz, Adelaida; Varela-López, Alfonso; García, Laura Carrera; Cianciosi, Danila; Forbes Hernández, Tamara Y.; Maurizio Battino; et al. (Food and Chemical Toxicology, 2021)
Edible flowers are being used as a new ingredient in modern gastronomy. Recently, these products have also gained interest as an important source of phenolic compounds with potential for biomedical applications. The present work studied a methanolic extract of Rosa x hybrida in which 35 individual phenolic compounds were identified. The extract has been evaluated for its antiproliferative properties in ovarian carcinoma cells. Results showed that the antiproliferative effect was associated with the induction of autophagy and apoptosis with the concomitant ROS increase probably related to mitochondria dysfunction. These antiproliferative effects might be associated with some components of the extract such as quercetin. The extract did not induce damage in healthy cells and that it was able to improve the wound healing activity. The present study also evaluated the properties of the mentioned extract in vivo in C. elegans. Tests demonstrated a lack of toxicity in the worm model. Promising results have been obtained in transgenic strains of C. elegans that produce human beta amyloid peptide, suggesting the possible utility of the extract from the point of view of Alzheimer disease. Altogether, results suggest that Rosa x hybrida extracts could be a new tool for the development of functional foods.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBPD%2F124612%2F2016/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FNAN-MAT%2F31100%2F2017/PT#
Keywords: Antioxidants; autophagy; Edible flowers; Flavonoids; Polyphenols; ROS

Square planar Au(III), Pt(II) and Cu(II) complexes with quinoline-substituted 2,2′:6′,2″-terpyridine ligands: From in vitro to in vivo biological properties - Choroba, Katarzyna; Machura, Barbara; Szlapa-Kula, Agata; Malecki, Jan G.; Luis Raposo; Catarina Roma-Rodrigues; Cordeiro, Sandra; Baptista, Pedro V; Fernandes, Alexandra R (European Journal of Medicinal Chemistry, 2021)
Cancer is the second leading cause of death worldwide. Cisplatin has challenged cancer treatment; however, resistance and side effects hamper its use. New agents displaying improved activity and more reduced side effects relative to cisplatin are needed. In this work we present the synthesis, characterization and biological activities of three complexes with quinoline-substituted 2,2′:6′,2″-terpyridine ligand: [Pt(4′-(2-quin)-terpy)Cl](SO3CF3) (1), [Au(4′-(2-quin)-terpy)Cl](PF6)2·CH3CN (2) and [Cu(4′-(2-quin)-terpy)Cl](PF6) (3). The three complexes displayed a high antiproliferative activity in ovarian carcinoma cell line (A2780) and even more noticeable in a colorectal carcinoma cell line (HCT116) following the order 3 > 2 > 1. The complexes IC50 are at least 20 × lower than the IC50 displayed by cisplatin (15.4 μM) in HCT116 cell line while displaying at the same time, much reduced cytotoxicity in a normal dermal fibroblast culture. These cytotoxic activities seem to be correlated with the inclination angles of 2-quin unit to the central pyridine. Interestingly, all complexes can interact with calf-thymus DNA (CT-DNA) in vitro via different mechanisms, although intercalation seems to be the preferred mechanism at least for 2 and 3 at higher concentrations of DNA. Moreover, circular dichroism (CD) data seems to indicate that complex 3, more planar, induces a high destabilization of the DNA double helix (shift from B-form to Z-form). Higher the deviation from planar, the lower the cytotoxicity displayed by the complexes. Cellular uptake may be also responsible for the different cytotoxicity exhibited by complexes with 3 > 2 >1. Complex 2 seems to enter cells more passively while complex 1 and 3 might enter cells via energy-dependent and -independent mechanisms. Complexes 1–3 were shown to induce ROS are associated with the increased apoptosis and autophagy. Moreover, all complexes dissipate the mitochondrial membrane potential leading to an increased BAX/BCL-2 ratio that triggered apoptosis. Complexes 2 and 3 were also shown to exhibit an anti-angiogenic effect by significantly reduce the number of newly formed blood vessel in a CAM model with no toxicity in this in vivo model. Our results seem to suggest that the increased cytotoxicity of complex 3 in HCT116 cells and its potential interest for further translation to pre-clinical mice xenografts might be associated with: 1) higher % of internalization of HCT116 cells via energy-dependent and -independent mechanisms; 2) ability to intercalate DNA and due to its planarity induced higher destabilization of DNA; 3) induce intracellular ROS that trigger apoptosis and autophagy; 4) low toxicity in an in vivo model of CAM; 5) potential anti-angiogenic effect.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBPD%2F124612%2F2016/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FCVT-EPI%2F6685%2F2014/PT#
Keywords: Apoptosis; Autophagy induction; Ex-ovo chorioallantoic membrane in vivo assay; Intracellular reactive oxygen species; Mitochondrial membrane potential changes; Square planar metal complexes

Benchtop X-ray fluorescence imaging as a tool to study gold nanoparticle penetration in 3D cancer spheroids - Pedro Pedrosa; Baptista, Pedro V; Fernandes, Alexandra R; Guerra, Mauro (RSC Advances, 2021)
The use of nanomaterials to improve medical diagnostics and therapeutics has been rapidly increasing. Among these materials are gold nanoparticles, which can be functionalized to target specific cells, acting as nanovectors for drug delivery, enhanced contrast agents as well as other targeted therapies. Au nanoparticles are very useful as they selectively accumulate in tumour sites due to the enhanced permeability-retention effect. There is however little information about the spatial distribution of the nanoparticles within tumours, which might hinder efficient therapies. In this study, X-ray fluorescence was used to investigate the diffusion of gold nanoparticles in cancer cell spheroids mimicking true tumour growth. Functionalization of the nanoparticles has the effect of allowing better diffusion into and out of the spheroid, while those nanoparticles that are only partially covered rapidly formed aggregates. This clustering led to size exclusion during transport within the tumour, changing its distribution profile while greatly increasing the nanoparticle concentration.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/157549/PT# UID/FIS/04559/2020

Ionogels Based on a Single Ionic Liquid for Electronic Nose Application - Gonçalves, Wellington B.; Cervantes, Evelyn P.; Pádua, Ana C. C. S.; Gonçalo Santos; Susana I C J Palma; Li, Rosamaria W. C.; Roque, Ana C A; Gruber, Jonas (Chemosensors, 2021)
Ionogel are versatile materials, as they present the electrical properties of ionic liquids and also dimensional stability, since they are trapped in a solid matrix, allowing application in electronic devices such as gas sensors and electronic noses. In this work, ionogels were designed to act as a sensitive layer for the detection of volatiles in a custom-made electronic nose. Ionogels composed of gelatin and a single imidazolium ionic liquid were doped with bare and functionalized iron oxide nanoparticles, producing ionogels with adjustable target selectivity. After exposing an array of four ionogels to 12 distinct volatile organic compounds, the collected signals were analyzed by principal component analysis (PCA) and by several supervised classification methods, in order to assess the ability of the electronic nose to distinguish different volatiles, which showed accuracy above 98%.
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Keywords: Composite; Electronic nose; Gas sensor; Ionic liquid; Ionogel; Volatile organic compound

Exploiting the antiproliferative potential of spiropyrazoline oxindoles in a human ovarian cancer cell line - Raposo, Luís R.; Silva, Ana Rute; Silva, Dário; Catarina Roma-Rodrigues; Espadinha, Margarida; Baptista, Pedro V; Santos, Maria M. M.; Fernandes, Alexandra R (Bioorganic and Medicinal Chemistry, 2021)
Cancer is still one of the deadliest diseases worldwide despite the efforts in its early detection and treatment strategies. However, most chemotherapeutic agents still present side effects in normal tissues and acquired resistance that limit their efficacy. Spiropyrazoline oxindoles might be good alternatives as they have shown antiproliferative activity in human breast and colon cancer cell lines, without eliciting cytotoxicity in healthy cells. However, their potential for ovarian cancer was never tested. In this work, the antiproliferative activity of five spiropyrazoline oxindoles was assessed in ovarian cancer cells A2780 and the biological targets and mechanism of action of the most promising compound evaluated. Compound 1a showed the highest antiproliferative effect, as well as the highest selectivity for A2780 cells compared to healthy fibroblasts. This antiproliferative effect results from the induction of cell death by mitochondria-mediated apoptosis and autophagy. In vitro DNA interaction studies demonstrated that 1a interacts with DNA by groove-binding, without triggering genotoxicity. In addition, 1a showed a strong affinity to bovine serum albumin that might be important for further inclusion in drug delivery platforms. Proteomic studies reinforced 1a role in promoting A2780 endoplasmatic reticulum (ER) stress by destabilizing the correct protein folding which triggers cell death via apoptosis and autophagy.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04138%2F2020/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FQUI-QOR%2F29664%2F2017/PT# info:eu-repo/grantAgreement/FCT/CEEC IND 2017/CEECIND%2F01772%2F2017%2FCP1476%2FCT0003/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBPD%2F124612%2F2016/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F117931%2F2016/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FNAN-MAT%2F31100%2F2017/PT#
Keywords: chemotherapy; Compound-DNA interaction; cytotoxicity; Ovarian cancer; Spiropyrazoline oxindoles

Specific Antiproliferative Properties of Proteinaceous Toxin Secretions from the Marine Annelid Eulalia sp. onto Ovarian Cancer Cells - Rodrigo, Ana P.; Mendes, Vera M.; Manadas, Bruno; Grosso, Ana R.; Alves de Matos, António P.; Baptista, Pedro V; Costa, Pedro M.; Fernandes, Alexandra R (Marine Drugs, 2021)
As Yondelis joins the ranks of approved anti-cancer drugs, the benefit from exploring the oceans' biodiversity becomes clear. From marine toxins, relevant bioproducts can be obtained due to their potential to interfere with specific pathways. We explored the cytotoxicity of toxin-bearing secretions of the polychaete Eulalia onto a battery of normal and cancer human cell lines and discovered that the cocktail of proteins is more toxic towards an ovarian cancer cell line (A2780). The secretions' main proteins were identified by proteomics and transcriptomics: 14-3-3 protein, Hsp70, Rab3, Arylsulfatase B and serine protease, the latter two being known toxins. This mixture of toxins induces cell-cycle arrest at G2/M phase after 3h exposure in A2780 cells and extrinsic programmed cell death. These findings indicate that partial re-activation of the G2/M checkpoint, which is inactivated in many cancer cells, can be partly reversed by the toxic mixture. Protein-protein interaction networks partake in two cytotoxic effects: cell-cycle arrest with a link to RAB3C and RAF1; and lytic activity of arylsulfatases. The discovery of both mechanisms indicates that venomous mixtures may affect proliferating cells in a specific manner, highlighting the cocktails' potential in the fine-tuning of anti-cancer therapeutics targeting cell cycle and protein homeostasis.
info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FBTA-BTA%2F28650%2F2017/PT# info:eu-repo/grantAgreement/FCT/OE/SFRH%2FBD%2F109462%2F2015/PT# info:eu-repo/grantAgreement/FCT/CEEC IND 2017/CEECIND%2F02699%2F2017%2FCP1462%2FCT0018/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UID%2FNEU%2F04539%2F2019/PT# info:eu-repo/grantAgreement/FCT/9471 - RIDTI/SAICTPAC%2F0010%2F2015/PT# info:eu-repo/grantAgreement/FCT/9471 - RIDTI/PTDC%2FBTM-TEC%2F29311%2F2017/PT# info:eu-repo/grantAgreement/FCT/9471 - RIDTI/PTDC%2FMEC-PSQ%2F30943%2F2017/PT# info:eu-repo/grantAgreement/FCT/3599-PPCDT/PTDC%2FMED-NEU%2F27946%2F2017/PT# POCI-01-0145-FEDER-402-022125 (ref.: ROTEIRO/0028/2013).
Keywords: Cancer; cell cycle; cytotoxicity; invertebrate; Ovarian cancer; programmed cell death; toxin

A Transcriptomic Approach to the Recruitment of Venom Proteins in a Marine Annelid - Rodrigo, Ana P.; Grosso, Ana R.; Baptista, Pedro V; Fernandes, Alexandra R; Costa, Pedro M. (Toxins, 2021)
The growing number of known venomous marine invertebrates indicates that chemical warfare plays an important role in adapting to diversified ecological niches, even though it remains unclear how toxins fit into the evolutionary history of these animals. Our case study, the Polychaeta Eulalia sp., is an intertidal predator that secretes toxins. Whole-transcriptome sequencing revealed proteinaceous toxins secreted by cells in the proboscis and delivered by mucus. Toxins and accompanying enzymes promote permeabilization, coagulation impairment and the blocking of the neuromuscular activity of prey upon which the worm feeds by sucking pieces of live flesh. The main neurotoxins ({"}phyllotoxins{"}) were found to be cysteine-rich proteins, a class of substances ubiquitous among venomous animals. Some toxins were phylogenetically related to Polychaeta, Mollusca or more ancient groups, such as Cnidaria. Some toxins may have evolved from non-toxin homologs that were recruited without the reduction in molecular mass and increased specificity of other invertebrate toxins. By analyzing the phylogeny of toxin mixtures, we show that Polychaeta is uniquely positioned in the evolution of animal venoms. Indeed, the phylogenetic models of mixed or individual toxins do not follow the expected eumetazoan tree-of-life and highlight that the recruitment of gene products for a role in venom systems is complex.
info:eu-repo/grantAgreement/FCT/3599-PPCDT/149867/PT# info:eu-repo/grantAgreement/FCT/OE/48636/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/157549/PT## CEECIND/02699/2017
Keywords: Annelida; marine environment; protein recruitment; selective pressure; toxins; whole-transcriptome sequencing

Cu(i) complexes as new antiproliferative agents against sensitive and doxorubicin resistant colorectal cancer cells: synthesis, characterization, and mechanisms of action - Sequeira, Diogo; Baptista, Pedro V; Valente, Ruben; Piedade, M. Fátima M. M.; Garcia, M. Helena; Morais, Tânia S.; Fernandes, Alexandra R (Dalton Transactions, 2021)
Cancer is one of the worst health issues worldwide, representing the second leading cause of death. Current chemotherapeutic drugs face some challenges like the acquired resistance of the tumoral cells and low specificity leading to unwanted side effects. There is an urgent need to develop new compounds that may target resistant cells. The synthesis and characterization of two Cu(i) complexes of general formula [Cu(PP)(LL)][BF4], where PP is a phosphane ligand (triphenylphosphine or 1,2-bis(diphenylphosphano) ethane) and LL = is a heteroaromatic bidentate ligand (4,4′-dimethyl-2,2′-bipyridine and 6,3-(2-pyridyl)-5,6-diphenyl-1,2,4-triazine). The new compounds were fully characterized by spectroscopic techniques (NMR, FTIR and UV-vis.), elemental analysis (C, H, N and S) and two structures were determined by single X-ray diffraction studies. The antiproliferative potential of the new Cu(i) complexes were studied in tumor (breast adenocarcinoma, ovarian carcinoma and in colorectal carcinoma sensitive and resistant to doxorubicin) and normal (fibroblasts) cell lines. Complexes1-4did not show any antiproliferative potential. Amongst the complexes5-8, complex8shows high cytotoxic potential against colorectal cancer sensitive and resistant to doxorubicin and low cytotoxicity towards healthy cells. We show that complexes5-8can cleave pDNA and, in particular, thein vitropDNA cleavage is due to an oxidative mechanism. This oxidative mechanism corroborates the induction of reactive oxygen species (ROS), that triggers HCT116 cell deathviaapoptosis, as proved by the increased expression of BAX protein relative to BCL-2 protein and the depolarization of mitochondrial membrane potential, andviaautophagy. Additionally, complex8can block the cell cycle in the G1 phase, also exhibiting a cytostatic potential. Proteomic analysis confirmed the apoptotic, autophagic and cytostatic potential of complex8, as well as its ability to produce ROS and cause DNA damage. The interference of the complex in folding and protein synthesis and its ability to cause post-translational modifications was also verified. Finally, it was observed that the complex causes a reduction in cellular metabolism. The results herein demonstrated the potential of Cu(i) complexes in targeting doxorubicin sensitive and resistant cells which is positive and must be further explored usingin vivoanimal models.
info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F00100%2F2020/PT# info:eu-repo/grantAgreement/FCT/CEEC IND 2017/CEECIND%2F00630%2F2017%2FCP1387%2FCT0012/PT#

Mapping Molecular Recognition of β1,3-1,4-Glucans by a Surface Glycan-Binding Protein from the Human Gut Symbiont Bacteroides ovatus - Correia, Viviana G.; Trovão, Filipa; Pinheiro, Benedita A.; Brás, Joana L A; Lisete M. Silva; Cláudia Nunes; Manuel A. Coimbra; Liu, Yan; Feizi, Ten; Fontes, Carlos M. G. A.; Mulloy, Barbara; Chai, Wengang; et al. (Microbiology spectrum, 2021)
A multigene polysaccharide utilization locus (PUL) encoding enzymes and surface carbohydrate (glycan)-binding proteins (SGBPs) was recently identified in prominent members of Bacteroidetes in the human gut and characterized in Bacteroides ovatus. This PUL-encoded system specifically targets mixed-linkage β1,3-1,4-glucans, a group of diet-derived carbohydrates that promote a healthy microbiota and have potential as prebiotics. The BoSGBPMLG-A protein encoded by the BACOVA_2743 gene is a SusD-like protein that plays a key role in the PUL's specificity and functionality. Here, we perform a detailed analysis of the molecular determinants underlying carbohydrate binding by BoSGBPMLG-A, combining carbohydrate microarray technology with quantitative affinity studies and a high-resolution X-ray crystallography structure of the complex of BoSGBPMLG-A with a β1,3-1,4-nonasaccharide. We demonstrate its unique binding specificity toward β1,3-1,4-gluco-oligosaccharides, with increasing binding affinities up to the octasaccharide and dependency on the number and position of β1,3 linkages. The interaction is defined by a 41-Å-long extended binding site that accommodates the oligosaccharide in a mode distinct from that of previously described bacterial β1,3-1,4-glucan-binding proteins. In addition to the shape complementarity mediated by CH-π interactions, a complex hydrogen bonding network complemented by a high number of key ordered water molecules establishes additional specific interactions with the oligosaccharide. These support the twisted conformation of the β-glucan backbone imposed by the β1,3 linkages and explain the dependency on the oligosaccharide chain length. We propose that the specificity of the PUL conferred by BoSGBPMLG-A to import long β1,3-1,4-glucan oligosaccharides to the bacterial periplasm allows Bacteroidetes to outcompete bacteria that lack this PUL for utilization of β1,3-1,4-glucans. IMPORTANCE With the knowledge of bacterial gene systems encoding proteins that target dietary carbohydrates as a source of nutrients and their importance for human health, major efforts are being made to understand carbohydrate recognition by various commensal bacteria. Here, we describe an integrative strategy that combines carbohydrate microarray technology with structural studies to further elucidate the molecular determinants of carbohydrate recognition by BoSGBPMLG-A, a key protein expressed at the surface of Bacteroides ovatus for utilization of mixed-linkage β1,3-1,4-glucans. We have mapped at high resolution interactions that occur at the binding site of BoSGBPMLG-A and provide evidence for the role of key water-mediated interactions for fine specificity and affinity. Understanding at the molecular level how commensal bacteria, such as prominent members of Bacteroidetes, can differentially utilize dietary carbohydrates with potential prebiotic activities will shed light on possible ways to modulate the microbiome to promote human health.
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Aggregation versus Biological Activity in Gold(I) Complexes. An Unexplored Concept - Pinto, Andrea; Catarina Roma-Rodrigues; Ward, Jas S.; Puttreddy, Rakesh; Kari Rissanen; Baptista, Pedro V; Fernandes, Alexandra R; Lima, João Carlos; Laura Rodríguez (Inorganic Chemistry, 2021)
The aggregation process of a series of mono- and dinuclear gold(I) complexes containing a 4-ethynylaniline ligand and a phosphane at the second coordination position (PR3-Au-CCC6H4-NH2, complexes 1-5, and (diphos)(Au-CCC6H4-NH2)2, complexes 6-8), whose biological activity was previously studied by us, has been carefully analyzed through absorption, emission, and NMR spectroscopy, together with dynamic light scattering and small-angle X-ray scattering. These experiments allow us to retrieve information about how the compounds enter the cells. It was observed that all compounds present aggregation in fresh solutions, before biological treatment, and thus they must be entering the cells as aggregates. Inductively coupled plasma atomic emission spectrometry measurements showed that mononuclear complexes are mainly found in the cytosolic fraction; the dinuclear complexes are mainly found in a subsequent fraction composed of nuclei and cytoskeleton. Additionally, dinuclear complex 8 affects the actin aggregation to a larger extent, suggesting a cooperative effect of dinuclear compounds.
Funding Information: info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F50006%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F50006%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDP%2F04378%2F2020/PT# info:eu-repo/grantAgreement/FCT/6817 - DCRRNI ID/UIDB%2F04378%2F2020/PT# The authors are grateful to the Spanish Ministerio de Ciencia, Innovación y Universidades (Project PID2019-104121GB-I00).LA/P/0140/2020 of the Associate Laboratory Institute for Health and Bioeconomy, i4HB. SAXS experiments were performed at the NCD-BL11 beamline of the ALBA Synchrotron Light Facility in collaboration with the ALBA staff. The authors also acknowledge COST Actions CA1740─Nano4clinics and CA18202─NECTAR. The Finnish Cultural Foundation (Grant 00201148 to J.S.W.) is also acknowledged for their support. Publisher Copyright: © 2021 American Chemical Society.

Exploring the Phytochemicals of Acacia melanoxylon R. Br. - Alves, Diana; Duarte, Sidonio; Arsenio, Pedro; Goncalves, Joana; Rodrigues, Cecilia M. P.; Lourenco, Ana; Maximo, Patricia (PLANTS-BASEL, 2021)
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Olfactory responses of Anaphes nitens (Hymenoptera, Mymaridae) to host and habitat cues - Branco, Sofia; Mateus, Eduardo Pires; Richter Gomes da Silva, Marco Diogo; Mendes, Davide; Araujo Pereira, Maria Manuela; Schuetz, Stefan; Paiva, Maria Rosa (JOURNAL OF APPLIED ENTOMOLOGY, 2021)
The eucalyptus weevil, Gonipterus spp. Schoenherr, 1833 (Coleoptera, Curculionidae) is considered a major pest of eucalyptus plantations. In regions where control is achieved, success is usually brought by the action of the solitary egg parasitoid Anaphes nitens (Girault, 1928) (Hymenoptera, Mymaridae). Research was conducted to ascertain which cues might mediate female wasp host location and selection. In Petri dish arenas, females were attracted to Gonipterus platensis Marelli, 1927 egg capsules, to G. platensis mated female faeces and to leaves of Eucalyptus globulus Labillardiere, 1799. Gas chromatography-mass spectrometry electroantennographic detection analysis was conducted using extracts obtained from leaves of E. globulus, from G. platensis egg capsules, as well as from adults of both sexes and their faeces, in order to detect and identify compounds perceived by the wasp's olfactory system. The parasitoids were shown to detect a wide range of compounds emitted by each one of these sources, and for 31 compounds, antennal response was confirmed by dose-response tests. Further behavioural trials were conducted in Petri dishes in order to decode the effect, on parasitoid behaviour, of selected compounds emitted by E. globulus and of the pheromones, emitted by the weevils on parasitoid behaviour. Attraction was observed to two compounds emitted by E. globulus, namely 1,8-cineole and gamma-terpinene, and to the main component of the male sex/aggregation pheromone, cis-verbenol. To our knowledge, this is the first report of attraction of a parasitoid from the family Mymaridae to a component of its host's sexual/aggregation pheromone. Similarly, to other egg parasitoid species, A. nitens females are likely to use the host plant volatiles as long-range host location cues and to adopt the `infochemical detour' strategy in order to get in the vicinity of their hosts.
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Drug delivery nanosystems targeted to hepatic ischemia and reperfusion injury - Ferreira-Silva, Margarida; Faria-Silva, Catarina; Baptista, Pedro Viana; Fernandes, Eduarda; Fernandes, Alexandra Ramos; Corvo, Maria Luísa (Drug delivery and translational research, 2021)
Abstract: Hepatic ischemia and reperfusion injury (IRI) is an acute inflammatory process that results from surgical interventions, such as liver resection surgery or transplantation, or hemorrhagic shock. This pathology has become a severe clinical issue, due to the increasing incidence of hepatic cancer and the high number of liver transplants. So far, an effective treatment has not been implemented in the clinic. Despite its importance, hepatic IRI has not attracted much interest as an inflammatory disease, and only a few reviews addressed it from a therapeutic perspective with drug delivery nanosystems. In the last decades, drug delivery nanosystems have proved to be a major asset in therapy because of their ability to optimize drug delivery, either by passive or active targeting. Passive targeting is achieved through the enhanced permeability and retention (EPR) effect, a main feature in inflammation that allows the accumulation of the nanocarriers in inflammation sites, enabling a higher efficacy of treatment than conventional therapies. These systems also can be actively targeted to specific compounds, such as inflammatory markers and overexpressed receptors in immune system intermediaries, allowing an even more specialized therapy that have already showed encouraging results. In this manuscript, we review drug delivery nanosystems designed for hepatic IRI treatment, addressing their current state in clinical trials, discussing the main hurdles that hinder their successful translation to the market and providing some suggestions that could potentially advance their clinical translation. Graphical abstract: [Figure not available: see fulltext.].
Funding Information: This work received financial support from PT national funds from Fundacao para a Ciencia e Tecnologia (FCT) through the FCT i3DU PhD programme (PD/BD/135264/2017), the grant FCT/MEC (UID/DTP/04138/2020 and UIDP/04138/2020) financing Research Institute for Medicines-iMed.ULisboa, the grant FCT/MEC (UIDP/04378/2020 and UIDB/04378/2020) financing Applied Molecular Biosciences Unit-UCIBIO, the grant by FCT/MCTES (UIDB/50006/2020) financing LAQV-REQUIMTE Associate Laboratory, from the European Union (FEDER funds through COMPETE POCI-01-0145-FEDER-029253), and by Phospholipid Research Center (project LCO-2017-052/1-1).
Keywords: Anti-inflammatory therapy; Drug delivery nanosystems; Inflammation; Ischemia and reperfusion injury; Liver transplant

Genetic predisposition for aggressive behaviour related with dopamine and serotonin pathways - an overview - Paulino, Cathy; Fernandes, Alexandra R; Baptista, Pedro V; Soeiro, Cristina; Grosso, Ana Rita; Quintas, Alexandre (Annals of Medicine, 2021)
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The genetic susceptibility linking preterm birth and periodontal disease a review - Joana Couceiroa; Grosso, Ana Rita; Baptista, Pedro V; Mendes, Jose J.; Fernandes, Alexandra R; Quintas, Alexandre (Annals of Medicine, 2021)
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Liposomal nanosystems in rheumatoid arthritis - Ferreira-Silva, Margarida; Faria-Silva, Catarina; Baptista, Pedro Viana; Fernandes, Eduarda; Fernandes, Alexandra Ramos; Corvo, Maria Luísa (Pharmaceutics, 2021)
Rheumatoid arthritis (RA) is an autoimmune disease that affects the joints and results in reduced patient quality of life due to its chronic nature and several comorbidities. RA is also associated with a high socioeconomic burden. Currently, several available therapies minimize symptoms and prevent disease progression. However, more effective treatments are needed due to current therapies’ severe side-effects, especially under long-term use. Drug delivery systems have demonstrated their clinical importance—with several nanocarriers present in the market—due to their capacity to improve therapeutic drug index, for instance, by enabling passive or active targeting. The first to achieve market authorization were liposomes that still represent a considerable part of approved delivery systems. In this manuscript, we review the role of liposomes in RA treatment, address preclinical studies and clinical trials, and discuss factors that could hamper a successful clinical translation. We also suggest some alterations that could potentially improve their progression to the market.
This work received financial support from PT national funds from Fundacao para a Ciencia e Tecnologia (FCT) through the grant FCT/MEC (UID/DTP/04138/2020 and UIDP/04138/2020) financing Research Institute for Medicines-iMed. ULisboa, grant FCT/MEC (UIDP/04378/2020 and UIDB/04378/2020) financing Applied Molecular Biosciences Unit-UCIBIO, a grant by FCT/MCTES (UIDB/50006/2020) financing LAQV-REQUIMTE Associate Laboratory, from the European Union (FEDER funds through COMPETE POCI-01-0145-FEDER-029253), and by Phospholipid Research Center (project LCO-2017-052/1-1). M.F.-S. acknowledges PhD grant PD/BD/135,264/2017 attributed by the FCT i3DU PhD program.
Keywords: Active targeting; Drug delivery nanosystems; Liposomes; Passive targeting; Rheumatoid arthritis

Comportamento sísmico da ligação laje-pilar considerando diferentes soluções de reforço - Brisid Isufi; Rui Marreiros; António Pinho Ramos; Válter Lúcio (Reabilitar & Betão Estrutural 2020, 2021)

Comportamento sísmico de ligações laje-pilar com variação da taxa de armadura de flexão - Mariana Rossi; Brisid Isufi; António Pinho Ramos (Reabilitar & Betão Estrutural 2020, 2021)

Cation-mediated gelation of the fucose-rich polysaccharide FucoPol: preparation and characterization of hydrogel beads and their cytotoxicity assessment - Fialho, Letícia; Araújo, Diana; Alves, Vitor D.; Catarina Roma-Rodrigues; Pedro V. Baptista; Alexandra R. Fernandes; Freitas, Filomena; Reis, Maria A M (International Journal of Polymeric Materials and Polymeric Biomaterials, 2021)
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Combined in silico and in vitro studies to identify novel antidiabetic flavonoids targeting glycogen phosphorylase - Brás, Natércia F.; Neves, Rui P.P.; Lopes, Filipa A.A.; Correia, Márcia A. S.; Palma, Angelina S.; Sousa, Sérgio F.; Ramos, Maria J. (2021)
Novel pharmacological strategies for the treatment of diabetic patients are now focusing on inhibiting glycogenolysis steps. In this regard, glycogen phosphorylase (GP) is a validated target for the discovery of innovative antihyperglycemic molecules. Natural products, and in particular flavonoids, have been reported as potent inhibitors of GP at the cellular level. Herein, free-energy calculations and microscale thermophoresis approaches were performed to get an in-depth assessment of the binding affinities and elucidate intermolecular interactions of several flavonoids at the inhibitor site of GP. To our knowledge, this is the first study indicating genistein, 8-prenylgenistein, apigenin, 8-prenylapigenin, 8-prenylnaringenin, galangin and valoneic acid dilactone as natural molecules with high inhibitory potency toward GP. We identified: i) the residues Phe285, Tyr613, Glu382 and/or Arg770 as the most relevant for the binding of the best flavonoids to the inhibitor site of GP, and ii) the 5-OH, 7-OH, 8-prenyl substitutions in ring A and the 4′-OH insertion in ring B to favor flavonoid binding at this site. Our results are invaluable to plan further structural modifications through organic synthesis approaches and develop more effective pharmaceuticals for Type 2 Diabetes treatment, and serve as the starting point for the exploration of food products for therapeutic usage, as well as for the development of novel bio-functional food and dietary supplements/herbal medicines.
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Keywords: Biophysical chemistry; Enzyme inhibition; Free-energy calculations; Microscale thermophoresis; Prenylflavonoids; Type 2 diabetes

Exploiting the antiproliferative potential of spiropyrazoline oxindoles in a human ovarian cancer cell line - Luís R. Raposo; Silva, Ana; Silva, Dário; Catarina Roma-Rodrigues; Espadinha, Margarida; Pedro V. Baptista; Santos, Maria M.M.; Alexandra R. Fernandes (Bioorganic & Medicinal Chemistry, 2021)
Cancer is still one of the deadliest diseases worldwide despite the efforts in its early detection and treatment strategies. However, most chemotherapeutic agents still present side effects in normal tissues and acquired resistance that limit their efficacy. Spiropyrazoline oxindoles might be good alternatives as they have shown antiproliferative activity in human breast and colon cancer cell lines, without eliciting cytotoxicity in healthy cells. However, their potential for ovarian cancer was never tested. In this work, the antiproliferative activity of five spiropyrazoline oxindoles was assessed in ovarian cancer cells A2780 and the biological targets and mechanism of action of the most promising compound evaluated. Compound 1a showed the highest antiproliferative effect, as well as the highest selectivity for A2780 cells compared to healthy fibroblasts. This antiproliferative effect results from the induction of cell death by mitochondria-mediated apoptosis and autophagy. In vitro DNA interaction studies demonstrated that 1a interacts with DNA by groove-binding, without triggering genotoxicity. In addition, 1a showed a strong affinity to bovine serum albumin that might be important for further inclusion in drug delivery platforms. Proteomic studies reinforced 1a role in promoting A2780 endoplasmatic reticulum (ER) stress by destabilizing the correct protein folding which triggers cell death via apoptosis and autophagy.
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Keywords: chemotherapy; Compound-DNA interaction; cytotoxicity; Ovarian cancer; Spiropyrazoline oxindoles

Implementation of Digital Monitoring Services During the COVID-19 Pandemic for Patients With Chronic Diseases: Design Science Approach - Luís Velez Lapão; Mariana Peyroteo; Maia, Melanie; Seixas, Jorge; Gregório, João; Mira da Silva, Miguel; Heleno, Bruno; Correia, Jorge César (JMIR, 2021)
Background: The COVID-19 pandemic is straining health systems and disrupting the delivery of health care services, in particular, for older adults and people with chronic conditions, who are particularly vulnerable to COVID-19 infection. Objective: The aim of this project was to support primary health care provision with a digital health platform that will allow primary care physicians and nurses to remotely manage the care of patients with chronic diseases or COVID-19 infections. Methods: For the rapid design and implementation of a digital platform to support primary health care services, we followed the Design Science implementation framework: (1) problem identification and motivation, (2) definition of the objectives aligned with goal-oriented care, (3) artefact design and development based on Scrum, (4) solution demonstration, (5) evaluation, and (6) communication. Results: The digital platform was developed for the specific objectives of the project and successfully piloted in 3 primary health care centers in the Lisbon Health Region. Health professionals (n=53) were able to remotely manage their first patients safely and thoroughly, with high degrees of satisfaction. Conclusions: Although still in the first steps of implementation, its positive uptake, by both health care providers and patients, is a promising result. There were several limitations including the low number of participating health care units. Further research is planned to deploy the platform to many more primary health care centers and evaluate the impact on patient’s health related outcomes.
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Keywords: chronic disease; COVID-19; design science research; digital health; elderly; implementation; Information Systems; Monitoring; primary healthcare; telemedicine

Interrogating the Inhibition Mechanisms of Human Aldehyde Oxidase by X-ray Crystallography and NMR Spectroscopy: The Raloxifene Case - Mota, Cristiano; Diniz, Ana; Coelho, Catarina; Santos-Silva, Teresa; Esmaeeli, Mariam; Leimkühler, Silke; Cabrita, Eurico J.; Marcelo, Filipa; Romão, Maria João (Journal of Medicinal ChemistryJournal of Medicinal Chemistry, 2021)
Human aldehyde oxidase (hAOX1) is mainly present in the liver and has an emerging role in drug metabolism, since it accepts a wide range of molecules as substrates and inhibitors. Herein, we employed an integrative approach by combining NMR, X-ray crystallography, and enzyme inhibition kinetics to understand the inhibition modes of three hAOX1 inhibitors—thioridazine, benzamidine, and raloxifene. These integrative data indicate that thioridazine is a noncompetitive inhibitor, while benzamidine presents a mixed type of inhibition. Additionally, we describe the first crystal structure of hAOX1 in complex with raloxifene. Raloxifene binds tightly at the entrance of the substrate tunnel, stabilizing the flexible entrance gates and elucidating an unusual substrate-dependent mechanism of inhibition with potential impact on drug–drug interactions. This study can be considered as a proof-of-concept for an efficient experimental screening of prospective substrates and inhibitors of hAOX1 relevant in drug discovery.Human aldehyde oxidase (hAOX1) is mainly present in the liver and has an emerging role in drug metabolism, since it accepts a wide range of molecules as substrates and inhibitors. Herein, we employed an integrative approach by combining NMR, X-ray crystallography, and enzyme inhibition kinetics to understand the inhibition modes of three hAOX1 inhibitors—thioridazine, benzamidine, and raloxifene. These integrative data indicate that thioridazine is a noncompetitive inhibitor, while benzamidine presents a mixed type of inhibition. Additionally, we describe the first crystal structure of hAOX1 in complex with raloxifene. Raloxifene binds tightly at the entrance of the substrate tunnel, stabilizing the flexible entrance gates and elucidating an unusual substrate-dependent mechanism of inhibition with potential impact on drug–drug interactions. This study can be considered as a proof-of-concept for an efficient experimental screening of prospective substrates and inhibitors of hAOX1 relevant in drug discovery.
doi: 10.1021/acs.jmedchem.1c01125

Molecular interactions of cefoperazone with bovine serum albumin: Extensive experimental and computational investigations - Ali, Mohd Sajid; Muthukumaran, Jayaraman; Jain, Monika; Santos-Silva, Teresa; Al-Lohedan, Hamad A.; Al-Shuail, Noura Saad (2021)
We investigated the binding of the cephalosporin-class drug cefoperazone (CFP) with bovine serum albumin (BSA) using spectroscopic techniques and in silico methods. The aim of this study was to (i) emphasize the importance of correcting for the inner filter effect in this type of study and (ii) understand the binding mechanism of CFP with BSA by addressing protein conformation and plausible binding sites. Formation of the complex was confirmed by UV–visible spectroscopy. Quenching of BSA fluorescence in the presence of CFP was also observed. Because of the high absorption of CFP in the fluorescence emission range of BSA, the fluorescence emission spectra were corrected for the inner filter effect. Fluorescence emission was studied at excitation wavelengths of 280 and 295 nm. The uncorrected data showed a significant contribution of tyrosine at the excitation wavelength of 280 nm; however, after correction, this contribution became negligible. The static-type mechanism was found to be involved in quenching, with almost 1:1 binding between BSA and CFP. Hydrogen bonding and hydrophobic forces were found to dominate the protein–ligand interactions with a slight decrease in the α-helical contents. Synchronous fluorescence spectral data (at Δλ = 15 and 60 nm) were also corrected for the inner filter effect, with the results being similar to those of excitation at 280 and 295 nm. Molecular docking and molecular dynamics (MD) simulation results suggest that, apart from the two known drug binding sites (drug site I and II), one putative binding site (binding site III) located between BSA domains 1 and 3 was also possible for CFP. MD simulations of the previously reported drug binding sites (drug site I and II) and putative binding site III revealed that binding site III showed excellent binding profiles and could be a target for future research related to BSA-drug binding.
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Keywords: Albumin binding; Cefoperazone; Inner filter effect; Molecular docking; molecular dynamics

Punching Shear Tests on RC Flat Slabs Strengthened with an UHPFRC Layer - Aurelio Sine; Mario Pimentel; Sandra Nunes (fib Symposium 2021, 2021)

Resistência ao punçoamento de lajes fungiformes reforçadas com UHPFRC - Mario Pimentel; Aurelio Sine; Sandra Nunes (Reabilitar & Betão Estrutural 2020, 2021)

The SOUL family of heme-binding proteins: Structure and function 15 years later - Goodfellow, Brian J.; Freire, Filipe; Ana Luísa Carvalho; Aveiro, Susana S.; Charbonnier, Peggy; Moulis, Jean-Marc; Delgado, Leonildo; Gloria C. Ferreira; Rodrigues, João E.; Poussin-Courmontagne, Pierre; Birck, Catherine; McEwen, Alastair; et al. (2021)
The SOUL, or heme-binding protein HBP/SOUL, family represents a group of evolutionary conserved putative heme-binding proteins that contains a number of members in animal, plant andbacterial species. The structures of the murine form of HEBP1, or p22HBP, and the human form of HEBP2, or SOUL, have been determined in 2006 and 2011 respectively. In this work we discuss the structures of HEBP1 and HEBP2 in light of new X-ray data for heme bound murine HEBP1. The interaction between tetrapyrroles and HEBP1, initially proven to be hydrophobic in nature, was thought to also involve electrostatic interactions between heme propionate groups and positively charged amino acid side chains. However, the new X-ray structure, and results from murine HEBP1 variants and human HEBP1, confirm the hydrophobic nature of the heme-HEBP1 interaction, resulting in Kd values in the low nanomolar range, and rules out any electrostatic stabilization. Results from NMR relaxation time measurements for human HEBP1 describe a rigid globular protein with no change in motional regime upon heme binding. X-ray structures deposited in the PDB for human HEBP2 are very similar to each other and to the new heme-bound murine HEBP1 X-ray structure (backbone rmsd ca. 1 Å). Results from a HSQC spectrum centred on the histidine side chain Nδ-proton region for HEBP2 confirm that HEBP2 does not bind heme via H42 as no chemical shift differences were observed upon heme addition for backbone NH and Nδ protons. A survey of the functions attributed to HEBP1 and HEBP2 over the last 20 years span a wide range of cellular pathways. Interestingly, many of them are specific to higher eukaryotes, particularly mammals and a potential link between heme release under oxidative stress and human HEBP1 is also examined using recent data. However, at the present moment, trying to relate function to the involvement of heme or tetrapyrrole binding, specifically, makes little sense with our current biological knowledge and can only be applied to HEBP1, as HEBP2 does not interact with heme. We suggest that it may not be justified to call this very small family of proteins, heme-binding proteins. The family may be more correctly called “the SOUL family of proteins related to cellular fate” as, even though only HEBP1 binds heme tightly, both proteins may be involved in cell survival and/or proliferation.
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Keywords: Function; HEBP1; HEBP2; NMR spectroscopy; SOUL protein; structure; X-ray crystallography

Structural insights into the molecular recognition mechanism of the cancer and pathogenic epitope, LacdiNAc by immune-related lectins - Lima, Carlos D. L.; Helena Coelho; Gimeno, Ana; Trovão, Filipa; Diniz, Ana; Dias, Jorge S.; Jesús Jiménez-Barbero; Francisco Corzana; Ana Luísa Carvalho; Cabrita, Eurico J.; Marcelo, Filipa (Chemistry – A European JournalChemistry – A European Journal, 2021)
Interactions of glycan-specific epitopes to human lectin receptors represent novel immune checkpoints for investigating cancer and infection diseases. By employing a multidisciplinary approach that combines isothermal titration calorimetry, NMR spectroscopy, molecular dynamics simulations, and X-ray crystallography, we disclosed the molecular determinants that govern the recognition of the tumour and pathogenic glycobiomarker LacdiNAc (GalNAc?1-4GlcNAc, LDN), including their comparison with the ubiquitous LacNAc epitope (Gal?1-4GlcNAc, LN), by two human immune-related lectins, galectin-3 (hGal-3) and the macrophage galactose C-type lectin (hMGL). A different mechanism of binding and interactions is observed for the hGal-3/LDN and hMGL/LDN complexes, which explains the remarkable difference in the binding specificity of LDN and LN by these two lectins. The new structural clues reported herein are fundamental for the chemical design of mimetics targeting hGal-3/hMGL recognition process.
https://doi.org/10.1002/chem.202100800
Keywords: glycan-protein interactions; hGal-3; hMGL; LacdiNAc; Molecular recognition

Travelling colourful patterns in self-organized cellulose-based liquid crystalline structures - Silva, Pedro E. S.; Chagas, Ricardo; Fernandes, Susete N.; Pieranski, Pawel; Selinger, Robin L. B.; Godinho, Maria Helena (2021)
Cellulose-based systems are useful for many applications. However, the issue of self-organization under non-equilibrium conditions, which is ubiquitous in living matter, has scarcely been addressed in cellulose-based materials. Here, we show that quasi-2D preparations of a lyotropic cellulose-based cholesteric mesophase display travelling colourful patterns, which are generated by a chemical reaction-diffusion mechanism being simultaneous with the evaporation of solvents at the boundaries. These patterns involve spatial and temporal variation in the amplitude and sign of the helix´s pitch. We propose a simple model, based on a reaction-diffusion mechanism, which simulates the observed spatiotemporal colour behaviour.
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Water Dynamics in Composite Aqueous Suspensions of Cellulose Nanocrystals and a Clay Mineral Studied through Magnetic Resonance Relaxometry - Kumar, Anant; Cruz, Carlos; Figueirinhas, João L.; Sebastião, Pedro J.; Trindade, Ana C.; Fernandes, Susete N.; Godinho, Maria H; Fossum, Jon Otto (The Journal of Physical Chemistry BThe Journal of Physical Chemistry B, 2021)
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doi: 10.1021/acs.jpcb.1c07331

Seismic behavior of slab-column connections with varying flexural reinforcement ratio - Mariana Rossi; Brisid Isufi; António Pinho Ramos (fib Symposium 2021, 2021)

Seismic behaviour of slab – column connections with various punching shear enhancement methods - Brisid Isufi; Rui Marreiros; António Pinho Ramos; Válter Lúcio (fib Symposium 2021, 2021)

Rational design of electron/proton transfer mechanisms in the exoelectrogenic bacteria Geobacter sulfurreducens - Silva, , Marta A.; Portela, Pilar C.; Salgueiro, , Carlos A. (Biochemical Journal, 2021)
{The redox potential values of cytochromes can be modulated by the protonation/deprotonation of neighbor groups (redox-Bohr effect), a mechanism that permits the proteins to couple electron/proton transfer. In the respiratory chains, this effect is particularly relevant if observed in the physiological pH range, as it may contribute to the electrochemical gradient for ATP synthesis. A constitutively produced family of five triheme cytochromes (PpcA−E) from the bacterium Geobacter sulfurreducens plays a crucial role in extracellular electron transfer, a hallmark that permits this bacterium to be explored for several biotechnological applications. Two members of this family (PpcA and PpcD) couple electron/proton transfer in the physiological pH range, a feature not shared with PpcB and PpcE. That ability is crucial for G. sulfurreducens’ growth in Fe(III)-reducing habitats since extra contributors to the electrochemical gradient are needed. It was postulated that the redox-Bohr effect is determined by the nature of residue 6, a leucine in PpcA/PpcD and a phenylalanine in PpcB/PpcE. To confirm this hypothesis, Phe6 was replaced by leucine in PpcB and PpcE. The functional properties of these mutants were investigated by NMR and UV–visible spectroscopy to assess their capability to couple electron/proton transfer in the physiological pH range. The results obtained showed that the mutants have an increased redox-Bohr effect and are now capable of coupling electron/proton transfer. This confirms the determinant role of the nature of residue 6 in the modulation of the redox-Bohr effect in this family of cytochromes, opening routes to engineer Geobacter cells with improved biomass production.}
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Development of conductive yarns for electronic textiles using the electrospinning technique - Frederico Ferreira Rodrigues (FCT NOVA, 2021)

1D Fiber-shaped supercapacitors - João Tiago Henriques (FCT NOVA, 2021)

3D-printed hybrid zeolitic/carbonaceous electrically conductive adsorbent structures - Diogo N D L Mendes; Ana Gaspar; Ferreira, Isabel; José P.B. Mota; Rui P.P.L. Ribeiro (Chemical Engineering Research and Design, 2021)

Affinity tags in protein purification and peptide enrichment: An overview, in Protein Downstream Processing: Design, Development, and Application of High and Low-Resolution Methods - Pina, A. S.; Batalha, IL; Dias, A M G C; Roque, A. C. A. (Methods in Molecular Biology, 2021)

Affinity-triggered hydrogels: Developments and prospects in biomaterials science - Fernandes, C.S.M.; Pina, A. S.; Roque, A. C. A. (Biomaterials, 2021)

Antibacterial bioglass in dental implants: a canine clinical study - Sílvia R. Gavinho; Manuel P. Graça; Prezas, Pedro; Borges, João Borges; Silva, Jorge Carvalho; Pires, Eduardo; Henrique Armês; José Coucelo (European Journal of Public Health, 2021)
Background Peri-implantitis is considered the most challenging biological complication in implantology, as untreated disease can progress and result in implant loss. Therefore, disease prevention is crucial in daily clinical practice. It has been reported that the use of bioactive glass, as an implant coating, can stimulate tissue integration and accelerate tissue regeneration. Besides these properties, it is possible to promote bacterial activity by inserting silver into the bioglass Methods Bioglass with composition 45S5 was synthesised by the fusion method, replacing the amount of Na2CO3 by AgNO3 (BG 2% wt). The implants were resealed by the CoBlast® technique. Clinical cases with pathology of the mandible/maxilla were selected and implants dimensioned for the canine bone structure were applied. Results Three months after implantation, imaging exams, namely CT scans, showed no signs of early rejection by septic or cytotoxic loss. No decrease or loss of peri-implant bone was observed. In all cases the implants remained without signs of instability, and with sufficient support for the application of the exo-prosthesis or dental crown. The results of histological analysis showed no signs of infection or osteolysis. The zone of peri-implant fibrosis was not observable in the samples, showing a good evolution in implant osteointegration. Conclusions The results show promising evidences for the use of this biomaterial as a coating, since aseptic rejection, later on, and that related to the shape and biomaterials used in the implant's design, usually begins during the first 3 months.

Artificial enzymes bringing together computational design and directed evolution - Mariz, BP; Carvalho, S.; Batalha, IL; Pina, A. S. (Organic & Biomolecular Chemistry, 2021)