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Sleep is a vital process that impacts biological functions such as cell renewal, bone regeneration, and immune system support. Disrupted sleep can interrupt erythropoiesis, leading to fewer red blood cells, reduced haemoglobin concentration, and decreased haematocrit levels, potentially contributing to haematological disorders. This is particularly concerning for shift workers for example firefighters. While previous studies have explored sleep's adverse effects on various professions, research specific to firefighters is limited. This study investigates the relationship between sleep quality and haematological parameters among firefighters in Northeast Portugal. From a sample of 201 firefighters, variations in red blood cells, haemoglobin, and haematocrit values were linked to sleep quality. The study utilised non-parametric tests (Wilcoxon-Mann-Whitney, Spearman's correlation) to explore the connection between sleep quality and haematological profile. The impact of covariates on haematological parameters was assessed using non-parametric ANCOVA (Quade's). A multiple regression analysis was employed to further understand how sleep quality and various confounding variables impact haematological levels. Findings suggest a negative link between sleep quality and haematological levels, meaning that as sleep quality deteriorates, there is a tendency for haematological levels to decrease, as indicated by Spearman's correlation (rRBC = -0.157, pRBC = 0.026; rHb = -0.158, pHb = 0.025; rHCT = -0.175, pHCT = 0.013). As observed in scientific literature, the correlation found suggests a possible inhibition of erythropoiesis, the process responsible for red blood cell production. Despite firefighters presenting a haematological profile within the reference range (RBC: 5.1 × 10 6 /mm 3 (SD ± 0.4), Hb: 15.6 g/dL (SD ± 1.3), 47% (SD ± 1.0), there is already an observable trend towards lower levels. The analysis of co-variables did not reveal a significant impact of sleep quality on haematological levels. In conclusion, this study underscores the importance of sleep quality in determining haematological parameters among firefighters. Future research should investigate the underlying mechanisms and long-term implications of poor sleep quality on firefighter health. Exploring interventions to enhance sleep quality is vital for evidence-based strategies promoting firefighter well-being.

Rett syndrome (RTT) is a rare neurodevelopmental disorder caused by mutation in the X-linked gene methyl-CpG-binding protein 2 (Mecp2), a ubiquitously expressed transcriptional regulator. RTT results in mental retardation and developmental regression that affects approximately 1 in 10,000 females. Currently, there is no curative treatment for RTT. Thus, it is crucial to develop new therapeutic approaches for children suffering from RTT. Several studies suggested that RTT is linked with defects in cholesterol homeostasis, but for the first time, therapeutic evaluation is carried out by modulating this pathway. Moreover, AAV-based CYP46A1 overexpression, the enzyme involved in cholesterol pathway, has been demonstrated to be efficient in several neurodegenerative diseases. Based on these data, we strongly believe that CYP46A1 could be a relevant therapeutic target for RTT. Herein, we evaluated the effects of intravenous AAVPHP.eB-hCYP46A1-HA delivery in male and female Mecp2 -deficient mice. The applied AAVPHP.eB-hCYP46A1 transduced essential neurons of the central nervous system (CNS). CYP46A1 overexpression alleviates behavioral alterations in both male and female Mecp2 knockout mice and extends the lifespan in Mecp2-deficient males. Several parameters related to cholesterol pathway are improved and correction of mitochondrial activity is demonstrated in treated mice, which highlighted the clear therapeutic benefit of CYP46A1 through the neuroprotection effect. IV delivery of AAVPHP.eB-CYP46A1 is perfectly well tolerated with no inflammation observed in the CNS of the treated mice. Altogether, our results strongly suggest that CYP46A1 is a relevant target and overexpression could alleviate the phenotype of Rett patients.

Colorectal cancer (CRC) is a leading cause of death worldwide. Conventional therapies are available with varying effectiveness. Acetate, a short-chain fatty acid produced by human intestinal bacteria, triggers mitochondria-mediated apoptosis preferentially in CRC but not in normal colonocytes, which has spurred an interest in its use for CRC prevention/therapy. We previously uncovered that acetate-induced mitochondrial-mediated apoptosis in CRC cells is significantly enhanced by the inhibition of the lysosomal protease cathepsin D (CatD), which indicates both mitochondria and the lysosome are involved in the regulation of acetate-induced apoptosis. Herein, we sought to determine whether mitochondrial function affects CatD apoptotic function. We found that enhancement of acetate-induced apoptosis by CatD inhibition depends on oligomycin A-sensitive respiration. Mechanistically, the potentiating effect is associated with an increase in cellular and mitochondrial superoxide anion accumulation and mitochondrial mass. Our results provide novel clues into the regulation of CatD function and the effect of tumor heterogeneity in the outcome of combined treatment using acetate and CatD inhibitors.

Among the many lysosomal storage disorders (LSDs) that would benefit from the establishment of novel cell models, either patient-derived or genetically engineered, is mucopolysaccharidosis type II (MPS II). Here, we present our results on the establishment and characterization of two MPS II patient-derived stem cell line(s) from deciduous baby teeth. To the best of our knowledge, this is the first time a stem cell population has been isolated from LSD patient samples obtained from the dental pulp. Taking into account our results on the molecular and biochemical characterization of those cells and the fact that they exhibit visible and measurable disease phenotypes, we consider these cells may qualify as a valuable disease model, which may be useful for both pathophysiological assessments and in vitro screenings. Ultimately, we believe that patient-derived dental pulp stem cells (DPSCs), particularly those isolated from human exfoliated deciduous teeth (SHEDs), may represent a feasible alternative to induced pluripotent stem cells (iPSCs) in many labs with standard cell culture conditions and limited (human and economic) resources.

Background and Objectives : Research on female soccer players that analyzes playing status is scarce and has previously only examined load monitoring, while other markers, such as physical (i.e., strength, power, and agility), physiological (i.e., maximal oxygen uptake), and body composition (i.e., body fat mass, fat-free mass, body water, and phase angle) markers, warrant further investigation. Thus, the study aims were to (a) compare physical, physiological, body composition, and load markers between starters and non-starters; (b) compare measurements pre- and post-training intervention (five weeks); and (c) analyze any relationships between physical, physiological, body composition, and load markers in an elite female soccer team. Materials and Methods : Fourteen first-team players participated in the study (age 23.29 ± 3.19 years, weight 59.14 ± 6.87 kg, height 1.66 ± 0.08 m). Several physical ( n = 15), physiological ( n = 1), body composition ( n = 11), and load markers ( n = 14) were collected. In addition, participants were sub-divided into starters ( n = 7) and non-starters ( n = 7). Results : No differences were revealed between starters and non-starters in any of the examined variables. Moreover, following the training intervention, a significantly lower value was found for total body water/fat-free mass ratio (p = 0.043; ES = 0.582). In addition, there were several correlations detected between load and physical/physiological markers ( n = 28); load and body composition markers ( n = 6); physical/physiological and body composition markers ( n = 34); and physical and physiological markers ( n = 42). Conclusions: In conclusion, only a slight tendency of higher load values for starters than non-starters was observed. In addition, no differences in physical, physiological, and body composition markers were found between starters and non-starters, possibly suggesting that five weeks were not enough to improve such variables. Finally, the present results provide novel information assessing the effects of the pre-season in elite female Portuguese soccer players and contribute to a better understanding of the associations between different types of measurements.

Canine atopic dermatitis (cAD) is a common and distressing skin condition in dogs, affecting up to 30% of the canine population. It not only impacts their quality of life but also that of their owners. Like human atopic dermatitis (hAD), cAD has a complex pathogenesis, including genetic and environmental factors. Current treatments focus on managing clinical signs, but they can be costly and have limitations. This article emphasizes the importance of preventing cAD from developing in the first place. Understanding the role of the skin's protective barrier is crucial, as its dysfunction plays a vital role in both hAD and cAD. hAD prevention studies have shown promising results in enhancing the skin barrier, but more research is needed to support more robust conclusions. While hAD primary prevention is currently a focal point of intensive investigation in human medicine, research on cAD primary prevention remains under-researched and almost non-existent. Pioneering effective prevention strategies for cAD holds immense potential to enhance the quality of life for both dogs and their owners. Additionally, it bears the promise of a translational impact on human research. Hence, further exploration of this crucial topic is not only relevant but also timely and imperative, warranting support and encouragement.

Niemann-Pick type C (NPC, ORPHA: 646) is a neuro-visceral, psychiatric disease caused predominantly by pathogenic variants in the NPC1 gene or seldom in NPC2 . The rarity of the disease, and its wide range of clinical phenotypes and ages of onset, turn the diagnosis into a significant challenge. Other than the detailed clinical history, the typical diagnostic work-up for NPC includes the quantification of pathognomonic metabolites. However, the molecular basis diagnosis is still of utmost importance to fully characterize the disorder. Here, the authors provide an overview of splicing variants in the NPC1 and NPC2 genes and propose a new workflow for NPC diagnosis. Splicing variants cover a significant part of the disease-causing variants in NPC. The authors used cDNA analysis to study the impact of such variants, including the collection of data to classify them as leaky or non-leaky pathogenic variants. However, the presence of naturally occurring spliced transcripts can misdiagnose or mask a pathogenic variant and make the analysis even more difficult. Analysis of the NPC1 cDNA in NPC patients in parallel with controls is vital to assess and detect alternatively spliced forms. Moreover, nonsense-mediated mRNA decay (NMD) analysis plays an essential role in evaluating the naturally occurring transcripts during cDNA analysis and distinguishing them from other pathogenic variants' associated transcripts.

Splicing of pre-mRNA is a crucial regulatory stage in the pathway of gene expression. The majority of human genes that encode proteins undergo alternative pre-mRNA splicing and mutations that affect splicing are more prevalent than previously thought. Targeting aberrant RNA(s) may thus provide an opportunity to correct faulty splicing and potentially treat numerous genetic disorders. To that purpose, the use of engineered U1 snRNA (either modified U1 snRNAs or exon-specific U1s-ExSpeU1s) has been applied as a potentially therapeutic strategy to correct splicing mutations, particularly those affecting the 5' splice-site (5'ss). Here we review and summarize a vast panoply of studies that used either modified U1 snRNAs or ExSpeU1s to mediate gene therapeutic correction of splicing defects underlying a considerable number of genetic diseases. We also focus on the pre-clinical validation of these therapeutic approaches both in vitro and in vivo, and summarize the main obstacles that need to be overcome to allow for their successful translation to clinic practice in the future.

Improving antigen presentation is crucial for the success of immunization strategies. Yeasts are classically used as biofactories to produce recombinant proteins and are efficient vehicles for antigen delivery, in addition to their adjuvant properties. Despite the absence of epidemic outbreaks, several vaccine approaches continue to be developed for Zika virus infection. The development of these prophylactic strategies is fundamental given the severity of clinical manifestations, mainly due to viral neurotropism. The present study aimed to evaluate in vivo the immune response induced by P. pastoris recombinant strains displaying epitopes of the envelope (ENV) and NS1 ZIKV proteins. Intramuscular immunization with heat-attenuated yeast enhanced the secretion of IL-6, TNF-α, and IFN-γ, in addition to the activation of CD4 + and CD8 + T cells, in BALB/c mice. P. pastoris displaying ENV epitopes induced a more robust immune response, increasing immunoglobulin production, especially IgG isotypes. Both proposed vaccines showed the potential to induce immune responses without adverse effects, confirming the safety of administering P. pastoris as a vaccine vehicle. Here, we demonstrated, for the first time, the evaluation of a vaccine against ZIKV based on a multiepitope construct using yeast as a delivery system and reinforcing the applicability of P. pastoris as a whole-cell vaccine.

Sleep research has grown over the past decades and investigators are deeply involved in studying sleep and its impact on human health and body regulation. Despite the understanding that insufficient sleep is strongly linked to the development of several disorders, unsatisfactory sleep exposes health and safety to innumerous risks. The present study aims to review and analyze the main results of clinical trials, registered at ClinicalTrials.gov and ICTRT databases, and developed construct strategies to improve sleep quality on firefighters and enhance professionals' sleep and health conditions. The protocol was registered in PROSPERO under number CRD42022334719. Trials registered between first registry and 2022 were included. We retrieved 11 registered clinical trials; seven met eligibility criteria and were included in the review. A relation between sleep disorders, shift work, and occupational health problems was found, and retrieved trials showed that sleep education programs can improve sleep quality and sleep hygiene. Science has already recognized sleep's importance for metabolic functioning and survivorship. Nevertheless, it continues to play a major role in discovering methods to diminish the problems faced. Strategies contemplating sleep education, or intervention, should be presented to fire services to tackle this problem and promote healthier and safer environments.

Falls are a major problem among older adults worldwide. Early detection of risk factors is important to decrease the burden of falls. The “Fall Risk Questionnaire” (FRQ) is a screening tool used to assess personal fall risk awareness in older populations, and it is also used as a behavior change tool. The aim of the present study was to undertake a cultural and linguistic adaptation of the FRQ to allow its use in the Portuguese population. To achieve this, we used the sequential method proposed by Beaton and colleagues. Statistical analysis was conducted by computing the intraclass correlation coefficient and Cronbach’s alpha score for intra-observer reliability. The panel revision demonstrated high concordance for all assessed items. The pretesting results indicated that, in general, the FRQ—Portuguese Version (FRQ-VP) was well accepted, and all items were adequate and easy to comprehend. The ICC and Cronbach’s alpha scores indicated high consistency between assessments (p < 0.01; alfa = 0.992). The FRQ-VP seemed to have good reliability and internal consistency. Because the definition of a fall experience may not be clear to the older population, a careful explanation of this item can lead to a better score computation.

The plasma membrane (PM) protects cells from extracellular threats and supports cellular homeostasis. Some pathogens produce pore-forming toxins (PFTs) that disrupt PM integrity by forming transmembrane pores. High PFT concentrations cause massive damage leading to cell death and facilitating infection. Sub-lytic PFT doses activate repair mechanisms to restore PM integrity, support cell survival and limit disease. Shedding of extracellular vesicles (EVs) has been proposed as a key mechanism to eliminate PFT pores and restore PM integrity. We show here that cholesterol-dependent cytolysins (CDCs), a specific family of PFTs, are at least partially eliminated through EVs release, and we hypothesize that proteins important for PM repair might be included in EVs shed by cells during repair. To identify new PM repair proteins, we collected EVs released by cells challenged with sub-lytic doses of two different bacterial CDCs, listeriolysin O and pneumolysin, and determined the EV proteomic repertoire by LC-MS/MS. Intoxicated cells release similar EVs irrespectively of the CDC used. Also, they release more and larger EVs than non-intoxicated cells. A cluster of 70 proteins including calcium-binding proteins, molecular chaperones, cytoskeletal, scaffold and membrane trafficking proteins, was detected enriched in EVs collected from intoxicated cells. While some of these proteins have well-characterized roles in repair, the involvement of others requires further study. As proof of concept, we show here that Copine-1 and Copine-3, proteins abundantly detected in EVs released by intoxicated cells, are required for efficient repair of CDC-induced PM damage. Additionally, we reveal here new proteins potentially involved in PM repair and give new insights into common mechanisms and machinery engaged by cells in response to PM damage., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Frequently, deep partial and full-thickness skin wounds do not spontaneously regenerate. To restore the normal function of skin, epidermal and dermal components have to be supplied to the wound bed by grafting various substrates. Available options are limited and frequently costly. Herein, authors present a possible approach using 3D skin scaffolds capable of mimicking structure and biological functions of the extracellular matrix, providing, in parallel, a good environment for cell attachment, proliferation and differentiation. Low-molecular weight chitosan-based membranes were prepared by freeze-drying and ionizing radiation techniques to be used as skin scaffolds. Poly (vinyl alcohol), PVA, vinyl pyrrolidone, VP, and gelatin from cold water fish were incorporated. Information regarding membranes' physical-chemical properties from SEM analysis, swelling and weight loss, together with biological response through in vitro assays (using Human Caucasian Fetal Foreskin Fibroblast) allowed the selection of an optimized batch of membranes that was used as skin scaffold in a dorsal rat model wound. The in vivo implantation assays (in Wistar rats) resulted in very promising results: (i) healing process faster than control; (ii) good vascularization; (iii) viable new tissues morphologically functional.

Background: Clinical management guidelines (CMGs) are decision support tools for patient care used by professionals, patients, and family caregivers. Since clinical experts develop numerous CMGs, their technical language hinders comprehension and access by nonmedical stakeholders. Additionally, the views of affected individuals and their families are often not incorporated into treatment guidelines. We developed an adequate methodology for addressing the needs and preferences of family and professional stakeholders regarding CMGs, a recently developed protocol for managing congenital disorders of glycosylation (CDG), a family of rare metabolic diseases. We used the CDG community and phosphomannomutase 2 (PMM2)-CDG CMGs as a pilot to test and implement our methodology., Results: We listened to 89 PMM2-CDG families and 35 professional stakeholders and quantified their CMG-related needs and preferences through an electronic questionnaire. Most families and professionals rated CMGs as relevant (86.5% and 94.3%, respectively), and valuable (84.3% and 94.3%, respectively) in CDG management. The most identified challenges were the lack of CMG awareness (50.6% of families) and the lack of plain language CMG (39.3% of professionals). Concordantly, among families, the most suggested solution was involving them in CMG development (55.1%), while professionals proposed adapting CMGs to include plain language (71.4%). Based on these results, a participatory framework built upon health literacy principles was created to improve CMG comprehension and accessibility. The outputs are six complementary CMG-related resources differentially adapted to the CDG community's needs and preferences, with a plain language PMM2-CDG CMG as the primary outcome. Additionally, the participants established a distribution plan to ensure wider access to all resources., Conclusions: This empowering, people-centric methodology accelerates CMG development and accessibility to all stakeholders, ultimately improving the quality of life of individuals living with a specific condition and raising the possibility of application to other clinical guidelines.

Effective psychological parenting interventions delivered to adoptive parents may prevent serious adjustment difficulties and promote a healthy functioning; however, less is known about adoptive parents' specific parental difficulties and help-seeking behaviors and perceptions, the understanding of which is deemed necessary to design well-informed interventions. This study aimed to describe parental difficulties, help-seeking behaviors, satisfaction with, and perceived barriers to seek, professional help, and acceptability of psychological parenting interventions among Portuguese adoptive parents. Comparisons with biological parents (Study 1) and between adoptive parents that requested adoption-specialized and non-specialized support (i.e., adoption-specialized vs. non-specialized help-seekers) (Study 2) were explored. A cross-sectional online survey was conducted. Participants were 471 adoptive and 552 biological parents of children aged under 18 years who were recruited through schools, adoption agencies, and social networks. They completed measures assessing parental difficulties, help-seeking behaviors, satisfaction with, and perceived barriers to seek, professional help, and acceptability of psychological parenting interventions. Comparison tests between the study groups, Pearson's correlations, analyses of covariance (ANCOVA) and multivariate logistic regressions (MLR) were performed. Parents' well-being and normative parenting challenges were the most frequent difficulties in both groups, but only associated with adoptive parents' help-seeking. Although difficulties related to a child's problems/parent-child relationship were more frequent among adoptive parents, adoption-related difficulties were rarely reported. Adoptive parents sought professional help more frequently, regardless of parental difficulties. Knowledge-related barriers to seeking help were the most frequent among adoptive parents. Adoption non-specialized help was less satisfactory. Acceptability of psychological parenting interventions was high, but dependent on parental difficulties. Implications for post-adoption services' development are discussed.

The present study about falls among the older adult population essentially explores bio-physiological factors. In light of the complexity of the factors that cause these accidents, it is important to identify the safety and self-care practices of institutionalized older adults and their relationship with falls in order to introduce prevention measures and personalized cognitive-behavioral strategies. The objective of this study was to: (a) determine the frequency of falls and their recurrence among institutionalized older adults, and (b) to associate the occurrence of falls among institutionalized adults with or without cognitive impairment to communication and safety practices. This was a quantitative, correlational, and longitudinal study with 204 institutionalized older adults living in two long-term care facilities in Portugal. The Scale of Practices and Behaviors for Institutionalized Elderly to Prevent Falls was administered to the sample. The prevalence of falls at a 12-month follow-up was 41.6%, of which 38.3% were recurring episodes. Older adults with cognitive decline showed lower mean scores for safety practices. Further research with larger samples should explore the relationships between communication and safety practices and falls, their recurrence, and fear of new falls.

Near-infrared spectroscopic (NIR) technique was used, for the first time, to predict volatile phenols content, namely guaiacol, 4-methyl-guaiacol, eugenol, syringol, 4-methyl-syringol and 4-allyl-syringol, of aged wine spirits (AWS). This study aimed to develop calibration models for the volatile phenol's quantification in AWS, by NIR, faster and without sample preparation. Partial least square regression (PLS-R) models were developed with NIR spectra in the near-IR region (12,500-4000 cm -1 ) and those obtained from GC-FID quantification after liquid-liquid extraction. In the PLS-R developed method, cross-validation with 50% of the samples along a validation test set with 50% of the remaining samples. The final calibration was performed with 100% of the data. PLS-R models with a good accuracy were obtained for guaiacol (r 2 = 96.34; RPD = 5.23), 4-methyl-guaiacol (r 2 = 96.1; RPD = 5.07), eugenol (r 2 = 96.06; RPD = 5.04), syringol (r 2 = 97.32; RPD = 6.11), 4-methyl-syringol (r 2 = 95.79; RPD = 4.88) and 4-allyl-syringol (r 2 = 95.97; RPD = 4.98). These results reveal that NIR is a valuable technique for the quality control of wine spirits and to predict the volatile phenols content, which contributes to the sensory quality of the spirit beverages.

Different ageing technology of wine spirits (WSs) has been investigated, but little has been published on the chemical evolution of aged WS during storage in bottle. The purpose of this study was to examine how 12 months of storage in bottle affected the evolution of antioxidant activity (DPPH, FRAP and ABTS assays), total phenolic index (TPI) and low molecular weight (LMW) compounds content of the WSs aged through alternative technology using three micro-oxygenation levels (MOX) and nitrogen control (N). Results revealed the ability of phenolic compounds from aged WSs to scavenge free radicals during storage in bottle. Among the in vitro antioxidant-activity methods, FRAP assay was the more effective to differentiate WSs according to the ageing technology. Concerning the overall influence of storage in bottle on antioxidant activity, and TPI and LMW compounds content, the higher results were obtained for the MOX modalities (O15, O30 and O60), which showed a similar evolution. In summary, this study provides innovative information, demonstrating that the differences between the aged WSs imparted throughout the ageing process (resulting from different MOX levels) were mostly retained, and only slight modifications during storage in bottle were found.

Using low-cost portable air quality (AQ) monitoring devices is a growing trend in personal exposure studies, enabling a higher spatio-temporal resolution and identifying acute exposure to high concentrations. Comprehension of the results by participants is not guaranteed in exposure studies. However, information on personal exposure is multiplex, which calls for participant involvement in information design to maximise communication output and comprehension. This study describes and proposes a model of a user-centred design (UCD) approach for preparing a final report for participants involved in a multi-sensor personal exposure monitoring study performed in seven cities within the EU Horizon 2020 ICARUS project. Using a combination of human-centred design (HCD), human-information interaction (HII) and design thinking approaches, we iteratively included participants in the framing and design of the final report. User needs were mapped using a survey ( n = 82), and feedback on the draft report was obtained from a focus group ( n = 5). User requirements were assessed and validated using a post-campaign survey ( n = 31). The UCD research was conducted amongst participants in Ljubljana, Slovenia, and the results report was distributed among the participating cities across Europe. The feedback made it clear that the final report was well-received and helped participants better understand the influence of individual behaviours on personal exposure to air pollution.

Use of a multi-sensor approach can provide citizens with holistic insights into the air quality of their immediate surroundings and their personal exposure to urban stressors. Our work, as part of the ICARUS H2020 project, which included over 600 participants from seven European cities, discusses the data fusion and harmonization of a diverse set of multi-sensor data streams to provide a comprehensive and understandable report for participants. Harmonizing the data streams identified issues with the sensor devices and protocols, such as non-uniform timestamps, data gaps, difficult data retrieval from commercial devices, and coarse activity data logging. Our process of data fusion and harmonization allowed us to automate visualizations and reports, and consequently provide each participant with a detailed individualized report. Results showed that a key solution was to streamline the code and speed up the process, which necessitated certain compromises in visualizing the data. A thought-out process of data fusion and harmonization of a diverse set of multi-sensor data streams considerably improved the quality and quantity of distilled data that a research participant received. Though automation considerably accelerated the production of the reports, manual and structured double checks are strongly recommended.

Falls are a complex problem, given their multifactorial nature, the comorbidities involved, and due to the dependency of older persons living in nursing homes. Risk, fear of falling, falls themselves, and their recurrence are the main factors behind fragility fractures, lack of independence, and increases in pain prevalence, and other comorbidities in older populations. The objectives of the present quantitative and longitudinal study were: (a) to characterize the cognitive state and fall frequency of older persons living in nursing homes; (b) to analyze the relationship between cognitive status and some fall risk factors; and (c) to associate cognitive decline, gait ability, and muscle strength of the examined institutionalized older persons with fall occurrence and recurrence over 12 months. The participants were 204 older persons who lived in Portuguese nursing homes, and data were collected from January 2019 to February 2020 by consulting medical records and applying the following instruments: the Mini-Mental State Examination, Timed Up and Go Test, and Medical Research Council Manual Muscle Testing Scale. Fall prevalence, assessed in two periods, 12 months apart, was similar in both samples (with and without cognitive decline) and close to 42%, and the annual recurrence rate was 38.3%. Older persons with no cognitive decline showed an association between gait speed and occurrence of first fall and recurrent fall ( p < 0.05). Muscle strength and use of gait aid devices were not related to falls and their recurrence, regardless of mental state.

The Notch-signaling ligand DLL1 has emerged as an important player and promising therapeutic target in breast cancer (BC). DLL1-induced Notch activation promotes tumor cell proliferation, survival, migration, angiogenesis and BC stem cell maintenance. In BC, DLL1 overexpression is associated with poor prognosis, particularly in estrogen receptor-positive (ER + ) subtypes. Directed therapy in early and advanced BC has dramatically changed the natural course of ER + BC; however, relapse is a major clinical issue, and new therapeutic strategies are needed. Here, we report the development and characterization of a novel monoclonal antibody specific to DLL1. Using phage display technology, we selected an anti-DLL1 antibody fragment, which was converted into a full human IgG1 (Dl1.72). The Dl1.72 antibody exhibited DLL1 specificity and affinity in the low nanomolar range and significantly impaired DLL1-Notch signaling and expression of Notch target genes in ER + BC cells. Functionally, in vitro treatment with Dl1.72 reduced MCF-7 cell proliferation, migration, mammosphere formation and endothelial tube formation. In vivo, Dl1.72 significantly inhibited tumor growth, reducing both tumor cell proliferation and liver metastases in a xenograft mouse model, without apparent toxicity. These findings suggest that anti-DLL1 Dl1.72 could be an attractive agent against ER + BC, warranting further preclinical investigation.

Stimuli-responsive polymer materials are used in smart nanocarriers to provide the stimuli-actuated mechanical and chemical changes that modulate cargo delivery. To take full advantage of the potential of stimuli-responsive polymers for controlled delivery applications, these have been grafted to the surface of mesoporous silica particles (MSNs), which are mechanically robust, have very large surface areas and available pore volumes, uniform and tunable pore sizes and a large diversity of surface functionalization options. Here, we explore the impact of different RAFT-based grafting strategies on the amount of a pH-responsive polymer incorporated in the shell of MSNs. Using a "grafting to" (gRAFT- to ) approach we studied the effect of polymer chain size on the amount of polymer in the shell. This was compared with the results obtained with a "grafting from" (gRAFT- from ) approach, which yield slightly better polymer incorporation values. These two traditional grafting methods yield relatively limited amounts of polymer incorporation, due to steric hindrance between free chains in "grafting to" and to termination reactions between growing chains in "grafting from." To increase the amount of polymer in the nanocarrier shell, we developed two strategies to improve the "grafting from" process. In the first, we added a cross-linking agent (gRAFT- cross ) to limit the mobility of the growing polymer and thus decrease termination reactions at the MSN surface. On the second, we tested a hybrid grafting process (gRAFT- hybrid ) where we added MSNs functionalized with chain transfer agent to the reaction media containing monomer and growing free polymer chains. Our results show that both modifications yield a significative increase in the amount of grafted polymer.

Lysosomal storage diseases (LSDs) are a heterogeneous group of genetic disorders with variable degrees of severity and a broad phenotypic spectrum, which may overlap with a number of other conditions. While individually rare, as a group LSDs affect a significant number of patients, placing an important burden on affected individuals and their families but also on national health care systems worldwide. Here, we present our results on the use of an in-house customized next-generation sequencing (NGS) panel of genes related to lysosome function as a first-line molecular test for the diagnosis of LSDs. Ultimately, our goal is to provide a fast and effective tool to screen for virtually all LSDs in a single run, thus contributing to decrease the diagnostic odyssey, accelerating the time to diagnosis. Our study enrolled a group of 23 patients with variable degrees of clinical and/or biochemical suspicion of LSD. Briefly, NGS analysis data workflow, followed by segregation analysis allowed the characterization of approximately 41% of the analyzed patients and the identification of 10 different pathogenic variants, underlying nine LSDs. Importantly, four of those variants were novel, and, when applicable, their effect over protein structure was evaluated through in silico analysis. One of the novel pathogenic variants was identified in the GM2A gene, which is associated with an ultra-rare (or misdiagnosed) LSD, the AB variant of GM2 Gangliosidosis. Overall, this case series highlights not only the major advantages of NGS-based diagnostic approaches but also, to some extent, its limitations ultimately promoting a reflection on the role of targeted panels as a primary tool for the prompt characterization of LSD patients.

More than two thirds of Lysosomal Storage Diseases (LSDs) present central nervous system involvement. Nevertheless, only one of the currently approved therapies has an impact on neuropathology. Therefore, alternative approaches are under development, either addressing the underlying enzymatic defect or its downstream consequences. Also under study is the possibility to block substrate accumulation upstream, by promoting a decrease of its synthesis. This concept is known as substrate reduction therapy and may be triggered by several molecules, such as small interfering RNAs (siRNAs). siRNAs promote RNA interference, a naturally occurring sequence-specific post-transcriptional gene-silencing mechanism, and may target virtually any gene of interest, inhibiting its expression. Still, naked siRNAs have limited cellular uptake, low biological stability, and unfavorable pharmacokinetics. Thus, their translation into clinics requires proper delivery methods. One promising platform is a special class of liposomes called stable nucleic acid lipid particles (SNALPs), which are characterized by high cargo encapsulation efficiency and may be engineered to promote targeted delivery to specific receptors. Here, we review the concept of SNALPs, presenting a series of examples on their efficacy as siRNA nanodelivery systems. By doing so, we hope to unveil the therapeutic potential of these nanosystems for targeted brain delivery of siRNAs in LSDs.

Unverricht-Lundborg disease (ULD) is a common form of progressive myoclonic epilepsy caused by mutations in the cystatin B gene ( CSTB ) that encodes an inhibitor of several lysosomal cathepsins. Presently, only pharmacological treatment and psychosocial support are available for ULD patients. To overcome the pathogenic effect of the ULD splicing mutation c.66G>A (exon 1), we investigated whether an antisense oligonucleotide therapeutic strategy could correct the defect in patient cells. A specific locked nucleic acid (LNA) antisense oligonucleotide was designed to block a cryptic 5'ss in intron 1. Overall, this approach allowed the restoration of the normal splicing pattern. Furthermore, the recovery was both sequence and dose-specific. In general, this work provides a proof of principle on the correction of a CSTB gene defect causing ULD through a mutation-specific antisense therapy. It adds evidence to the feasibility of this approach, joining the many studies that are paving the way for translating antisense technology into the clinical practice. The insights detailed herein make mutation-based therapy a clear candidate for personalized treatment of ULD patients, encouraging similar investigations into other genetic diseases.

The exploitation of the yeast Saccharomyces cerevisiae as a biological model for the investigation of complex molecular processes conserved in multicellular organisms, such as humans, has allowed fundamental biological discoveries. When comparing yeast and human proteins, it is clear that both amino acid sequences and protein functions are often very well conserved. One example of the high degree of conservation between human and yeast proteins is highlighted by the members of the RAS family. Indeed, the study of the signaling pathways regulated by RAS in yeast cells led to the discovery of properties that were often found interchangeable with RAS proto-oncogenes in human pathways, and vice versa. In this work, we performed an updated critical literature review on human and yeast RAS pathways, specifically highlighting the similarities and differences between them. Moreover, we emphasized the contribution of studying yeast RAS pathways for the understanding of human RAS and how this model organism can contribute to unveil the roles of RAS oncoproteins in the regulation of mechanisms important in the tumorigenic process, like autophagy., Competing Interests: The authors declare no conflict of interest.

Secondary and tertiary alcohols synthesized via allylation of aldehydes and ketones are important compounds in bioactive natural products and industry, including pharmaceuticals. Development of a mechanochemical method using potassium allyltrifluoroborate salt and water, to successfully perform the allylation of aromatic and aliphatic carbonyl compounds is reported for the first time. By controlling the grinding parameters, the methodology can be selective, namely, very efficient for aldehydes and ineffective for ketones, but by employing lanthanide catalysts, the reactions with ketones can become practically quantitative. The catalyzed reactions can also be performed under mild aqueous stirring conditions. Considering the allylation agent and its by-products, aqueous media, energy efficiency and use of catalyst, the methodology meets most of the green chemistry principles.

Lysosomal storage diseases (LSDs) are a group of rare, life-threatening genetic disorders, usually caused by a dysfunction in one of the many enzymes responsible for intralysosomal digestion. Even though no cure is available for any LSD, a few treatment strategies do exist. Traditionally, efforts have been mainly targeting the functional loss of the enzyme, by injection of a recombinant formulation, in a process called enzyme replacement therapy (ERT), with no impact on neuropathology. This ineffectiveness, together with its high cost and lifelong dependence is amongst the main reasons why additional therapeutic approaches are being (and have to be) investigated: chaperone therapy; gene enhancement; gene therapy; and, alternatively, substrate reduction therapy (SRT), whose aim is to prevent storage not by correcting the original enzymatic defect but, instead, by decreasing the levels of biosynthesis of the accumulating substrate(s). Here we review the concept of substrate reduction, highlighting the major breakthroughs in the field and discussing the future of SRT, not only as a monotherapy but also, especially, as complementary approach for LSDs.

Conventionally, in its decision-making, the U.S. EPA has evaluated the effects and risks associated with a single pollutant in a single exposure medium. In reality, people are exposed to mixtures of pollutants or to the same pollutant through a variety of media, including the air, water, and food. It is now more recognized than before that environmental exposure to pollutants occurs via multiple exposure routes and pathways, including inhalation, ingestion, and dermal absorption. Moreover, chemical, biologic, radiologic, physical, and psychologic stressors are all acknowledged as affecting human health. Although many EPA offices attempt to consider cumulative risk assessment and cumulative effects in various ways, there is no Agency-wide policy for considering these risks and the effects of exposure to these risks when making environmental decisions. This article examines how U.S. courts might assess EPA's general authority and discretion to use cumulative risk assessment as the basis for developing data in support of environmental decision-making, and how courts might assess the validity of a cumulative risk assessment methodology itself.