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Multi-Purpose Concept of Design: A Sustainable Strategy in Addressing the Widespread Redundancy of the “White Wedding Gowns” in the Ghanaian Culture
Publication . Ibrahim, Haruna; Romãozinho, Ana Mónica Pereira Reis de Matos; Silva, Fernando José Carneiro Moreira da
Sustainable best practices and material culture are at the forefront in this 21st century, and as a result, sustainability and circularity have become very crucial for global industrial growth and development, and the clothing and textile industry cannot be left out. The clothing and textile industry is considered the second largest contributor to environmental pollution, after the oil industry, and this is due to the fashion industry's adherence to a linear economic system. The alarming rate of this disposable behaviour in the clothing industry requires the adoption of a multi-purpose design concept as a strategy to curb this design stance that may have the potential to extend the life of clothing, encourage sustainable consumption, minimise greenhouse gases and facilitate the transition to a circular economy. The wedding dress is perhaps one of the most common clothing items to be discarded prematurely or become redundant in many wardrobes once the wedding ceremonies are over, meaning that it has become a one-of-a-kind outfit regardless of the time, money and other resources invested in its acquisition. This consumer behaviour is attributed to the lack of innovative design thinking and also the failure to incorporate the needs and values of clothing consumers through the adoption of a co-design strategy. This study is qualitative in nature and therefore employs a mixed-methodological approach, i.e., a combination of interventionist and non-interventionist research methods, to address the study objectives. The thesis has two main objectives (1) to give wedding dresses an extended life within the wedding culture in Ghana by exploring on the one hand the multi-purpose concept of design and on the other hand, product development through co-design with wedding dress wearers; (2) propose a new conceptual model and produce a sample wedding dress through a sustainable prototype to be tested and validated.
Non-canonical DNA secondary structures as a therapeutic strategy for lung cancer
Publication . Miranda, André Filipe Rodrigues ; Cruz, Carla Patrícia Alves Freire Madeira; Mergny, Jean-Louis; Oliveira, Paula Alexandra Martins de
Nucleic acids store and control genetic information and, beyond the canonical B-form DNA double helix, can adopt a variety of non-canonical architectures, including G-quadruplex (G4s), i-motifs (iMs), triplexes, R-loops, and Z-DNA/Z-RNA. These structures are widespread across genomes and transcriptomes and play important biological roles (transcription, replication, translation, and genome stability). They are recognized as regulatory hotspots enriched at promoters, enhancers, telomeres, untranslated regions, and viral genomes and their involvement in cancer, viral infection, neurodegeneration, and inflammatory disorders. Their in vivo functions have also opened therapeutic avenues both as drug targets and therapeutic tools. As drug targets, these non-canonical structures can be modulated to alter cellular behavior, while as therapeutic agents, nucleic acids can be engineered to adopt non-canonical folds that bind key cellular partners and exert biological effects. This dual role is unusual for other approaches and positions non-canonical structures at the forefront of modern drug discovery.
These structures are emerging as important tools in oncological research and therapeutic development. Proto-oncogenes encode proteins essential to normal physiology; however, when mutated or amplified become oncogenes, they drive unchecked growth and survival. The transcription factors (TFs) are a particularly important class of proto-oncogenes that bind to DNA and recruit co-regulators, orchestrating broad gene-expression programs. Their sweeping control of transcription makes them powerful cancer drivers but also challenging drug targets. Unlike enzymes with well-defined catalytic pockets, TFs often rely on flexible, intrinsically disordered regions for interactions, contributing to their reputation as “undruggable.” Numerous TFs have been reported as dysregulated in cancer and are frequently correlated with poor prognosis and resistance to chemotherapy. Other TF are gaining relevance, such as B-MYB (MYBL2), a transcription factor from the MYB family required for normal cell-cycle progression, which acts as an oncogene when overexpressed or deregulated.
Functionally, the B-MYB protein serves as a master cell-cycle regulator by integrating into multiprotein assemblies, most notably the DREAM and MMB complexes, that coordinate cell-cycle gene expression. Beyond these physiological functions, B-MYB is upregulated in multiple cancers, such as breast and lung, and its overexpression is correlated with aggressive disease, treatment resistance, and unfavorable prognosis. The molecular mechanisms linking B-MYB to tumorigenesis include gene amplification, cell cycle deregulation, genomic instability, apoptosis suppression, post-transcriptional and post-translational modifications, or can contribute to epithelial-to-mesenchymal transition. Thus, B-MYB TF can be seen as a relevant clinical biomarker and one of the main orchestrators of carcinogenesis; however, until now, no pharmacological therapy against B-MYB has been developed, also related to its “undruggable” profile.
Also, it’s known, by bioinformatic analysis of the human genome, that oncogene promoter regions are G/C rich around transcription start sites (TSS), which allows the formation of structures called G4 or iM. The G4s are formed by the self-association of four guanine bases in a quasi-planar arrangement via Hoogsteen bonds and are described as having a gene regulatory function, namely at the transcriptional level, while the iM is formed by Hoogsteen bonds between protonated and deprotonated cytosines. Motivated by these observations, the central objective of this thesis was to explore alternative routes to target the B-MYB oncogene using non-canonical nucleic-acid structures: first as drug targets and later as therapeutics.
The thesis started by the identification of G-rich sequences at the B-MYB promoter capable of forming G4 structures. The identification of G-rich sequences was performed using the G4Hunter algorithm, and their conservation across mammalian species was also verified. The experimental validation of their formation was performed by combining 10 biophysical and biochemical methods. Later, the in-cell relevance of G4 structures was evaluated employing the G4access method, which reveals that from the predicted sequences, only the most stable one (B-MYB 43R) was shown to be significantly formed in cells, evidencing a potential impact on the transcription of this gene to its location closest to the TSS.
Next, the ability of C-rich sequences to form iM structure was evaluated. Again, the iM-forming sequences were predicted using the G4Hunter algorithm and the experimental validation started by circular dichroism (CD) and nuclear magnetic resonance (NMR) to determine if the sequences fold into an iM structure. Then, stability parameters such as pHT (pH transitional midpoint) were determined by acquiring spectra at different pH values (between pH=5 and pH=8; 0.25-unit increments). The thermal stability and thermodynamic parameters were also calculated using the denaturation and renaturation melting curves. Then, the in-cell formation was assessed using iM-CUT&Tag experiments in HEK293T, which revealed the formation of the iB-MYB 43 that was characterized to have the highest Tm and the highest pHT among the studied sequences. After this characterization, the interaction of small molecules with iB-MYB structures was assessed. As a general tendency, the ligands did not affect the CD spectral shape; however, some of them evidenced changes in secondary structure or thermal destabilization. After evaluating the capacity to form G4 or iM, we continued to validate the G4 formation within a cellular environment using a G4-triggered fluorogenic hybridization probe. This strategy circumvents the limitations of the antibodies and small-molecule probes, which show a lack of structural selectivity labeling diverse DNA and RNA G4s indiscriminately, and cannot specifically target the desired G4. The molecular probe was composed of G4-recognizing light-up ligand (acridine derivative) with an antisense oligonucleotide that hybridizes adjacent region of B-MYB G4. Before probe synthesis using a click chemistry approach, the ligands were photophysically characterized, biophysically evaluated against G4, and further validated spectroscopically and in cells. Cellular studies confirmed the co-localization between the molecular probe and B-MYB G4 in-cell, offering promise for future applications in cancer research in terms of targeted therapies and monitoring of G4.
Although G4s are highly dynamic and topologically diverse, promoter G4s are commonly parallel, whereas antiparallel forms are underreported and less characterized. In earlier work, we identified a B-MYB promoter sequence (B-MYB 26RA), that forms an antiparallel G4. Thus, a biophysical pipeline, starting in solution NMR spectroscopy alongside in-cell NMR studies, was made to predict a three-dimensional model (antiparallel quadruplex-duplex junction) and to assess the formation in a live complex environment of B-MYB 26RA. Interestingly, B-MYB 26RA evidenced an ionic sensitivity to K+ and Na+, increasing their topological dynamics, and demonstrated that, when associated with PhenDC3 ligand, a conformational shift from hybrid to antiparallel topology happens.
Then, we moved to B-MYB 43R G4 to discuss the therapeutic relevance and the capacity to be targeted by small molecules. Thus, using spectroscopic methods, the G4 formation and its interaction with a panel of G4-stabilizing ligands were confirmed. From the tested ligands, PhenDC3 and TMPyP4 demonstrated the highest binding affinity and stabilization and revealed that both ligands inhibited proliferation and migration in two lung cancer cell lines (A549 and H1299), with PhenDC3 showing potent cytotoxic and cytostatic effects. Furthermore, PhenDC3 upregulated B-MYB expression despite its strong phenotypic effects, highlighting the complexity of G4-targeting mechanisms and supporting the growing evidence that ligands do not always downregulate their target genes.
Finally, we explored the use of Polypurine Reverse-Hoogsteen (PPRH) hairpins as an alternative therapeutic approach to target the B-MYB promoter. PPRH are DNA oligonucleotides that fold into intramolecular hairpins and bind complementary polypyrimidine, forming stable DNA triplexes, impeding transcription, and displacing the G-rich strand, inducing G4 formation, enabling dual-level regulation of oncogene expression. Thus, using a bioinformatic tool (TFO Searching Tool), a PPRH was designed against the G4-forming region at the B-MYB promoter (B-MYB 43R), and then experimentally assessed the triplex formation using biophysical methods. The biological effects of designed PPRH, evaluated in lung cellular models (A549, H1299 and MRC-5), demonstrated a low cell viability and clonogenic capacity accompanied by mRNA expression reduction and proteomic profile alterations. Thus, the combination of triplex-based approaches with G4 biology could be a future venue for therapy.
Overall, this thesis establishes non-canonical nucleic-acid structures as both actionable targets and therapeutic agents. As a target, the formation of G4 and iM structures at the promoter region of the B-MYB oncogene; was provided new insights about structure and topological dynamics according to the surrounding environment; was validated their formation in-cell, as well as explored the interaction with small molecules. On the other side, the therapeutic potential was explored using PPRH, which revealed a selective and powerful tool to target B-MYB.
Together, these findings provide fundamental insights and open new avenues for translational research on the B-MYB oncogene.
Production and Characterization of a Pullulan-Based Facial Mask Incorporating Grape Seed Flour Extract for Cosmeceutical Applications
Publication . Ferreira, Ester; Encarnação, Bárbara; Cascalheira, José Francisco; Ferreira, Susana; Ramos, Ana; Luís, Ângelo; Gallardo, Eugenia; Domingues, Fernanda; Magni, Chiara
This study aimed to develop and characterize an eco-friendly facial mask based on the pullulan biopolymer incorporating grape seed flour extract, a sustainable source of polyphenols. The extract was characterized by its phenolic content, antioxidant capacity, enzyme inhibition and antimicrobial activity. High total phenolic and flavonoid contents, along with the presence of -resveratrol, conferred strong antioxidant activity. The extract effectively inhibited tyrosinase and elastase enzymes, indicating its anti-aging potential, and exhibited antimicrobial effects particularly against . The incorporation of the extract in pullulan films increased thickness and coloration while maintaining transparency and improving barrier properties. The bioactive films showed strong antioxidant activity and displayed selective antibacterial activity against , including MRSA strains. A facial mask prototype was successfully produced, demonstrating flexibility, solubility, and potential for topical applications. Overall, the developed pullulan-grape seed extract films exhibit multifunctional cosmeceutical potential combining antioxidant, antimicrobial, and anti-aging effects with sustainable valorization of wine industry by-products.
MicroRNAs in Takotsubo Syndrome: A Systematic Review of Regulatory Networks in Stress-Induced Cardiomyopathy
Publication . Sousa, Domingos; Martins, Filipa Abreu; Luís, Ângelo; Serralheiro, Pedro
MicroRNAs (miRNAs) have emerged as crucial regulators of gene expression and have been implicated in various physiological and pathological processes, including cardiovascular diseases. The clinical presentation, diagnostic criteria, and proposed pathophysiological mechanisms of Takotsubo Syndrome (TTS) are discussed, with an emphasis on the emerging evidence implicating miRNAs in its etiology and progression. A systematic review following the PRISMA guidelines was performed on the evidence regarding the interplay between miRNAs and TTS. A search of the Pubmed, Web of Science, and Scopus databases was conducted and resulted in 584 articles. Of these, 14 full-text articles were eligible for inclusion in the qualitative analysis. The reviewed studies suggest that multiple miRNAs are involved in the processes associated with TTS pathophysiology, including acute and chronic myocardial inflammation, oxidative stress, apoptosis, microvascular dysfunction, hypertrophy, and, ultimately, maladaptive cardiac remodelling. This review provides an overview of the current understanding of miRNAs in cardiovascular pathophysiology, with a specific focus on their potential roles in TTS. To the best of our knowledge, this is the first systematic exploration of the miRNAs involved in TTS and its modulation as potential biomarkers or therapeutic targets.
Multimodal ionic liquid-based chromatographic supports for an effective RNA purification
Publication . Carapito, Ana Rita; Bernardo, Sandra C.; Pereira, Matheus M.; Neves, Márcia C.; Freire, Mara; Sousa, Fani
Nucleic acids have been considered interesting molecules to be used as biopharmaceuticals for the treatment of various diseases, in gene therapy strategies. In particular, RNA arises as the most promising approach because it does not require access to the nucleus of cells to exert its function; however, it is quite challenging due to its labile nature. To increase the possibility of translating RNA-based technology to clinical protocols, the biomanufacturing of RNAs has been intensively exploited in the last few years. However, the standard RNA purification processes remain time-consuming and present limitations regarding recovery yield and purity. This work describes the functionalization of chromatographic silica-based supports with four ionic liquids (ILs) composed of functional moieties that can promote distinct interactions with nucleic acids. After an initial screening to evaluate the binding and elution behavior of nucleic acids in the IL-based supports, SSi[C3C3NH2Im]Cl has shown to be the most promising for further purification assays. This support was studied for the RNA purification from different samples (clarified or more complex) and has shown to be highly effective, for all the conditions studied. Generally, it is here presented a new method for RNA isolation in a single step, using an IL-based chromatographic support, able to eliminate the usage of hazardous compounds often included in standard RNA extraction protocols.