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Access to better health care anticipates that more medical devices can be found alongside skeletal remains. Those employed in oral rehabilitation, with available brands or batch/series, can prove useful in the identification process. A previous study in the Colecção de Esqueletos Identificados Século XXI described macroscopically the dental prostheses. An unusual case of a dental device with chromatic alterations demonstrated to require a more detailed analysis. The individual, a 53-year-old male, exhibited, at both arches, a fixed tooth-supported rehabilitation, with gold colouring classified initially as a gold-palladium alloy. Simultaneously, a green pigmentation deposit was observable in bone and prosthesis. This investigation aimed to verify the elemental composition of the dental prosthesis alloy. Elemental analysis was performed by X-ray fluorescence in two regions (labial surface of the prosthetic crown and the root surface of the lower right lateral incisor). Both the spectra and the qualitative results found higher levels of copper and aluminium, followed by nickel, iron, zinc, and manganese. No gold or palladium was detected. The most probable assumption is that a copper-aluminium alloy was used, as its elemental concentration corresponds to those measured in similar devices. Dental prostheses of copper-aluminium alloys have been made popular since the 1980s, particularly in the USA, Japan, and Eastern Europe. Apart from the biographical information, it was also known that the individual's place of birth was an Eastern European country, which highlighted the usefulness of this type of information when dealing with missing people cases.
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Prótese Dentária , Paládio , Masculino , Humanos , Pessoa de Meia-Idade , Raios X , Portugal , Paládio/análise , Cobre/análise , Alumínio/análise , Fluorescência , Incisivo , Ligas de Ouro/análiseRESUMO
PURPOSE: To evaluate the impact of the pandemic on the consumption of antidepressive agents in Central Portugal. METHODS: To estimate the causal effect of the pandemic an interrupted time series analysis was conducted. Data of antidepressant drugs monthly dispensed in community pharmacies between Jan-2010 and Dec-2021 were provided by the regional Health Administration. Anti-Parkinson dopaminergic agents and statins, theoretically not influenced by COVID-19 pandemics, were used as comparator series. The number of packages was converted into defined daily doses and presented as defined daily doses/1000 inhabitants/day. A Bayesian structural time-series model with CausalImpact on R/RStudio was used to predict the counterfactual. Analyses with different geographical granularity (9 sub-regions and 78 municipalities) were performed. RESULTS: When compared to counterfactual, regional consumption non-significantly increased after the pandemic declaration, with a relative effect of + 1.30% [95%CI -1.6%:4.2%]. When increasing the granularity, differences appeared between sub-region with significant increases in Baixo Mondego + 6.5% [1.4%:11.0%], Guarda + 4.4% [1.1%:7.7%] or Cova da Beira + 4.1% [0.17%:8.3%], but non-significant variation in the remaining 6 sub-regions. Differences are more obvious at municipality level, ranging from increases of + 37.00% [32.00%:42.00%] to decreases of -11.00% [-17.00%:-4.20%]. Relative impact positively correlated with percentage of elderly in the municipality (r = 0.301; p = 0.007), and negatively with population density (r=-0.243; p = 0.032). No other predicting variables were found. CONCLUSION: Antidepressant consumption suffered very slight variations at regional level after the COVID-19 pandemic declaration. Analysis with higher granularity allowed identifying municipalities with higher impact (increase or decrease). The absence of clear association patterns suggests other causal hypotheses of the differences.
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Eugenol, 4-allyl-2-methoxyphenol, is the main constituent of clove essential oil and has demonstrated relevant biological activity, namely anticancer activity. Aiming to increase this activity, we synthesized a series of eugenol ß-amino alcohol and ß-alkoxy alcohol derivatives, which were then tested against two human cancer cell lines, namely gastric adenocarcinoma cells (AGS) and lung adenocarcinoma cells (A549). An initial screening was performed to identify the most cytotoxic compounds. The results demonstrated that three ß-amino alcohol derivatives had anticancer activity that justified subsequent studies, having been shown to trigger apoptosis. Importantly, the most potent molecules displayed no appreciable toxicity towards human noncancer cells. Structure-activity relationships show that changes in eugenol structure led to enhanced cytotoxic activity and can contribute to the future design of more potent and selective drugs.
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Antineoplásicos , Eugenol , Álcoois , Amino Álcoois , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose , Óleo de Cravo/química , HumanosRESUMO
Crimean-Congo hemorrhagic fever virus (CCHFV) is a widespread zoonotic pathogen that can cause mild to severe hemorrhagic disease in humans. CCHFV may be transmitted through direct contact with tissue or blood of viremic animals; however, the primary transmission route is through infected tick bites. CCHFV RNA has been detected in ticks feeding on domestic and wild animals in western Spain, suggesting an established circulation of CCHFV in Western Europe. Ruminants have been recognized as important CCHFV reservoirs and have been linked to human cases in endemic regions. Given the emergence of CCHF in neighboring Spain, and a report of two CCHFV seropositive humans in southern Portugal in 1985, we investigated the potential circulation of this virus in the country by performing a nationwide anti-CCHFV IgG serosurvey in sentinel sheep of Portugal. Sera (n = 459) randomly selected from widely distributed farms (n = 20) of Portugal were tested using a commercial double-antigen enzyme-linked immunosorbent assay, yielding an overall seroprevalence of 0.4% (95% confidence interval [CI] 0.04-1.56%). Positive sheep were from the southern region of Portugal (Alentejo region), which raise the seroprevalence of this region to 0.74% (95% CI 0.09-2.66%). This is the first study reporting the presence of CCHFV antibodies in sheep of Portugal, thus suggesting a geographical expansion of CCHFV to this country. It seems likely that CCHFV may exist focally in southern Portugal.
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Vírus da Febre Hemorrágica da Crimeia-Congo , Febre Hemorrágica da Crimeia , Doenças dos Ovinos , Carrapatos , Animais , Anticorpos Antivirais , Vírus da Febre Hemorrágica da Crimeia-Congo/genética , Febre Hemorrágica da Crimeia/epidemiologia , Febre Hemorrágica da Crimeia/veterinária , Humanos , Portugal/epidemiologia , Estudos Soroepidemiológicos , Ovinos , Doenças dos Ovinos/epidemiologiaRESUMO
For identification of the unknown, the analysis of individualizing characteristics is a paramount procedure, which allows the reconciliation of antemortem with postmortem data. Worldwide, populations are commonly affected by tooth loss, leading to procedures of oral rehabilitation, such as dental prostheses. Although the potential of these devices for human identification is well known, the lack of marks or serial numbers as well as scarce systematic register on databases may difficult its use. The objective of this paper is to present and describe the cases with dentures and bridges of an identified osteological collection, the Colecção de Esqueletos Identificados Século XXI. All the individuals were macroscopically observed, and the cases presenting dental prostheses were registered. A total of thirty-one individuals (20 females; 11 males), ages 38 to 91 years, presented 49 dental prosthesis (removable or fixed), of various materials. The removable acrylic dentures (n = 42) were the most frequent. No identifying marks or production labels were found. This research reinforces two focal points: the value of studying and recognizing medical devices and their application for identification, and the relevance of identified skeletal collections for the advance of forensic science in the current multidisciplinary approach.
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Prótese Dentária , Adulto , Idoso , Idoso de 80 Anos ou mais , Coleções como Assunto , Feminino , Antropologia Forense , Odontologia Legal , Humanos , Masculino , Pessoa de Meia-Idade , PortugalRESUMO
After the publication of the first article in 2014, 21st Century Identified Skeletal Collection, housed in the Laboratory of Forensic Anthropology, Department of Life Sciences at the University of Coimbra, Portugal, has been growing. Currently, the collection is composed of 302 complete adult skeletons of both sexes, which means that in 5 years it has doubled. The collection consists mostly of elderly individuals, with only 12.25% of the individuals aged less than 61 years old. All individuals are Portuguese nationals who died between 1982 and 2012. Ninety individuals exhibit prostheses, other medical devices and signs of surgical procedures. Moreover, a sub-collection of experimentally burned skeletons is under development, and currently includes 56 individuals (18.54% of the collection). The 21st Century Identified Skeletal Collection constitutes a fundamental tool for forensic anthropology research, including the development and validation studies of methods that focus on elderly individuals, as can be ascertained by the numerous scientific publications and academic scholarship that have been produced in previous years.
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Restos Mortais , Osso e Ossos , Coleções como Assunto , Antropologia Forense , Feminino , Humanos , Laboratórios , Masculino , Portugal , Pesquisa , UniversidadesRESUMO
Osteoporosis is a metabolic disorder characterized by a loss of bone mass and structure and increasing the risk of fragility fractures, mostly among postmenopausal women. Sheep is a recognized large animal model for osteoporosis research. An experimental group of ewes (3-4 years old) was subjected to ovariectomy (OVX) and weekly glucocorticoid (GC) application for 24 weeks and compared with a sham control group. Blood and bone marrow parameters were analyzed before and 24 weeks after OVX and GC administration. Osteopenia was confirmed through micro-computed tomography and histomorphometric analysis of L4 vertebra in the study end. A statistically significant increase was observed in mean corpuscular volume, mean cell hemoglobin and monocytes and a decrease in red blood count and eosinophils (p<0.05). Alkaline phosphatase (ALP), gamma-glutamyl transpeptidase, magnesium and α1-globulin increased, and creatinine, albumin, sodium and estradiol decreased (p<0.05). A slight decrease of bone formation markers (bone ALP and osteocalcin) and an increase of bone resorption markers (C-terminal telopeptides of collagen type 1 and tartrate-resistant acid phosphatase) were observed, but without statistical significance. This study aims to contribute to better knowledge of sheep as a model for osteoporosis research and the consequences that a performed induction protocol may impose on organic metabolism.
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Hematologia , Osteoporose , Animais , Medula Óssea , Remodelação Óssea , Pré-Escolar , Feminino , Glucocorticoides , Humanos , Ovariectomia , Pesquisa , Ovinos , Microtomografia por Raio-XRESUMO
Aldehyde oxidase (AOX) is a xanthine oxidase (XO)-related enzyme with emerging importance due to its role in the metabolism of drugs and xenobiotics. We report the first crystal structures of human AOX1, substrate free (2.6-Å resolution) and in complex with the substrate phthalazine and the inhibitor thioridazine (2.7-Å resolution). Analysis of the protein active site combined with steady-state kinetic studies highlight the unique features, including binding and substrate orientation at the active site, that characterize human AOX1 as an important drug-metabolizing enzyme. Structural analysis of the complex with the noncompetitive inhibitor thioridazine revealed a new, unexpected and fully occupied inhibitor-binding site that is structurally conserved among mammalian AOXs and XO. The new structural insights into the catalytic and inhibition mechanisms of human AOX that we now report will be of great value for the rational analysis of clinical drug interactions involving inhibition of AOX1 and for the prediction and design of AOX-stable putative drugs.
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Aldeído Oxidase/química , Aldeído Oxidase/metabolismo , Inibidores Enzimáticos/metabolismo , Xenobióticos/metabolismo , Aldeído Oxidase/antagonistas & inibidores , Aldeído Oxidase/genética , Aldeído Oxirredutases/antagonistas & inibidores , Aldeído Oxirredutases/química , Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Animais , Domínio Catalítico , Cristalografia por Raios X , Inibidores Enzimáticos/química , Escherichia coli/genética , Humanos , Camundongos , Modelos Moleculares , Ligação Proteica , Conformação Proteica , Especificidade da Espécie , Especificidade por Substrato , Xenobióticos/químicaRESUMO
Aldehyde oxidase (AOX1) is an enzyme with broad substrate specificity, catalyzing the oxidation of a wide range of endogenous and exogenous aldehydes as well as N-heterocyclic aromatic compounds. In humans, the enzyme's role in phase I drug metabolism has been established and its importance is now emerging. However, the true physiologic function of AOX1 in mammals is still unknown. Further, numerous single-nucleotide polymorphisms (SNPs) have been identified in human AOX1. SNPs are a major source of interindividual variability in the human population, and SNP-based amino acid exchanges in AOX1 reportedly modulate the catalytic function of the enzyme in either a positive or negative fashion. For the reliable analysis of the effect of amino acid exchanges in human proteins, the existence of reproducible expression systems for the production of active protein in ample amounts for kinetic, spectroscopic, and crystallographic studies is required. In our study we report an optimized expression system for hAOX1 in Escherichia coli using a codon-optimized construct. The codon-optimization resulted in an up to 15-fold increase of protein production and a simplified purification procedure. The optimized expression system was used to study three SNPs that result in amino acid changes C44W, G1269R, and S1271L. In addition, the crystal structure of the S1271L SNP was solved. We demonstrate that the recombinant enzyme can be used for future studies to exploit the role of AOX in drug metabolism, and for the identification and synthesis of new drugs targeting AOX when combined with crystallographic and modeling studies.
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Aldeído Oxidase/biossíntese , Aldeído Oxidase/genética , Polimorfismo de Nucleotídeo Único , Engenharia de Proteínas/métodos , Aldeído Oxidase/química , Códon , Cristalografia por Raios X , Escherichia coli/enzimologia , Escherichia coli/genética , Regulação Enzimológica da Expressão Gênica , Genótipo , Humanos , Cinética , Modelos Moleculares , Fenótipo , Conformação Proteica , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Relação Estrutura-Atividade , Especificidade por SubstratoRESUMO
Porcine hokovirus (PHoV), also referred to as porcine parvovirus 4 (P-PARV4), a recently discovered parvovirus of swine that is closely related to human parvovirus 4/5 (H-PARV4/5), was first described in Hong Kong. To evaluate the occurrence of P-PARV4 in Portuguese wild boars in the hunting season of 2011/2012, liver and serum samples were tested. P-PARV4 was detected in 24 % of the wild boars analyzed. Phylogenetic analysis showed a close relationship between the P-PARV4 isolates and other P-PARV4 reference strains. This virus appears to be emerging, with yet unknown implications for public health.
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Infecções por Parvoviridae/veterinária , Parvovirus Suíno/isolamento & purificação , Sus scrofa/virologia , Doenças dos Suínos/virologia , Animais , Feminino , Masculino , Infecções por Parvoviridae/virologia , Parvovirus Suíno/genética , Filogenia , Portugal/epidemiologia , Suínos , Doenças dos Suínos/epidemiologiaRESUMO
OBJECTIVES: Complete and accurate human skeletal inventory is seldom possible in archaeological and forensic cases involving severe fragmentation. In such cases, skeletal mass comparisons with published references may be used as an alternative to assess skeletal completeness but they are too general for a case-by-case routine analysis. The objective is to solve this issue by creating linear regression equations to estimate the total mass of a skeleton based on the mass of individual bones. MATERIALS AND METHODS: Total adult skeletal mass and individual mass of the clavicle, humerus, femur, patella, carpal, metacarpal, tarsal, and metatarsal bones were recorded in a sample of 60 skeletons from the 21st century identified skeletal collection (University of Coimbra). The sample included 32 females and 28 males with ages ranging from 31 to 96 years (mean = 76.4; sd = 14.8). Skeletal mass linear regression equations were calculated based on this sample. RESULTS: The mass of individual bones was successfully used to predict the approximate total mass of the adult skeleton. The femur, humerus, and second metacarpal were the best predictors of total skeletal mass with root mean squared errors ranging from 292.9 to 346.1 g. DISCUSSION: Linear regression was relatively successful at estimating adult skeletal mass. The non-normal distribution of the sample in terms of mass may have reduced the predictive power of the equations. These results have clear impact for bioanthropology, especially forensic anthropology, since this method may provide better estimates of the completeness of the skeleton or the minimum number of individuals. Am J Phys Anthropol 160:427-432, 2016. © 2016 Wiley Periodicals, Inc.
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Osso e Ossos/anatomia & histologia , Osso e Ossos/fisiologia , Antropologia Forense/métodos , Modelos Lineares , Esqueleto/anatomia & histologia , Esqueleto/fisiologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antropologia Física , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Tamanho do Órgão/fisiologiaRESUMO
Mammalian aldehyde oxidases (AOXs; EC1.2.3.1) are a group of conserved proteins belonging to the family of molybdo-flavoenzymes along with the structurally related xanthine dehydrogenase enzyme. AOXs are characterized by broad substrate specificity, oxidizing not only aromatic and aliphatic aldehydes into the corresponding carboxylic acids, but also hydroxylating a series of heteroaromatic rings. The number of AOX isoenzymes expressed in different vertebrate species is variable. The two extremes are represented by humans, which express a single enzyme (AOX1) in many organs and mice or rats which are characterized by tissue-specific expression of four isoforms (AOX1, AOX2, AOX3, and AOX4). In vertebrates each AOX isoenzyme is the product of a distinct gene consisting of 35 highly conserved exons. The extant species-specific complement of AOX isoenzymes is the result of a complex evolutionary process consisting of a first phase characterized by a series of asynchronous gene duplications and a second phase where the pseudogenization and gene deletion events prevail. In the last few years remarkable advances in the elucidation of the structural characteristics and the catalytic mechanisms of mammalian AOXs have been made thanks to the successful crystallization of human AOX1 and mouse AOX3. Much less is known about the physiological function and physiological substrates of human AOX1 and other mammalian AOX isoenzymes, although the importance of these proteins in xenobiotic metabolism is fairly well established and their relevance in drug development is increasing. This review article provides an overview and a discussion of the current knowledge on mammalian AOX.
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Aldeído Oxidase/química , Aldeído Oxidase/metabolismo , Evolução Molecular , Aldeído Oxidase/genética , Aldeído Oxirredutases/química , Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Animais , Catálise , Cristalografia por Raios X , Escherichia coli/genética , Humanos , Inativação Metabólica , Mamíferos , Polimorfismo de Nucleotídeo Único , Xenobióticos/metabolismo , Xenobióticos/farmacocinéticaRESUMO
We report an outbreak of dermatophytoses in rabbits, which was the origin of a dermatophytose epidemic in an agricultural school in central Portugal, affecting 15 people. Both the phenotypic characteristics and internal transcribed spacer (ITS) sequence of the dermatophytes isolated from the rabbits and patients were identical, suggesting that a single strain was responsible for both the epizootic and epidemic dermatophytoses and confirming that these two outbreaks were linked. The ITS sequences were also 100% identical to the ITS sequence of five strains isolated from rabbits in Greece and Italy, but different from that of Trichophyton mentagrophytes commonly isolated from dogs and cats. These results suggest that a particular T. mentagrophytes genotype could be prevalent in rabbits in southern Europe.
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Coelhos/microbiologia , Tinha/microbiologia , Tinha/transmissão , Trichophyton/genética , Trichophyton/isolamento & purificação , Animais , DNA Fúngico , DNA Espaçador Ribossômico , Surtos de Doenças , Epidemias , Europa (Continente) , Genótipo , Grécia/epidemiologia , Humanos , Itália/epidemiologia , Portugal , Tinha/epidemiologia , Trichophyton/crescimento & desenvolvimento , Trichophyton/ultraestruturaRESUMO
In this work, a combination of homology modeling and molecular dynamics (MD) simulations was used to investigate the factors that modulate substrate specificity and activity of the mouse AOX isoforms: mAOX1, mAOX2 (previously mAOX3l1), mAOX3 and mAOX4. The results indicate that the AOX isoform structures are highly preserved and even more conserved than the corresponding amino acid sequences. The only differences are at the protein surface and substrate-binding site region. The substrate-binding site of all isoforms consists of two regions: the active site, which is highly conserved among all isoforms, and a isoform-specific region located above. We predict that mAOX1 accepts a broader range of substrates of different shape, size and nature relative to the other isoforms. In contrast, mAOX4 appears to accept a more restricted range of substrates. Its narrow and hydrophobic binding site indicates that it only accepts small hydrophobic substrates. Although mAOX2 and mAOX3 are very similar to each other, we propose the following pairs of overlapping substrate specificities: mAOX2/mAOX4 and mAOX3/mAXO1. Based on these considerations, we propose that the catalytic activity between all isoforms should be similar but the differences observed in the binding site might influence the substrate specificity of each enzyme. These results also suggest that the presence of several AOX isoforms in mouse allows them to oxidize more efficiently a wider range of substrates. This contrasts with the same or other organisms that only express one isoform and are less efficient or incapable of oxidizing the same type of substrates.
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Aldeído Oxidase/química , Aldeído Oxirredutases/química , Flavoproteínas/química , Conformação Proteica , Aldeído Oxidase/metabolismo , Aldeído Oxirredutases/metabolismo , Sequência de Aminoácidos , Animais , Sítios de Ligação , Domínio Catalítico , Cinética , Camundongos , Simulação de Dinâmica Molecular , Oxirredução , Especificidade por SubstratoRESUMO
In forensic anthropology, ancestry estimation is essential in establishing the individual biological profile. The aim of this study is to present a new program--AncesTrees--developed for assessing ancestry based on metric analysis. AncesTrees relies on a machine learning ensemble algorithm, random forest, to classify the human skull. In the ensemble learning paradigm, several models are generated and co-jointly used to arrive at the final decision. The random forest algorithm creates ensembles of decision trees classifiers, a non-linear and non-parametric classification technique. The database used in AncesTrees is composed by 23 craniometric variables from 1,734 individuals, representative of six major ancestral groups and selected from the Howells' craniometric series. The program was tested in 128 adult crania from the following collections: the African slaves' skeletal collection of Valle da Gafaria; the Medical School Skull Collection and the Identified Skeletal Collection of 21st Century, both curated at the University of Coimbra. The first step of the test analysis was to perform ancestry estimation including all the ancestral groups of the database. The second stage of our test analysis was to conduct ancestry estimation including only the European and the African ancestral groups. In the first test analysis, 75% of the individuals of African ancestry and 79.2% of the individuals of European ancestry were correctly identified. The model involving only African and European ancestral groups had a better performance: 93.8% of all individuals were correctly classified. The obtained results show that AncesTrees can be a valuable tool in forensic anthropology.
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Cefalometria , Bases de Dados como Assunto , Árvores de Decisões , Antropologia Forense/métodos , Grupos Raciais , Adulto , Algoritmos , Etnicidade , Feminino , Humanos , Aprendizado de Máquina , Masculino , Determinação do Sexo pelo EsqueletoRESUMO
The in vitro evaluation of 3D scaffolds for bone tissue engineering in mono-cultures is a common practice; however, it does not represent the native complex nature of bone tissue. Co-cultures of osteoblasts and osteoclasts, without the addition of stimulating agents for monitoring cellular cross-talk, remains a challenge. In this study, a growth factor-free co-culture of human bone marrow-derived mesenchymal stem cells (hBM-MSCs) and human peripheral blood mononuclear cells (hPBMCs) has been established and used for the evaluation of 3D-printed scaffolds for bone tissue engineering. The scaffolds were produced from PLLA/PCL/PHBV polymeric blends, with two composite materials produced through the addition of 2.5% w/v nanohydroxyapatite (nHA) or strontium-substituted nanohydroxyapatite (Sr-nHA). Cell morphology data showed that hPBMCs remained undifferentiated in co-culture, while no obvious differences were observed in the mono- and co-cultures of hBM-MSCs. A significantly increased alkaline phosphatase (ALP) activity and osteogenic gene expression was observed in co-culture on Sr-nHA-containing scaffolds. Tartrate-resistant acid phosphatase (TRAP) activity and osteoclastogenic gene expression displayed significantly suppressed levels in co-culture on Sr-nHA-containing scaffolds. Interestingly, mono-cultures of hPBMCs on Sr-nHA-containing scaffolds indicated a delay in osteoclasts formation, as evidenced from TRAP activity and gene expression, demonstrating that strontium acts as an osteoclastogenesis inhibitor. This co-culture study presents an effective 3D model to evaluate the regenerative capacity of scaffolds for bone tissue engineering, thus minimizing time-consuming and costly in vivo experiments.
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The production of small ruminant autochthonous breeds in the Centre region of Portugal is practiced in a semi-extensive husbandry system, exposing animals to parasitic infections. The main objective of this study was to estimate the prevalence of lungworm infection and identify risk factors. Fecal samples of 203 goats and 208 sheep from 30 herds were collected per rectum and subjected to the modified Baermann test. The overall prevalence of infection was 57.7%, significantly higher in goats (95.6%) than in sheep (20.7%) (p < 0.001). According to the binary logistic regression model, sheep dewormed with albendazole, mebendazole plus closantel, or ivermectin plus clorsulon presented a risk of Protostrongylidae infection 29.702, 7.426, or 8.720 times higher, respectively, than those dewormed with eprinomectin. Additionally, the presence of gastrointestinal parasites was investigated in 307 fecal samples using Mini-FLOTAC®. The overall prevalence of infection was 86.3%, also significantly higher in goats (93.2%) than in sheep (79.9%) (p < 0.001). Strongyle-type eggs were the most frequently identified, both in sheep (69.8%) and goats (87.8%), followed by Eimeria oocysts (40.3% in sheep and 68.9% in goats). Considering the high prevalence and the burden of lungworm parasitic infection, it is urgent to determine its economic impact and the repercussions in animal health in the Centre region of Portugal to establish appropriate therapeutic guidelines.
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BACKGROUND: Aldehyde oxidases have pharmacological relevance, and AOX3 is the major drug-metabolizing enzyme in rodents. RESULTS: The crystal structure of mouse AOX3 with kinetics and molecular docking studies provides insights into its enzymatic characteristics. CONCLUSION: Differences in substrate and inhibitor specificities can be rationalized by comparing the AOX3 and xanthine oxidase structures. SIGNIFICANCE: The first aldehyde oxidase structure represents a major advance for drug design and mechanistic studies. Aldehyde oxidases (AOXs) are homodimeric proteins belonging to the xanthine oxidase family of molybdenum-containing enzymes. Each 150-kDa monomer contains a FAD redox cofactor, two spectroscopically distinct [2Fe-2S] clusters, and a molybdenum cofactor located within the protein active site. AOXs are characterized by broad range substrate specificity, oxidizing different aldehydes and aromatic N-heterocycles. Despite increasing recognition of its role in the metabolism of drugs and xenobiotics, the physiological function of the protein is still largely unknown. We have crystallized and solved the crystal structure of mouse liver aldehyde oxidase 3 to 2.9 Å. This is the first mammalian AOX whose structure has been solved. The structure provides important insights into the protein active center and further evidence on the catalytic differences characterizing AOX and xanthine oxidoreductase. The mouse liver aldehyde oxidase 3 three-dimensional structure combined with kinetic, mutagenesis data, molecular docking, and molecular dynamics studies make a decisive contribution to understand the molecular basis of its rather broad substrate specificity.
Assuntos
Aldeído Oxirredutases/química , Aldeído Oxidase/química , Aldeído Oxidase/genética , Aldeído Oxidase/metabolismo , Aldeído Oxirredutases/genética , Aldeído Oxirredutases/metabolismo , Sequência de Aminoácidos , Animais , Domínio Catalítico , Bovinos , Cristalografia por Raios X , Dimerização , Humanos , Cinética , Mamíferos/genética , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Estrutura Terciária de Proteína , Alinhamento de Sequência , Especificidade por SubstratoRESUMO
Nanohydroxyapatite (nanoHA) is the major mineral component of bone. It is highly biocompatible, osteoconductive, and forms strong bonds with native bone, making it an excellent material for bone regeneration. However, enhanced mechanical properties and biological activity for nanoHA can be achieved through enrichment with strontium ions. Here, nanoHA and nanoHA with a substitution degree of 50 and 100% of calcium with strontium ions (Sr-nanoHA_50 and Sr-nanoHA_100, respectively) were produced via wet chemical precipitation using calcium, strontium, and phosphorous salts as starting materials. The materials were evaluated for their cytotoxicity and osteogenic potential in direct contact with MC3T3-E1 pre-osteoblastic cells. All three nanoHA-based materials were cytocompatible, featured needle-shaped nanocrystals, and had enhanced osteogenic activity in vitro. The Sr-nanoHA_100 indicated a significant increase in the alkaline phosphatase activity at day 14 compared to the control. All three compositions revealed significantly higher calcium and collagen production up to 21 days in culture compared to the control. Gene expression analysis exhibited, for all three nanoHA compositions, a significant upregulation of osteonectin and osteocalcin on day 14 and of osteopontin on day 7 compared to the control. The highest osteocalcin levels were found for both Sr-substituted compounds on day 14. These results demonstrate the great osteoinductive potential of the produced compounds, which can be exploited to treat bone disease.
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The combination of biomaterials and bioactive particles has shown to be a successful strategy to fabricate electrospun scaffolds for bone tissue engineering. Among the range of bioactive particles, hydroxyapatite and mesoporous bioactive glasses (MBGs) have been widely used for their osteoconductive and osteoinductive properties. Yet, the comparison between the chemical and mechanical characteristics as well as the biological performances of these particle-containing scaffolds have been characterized to a limited extent. In this work, we fabricated PEOT/PBT-based composite scaffolds incorporating either nanohydroxyapatite (nHA), strontium-containing nanohydroxyapatite (nHA_Sr) or MBGs doped with strontium ions up to 15 wt./vol% and 12,5 wt./vol% for nHA and MBG, respectively. The composite scaffolds presented a homogeneous particle distribution. Morphological, chemical and mechanical analysis revealed that the introduction of particles into the electrospun meshes caused a decrease in the fiber diameter and mechanical properties, yet maintaining the hydrophilic nature of the scaffolds. The Sr2+ release profile differed according to the considered system, observing a 35-day slowly decreasing release from strontium-containing nHA scaffolds, whereas MBG-based scaffolds showed a strong burst release in the first week. In vitro, culture of human bone marrow-derived mesenchymal stromal cells (hMSCs) on composite scaffolds demonstrated excellent cell adhesion and proliferation. In maintenance and osteogenic media, all composite scaffolds showed high mineralization as well as expression of Col I and OCN compared to PEOT/PBT scaffolds, suggesting their ability to boost bone formation even without osteogenic factors. The presence of strontium led to an increase in collagen secretion and matrix mineralization in osteogenic medium, while gene expression analysis showed that hMSCs cultured on nHA-based scaffolds had a higher expression of OCN, ALP and RUNX2 compared to cells cultured on nHA_Sr scaffolds in osteogenic medium. Yet, cells cultured on MBGs-based scaffolds showed a higher gene expression of COL1, ALP, RUNX2 and BMP2 in osteogenic medium compared to nHA-based scaffolds, which is hypothesized to lead to high osteoinductivity in long term cultures.