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CRISPR/Cas9 has become a powerful tool for genome editing in zebrafish that permits the rapid generation of loss of function mutations and the knock-in of specific alleles using DNA templates and homology directed repair (HDR). We examined the efficiency of synthetic, chemically modified gRNAs and demonstrate induction of indels and large genomic deletions in combination with recombinant Cas9 protein. We developed an in vivo genetic assay to measure HDR efficiency and we utilized this assay to test the effect of altering template design on HDR. Utilizing synthetic gRNAs and linear dsDNA templates, we successfully performed knock-in of fluorophores at multiple genomic loci and demonstrate transmission through the germline at high efficiency. We demonstrate that synthetic HDR templates can be used to knock-in bacterial nitroreductase (ntr) to facilitate lineage ablation of specific cell types. Collectively, our data demonstrate the utility of combining synthetic gRNAs and dsDNA templates to perform homology directed repair and genome editing in vivo.
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Proteína 9 Asociada a CRISPR , Sistemas CRISPR-Cas , Edición Génica , Reparación del ADN por Recombinación , Animales , Proteína 9 Asociada a CRISPR/genética , Colorantes Fluorescentes , Proteínas Fluorescentes Verdes/genética , Mutación INDEL , Indicadores y Reactivos , Melanocitos , Nitrorreductasas/genética , ARN/química , Moldes Genéticos , Pez Cebra/embriología , Pez Cebra/genéticaRESUMEN
Mechanisms of cellular and nuclear mechanosensation are unclear, partially because of a lack of methods that can reveal dynamic processes. Here, we present a new concept for a low-cost, three-dimensionally printed device that enables high-magnification imaging of cells during stretch. We observed that nuclei of mouse embryonic skin fibroblasts underwent rapid (within minutes) and divergent responses, characterized by nuclear area expansion during 5% strain but nuclear area shrinkage during 20% strain. Only responses to low strain were dependent on calcium signaling, whereas actin inhibition abrogated all nuclear responses and increased nuclear strain transfer and DNA damage. Imaging of actin dynamics during stretch revealed similar divergent trends, with F-actin shifting away from (5% strain) or toward (20% strain) the nuclear periphery. Our findings emphasize the importance of simultaneous stimulation and data acquisition to capture mechanosensitive responses and suggest that mechanical confinement of nuclei through actin may be a protective mechanism during high mechanical stretch or loading.
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Citoesqueleto de Actina , Actinas , Animales , Núcleo Celular , Células Cultivadas , Ratones , Estrés MecánicoRESUMEN
Co-flowering plant species commonly share flower visitors, and thus have the potential to influence each other's pollination. In this study we analysed 750 quantitative plant-pollinator networks from 28 studies representing diverse biomes worldwide. We show that the potential for one plant species to influence another indirectly via shared pollinators was greater for plants whose resources were more abundant (higher floral unit number and nectar sugar content) and more accessible. The potential indirect influence was also stronger between phylogenetically closer plant species and was independent of plant geographic origin (native vs. non-native). The positive effect of nectar sugar content and phylogenetic proximity was much more accentuated for bees than for other groups. Consequently, the impact of these factors depends on the pollination mode of plants, e.g. bee or fly pollinated. Our findings may help predict which plant species have the greatest importance in the functioning of plant-pollination networks.
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Flores/genética , Magnoliopsida/genética , Filogenia , Polinización , Animales , Abejas , Dípteros , Modelos Biológicos , Néctar de las Plantas/químicaRESUMEN
OBJECTIVES: Growing evidence suggests that religiosity is an important social determinant of health, including cognitive health. Yet most prior work focused on older adults or was conducted in racially and denominationally homogeneous regional samples. This study investigates the association of religious service attendance in midlife with cognitive function later in midlife. METHODS: Using data from the Coronary Artery Risk Development in Young Adults (CARDIA) study, a racially and geographically diverse prospective cohort study, we explored the association of religious service attendance in midlife with cognitive function 5 years later. Cognitive function was measured using four cognitive tests administered by CARDIA technicians. Multivariable linear regression was used for analyses. Primary analyses controlled for sociodemographics, physical health, depression, and prior religious involvement. Sensitivity analyses additionally controlled for baseline cognition and social support. RESULTS: Our study population included 2,716 participants (57.2% female, 44.9% Black, and mean age 50). In primary analyses, attending services more than weekly (compared to never) in midlife was associated with better global cognition (ß = 0.14 standard deviations, 95% [confidence interval] CI = 0.02, 0.26) and verbal memory (ß = 0.17 standard deviations, 95% CI = 0.04, 0.30), but not with processing speed (ß = 0.04 standard deviations, 95% CI = -0.08, 0.16). A reverse association was observed with executive function (ß = -0.16 standard deviations, 95% CI = -0.30, -0.02). Most findings persisted in analyses accounting for loss to follow-up via inverse probability weighting. DISCUSSION: Our findings suggest that frequent involvement in religious services at midlife is associated with better global cognition and verbal memory but worse executive function. There was no association with processing speed.
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Cognición , Vasos Coronarios , Humanos , Femenino , Anciano , Masculino , Estudios Prospectivos , Factores de Riesgo , Función EjecutivaRESUMEN
Stakeholders need data on health and drivers of health parsed to the boundaries of essential policy-relevant geographies. US Congressional Districts are an example of a policy-relevant geography which generally lack health data. One strategy to generate Congressional District heath data metric estimates is to aggregate estimates from other geographies, for example, from counties or census tracts to Congressional Districts. Doing so requires several methodological decisions. We refine a method to aggregate health metric estimates from one geography to another, using a population weighted approach. The method's accuracy is evaluated by comparing three aggregated metric estimates to metric estimates from the US Census American Community Survey for the same years: Broadband Access, High School Completion, and Unemployment. We then conducted four sensitivity analyses testing: the effect of aggregating counts vs. percentages; impacts of component geography size and data missingness; and extent of population overlap between component and target geographies. Aggregated estimates were very similar to estimates for identical metrics drawn directly from the data source. Sensitivity analyses suggest the following best practices for Congressional district-based metrics: utilizing smaller, more plentiful geographies like census tracts as opposed to larger, less plentiful geographies like counties, despite potential for less stable estimates in smaller geographies; favoring geographies with higher percentage population overlap.
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Perturbations to the epigenome are known drivers of tumorigenesis. In melanoma, alterations in histone methyltransferases that catalyze methylation at histone 3 lysine 9 and histone 3 lysine 27-two sites of critical post-translational modification-have been reported. To study the function of these methyltransferases in melanoma, we engineered melanocytes to express histone 3 lysine-to-methionine mutations at lysine 9 and lysine 27, which are known to inhibit the activity of histone methyltransferases, in a zebrafish melanoma model. Using this system, we found that loss of histone 3 lysine 9 methylation dramatically suppressed melanoma formation and that inhibition of histone 3 lysine 9 methyltransferases in human melanoma cells increased innate immune response signatures. In contrast, loss of histone 3 lysine 27 methylation significantly accelerated melanoma formation. We identified FOXD1 as a top target of PRC2 that is silenced in melanocytes and found that aberrant overexpression of FOXD1 accelerated melanoma onset. Collectively, these data demonstrate how histone 3 lysine-to-methionine mutations can be used to uncover critical roles for methyltransferases.
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OBJECTIVES: To measure pain during first trimester medication abortion using auricular acupressure or auricular acupuncture as an adjunct to pain management. We measured anxiety as a secondary outcome. STUDY DESIGN: This randomized, double-blinded, 3-arm trial enrolled women seeking medication abortion with mifepristone and misoprostol. Participants received auricular acupressure, auricular acupuncture, or inert auricular placebo patches immediately after receiving mifepristone. In addition, all participants received ibuprofen to use at home as needed. The study started with 1:1:1 randomization, but later overenrolled into the acupressure group after retraining for greater fidelity to that intervention. Participants reported pain and anxiety using numeric rating scales via text message for 4 days, and using a visual analog scale at follow-up. Analyses compared median pain scores of those receiving acupressure, acupuncture, or placebo. RESULTS: We randomized 136 participants of whom 57 received acupressure, 40 received acupuncture, and 39 received placebo. Groups had similar baseline characteristics. One hundred thirty-two participants (97%) reported outcomes by text message and 120 (88%) completed a follow-up interview. For acupressure, acupuncture and placebo groups the median maximum pain scores reported via text message were 60.0, 75.0, and 55.0 (p = 0.38); median maximum pain scores reported at follow-up were 76.5, 60.0, and 71.0 (p = 0.97), respectively. Acupressure results were similar before and after retraining. Maximum anxiety scores reported via text message were 10.0, 45.0, and 30.0 (p = 0.57). Maximum anxiety scores reported at follow-up were 10.5, 20.0, and 13.0 (p = 0.59). CONCLUSIONS: Pain and anxiety during medication abortion were similar among women receiving acupressure, acupuncture or placebo. IMPLICATIONS: We found no benefit in administering auricular acupressure or auricular acupuncture during medication abortion. These modalities are intended to be simple to use, but perhaps the brief provider training for this study was insufficient. These modalities should not be used in clinical practice without further study.
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Acupresión , Acupuntura Auricular , Femenino , Humanos , Dolor , Manejo del Dolor , Embarazo , Primer Trimestre del EmbarazoRESUMEN
Objetivo: Identificar o conhecimento e a aplicação das práticas integrativas e complementares pelos enfermeiros da atenção básica. Métodos: É uma pesquisa descritiva de abordagem qualitativa envolvendo 19 enfermeiros da atenção básica do município de Mossoró situado no estado do Rio Grande do Norte, aprovada pelo comitê de ética e pesquisa sob o parecer 2.113.411. Resultados: Identificou-se quatro categorias: conhecimento dos enfermeiros sobre práticas integrativas; a não implementação das práticas integrativas e complementares na assistência do enfermeiro; implementação das práticas integrativas e complementares na assistência do enfermeiro; e as práticas são importantes para o cuidado em saúde. Conclusão: O conhecimento sobre a politica nacional de práticas integrativas e complementares, a variabilidade e a finalidade de tais práticas ainda se apresenta limitado, a solução para o conhecimento e implementação deficitários seria o ensino, seja ele na graduação ou na vida profissional
Objective: The study's purpose has been to identify the understanding and implementation of integrative and complementary practices by primary care nurses. Methods: It is a descriptive research with a qualitative approach that counted with the participation of 19 primary care nurses. This study took place in the municipality of Mossoró located in the Rio Grande do Norte State, being approved by the Research Ethics Committee under the Legal Opinion No. 2.113.411. Results: The following four categories were identified: nurses' knowledge regarding integrative practices; failure to implement integrative and complementary practices in nursing care; implementation of integrative and complementary practices in nursing care; and, the professional practices are important for health care. Conclusion: The knowledge concerning the national policy for integrative and complementary practices, the variability and the purpose of such practices is still limited. Bearing this in mind, the solution for such understanding and deficient implementation would be through teaching, being it during either graduation or professional life
Objetivo: Identificar el conocimiento e implementación de las prácticas integrativas y complementarias por los enfermeros de la atención básica. Métodos: Es una investigación descriptiva de abordaje cualitativo involucrando a 19 enfermeras de la atención básica del municipio de Mossoró, localizado en el estado de Rio Grande do Norte, aprobado por el comité de ética e investigación bajo la opinión 2.111.411. Resultados: Se identificaron cuatro categorías: conocimiento de enfermeros sobre prácticas integrativas; la no implementación de prácticas integrativas y complementarias en cuidados de enfermería; implementación de prácticas integradoras y complementarias en cuidados; y prácticas de enfermería son importantes para la atención de salud. Conclusión: El conocimiento sobre la política nacional de prácticas integrativas y complementarias, la variabilidad y el propósito de las mismas todavía está limitado, de esa forma, la solución sería la enseñanza
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Humanos , Masculino , Femenino , Adulto , Persona de Mediana Edad , Anciano , Terapias Complementarias/enfermería , Educación Continua en Enfermería , Enfermería de Atención Primaria/métodos , Investigación Cualitativa , Capacitación de Recursos Humanos en Salud , Integralidad en SaludRESUMEN
Introdução: O Transplante de Células-Tronco Hematopoéticas (TCTH) é um procedimento que substitui células doentes por células sadias, por meio da infusão de células-tronco hematopoéticas. Objetivo: Identificar na literatura as publicações da enfermagem que abordem o TCTH de Sangue de Cordão Umbilical (SCU). Método: Trata-se de uma revisão integrativa, para qual se realizou buscas por meio das bases de dados eletrônicas: Biblioteca Virtual em Saúde, National Library of Medicine (PUBMED/MEDLINE), Science Direct e SCOPUS. Foram estabelecidos os seguintes critérios de inclusão: estudos disponíveis em qualquer idioma, estudos que abordassem o TCTH de sangue de cordão umbilical e estudo publicado na íntegra. Resultados: O processo de busca eletrônica dos estudos resultou em um total de 978 estudos, dos quais cinco compuseram a amostra da revisão. Os estudos selecionados abordaram o conhecimento dos enfermeiros sobre o transplante de SCU, a coleta e armazenamento do SCU e o banco de SCU. Conclusão: O presente estudo proporcionou a percepção da importância do papel do enfermeiro no TCTH de SCU, visto que seu empenho junto à equipe dos bancos públicos de SCU é importante para alcançar melhores resultados, com a ampliação do número de doadores e maior satisfação dos clientes e seus familiares
Introducción: El trasplante de células madre hematopoyéticas (TCMH) es un procedimiento que reemplaza las células enfermas a las células sanas, a través de la infusión de células madre hematopoyéticas. Objetivo: Identificar en la literatura las publicaciones de enfermería que se ocupan de la sangre de cordón umbilical La sangre (UCB) TCMH. Método: Se trata de una revisión integradora, para lo cual allanó a través de bases de datos electrónicas: Biblioteca Virtual en Salud, Biblioteca Nacional de Medicina (PubMed/MEDLINE), Science Direct y Scopus. Se establecieron los siguientes criterios de inclusión: los estudios disponibles en cualquier idioma, los estudios que abordaron la sangre del cordón umbilical TPH y el estudio en su totalidad. Resultados: El proceso de búsqueda electrónica de los estudios dieron como resultado un total de 978 estudios, de los cuales cinco fueron incluidas en la muestra de revisión. Los estudios seleccionados abordan el conocimiento del personal de enfermería en el trasplante de SCU, la recogida y almacenamiento de sangre del cordón umbilical y el Banco de SCU. Conclusión: Este estudio proporciona la percepción de la importancia del papel de la enfermera en TPH de SCU, como su compromiso con el equipo de los bancos públicos UCB es importante para lograr mejores resultados al aumentar el número de donantes y el aumento de la satisfacción del cliente y sus familias
Introduction: Hematopoietic stem cell transplantation (HSCT) is a procedure that replaces diseased cells with healthy cells by infusing hematopoietic stem cells. Objective: To identify in the literature the nursing publications that addresses HSCT of Umbilical Cord Blood (UCB). Method: This is an integrative review which has been searched through electronic databases: Virtual Health Library, National Library of Medicine (PUBMED/MEDLINE), Science Direct and SCOPUS. The following inclusion criteria were established: studies available in any language, studies addressing HSCT of umbilical cord blood and a study published in its entirety. Results: The electronic search process of the studies resulted in a total of 978 studies, of which five comprised the review sample. The selected studies addressed the nurses' knowledge about SCU transplantation, the collection and storage of SCU and the SCU database. Conclusion: The present study provided an insight into the importance of the role of nurses in HSCTWC, since their commitment to the team of SCU public banks is important to achieve better results, with a larger number of donors and greater customer satisfaction And their families