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1.
Methods Mol Biol ; 2351: 165-179, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34382189

RESUMO

Targeted chromatin capture (T2C) is a 3C-based method and is used to study the 3D chromatin organization, interactomes and structural changes associated with gene regulation, progression through the cell cycle, and cell survival and development. Low input targeted chromatin capture (low-T2C) is an optimized version of the T2C protocol for low numbers of cells. Here, we describe the protocol for low-T2C, including all experimental steps and bioinformatics tools in detail.


Assuntos
Montagem e Desmontagem da Cromatina , Cromatina/genética , Biologia Computacional/métodos , Cromatina/química , Cromatina/metabolismo , Mapeamento Cromossômico , Regulação da Expressão Gênica , Biblioteca Gênica , Genômica/métodos , Reprodutibilidade dos Testes
2.
Artigo em Inglês | MEDLINE | ID: mdl-34360162

RESUMO

Clear role descriptions promote the quality of interprofessional collaboration. Currently, it is unclear to what extent healthcare professionals consider pharmaceutical care (PC) activities to be nurses' responsibility in order to obtain best care quality. This study aimed to create and evaluate a framework describing potential nursing tasks in PC and to investigate nurses' level of responsibility. A framework of PC tasks and contextual factors was developed based on literature review and previous DeMoPhaC project results. Tasks and context were cross-sectionally evaluated using an online survey in 14 European countries. A total of 923 nurses, 240 physicians and 199 pharmacists responded. The majority would consider nurses responsible for tasks within: medication self-management (86-97%), patient education (85-96%), medication safety (83-95%), monitoring adherence (82-97%), care coordination (82-95%), and drug monitoring (78-96%). The most prevalent level of responsibility was 'with shared responsibility'. Prescription management tasks were considered to be nurses' responsibility by 48-81% of the professionals. All contextual factors were indicated as being relevant for nurses' role in PC by at least 74% of the participants. No task nor contextual factor was removed from the framework after evaluation. This framework can be used to enable healthcare professionals to openly discuss allocation of specific (shared) responsibilities and tasks.


Assuntos
Enfermeiras e Enfermeiros , Assistência Farmacêutica , Estudos Transversais , Europa (Continente) , Humanos , Papel do Profissional de Enfermagem , Farmacêuticos
3.
Nurse Educ Today ; 104: 104926, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34274774

RESUMO

BACKGROUND: Nurses play an important role in pharmaceutical care. They are involved in: detecting clinical change; communicating/discussing pharmacotherapy with patients, their advocates, and other healthcare professionals; proposing and implementing medication-related interventions; and ensuring follow-up of patients and medication regimens. To date, a framework of nurses' competences on knowledge, skills, and attitudes as to interprofessional pharmaceutical care tasks is missing. OBJECTIVES: To reach agreement with experts about nurses' competences for tasks in interprofessional pharmaceutical care. METHODS: A two-phase study starting with a scoping review followed by five Delphi rounds was performed. Competences extracted from the literature were assessed by an expert panel on relevance by using the RAND/UCLA method. The experts (n = 22) involved were healthcare professionals, nurse researchers, and educators from 14 European countries with a specific interest in nurses' roles in interprofessional pharmaceutical care. Descriptive statistics supported the data analysis. RESULTS: The expert panel reached consensus on the relevance of 60 competences for 22 nursing tasks. Forty-one competences were related to 15 generic nursing tasks and 33 competences were related to seven specific nursing tasks. CONCLUSIONS: This study resulted in a competence framework for competency-based nurse education. Future research should focus on imbedding these competences in nurse education. A structured instrument should be developed to assess students' readiness to achieve competence in interprofessional pharmaceutical care in clinical practice.


Assuntos
Enfermeiras e Enfermeiros , Assistência Farmacêutica , Competência Clínica , Técnica Delfos , Europa (Continente) , Humanos , Papel do Profissional de Enfermagem
4.
PLoS One ; 16(5): e0251982, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34043650

RESUMO

OBJECTIVES: To understand healthcare professionals' experiences and perceptions of nurses' potential or ideal roles in pharmaceutical care (PC). DESIGN: Qualitative study conducted through semi-structured in-depth interviews. SETTING: Between December 2018 and October 2019, interviews were conducted with healthcare professionals of 14 European countries in four healthcare settings: hospitals, community care, mental health and long-term residential care. PARTICIPANTS: In each country, pharmacists, physicians and nurses in each of the four settings were interviewed. Participants were selected on the basis that they were key informants with broad knowledge and experience of PC. DATA COLLECTION AND ANALYSIS: All interviews were conducted face to face. Each country conducted an initial thematic analysis. Consensus was reached through a face-to-face discussion of all 14 national leads. RESULTS: 340 interviews were completed. Several tasks were described within four potential nursing responsibilities, that came up as the analysis themes, being: 1) monitoring therapeutic/adverse effects of medicines, 2) monitoring medicines adherence, 3) decision making on medicines, including prescribing 4) providing patient education/information. Nurses' autonomy varied across Europe, from none to limited to a few tasks and emergencies to a broad range of tasks and responsibilities. Intended level of autonomy depended on medicine types and level of education. Some changes are needed before nursing roles can be optimised and implemented in practice. Lack of time, shortage of nurses, absence of legal frameworks and limited education and knowledge are main threats to European nurses actualising their ideal role in PC. CONCLUSIONS: European nurses have an active role in PC. Respondents reported positive impacts on care quality and patient outcomes when nurses assumed PC responsibilities. Healthcare professionals expect nurses to report observations and assessments. This key patient information should be shared and addressed by the interprofessional team. The study evidences the need of a unique and consensus-based PC framework across Europe.

5.
Genome Res ; 30(8): 1119-1130, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32747411

RESUMO

Polycomb group proteins are important for maintaining gene expression patterns and cell identity in metazoans. The mammalian Polycomb repressive deubiquitinase (PR-DUB) complexes catalyze removal of monoubiquitination on lysine 119 of histone H2A (H2AK119ub1) through a multiprotein core comprised of BAP1, HCFC1, FOXK1/2, and OGT in combination with either of ASXL1, 2, or 3. Mutations in PR-DUB components are frequent in cancer. However, mechanistic understanding of PR-DUB function in gene regulation is limited. Here, we show that BAP1 is dependent on the ASXL proteins and FOXK1/2 in facilitating gene activation across the genome. Although PR-DUB was previously shown to cooperate with PRC2, we observed minimal overlap and functional interaction between BAP1 and PRC2 in embryonic stem cells. Collectively, these results demonstrate that PR-DUB, by counteracting accumulation of H2AK119ub1, maintains chromatin in an optimal configuration ensuring expression of genes important for general functions such as cell metabolism and homeostasis.

6.
Hum Mol Genet ; 29(15): 2535-2550, 2020 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-32628253

RESUMO

The transcription factor zinc finger E-box binding protein 2 (ZEB2) controls embryonic and adult cell fate decisions and cellular maturation in many stem/progenitor cell types. Defects in these processes in specific cell types underlie several aspects of Mowat-Wilson syndrome (MOWS), which is caused by ZEB2 haplo-insufficiency. Human ZEB2, like mouse Zeb2, is located on chromosome 2 downstream of a ±3.5 Mb-long gene-desert, lacking any protein-coding gene. Using temporal targeted chromatin capture (T2C), we show major chromatin structural changes based on mapping in-cis proximities between the ZEB2 promoter and this gene desert during neural differentiation of human-induced pluripotent stem cells, including at early neuroprogenitor cell (NPC)/rosette state, where ZEB2 mRNA levels increase significantly. Combining T2C with histone-3 acetylation mapping, we identified three novel candidate enhancers about 500 kb upstream of the ZEB2 transcription start site. Functional luciferase-based assays in heterologous cells and NPCs reveal co-operation between these three enhancers. This study is the first to document in-cis Regulatory Elements located in ZEB2's gene desert. The results further show the usability of T2C for future studies of ZEB2 REs in differentiation and maturation of multiple cell types and the molecular characterization of newly identified MOWS patients that lack mutations in ZEB2 protein-coding exons.

7.
BMJ Open ; 10(6): e036269, 2020 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-32499269

RESUMO

OBJECTIVES: Safe pharmaceutical care (PC) requires an interprofessional team approach, involving physicians, nurses and pharmacists. Nurses' roles however, are not always explicit and clear, complicating interprofessional collaboration. The aim of this study is to describe nurses' practice and interprofessional collaboration in PC, from the viewpoint of nurses, physicians and pharmacists. DESIGN: A cross-sectional survey. SETTING: The study was conducted in 17 European countries, each with their own health systems. PARTICIPANTS: Pharmacists, physicians and nurses with an active role in PC were surveyed. MAIN OUTCOME MEASURES: Nurses' involvement in PC, experiences of interprofessional collaboration and communication and views on nurses' competences. RESULTS: A total of 4888 nurses, 974 physicians and 857 pharmacists from 17 European countries responded. Providing patient education and information (PEI), monitoring medicines adherence (MMA), monitoring adverse/therapeutic effects (ME) and prescribing medicines were considered integral to nursing practice by 78%, 73%, 69% and 15% of nurses, respectively. Most respondents were convinced that quality of PC would be improved by increasing nurses' involvement in ME (95%), MMA (95%), PEI (91%) and prescribing (53%). Mean scores for the reported quality of collaboration between nurses and physicians, collaboration between nurses and pharmacists and interprofessional communication were respectively <7/10, ≤4/10, <6/10 for all four aspects of PC. CONCLUSIONS: ME, MMA, PEI and prescribing are part of nurses' activities, and most healthcare professionals felt their involvement should be extended. Collaboration between nurses and physicians on PC is limited and between nurses and pharmacists even more.


Assuntos
Estudos Transversais , Relações Interprofissionais , Papel do Profissional de Enfermagem , Equipe de Assistência ao Paciente/estatística & dados numéricos , Assistência Farmacêutica/estatística & dados numéricos , Comparação Transcultural , Europa (Continente) , Humanos , Comunicação Interdisciplinar , Inquéritos e Questionários
8.
Haematologica ; 105(7): 1802-1812, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-31582556

RESUMO

GATA1 is an essential transcriptional regulator of myeloid hematopoietic differentiation towards red blood cells. During erythroid differentiation, GATA1 forms different complexes with other transcription factors such as LDB1, TAL1, E2A and LMO2 ("the LDB1 complex") or with FOG1. The functions of GATA1 complexes have been studied extensively in definitive erythroid differentiation; however, the temporal and spatial formation of these complexes during erythroid development is unknown. We applied proximity ligation assay (PLA) to detect, localize and quantify individual interactions during embryonic stem cell differentiation and in mouse fetal liver (FL) tissue. We show that GATA1/LDB1 interactions appear before the proerythroblast stage and increase in a subset of the CD71+/TER119- cells to activate the terminal erythroid differentiation program in 12.5 day FL. Using Ldb1 and Gata1 knockdown FL cells, we studied the functional contribution of the GATA1/LDB1 complex during differentiation. This shows that the active LDB1 complex appears quite late at the proerythroblast stage of differentiation and confirms the power of PLA in studying the dynamic interaction of proteins in cell differentiation at the single cell level. We provide dynamic insight into the temporal and spatial formation of the GATA1 and LDB1 transcription factor complexes during hematopoietic development and differentiation.


Assuntos
Células-Tronco Embrionárias/citologia , Fator de Transcrição GATA1 , Proteínas com Domínio LIM , Animais , Diferenciação Celular , Proteínas de Ligação a DNA , Fator de Transcrição GATA1/genética , Fígado , Camundongos , Fatores de Transcrição
9.
Int J Health Care Qual Assur ; 32(6): 1004-1012, 2019 Jul 08.
Artigo em Inglês | MEDLINE | ID: mdl-31282261

RESUMO

PURPOSE: The purpose of this paper is to investigate the effect of nurse staffing, nurse education and work experience on patients' length of stay (LOS) in the Greek public hospitals. DESIGN/METHODOLOGY/APPROACH: A cross-sectional study, with retrospective administrative data, was implemented. From all seven Regional Health Authorities of Greece, 25 general surgical units in 17 public hospitals participated in the study. FINDINGS: All over the hospitals were studied, 32,287 patients ⩾17 years old and 203 nursing staff, who were working in the study units, were included in the analysis. According to the multivariate linear regression model, increased years of experience as a nurse (b= -0.04, 95% CI= -0.06 to -0.02, p=0.001) and increased percentage of registered nurse to the total nursing staff (b= -1.18, CI= -1.88 to -0.47, p=0.03) were associated with decreased patient LOS. ORIGINALITY/VALUE: This was the first extended study in Greece, which explored the relationship between nurse staffing, nurse education, work experience and the LOS. The role that nurse staffing play together with its characteristics in the provision toward the quality healthcare services has already been recognized worldwide. The findings revealed the great shortage of nursing staff and the significant correlation between the work experience and educational level to patients' LOS.


Assuntos
Tempo de Internação/estatística & dados numéricos , Recursos Humanos de Enfermagem no Hospital/organização & administração , Admissão e Escalonamento de Pessoal/organização & administração , Qualidade da Assistência à Saúde , Adulto , Idoso , Competência Clínica , Estudos Transversais , Feminino , Grécia , Hospitais Públicos , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
10.
Cell Stem Cell ; 24(5): 736-752.e12, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-30982769

RESUMO

The pathological consequences of structural variants disrupting 3D genome organization can be difficult to elucidate in vivo due to differences in gene dosage sensitivity between mice and humans. This is illustrated by branchiooculofacial syndrome (BOFS), a rare congenital disorder caused by heterozygous mutations within TFAP2A, a neural crest regulator for which humans, but not mice, are haploinsufficient. Here, we present a BOFS patient carrying a heterozygous inversion with one breakpoint located within a topologically associating domain (TAD) containing enhancers essential for TFAP2A expression in human neural crest cells (hNCCs). Using patient-specific hiPSCs, we show that, although the inversion shuffles the TFAP2A hNCC enhancers with novel genes within the same TAD, this does not result in enhancer adoption. Instead, the inversion disconnects one TFAP2A allele from its cognate enhancers, leading to monoallelic and haploinsufficient TFAP2A expression in patient hNCCs. Our work illustrates the power of hiPSC differentiation to unveil long-range pathomechanisms.


Assuntos
Síndrome Brânquio-Otorrenal/genética , Variação Estrutural do Genoma/genética , Mutação/genética , Crista Neural/fisiologia , Fator de Transcrição AP-2/metabolismo , Adolescente , Alelos , Animais , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Elementos Facilitadores Genéticos/genética , Haploinsuficiência , Humanos , Masculino , Camundongos , Análise de Célula Única , Fator de Transcrição AP-2/genética
11.
J Pediatr Nurs ; 40: e2-e8, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29402659

RESUMO

PURPOSE: To investigate the effect of lifestyle habits in childhood Metabolic Syndrome (MTS). DESIGN AND METHODS: Descriptive correlation study with 480 participants (5-12 years old) using a specially designed questionnaire was conducted. Anthropometric and biochemical analyses were performed. RESULTS: Fifteen percent of children exhibited predisposition for MTS. Regarding sleep habits, logistic regression analysis (LRA) showed that hour of sleep -before 22:00- was associated with decreased waist circumference (WC%) (p = .026). Midday siesta was negatively correlated with systolic (SBP) (p = .001) and diastolic blood pressure (DBP) (p = .046). In children without MTS, lack of sleep and night time sleep was positively correlated with DBP (p = .044) and fasting blood glucose (FBG) (p = .005). Regarding nutrition habits, fast food consumption was positively correlated with SBP (p = .006) and meat consumption was positively correlated with both Body Mass Index% (BMI%) (p = .038) and WC% (p = .023). LRA showed that fruit (p = .001) and legume (p = .040) consumption was associated with decreased FBG; fish consumption with decreased Low Density Lipoprotein (LDL) cholesterol (p = .031), vegetable (p = .054) and cereal consumption (p = .012) with decreased DBP. In children with MTS, fruits were associated with increased FBG (p = .034). In children without MTS, meat consumption was associated with increased LDL (p = .024), cereal with increased WC% (p = .002) and olive products with increased High Density Lipoprotein (HDL) cholesterol and BMI% (p = .037). CONCLUSIONS: The adoption of both balanced diet and sleep habits seemed to be crucial for the prevention of MTS. PRACTICE IMPLICATIONS: Clinical health nurses could develop and implement preventive intervention programs in order to avoid metabolic complications in adulthood.


Assuntos
Proteção da Criança/estatística & dados numéricos , Promoção da Saúde/organização & administração , Síndrome Metabólica/prevenção & controle , Estado Nutricional , Obesidade Pediátrica/prevenção & controle , Atitude Frente a Saúde , Índice de Massa Corporal , Criança , Pré-Escolar , Feminino , Preferências Alimentares/psicologia , Grécia , Humanos , Modelos Logísticos , Masculino , Síndrome Metabólica/enfermagem , Obesidade Pediátrica/enfermagem , Fatores de Risco
12.
Nat Protoc ; 13(3): 459-477, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29419817

RESUMO

Chromosome conformation capture (3C) and its derivatives (e.g., 4C, 5C and Hi-C) are used to analyze the 3D organization of genomes. We recently developed targeted chromatin capture (T2C), an inexpensive method for studying the 3D organization of genomes, interactomes and structural changes associated with gene regulation, the cell cycle, and cell survival and development. Here, we present the protocol for T2C based on capture, describing all experimental steps and bio-informatic tools in full detail. T2C offers high resolution, a large dynamic interaction frequency range and a high signal-to-noise ratio. Its resolution is determined by the resulting fragment size of the chosen restriction enzyme, which can lead to sub-kilobase-pair resolution. T2C's high coverage allows the identification of the interactome of each individual DNA fragment, which makes binning of reads (often used in other methods) basically unnecessary. Notably, T2C requires low sequencing efforts. T2C also allows multiplexing of samples for the direct comparison of multiple samples. It can be used to study topologically associating domains (TADs), determining their position, shape, boundaries, and intra- and inter-domain interactions, as well as the composition of aggregated loops, interactions between nucleosomes, individual transcription factor binding sites, and promoters and enhancers. T2C can be performed by any investigator with basic skills in molecular biology techniques in ∼7-8 d. Data analysis requires basic expertise in bioinformatics and in Linux and Python environments.


Assuntos
Biologia Computacional/métodos , Mapeamento Físico do Cromossomo/métodos , Análise de Sequência de DNA/métodos , Animais , Cromatina/ultraestrutura , Montagem e Desmontagem da Cromatina/fisiologia , Mapeamento Cromossômico/métodos , DNA , Regulação da Expressão Gênica , Genoma/genética , Genoma Humano/genética , Genoma Humano/fisiologia , Genômica , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Camundongos , Nucleossomos , Software
13.
PLoS Genet ; 13(12): e1007137, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-29261648

RESUMO

Cohesin is crucial for genome stability, cell division, transcription and chromatin organization. Its functions critically depend on NIPBL, the cohesin-loader protein that is found to be mutated in >60% of the cases of Cornelia de Lange syndrome (CdLS). Other mutations are described in the cohesin subunits SMC1A, RAD21, SMC3 and the HDAC8 protein. In 25-30% of CdLS cases no mutation in the known CdLS genes is detected. Until now, functional elements in the noncoding genome were not characterized in the molecular etiology of CdLS and therefore are excluded from mutation screening, although the impact of such mutations has now been recognized for a wide range of diseases. We have identified different elements of the noncoding genome involved in regulation of the NIPBL gene. NIPBL-AS1 is a long non-coding RNA transcribed upstream and antisense to NIPBL. By knockdown and transcription blocking experiments, we could show that not the NIPBL-AS1 gene product, but its actual transcription is important to regulate NIPBL expression levels. This reveals a possibility to boost the transcriptional activity of the NIPBL gene by interfering with the NIPBL-AS1 lncRNA. Further, we have identified a novel distal enhancer regulating both NIPBL and NIPBL-AS1. Deletion of the enhancer using CRISPR genome editing in HEK293T cells reduces expression of NIPBL, NIPBL-AS1 as well as genes found to be dysregulated in CdLS.


Assuntos
Elementos Facilitadores Genéticos , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Proteínas/genética , Proteínas/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Segregação de Cromossomos , Síndrome de Cornélia de Lange/genética , Regulação da Expressão Gênica , Genoma Humano , Células HEK293 , Humanos , Mutação , Fenótipo , Regiões Promotoras Genéticas , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Análise de Sequência de DNA
14.
Hum Genomics ; 11(1): 24, 2017 10 23.
Artigo em Inglês | MEDLINE | ID: mdl-29061162

RESUMO

BACKGROUND: Human erythropoiesis is characterized by distinct gene expression profiles at various developmental stages. Previous studies suggest that fetal-to-adult hemoglobin switch is regulated by a complex mechanism, in which many key players still remain unknown. Here, we report our findings from whole transcriptome analysis of erythroid cells, isolated from erythroid tissues at various developmental stages in an effort to identify distinct molecular signatures of each erythroid tissue. RESULTS: From our in-depth data analysis, pathway analysis, and text mining, we opted to focus on the VEGFA gene, given its gene expression characteristics. Selected VEGFA genomic variants, identified through linkage disequilibrium analysis, were explored further for their association with elevated fetal hemoglobin levels in ß-type hemoglobinopathy patients. Our downstream analysis of non-transfusion-dependent ß-thalassemia patients, ß-thalassemia major patients, compound heterozygous sickle cell disease/ß-thalassemia patients receiving hydroxyurea as fetal hemoglobin augmentation treatment, and non-thalassemic individuals indicated that VEGFA genomic variants were associated with disease severity in ß-thalassemia patients and hydroxyurea treatment efficacy in SCD/ß-thalassemia compound heterozygous patients. CONCLUSIONS: Our findings suggest that VEGFA may act as a modifier gene of human globin gene expression and, at the same time, serve as a genomic biomarker in ß-type hemoglobinopathy disease severity and hydroxyurea treatment efficacy.


Assuntos
Células Eritroides , Hemoglobina Fetal/genética , Hidroxiureia/farmacologia , Fator A de Crescimento do Endotélio Vascular/genética , Talassemia beta/tratamento farmacológico , Biomarcadores Farmacológicos , Simulação por Computador , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Heterozigoto , Humanos , Desequilíbrio de Ligação , Farmacogenética , Polimorfismo de Nucleotídeo Único , Talassemia beta/genética
15.
Adv Exp Med Biol ; 989: 129-139, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28971422

RESUMO

The purpose of this research is to study expectations regarding ageing (ERA) among individuals who participate in running events as well as to explore personality and demographic features as potential variables that influence ERA values. A quantitative questionnaire was selected as the predominant means of collecting the data and 196 successfully completed questionnaires were analyzed by means of the SPSS. Results indicate a positive correlation between ERA values and Change Seeker Index in our sample. Moreover gender and frequency of exercise found to have no significant effect on ERA score. Finally, ERA was examined among three generational cohorts and differences were noticed to physical subscale.


Assuntos
Envelhecimento , Corrida , Exercício Físico , Humanos , Personalidade , Inquéritos e Questionários
16.
Adv Exp Med Biol ; 987: 23-34, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28971444

RESUMO

BACKGROUND: Anxiety combined with nervousness and apprehension consist a focal response to different life conditions. Lifestyle habits, anxiety and biochemical markers are in a constant interaction. AIM: To investigate the prevalence of anxiety in healthy adults and its possible association with biochemical factors-lipid profile, liver markers, thyroid hormones-and lifestyle habits. METHODS: Quantitative descriptive correlation study. A total of 100 healthy adults participated in the research. A specially designed questionnaire and Hamilton's scale were used. Anthropometric and biochemical analyses were performed. FINDINGS: Overall, 61% of the participants presented moderate to very serious anxiety. The average score on the Hamilton scale was 13.82 (±9.000), with men exhibiting less stress than women. For p ≤ 0.05: Stress was positively correlated with impaired thyroid and hepatic function. Hepatic function was affected by both sugar products and water melon, which were positively correlated with total bilirubin and AST/SGOT respectively. Tomato, peppers and legumes were negatively correlated with AST/SGOT. Deep fried food was positively correlated with GGT and triglycerides. Legumes and fish were negatively correlated with CPK. Regarding the lipid metabolism, it was found that food cooked with oil was positively associated with uric acid, but non-cooked olive oil was negatively correlated with the risk for CAD. Thyroid function was negatively correlated with non-homemade food and pasta consumption and positively correlated with consumption of whole grains and green tea. Participants with subclinical hypothyroidism seemed to consume less vitamin B12, folic acid and vegetables. CONCLUSION: No direct correlation between lifestyle habits and anxiety was found. Nevertheless, eating habits influenced biochemical markers-especially the thyroid hormones-which may be indirectly responsible for anxiety and related moods.


Assuntos
Ansiedade/fisiopatologia , Metabolismo Energético/fisiologia , Comportamento Alimentar , Estilo de Vida , Estado Nutricional , Adulto , Ansiedade/epidemiologia , Ansiedade/psicologia , Índice de Massa Corporal , Ingestão de Alimentos/fisiologia , Feminino , Grécia/epidemiologia , Hábitos , Humanos , Masculino , Pessoa de Meia-Idade , Prevalência , Inquéritos e Questionários
17.
Cell Stem Cell ; 20(5): 689-705.e9, 2017 05 04.
Artigo em Inglês | MEDLINE | ID: mdl-28285903

RESUMO

Poised enhancers marked by H3K27me3 in pluripotent stem cells have been implicated in the establishment of somatic expression programs during embryonic stem cell (ESC) differentiation. However, the functional relevance and mechanism of action of poised enhancers remain unknown. Using CRISPR/Cas9 technology to engineer precise genetic deletions, we demonstrate that poised enhancers are necessary for the induction of major anterior neural regulators. Interestingly, circularized chromosome conformation capture sequencing (4C-seq) shows that poised enhancers already establish physical interactions with their target genes in ESCs in a polycomb repressive complex 2 (PRC2)-dependent manner. Loss of PRC2 does not activate poised enhancers or induce their putative target genes in undifferentiated ESCs; however, loss of PRC2 in differentiating ESCs severely and specifically compromises the induction of major anterior neural genes representing poised enhancer targets. Overall, our work illuminates an unexpected function for polycomb proteins in facilitating neural induction by endowing major anterior neural loci with a permissive regulatory topology.


Assuntos
Células-Tronco Pluripotentes/citologia , Células-Tronco Pluripotentes/metabolismo , Complexo Repressor Polycomb 2/metabolismo , Animais , Diferenciação Celular/genética , Diferenciação Celular/fisiologia , Linhagem Celular , Imunoprecipitação da Cromatina , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Imunofluorescência , Camundongos , Complexo Repressor Polycomb 2/genética , Reação em Cadeia da Polimerase
18.
Mol Syst Biol ; 12(12): 891, 2016 Dec 09.
Artigo em Inglês | MEDLINE | ID: mdl-27940490

RESUMO

Mammalian interphase chromosomes fold into a multitude of loops to fit the confines of cell nuclei, and looping is tightly linked to regulated function. Chromosome conformation capture (3C) technology has significantly advanced our understanding of this structure-to-function relationship. However, all 3C-based methods rely on chemical cross-linking to stabilize spatial interactions. This step remains a "black box" as regards the biases it may introduce, and some discrepancies between microscopy and 3C studies have now been reported. To address these concerns, we developed "i3C", a novel approach for capturing spatial interactions without a need for cross-linking. We apply i3C to intact nuclei of living cells and exploit native forces that stabilize chromatin folding. Using different cell types and loci, computational modeling, and a methylation-based orthogonal validation method, "TALE-iD", we show that native interactions resemble cross-linked ones, but display improved signal-to-noise ratios and are more focal on regulatory elements and CTCF sites, while strictly abiding to topologically associating domain restrictions.


Assuntos
Núcleo Celular/genética , Cromossomos Humanos/química , Cromossomos Humanos/genética , Animais , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Células Endoteliais da Veia Umbilical Humana , Humanos , Interfase , Células K562 , Mamíferos/genética , Modelos Genéticos , Conformação de Ácido Nucleico , Análise de Sequência com Séries de Oligonucleotídeos , Análise de Sequência de DNA/métodos
19.
Artigo em Inglês | MEDLINE | ID: mdl-28035242

RESUMO

BACKGROUND: The dynamic three-dimensional chromatin architecture of genomes and its co-evolutionary connection to its function-the storage, expression, and replication of genetic information-is still one of the central issues in biology. Here, we describe the much debated 3D architecture of the human and mouse genomes from the nucleosomal to the megabase pair level by a novel approach combining selective high-throughput high-resolution chromosomal interaction capture (T2C), polymer simulations, and scaling analysis of the 3D architecture and the DNA sequence. RESULTS: The genome is compacted into a chromatin quasi-fibre with ~5 ± 1 nucleosomes/11 nm, folded into stable ~30-100 kbp loops forming stable loop aggregates/rosettes connected by similar sized linkers. Minor but significant variations in the architecture are seen between cell types and functional states. The architecture and the DNA sequence show very similar fine-structured multi-scaling behaviour confirming their co-evolution and the above. CONCLUSIONS: This architecture, its dynamics, and accessibility, balance stability and flexibility ensuring genome integrity and variation enabling gene expression/regulation by self-organization of (in)active units already in proximity. Our results agree with the heuristics of the field and allow "architectural sequencing" at a genome mechanics level to understand the inseparable systems genomic properties.

20.
Genome Res ; 26(11): 1478-1489, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27633323

RESUMO

Mammalian cells have developed intricate mechanisms to interpret, integrate, and respond to extracellular stimuli. For example, tumor necrosis factor (TNF) rapidly activates proinflammatory genes, but our understanding of how this occurs against the ongoing transcriptional program of the cell is far from complete. Here, we monitor the early phase of this cascade at high spatiotemporal resolution in TNF-stimulated human endothelial cells. NF-κB, the transcription factor complex driving the response, interferes with the regulatory machinery by binding active enhancers already in interaction with gene promoters. Notably, >50% of these enhancers do not encode canonical NF-κB binding motifs. Using a combination of genomics tools, we find that binding site selection plays a key role in NF-κΒ-mediated transcriptional activation and repression. We demonstrate the latter by describing the synergy between NF-κΒ and the corepressor JDP2. Finally, detailed analysis of a 2.8-Mbp locus using sub-kbp-resolution targeted chromatin conformation capture and genome editing uncovers how NF-κΒ that has just entered the nucleus exploits pre-existing chromatin looping to exert its multimodal role. This work highlights the involvement of topology in cis-regulatory element function during acute transcriptional responses, where primary DNA sequence and its higher-order structure constitute a regulatory context leading to either gene activation or repression.


Assuntos
Sequência Consenso , NF-kappa B/metabolismo , Regiões Promotoras Genéticas , Ativação Transcricional , Células Cultivadas , Cromatina/metabolismo , Edição de Genes , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Humanos , Inflamação/genética , Inflamação/metabolismo , NF-kappa B/genética , Ligação Proteica , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Fator de Necrose Tumoral alfa/farmacologia
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