National Institute of Health Dr. Ricardo Jorge
Repositório Científico do Instituto Nacional de SaúdeNot a member yet
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Effect of pomegranate peels and extract in barrier, optical and mechanical properties of polylactic acid-based active packaging
Get PDFBeing more than 50 % of pomegranate (Punica granatum L.) constituted by non-edible parts, namely peels (50%) and seeds (10%), pomegranate is an excellent source of by-products. Its peels and seeds present excellent antioxidant and antimicrobial activities and a high content of phenolic compounds, namely ellagitannins.
This work aimed to evaluate the mechanical and optical properties of two polylactic acid (PLA)-based active packaging with 3 wt.% pomegranate peels (3PP) or 3 wt.% pomegranate peel extract (3PPE). All the samples were produced on a laboratory scale with techniques and processing conditions used in industry. The production of packaging with flexible films is mainly carried out by tubular film extrusion. With this processing method it is possible to produce samples with molecular orientation and reduced thickness equal to that of the packages currently on the market. The structural and morphological characterization of the films were evaluated by FTIR and SEM, and the color by UV-vis. Water vapor transmission and mechanical properties were also measured. The color was measured by Shimadzu UV2401PC reflectance spectrophotometer. Water vapor transmission, oxygen permeability and mechanical properties were also measured.
The FTIR and SEM results indicate the incorporation of the pomegranate peels and peels extract in the PLA matrix, where PLA/3PPE showed better particle homogenization than the PLA/3PP. Regarding the color variations, the PLA/3PPE presented higher variations in terms of L*, a*, and b*. The incorporation of pomegranate derivates has a negative effect on the tensile strength and Young modulus, but a significant increase of the elongation at break for PLA/3PPE. The PLA film's water vapor barrier properties do not suffer any alteration with the incorporation of pomegranate extract.Cássia H. Barbosa thanks the Fundação para a Ciência e Tecnologia (FCT), Portugal, for
the PhD. Grant 2021.08154.BD. This work was carried out under the MobFOOD Project (POCI-01-0247-FEDER-024524 and LISBOA-01-0247-FEDER-024524), funded by POCI (Operational Programme “Competitiveness and Internationalization”) and POR Lisboa (Lisbon Regional Operational Programme), through ANI, and by the Programa de Cooperación Interreg-A España–Portugal (POCTEP) 2014–2020 (project 0377_IBERPHENOL_6_E). This research was also funded by PT national funds (FCT/MCTES, Fundação para a Ciência e Tecnologia and Ministério da Ciência, Tecnologia e Ensino Superior) through the grant UIDB/00211/2020).info:eu-repo/semantics/publishedVersio
The complexity of identification of pathogenic variants
Get PDFIntroduction: Natural occurring genomic variant, from single nucleotide to balanced, unbalanced and complex rearrangements, spanning large chromosomal regions, has been reported to cause human pathologies. As such, we present cases with neurodevelopmental disorder, infertility, and recurrent miscarriage, which reflect the complexity of the identification of pathogenic variants, considering the variation spectrum, the underlying pathogenic mechanisms, and the heterogeneous clinical presentations.
Methods: Long and small insert genomic sequencing (GS) was applied to four cases. Variants were identified from GS data mapped against the reference human genome and confirmed through Sanger sequencing. Results were interpreted using SVInterpreter, Exomiser, genotype-phenotype correlation and convergent genomic data analysis.
Results: Although the first case is a carrier of a t(17;19)(p13.1;p13.3)mat, disrupting GSG1L2, and of a presumably paternally inherited dup(2)(q14.3q21.1), encompassing the autosomal dominant (AD) phenotype-associated PROC and HS6ST1 genes, the identified novel frameshift c.4442del, p.(Gly1481Valfs*21) variant of CHD4, was considered the disease-causing variant, since the proband’s phenotype fits the CHD4-associated Sifrim-Hitz-Weiss syndrome (Da Silva et al., 2022). Cases 2 and 3 were both reported with infertility, and carriers of t(5;9)(q31.3;p13) and t(4;21)(p14;q21.3), respectively. Our study revealed that the phenotype most plausibly resulted from a chromosomal position effect over YIPF5 and SPATC1L. The last case, presented intellectual disability and recurrent miscarriage, associated to t(7;22)(p13;q13.1). The 7p13 breakpoint disrupts the brain specific CAMK2B, causing AD mental retardation 54 (OMIM #617799), whereas increased meiotic segregation of der(22), during gametogenesis, most likely explain the reported miscarriage (David et al., 2023).
Conclusions: These cases highlight the intricacy of pathogenic mechanisms leading to human disorders, the necessity for identification and evaluation of the “full” spectrum of genomic and genetic variants, of comparative reverse phenotyping, including patients with pathogenic variants affecting the same genes. Finally, highlight the need of introducing a more precise genomic medicine in clinical practice.
This research was supported by FCT—Fundação para a Ciência e a Tecnologia, Research Grant HMSP-ICT/0016/2013 of the Harvard Medical School—Portugal Program.Study supported by Fundação para a Ciência e Tecnologia (HMSP-ICT/0016/2013).info:eu-repo/semantics/publishedVersio
ABCG5 and ABCG8 variants associated to sitosterolemia in ClinVar – state of art
No full textSitosterolemia is a rare lipid disorder characterized by the accumulation of plant sterols in the blood, which can lead to several cardiovascular complications. This autosomal recessive disorder is due to pathogenic alterations in ABCG5 and ABCG8 genes. We aimed to assess ClinVar information regarding ABCG5/8 variants possibly causing sitosterolemia. ClinVar was consulted, and all variants submitted as related to “sitosterolemia” were considered (even if some of these were also associated with other phenotypes such as “cardiovascular phenotype”). ClinVar review status of two stars (multiple submissions, no conflicts), one star (single submissions or conflicting interpretations) and no stars (no assertion criteria provided) were considered. The file extracted for ABCG5 analysis contained 295 variants and the file extracted for ABCG8 contained 138 variants. In both files appeared variants mentioning both genes ABCG5/ABCG8 (=11). These were not considered for the descriptive analysis to avoid overcounting. Concerning ABCG5 variants (n=295), 24 are classified as pathogenic/likely pathogenic (3 with review status of two stars, 7 with one, and 6 with no stars); 67 as benign/likely benign (25 with two stars and 42 with one star). Additionally, 160 were classified as variants of uncertain significance (VUS) (61 with two stars, 98 with one, and 1 with no stars), and 44 have conflicting interpretations of pathogenicity (review status of one star). In the ABCG8 variants file (n=138), 16 are classified as pathogenic/likely pathogenic (7 with review status of two stars, 5 with one star, and 4 with no stars); and 26 as benign/likely benign (all with two stars). Additionally, 59 were found classified as VUS (28 with two stars and 31 with one star), and 37 have conflicting interpretations of pathogenicity (review status of one star). Despite there is information in ClinVar about these variants being found in homozygosity/compound heterozygosity in sitosterolemia patients (enabling a genotype-phenotype analysis), the great majority of variants in both genes lack other kind of information as functional characterization and in silico prediction analysis. ClinVar constitutes a fundamental tool for data sharing and to be consulted to know if a specific variant has been reported elsewhere but lacks other important information for variant classification. Although the majority are nonsense variants there are still many missense variants that need other type of information to be classified as pathogenic and this poses an important gap in sitosterolemia diagnosis. It remains crucial to improve the classification and knowledge of ABCG5/8 variants since the correct genetic diagnosis of sitosterolemia is vital for a personalized treatment.N/
Epidemiology of invasive meningococcal disease in Portugal from 2012 to 2022
No full textBackroud and aim: Surveillance of invasive meningococcal disease (IMD) is essential to monitor changes in the epidemiology of the disease and for
more effective infection control.
In Portugal, a surveillance system based on mandatory clinical and laboratory notifications was implemented in late 2002, with
the laboratory component under the responsibility of the National Reference Laboratory (NRL) for Neisseria meningitidis. In
addition, since 2017, whole genome sequencing (WGS) has been routinely implemented at the NRL as a reference typing method
for IMD surveillance. This work aims to analyze both the epidemiology of IMD and the genetic diversity of Neisseria meningitidis strains during the
last 11 years (2012-2022) in Portugal. Conclusion: Although the incidence of meningococcal disease has decreased over the past 11 years in Portugal, MenB meningococci are still an important cause of meningitis and septicaemia. While serogroup C IMD was rare and restricted to adults, serogroup Y IMD affected all age groups. On the other hand, serogroup W IMD has been increasing since 2017, initially in adults and later (2019) in children under 4 years of age.
WGS analysis revealed a high genetic diversity among N. meningitidis isolates, especially in the population of MenB isolates. The increased
circulation of MenB cc213 strains observed over these nine years is particularly important and deserves close monitoring, given the predicted
low coverage of this strain by the 4CMenB vaccine [6]. In addition, the slight increase in IMD cases due to the MenW cc11 strains, previously
identified as clustered into two sublineages the “Original UK” and “UK 2013” strains [7], should also be monitored, as these strains are
associated with an unusual clinical presentation and a higher lethality rate compared to other IMD serogroups [8, 9].
Our results underline the need for continuous surveillance of N. meningitidis infections susceptible to changes in their pattern, in order to
promptly adapt IMD control strategies in Portugal.info:eu-repo/semantics/draf
Improving data on overweight, obesity and undernutrition among children under the age of 5 years in the WHO European Region
No full textIntroduction:
It is important for countries to be able to examine their progress toward the Sustainable Development
Goals on malnutrition. Unfortunately, in the WHO European Region, there is limited and sparse crosssectional anthropometric measured data at national level from children under five years of age. The
WHO European Office for the Prevention and Control of Noncommunicable Diseases in collaboration
with WHO Headquarters and as part of its participation in the European Union funded project
“Science Technology Obesity Policy” (STOP), is exploring to address these data gaps. Therefore, in
October 2022, the WHO Regional Office for Europe convened an expert meeting to discuss the
current overview of data availability, data collection, and next steps to move forward.
Methods:
Key stakeholders working in the areas related to childhood obesity surveillance were invited to this
meeting to discuss and explore the availability of data, the feasibility, generalisability and practicality
of anthropometric data collection in children under five and suggest next steps to move forward.
Results:
Addressing the challenge of childhood obesity in Europe was discussed, as well as the importance of
anthropometric data on children under five years of age. Results of a survey from 31 European
countries on the availability of anthropometric data in this age group were presented, focusing on data
available from “routine health checks”. Future work is needed to identify the feasibility of accessing
this data for surveillance and research purposes. Three Member States, namely Italy, Portugal and
Latvia, presented their experience in data collection and perspectives on how to improve data on
children under five. Discussions took place on the feasibility, generalisability and practicality of
anthropometric data collection, and expected challenges and solutions. It was discussed that further
explorations need to be done to harmonize joint data collection efforts coming from different sources
within national health information systems.
Conclusion:
We concluded that it is important to move on from the idea of a perfect, ideal data source. All data
sources — and the possibility of combining data from different sources — should be explored. Latvia
has demonstrated the feasibility of a kindergarten-based survey and several countries have indicated
willingness to participate in similar surveys. Other data sources should be further explored — it is
important to gather enough information from routine data sources to be able to use and interpret
these data (and combine them with other data). This is particularly important because kindergarten-based data will reach the older children in the under-five age group, while there tends to be higher
coverage of younger children through routine data from paediatric systems.This study was supported by European Union’s Horizon 2020 Research and Innovation programme
(Grant No. 774548) and the WHO European Office for the Prevention and Control of NCDs and
selected Member States from WHO/EURO.info:eu-repo/semantics/publishedVersio
LDLR variant classification by the ClinGen Familial Hypercholesterolemia Variant Curation Expert Panel - first 10 rounds of curation
No full textFamilial hypercholesterolemia (FH) is the most common monogenic disorder of lipid metabolism associated with increased cardiovascular risk. FH is caused by pathogenic variants in three main FH genes, of which LDLR is the most frequent. Genetic testing can confirm the clinical diagnosis, but there are currently over 3700 different LDLR variants deposited in ClinVar.
In 2022 the Clinical Genome Resource FH Variant Curation Expert Panel (VCEP) recommendations to classify LDLR variants were published. Here, we present the results of the first ten rounds of LDLR variant classification by the FH VCEP.
The FH VCEP is currently composed of 12 reviewers, 19 curators, and 10 associated laboratories. Training of biocurators started in August/2021, sustained curation in November/2021, and the most recent round (10th) will end in June/2023.
Variants with conflicting classifications in ClinVar and other variants in the same codon (required to properly classify variants) are being prioritized.
The FH VCEP preparation team is composed of one alternating curator that uploads de-identified case data sent from associated labs, and one dedicated curator that screens and uploads published papers with functional studies into ClinGen’s Variant Curation Interface (VCI) prior to the curation round. Each variant is assessed using the VCI independently by two curators or one senior curator and approved by three reviewers before being officially published to the Evidence Repository and ClinVar.
Once we complete our first ten curation rounds, we will have evaluated 451 LDLR variants: 25 with previous ClinVar review status of no stars (criteria not provided), 333 with one-star (conflicting and single submitter) and 93 with two-stars (multiple submitters, no conflicts). Prior ClinVar classifications were 246 conflicting, 148 Pathogenic/Likely pathogenic (P/LP), 45 Uncertain Significance (VUS), 10 Benign/Likely benign (B/LB) and 2 not provided, totalling only 35% of variants with definite classifications.
Currently, 231 variants are published in ClinVar at three-star status, 85 are approved and in the process of being published, 64 are nearly approved and 71 are under evaluation.
Among the 374 variants already with an FH VCEP classification, 157 were classified as P/LP, 186 as VUS, 24 as B/LB and only 7 remained conflicting, improving the number of definite classifications to 48% and decreasing the number of conflicting classifications by ~35-fold.
Accurate genetic diagnosis relies on correct variant interpretation. FH VCEP guidelines significantly decrease conflicting classifications and provide expert-level consensus data that will facilitate genetic diagnosis, ultimately improving patient management and prognosis.N/
Remote monitoring of disease vector mosquitoes with a new optical sensor system for automatic classification
Get PDFIntroduction:
- Mosquitoes represent a major threat to public health given their ability to transmit several pathogens. Some species of Aedes can
transmit viruses such as dengue, Zika, or chikungunya.
- Prevention of vector-borne diseases largely depends on effective and sustainable vector surveillance.
Objectives:
- Help to develop a novel bioacustic sensor that is able to identify the
mosquitoes' species in real time
- Deploy the sensor in the field in Madeira and Algarve.N/
Effects and Mechanisms of Action of Preussin, a Marine Fungal Metabolite, against the Triple-Negative Breast Cancer Cell Line, MDA-MB-231, in 2D and 3D Cultures
No full textTriple-negative breast cancer (TNBC) represents an aggressive subtype of breast cancer (BC) with a typically poorer prognosis than other subtypes of BC and limited therapeutic options. Therefore, new drugs would be particularly welcome to help treat TNBC. Preussin, isolated from the marine sponge-associated fungus, Aspergillus candidus, has shown the potential to reduce cell viability and proliferation as well as to induce cell death and cell cycle arrest in 2D cell culture models. However, studies that better mimic the tumors in vivo, such as 3D cell cultures, are needed. Here, we studied the effects of preussin in the MDA-MB-231 cell line, comparing 2D and 3D cell cultures, using ultrastructural analysis and the MTT, BrdU, annexin V-PI, comet (alkaline and FPG modified versions), and wound healing assays. Preussin was found to decrease cell viability, both in 2D and 3D cell cultures, in a dose-dependent manner, impair cell proliferation, and induce cell death, therefore excluding the hypothesis of genotoxic properties. The cellular impacts were reflected by ultrastructural alterations in both cell culture models. Preussin also significantly inhibited the migration of MDA-MB-231 cells. The new data expanded the knowledge on preussin actions while supporting other studies, highlighting its potential as a molecule or scaffold for the development of new anticancer drugs against TNBC.This research was partially supported by the strategic funding UIDB/04423/2020 and UIDP/04423/2020 through national funds provided by FCT—Fundação para a Ciência e a Tecnologia to CIIMAR. The Master in Oncology of the ICBAS—U.Porto offered additional support.info:eu-repo/semantics/publishedVersio
How can multi-national occupational studies support policy making in Europe? Experiences from HBM4EU and PARC occupational studies
No full textProviding policy relevant data on chemical exposures was a major aim of the EU biomonitor-ing initiative, HBM4EU. Similarly, the recently launched “Partnership for the risk assessment of chemicals” (PARC) emphasizes the relevance of the research findings to regulatory deci-sion making. Occupational biomonitoring studies performed under HBM4EU were focused on HBM4EU priority substances with specific regulatory relevance. Chromates and diisocya-nate studies were developed since these substances are of high concern for workers and to support regulatory measures under both EU chemicals regulation (REACH) and occupational safety and health (OSH) legislation. E-waste study was targeted to support European circular economy strategies and identify issues e.g. under OSH legislation. Based on the results of the chromates study, we drew several policy relevant conclusions which support the future up-dating of occupational exposure limits and the development of monitoring and risk man-agement practices in industry. A survey to policy makers on the usefulness of the study re-sults gave us confidence. We emphasise the importance of early and continued engagement of policy makers to ensure the usability of the results. Early communication with regulators of the anticipated benefits and formatting outputs appropriately are important and have been considered and improved within the EU PARC project.This project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No 733032 and received co-funding from the author’s organizations and/or Ministries.info:eu-repo/semantics/publishedVersio
Effects of TiO2 Nanoparticles on the Genome-Wide Methylation of Human Epithelial Intestinal Cells
No full textIntroduction: Titanium dioxide nanoparticles (TiO2NP) have multiple applications in industry (e.g., engineering, cosmetics, food additives), and biomedicine (e.g., targeted drug delivery and biosensing). Food-grade TiO2 (E171) is applied as a food additive to whiten and improve the opacity of food products, while also having the ability to enhance its flavour. In 2021, EU member states banned E171 from all food products, since there is doubts about its genotoxicity. Nevertheless, the ingestion of TiO2NPs may still occur through to other sources, such as contaminated food or water, consumer products (e.g., toothpaste and lipstick) or pharmaceutics. To date, there is some in vitro evidence that TiO2NP may induce changes in DNA methylation. However, very few studies were performed, and none used genome-wide approaches to identify possible differentially methylated genes induced by TiO2NP exposure, and its impact on molecular pathways.
Methodology: Caco-2 epithelial intestinal cells were exposed to 14 μg/ml of anatase, rutile or brookite phase TiO2NP for 24h. Genomic DNA was extracted from exposed and non-exposed cells. DNA libraries were generated using the Premium Reduced Representative Bisulfite Sequencing (RRBS) kit (Diagenode) and sequenced on the NextSeq 550 system (Illumina) using 100 bp paired reads. The Galaxy platform was used for read treatment and mapping, methylation calling and assessing of differentially methylated regions between exposed and non-exposed cells. Pathway analysis was performed using Reactome, and gene ontology analysis with the ClueGO plugin in Cytoscape.
Results: Significant differential methylation (p ≤ 0.05) of 92 genes (21 hyper- and 71 hypo-methylated), 70 genes (12 hyper- and 58 hypo-methylated) and 88 genes (21 hyper- and 67 hypo-methylated) was observed for the anatase, rutile and brookite phase TiO2NP, respectively. Functional pathway analysis of these methylation changes identified several relevant cellular pathways that may be altered by exposure, such as G alpha signalling events, being some associated to colon cancer.
Conclusions: All types of TiO2NP induce changes in genome methylation of intestinal cells, which may affect cell proliferation, differentiation and survival. Moreover, although some dysfunctional pathways are shared between the three TiO2NP, many are type-specific, suggesting different molecular mechanisms of action for each TiO2NP.Funding: FCT (PTDC/SAU-PUB/29481/2017), FCT UIDB/00009/2020 UIDP/00009/2020.info:eu-repo/semantics/publishedVersio