National Institute of Health Dr. Ricardo Jorge
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Lactate-coated polyurea-siRNA dendriplex: a gene therapy-directed and metabolism-based strategy to impair glioblastoma (GBM)
No full textCorrection: Cancer Gene Ther. 2025 Aug;32(8):911. Epub 2025 Jul 10. https://doi.org/10.1038/s41417-025-00933-5Glioblastoma (GBM) is a highly lethal disease with limited treatment options due to its infiltrative nature and the lack of efficient therapy able to cross the protective blood-brain barrier (BBB). GBMs are metabolically characterized by increased glycolysis and glutamine dependence. This study explores a novel metabolism-based therapeutic approach using a polyurea generation 4 dendrimer (PURE) surface functionalized with lactate (LA) (PURE-LA), to take advantage of glucose-dependent monocarboxylate transporters (MCTs) overexpression, loaded with selenium-chrysin (SeChry) and temozolomide (TMZ) or complexed with anti-glutaminase (GLS1) siRNAs to abrogate glutamine dependence. The nanoparticles (PURE-LA) were efficient vehicles for cytotoxic compounds delivery, since SeChry@PURE-LA and TMZ@PURE-LA induced significant cell death in GBM cell lines, particularly in U251, which exhibits higher MCT1 expression. The anti-GLS1 siRNA-dendriplex with PURE-LA (PURE-LA-anti-GLS1-siRNA) knocked down GLS1 in the GBM cell lines. In two in vitro BBB models, these dendriplexes successfully crossed the BBB, decreased GLS1 expression and altered the exometabolome of GBM cell lines, concomitantly with autophagy activation. Our findings highlight the potential of targeting glucose and glutamine pathways in GBM using dendrimer-based nanocarriers, overcoming the BBB and disrupting key metabolic processes in GBM cells. PURE-LA-anti-GLS1-siRNA dendriplexes cross the blood-brain barrier (BBB) and impair glioblastoma (GBM) metabolism. The BBB is formed by a thin monolayer of specialized brain microvascular endothelial cells joined together by tight junctions that selectively control the passage of substances from the blood to the brain. It is a major obstacle in the treatment of GBM, since many chemotherapeutic drugs are unable to penetrate the brain. Therefore, we developed a strategy to overcome this obstacle: a lactate-coated polyurea dendrimer generation 4 (PURE) able to cross the BBB in vitro, that act as a nanocarrier of drugs and siRNA to the GBM cells. PURE-LA are nanoparticles functionalized with lactate (LA) to target MCT1, a lactate transporter highly expressed by GBM cells. Moreover, a complex of this nanoparticle with anti-GLS1 (glutaminase) siRNA (PURE-LA-anti-GLS1-siRNA) was made, to target glutamine metabolism. It efficiently knocked down GLS1. Moreover, PURE-LA loaded with SeChry led to BBB disruption.The institutions are funded by Fundação para a Ciência e a Tecnologia/Ministério da Ciência, Tecnologia e Ensino Superior (FCT/MCTES, Portugal) through national funds to iNOVA4Health (UIDB/04462/2020 and UIDP/04462/2020), to MOSTMICRO-ITQB (UIDB/04612/2020 and UIDP/04612/2020), to BioISI (Center Grant doi 10.54499/UIDB/ 04046/2020), and the Associated Laboratory LS4FUTURE (LA/P/0087/2020). Filipa Martins was funded by a FCT individual Ph.D. fellowship (2020.04780.BD). Luis G. Gonçalves was financed by a FCT contract according to DL57/2016, [SFRH/BPD/ 111100/2015]. Data availability in a public repository is supported by NIH U2C DK119886 and OT2-OD030544 grants
Re‐evaluation of oxygen (E 948) and hydrogen (E 949) as food additives
No full textThe Panel on Food Additives and Flavourings (FAF) provides a scientific opinion re‐evaluating the safety of oxygen (E 948) and hydrogen (E 949) as food additives. Their currently permitted use in food in the European Union (EU) is in all food categories, including in foods for infants and young children at quantum satis (QS). They can also be used in food additive preparations, food enzymes and nutrients also at QS. No interested business operators (IBOs) provided information in response to the call for data published by EFSA to support their re‐evaluation. The original evaluation by the EU in 1990 indicated their use as packaging gases, and in the case of oxygen (E 948), also as propellant. The Panel considered the two gases to be of low toxicological concern when used as food additives and their dietary exposure very low. The Panel concluded that the use of oxygen (E 948) and hydrogen (E 949) as food additives does not raise a safety concern. The Panel made some recommendations for amending existing EU specifications for both oxygen (E 948) and hydrogen (E 949)
Ten winters of surveillance: 10 years of the FRIESA system (Extreme Cold in Health)
No full textO Sistema FRIESA foi criado em 2014 para monitorizar o impacto do frio extremo na mortalidade nos distritos de Lisboa e do Porto. Após uma década de atividade, este artigo sintetiza os principais resultados e seu contributo na definição e implementação das medidas de saúde pública.
Para este trabalho foram analisados os resultados dos relatórios anuais do FRIESA, tendo sido demonstrada a utilidade do sistema na identificação precoce de períodos críticos e na produção de evidência científica para apoio à decisão em saúde pública.The FRIESA System was created in 2014 to monitor the effects of extreme cold on mortality in the districts of Lisbon and Porto. After a decade of activity, this article summarizes the main results and their contribution to the definition and implementation of public health measures.
For this work, the results of FRIESA's annual reports were analysed, demonstrating the usefulness of the system in the early identification of critical periods and in the production of scientific evidence to support public health decision-making
Molecular genetics of hemoglobinopathies in the Department of Human Genetics of INSA: a trajectory of contributions to science and society
No full textComparative analysis of hybrid‑SNP microarray and nanopore sequencing for detection of large‑sized copy number variants in the human genome
No full textBackground: Nanopore sequencing is a technology that holds great promise for identifying all types of human genome variations, particularly structural variations. In this work, we used nanopore sequencing technology to sequence 2 human cell lines at low depth of coverage to call copy number variations (CNV), and compared the results variant by variant with chromosomal microarray (CMA) results.
Results: We analysed sequencing data using CuteSV and Sniffles2 variant callers, compared breakpoints based on hybrid-SNP microarray, nanopore sequencing and Sanger sequencing, and analysed CNV coverage. From a total of 48 high confidence variants (truth set), variant calling detected 79% of the truth set variants, increasing to 86% for interstitial CNV. Simultaneous use of the 2 callers slightly increased variant calling. Both callers performed better when calling CNV losses than gains. Variant sizes from CMA and nanopore sequencing showed an excellent correlation, with breakpoints determined by nanopore sequencing differing by only 20 base pairs on average from Sanger sequencing. Nanopore sequencing also revealed that four variants concealed genomic inversions undetectable by CMA. In the 10 CNV not called in nanopore sequencing, 8 showed coverage evidence of genomic loss or gain, highlighting the need to improve SV calling algorithms performance.
Conclusions: Nanopore sequencing offers advantages over CMA for structural variant detection, including the identification of multiple variant types and their breakpoints with increased precision. However, further improvements in variant calling algorithms are still needed for nanopore sequencing to become a highly robust and standardized approach for a comprehensive analysis of genomic structural variation.This work is a result of the GenomePT project (POCI-01–0145-FEDER-022184), supported by COMPETE 2020—Operational Programme for Competitiveness and Internationalization (POCI), Lisboa Portugal Regional Operational Programme (Lisboa2020), Algarve Portugal Regional Operational Programme (CRESC Algarve2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF), and by Fundação para a Ciência e a Tecnologia (FCT). This work was also supported by Fundos FEDER through the Programa Operacional Factores de Competitividade – COMPETE and by Fundos Nacionais through the Fundação para a Ciência e a Tecnologia within the scope of the project UID/BIM/00009/2019 (Centre for Toxicogenomics and Human Health -ToxOmics)
Assessing the impact of TiO2 nanomaterials on intestinal cells: New evidence for epithelial translocation and potential pro-inflammatory effects
No full textUnderstanding the potential impact of nanomaterials (NMs) on human health requires further investigation into the organ-specific nano-bio interplay at the cellular and molecular levels. We showed increased chromosomal damage in intestinal cells exposed to some of in vitro digested Titanium dioxide (TiO2) NMs. The present study
aimed to explore possible mechanisms linked to the uptake, epithelial barrier integrity, cellular trafficking, as well as activation of pro-inflammatory pathways, after exposure to three TiO2-NMs (NM-102, NM-103, and NM-105). Using confocal microscopy, we show that all NMs, digested or not, were able to enter different types of intestinal cells. At the physiologically relevant concentration of 14 μg/mL, the digested TiO2-NMs did not compromise the transepithelial resistance, nor the levels of epithelial markers E-cadherin and Zonula occludens protein 1 (ZO-1), of polarized enterocyte monolayers. Nonetheless, all NMs were internalized by intestinal cells and, while NM-102 was retained in lysosomes, NM-103 and NM-105 were able to transverse the epithelial barrier through transcytosis. Moreover, 24 h exposure of 14 and 1.4 μg/mL digested NM-105, promoted interleukin IL-1β expression in activated M1 macrophages, indicating a potential pro-inflammatory action in the gut. Taken together, our findings shed light on the cell-specific nano-bio interplay of TiO2-NMs in the context of the intestinal tract and highlight transcytosis as a potential gateway for their systemic distribution. The potential proinflammatory action of digested NM-105 emphasizes the importance of pursuing research into the potential impact of NMs on human health and contribute to the weight of evidence to limit their use in food.This work was funded by national funds through the FCT - Foundation for Science and Technology, I.P., under the project PTDC/SAU- PUB/29481/2017. Research was co-funded by ToxOmics (UIDB/00009/ 2020 and UIDP/00009/2020), BioISI (UID/MULTI/04046/2019), iMed. ULisboa (UIDB/04138/2020 and UIDP/04138/2020), and principal investigator contract CEECIND/03143/2017 (L.G.)
Proposal of a Familial Hypercholesterolemia Pediatric Diagnostic Score (FH-PeDS)
No full textFH-PeDS Collaborators: Barbara Cugalj Kern, Jernej Kovač, Ana Drole Torkar, Maja Filipič, Mia Becker, Žiga Iztok Remec, Barbka Repič Lampret, Maruša Debeljak, Katarina Trebušak Podkrajšek, Zlatko Fras, Borut Jug, Fouzia Sadiq, António Guerra, Ana Gaspar, Henedina Antunes, Sílvia Sequeira, Susana Correia, Paula Garcia, Luísa Diogo Matos, Goreti Lobarinhas, Paula Martins, Guida Gama, Mónica Tavares, Eduard Ostarijas, Zala Jelinčič, Dimitar Trifunoski, Luka PesjakBackground and aims: Familial hypercholesterolemia (FH) significantly increases cardiovascular risk from childhood yet remains widely underdiagnosed. This cross-sectional study aimed to evaluate existing pediatric FH diagnostic criteria in real-world cohorts and to develop two novel diagnostic tools: a semi-quantitative scoring system (FH-PeDS) and a machine learning model (ML-FH-PeDS) to enhance early FH detection.
Methods: Five established FH diagnostic criteria were assesed (Dutch Lipid Clinics Network [DLCN], Simon Broome, EAS, Simplified Canadian, and Japanese Atherosclerosis Society) in Slovenian (N=1,360) and Portuguese (N=340) pediatric hypercholesterolemia cohorts, using FH-causing variants as the reference standard. FH-PeDS was developed from the Slovenian cohort, and ML-FH-PeDS was trained and tested using a 60%/40% split before external validation in the Portuguese cohort.
Results: Only 47.4% of genetically confirmed FH cases were identified by all established criteria, while 10.9% were missed entirely. FH-PeDS outperformed DLCN in the combined cohort (AUC 0.897 vs. 0.857; p<0.01). ML-FH-PeDS showed superior predictive power (AUC 0.932 in training, 0.904 in testing vs. 0.852 for DLCN; p<0.01) and performed best as a confirmatory test in the testing subgroup (39.7% sensitivity, 87.7% PPV at 98% specificity). In the Portuguese cohort, ML-FH-PeDS maintained strong predictive performance (AUC 0.867 vs. 0.815 for DLCN; p<0.01) despite population differences.
Conclusions: Current FH diagnostic criteria perform suboptimally in children. FH-PeDS and ML-FH-PeDS provide tools to improve FH detection, particularly where genetic testing is limited. They also help guide genetic testing decisions for hypercholesterolemic children. By enabling earlier diagnosis and intervention, these tools may reduce long-term cardiovascular risk and improve outcomes.Lay Summary: - Familial hypercholesterolemia (FH) is a common inherited condition that causes high cholesterol from childhood and increases heart disease risk, but it is often missed early in life; - We developed two new tools—FH-PeDS and ML-FH-PeDS—that identify children with FH more accurately than current diagnostic scores; - These tools can help clinicians decide which children need genetic testing, especially in countries where such testing is limited.This work was supported by the Slovenian Research and Innovation Agency (grants: P3-0343 and J3-2536). The Portuguese FH Study has been funded by Science and Technology Foundation and Portuguese Cardiology Society.
B.M. acknowledges funding support from ‘la Caixa’ Foundation and Fundação para a Ciência e Tecnologia under the grant agreement LCF/PR/HP23/52330032. S.E.H. was supported by a grant from the British Heart Foundation (BHF grant PG 08/008) and received additional support from the National Institute for Health Research University College LondonHospitals Biomedical Research Centre. T.F. was supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (programme EXCELES, ID Project No. LX22NPO5104)—Funded by the European Union—Next Generation EU
Oral health behavior associated with cardiometabolic outcomes: A nationally representative cross-sectional study in Portugal
No full textBackground: Oral diseases are associated with the development of cardiometabolic diseases. This study aimed to evaluate the relationship between oral health behaviors (tooth brushing and oral health appointments) with cardiometabolic diseases.
Methods: Data from the First National Health Examination Survey were used. Participants aged 25-74 years and diagnosis of acute myocardial infarction (n = 4442), stroke (n = 4441), hypertension (n = 4450) and diabetes (n = 4327) were analyzed. A fifth subsample (n = 2555) included participants aged 40-69 for calculating cardiovascular risk. Poor oral health behavior was defined as brushing once a day or less and having the last oral health appointment at 12 months or more. Poisson regression models assessed the relationship between poor oral health behavior and these cardiometabolic outcomes.
Results: Among 4.450 participants, 20.5 % had poor oral health behavior. A statistically significant association was found between poor oral health behavior with diabetes (PR: 1.44 [95 % CI: 1.10-1.98], and high/very high cardiovascular risk (PR: 1.42 [95 % CI: 1.25-1.62]). In the sensitivity analysis the association with diabetes and high/very high cardiovascular risk persisted when considering only brushing behavior but not when considering only oral health appointments at 12 months or more.
Conclusions: Individuals with poor oral health behavior had a higher prevalence of diabetes and high/very high cardiovascular risk. Sensitivity analysis suggested that regular tooth brushing may be the main behavior for preventing diabetes and cardiovascular risk. The results suggest that regular tooth brushing may act in prevention for diabetes and cardiovascular risk.Highlights: - Oral health is associated with cardiovascular outcomes; - Poor oral health increased the prevalence of diabetes and cardiovascular risk; - Regular tooth brushing may help prevent diabetes and cardiovascular risk.INSEF development was part of the project “Improvement of epide miological health information to support public health decision and management in Portugal. Towards reduced inequalities, improved health, and bilateral cooperation”, benefiting a €1,500,000 Grant from Iceland, Liechtenstein and Norway through the EEA Grants. This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors
Surveying genetic markers of antibiotic resistance and genomic background in Chlamydia trachomatis: insights from a multiplex NGS-based approach in clinical strains from Portugal
No full textObjectives: To survey genetic markers of potential antimicrobial resistance (AMR) to macrolides and fluoroquinolones among Chlamydia trachomatis–positive samples from the collection of the Portuguese National Reference Laboratory for Sexually Transmitted Infections (STIs), and explore a multiplex PCR approach coupled with NGS to provide complementary information regarding a strain’s genomic backbone.
Methods: A total of 502 C. trachomatis–positive samples, mostly anorectal exudates, were subjected to PCR and sequencing of five targets, including loci potentially driving AMR (23S rRNA, gyrA and parC) and loci potentially informative about a strain’s genomic backbone with emphasis on differentiation of lymphogranuloma venereum (LGV)/non-LGV and L2/L2b (a 9 bp insertion in pmpH, a 74 bp insertion upstream from CT105 and the polymorphic CT442).
Results: No samples evidenced 23S rRNA mutations recognizably linked to macrolide resistance. Three samples harboured the Ser83Ile mutation in GyrA putatively driving fluoroquinolone resistance: two recombinant L2-L2b/D-Da (0.4%) and one L2 (0.2%). The screened regions in pmpH, upstream CT105 and CT442 were fully concordant with LGV/non-LGV differentiation. As expected, the pmpH L2b-specific genetic trait locus was detected in all L2b and recombinant L2-L2b/D-Da ompA genotypes, but also in 96.0% of L2 specimens, which also likely possess an L2b genomic backbone. The insertion upstream from CT105 exhibited full LGV specificity, constituting a promising target for the development of rapid LGV diagnostic assays.
Conclusions: This study contributes to enhancing the knowledge of C. trachomatis molecular epidemiology, suggesting that the known genetic determinants of AMR are not disseminated in clinical C. trachomatis strains, and presents an exploratory approach that can be suitable for LGV/non-LGV and L2/L2b genomic background differentiation
Boletim Epidemiológico Observações: Vol. 14, Nº 38, mai-ago 2025
No full textObservações é uma publicação científica do INSA, IP, que visa contribuir para o conhecimento da saúde da população, os fatores que a influenciam, a decisão e a intervenção em Saúde Pública, assim como a avaliação do seu impacte na população portuguesa. Através do acesso público e gratuito a resultados científicos gerados por atividades de observação em saúde, monitorização e vigilância epidemiológica nas áreas de atuação do Instituto - Alimentação e Nutrição, Doenças Infeciosas, Genética Humana, Saúde Ambiental, Promoção da Saúde e Prevenção de Doenças Não Transmissíveis, Epidemiologia, Investigação em Serviços e Políticas de Saúde - é dada especial atenção à disseminação rápida de informação relevante para a resposta a temas de relevo para a saúde da população portuguesa, tendo como principal alvo todos os profissionais, investigadores e decisores intervenientes na área da Saúde Pública em Portugal