RESUMO
A major challenge in systems biology is to understand the gene regulatory networks that drive development, physiology and pathology. Interactions between transcription factors and regulatory genomic regions provide the first level of gene control. Gateway-compatible yeast one-hybrid (Y1H) assays present a convenient method to identify and characterize the repertoire of transcription factors that can bind a DNA sequence of interest. To delineate genome-scale regulatory networks, however, large sets of DNA fragments need to be processed at high throughput and high coverage. Here we present enhanced Y1H (eY1H) assays that use a robotic mating platform with a set of improved Y1H reagents and automated readout quantification. We demonstrate that eY1H assays provide excellent coverage and identify interacting transcription factors for multiple DNA fragments in a short time. eY1H assays will be an important tool for mapping gene regulatory networks in Caenorhabditis elegans and other model organisms as well as in humans.
Assuntos
Redes Reguladoras de Genes , Ensaios de Triagem em Larga Escala , Técnicas do Sistema de Duplo-Híbrido , Animais , Caenorhabditis elegans/genética , Caenorhabditis elegans/metabolismo , DNA/genética , Regulação da Expressão Gênica , Humanos , Reprodutibilidade dos Testes , Biologia de Sistemas , Fatores de Transcrição/metabolismoRESUMO
The neural control of hand movement involves coordination of the sensory, motor, and memory systems. Recent studies have documented the motor coordinates for hand shape, but less is known about the corresponding patterns of somatosensory activity. To initiate this line of investigation, the present study characterized the sense of hand shape by evaluating the influence of differences in the amount of grasping or twisting force, and differences in forearm orientation. Human subjects were asked to use the left hand to report the perceived shape of the right hand. In the first experiment, six commonly grasped items were arranged on the table in front of the subject: bottle, doorknob, egg, notebook, carton, and pan. With eyes closed, subjects used the right hand to lightly touch, forcefully support, or imagine holding each object, while 15 joint angles were measured in each hand with a pair of wired gloves. The forces introduced by supporting or twisting did not influence the perceptual report of hand shape, but for most objects, the report was distorted in a consistent manner by differences in forearm orientation. Subjects appeared to adjust the intrinsic joint angles of the left hand, as well as the left wrist posture, so as to maintain the imagined object in its proper spatial orientation. In a second experiment, this result was largely replicated with unfamiliar objects. Thus, somatosensory and motor information appear to be coordinated in an object-based, spatial-coordinate system, sensitive to orientation relative to gravitational forces, but invariant to grasp forcefulness.
Assuntos
Força da Mão/fisiologia , Mãos/fisiologia , Movimento/fisiologia , Desempenho Psicomotor/fisiologia , Tato/fisiologia , Adulto , Análise de Variância , Fenômenos Biomecânicos/fisiologia , Feminino , Humanos , MasculinoRESUMO
BACKGROUND: Surgical reconstruction of congenital heart defects is often limited by the nonresorbable material used to approximate normal anatomy. In contrast, biologic scaffold materials composed of resorbable non-cross-linked extracellular matrix (ECM) have been used for tissue reconstruction of multiple organs and are replaced by host tissue. Preparation of whole organ ECM by decellularization through vascular perfusion can maintain much of the native three-dimensional (3D) structure, strength, and tissue-specific composition. A 3D cardiac ECM (C-ECM) biologic scaffold material would logically have structural and functional advantages over materials such as Dacron™ for myocardial repair, but the in vivo remodeling characteristics of C-ECM have not been investigated to date. METHODS AND RESULTS: A porcine C-ECM patch or Dacron patch was used to reconstruct a full-thickness right ventricular outflow tract (RVOT) defect in a rat model with end points of structural remodeling function at 16 weeks. The Dacron patch was encapsulated by dense fibrous tissue and showed little cellular infiltration. Echocardiographic analysis showed that the right ventricle of the hearts patched with Dacron were dilated at 16 weeks compared to presurgery baseline values. The C-ECM patch remodeled into dense, cellular connective tissue with scattered small islands of cardiomyocytes. The hearts patched with C-ECM showed no difference in the size or function of the ventricles as compared to baseline values at both 4 and 16 weeks. CONCLUSIONS: The C-ECM patch was associated with better functional and histomorphological outcomes compared to the Dacron patch in this rat model of RVOT reconstruction.
Assuntos
Matriz Extracelular/química , Ventrículos do Coração/citologia , Ventrículos do Coração/cirurgia , Engenharia Tecidual/métodos , Alicerces Teciduais/química , Animais , Ecocardiografia , Feminino , Cardiopatias Congênitas/patologia , Cardiopatias Congênitas/cirurgia , Ventrículos do Coração/patologia , Polietilenotereftalatos/química , Ratos , Ratos Endogâmicos Lew , SuínosRESUMO
Physicians underutilize adverse event reporting systems. Web-based platforms have increased participation; thus, it was hypothesized that a mobile application would increase adverse event reporting. The authors developed a mobile reporting application for iOS and Android operating systems and performed a retrospective review on reporting rates by clinicians in the Department of Anesthesia and Critical Care. Monthly reporting rates were calculated for the intervention year and for the 2 prior years (2013-2016). The Wilcoxon rank sum test and χ2 test were used to evaluate significance. Overall monthly reporting rates for all clinicians were 15.3 ± 7 for the first time period, 17.3 ± 6 for the second time period, and 27.9 ± 7 for the third time period ( P = .0035). The majority of reports in the third time period were submitted using the mobile application (193/337, 57%, P = .026). Deployment of a mobile application reduced barriers to adverse event reporting and increased monthly reporting rates for all clinicians.
Assuntos
Sistemas de Notificação de Reações Adversas a Medicamentos , Aplicativos Móveis , Anestesia , Humanos , Avaliação de Programas e Projetos de Saúde , Estudos RetrospectivosRESUMO
BACKGROUND: Genome-wide sensitivity screens in yeast have been immensely popular following the construction of a collection of deletion mutants of non-essential genes. However, the auxotrophic markers in this collection preclude experiments on minimal growth medium, one of the most informative metabolic environments. Here we present quantitative growth analysis for mutants in all 4,772 non-essential genes from our prototrophic deletion collection across a large set of metabolic conditions. RESULTS: The complete collection was grown in environments consisting of one of four possible carbon sources paired with one of seven nitrogen sources, for a total of 28 different well-defined metabolic environments. The relative contributions to mutants' fitness of each carbon and nitrogen source were determined using multivariate statistical methods. The mutant profiling recovered known and novel genes specific to the processing of nutrients and accurately predicted functional relationships, especially for metabolic functions. A benchmark of genome-scale metabolic network modeling is also given to demonstrate the level of agreement between current in silico predictions and hitherto unavailable experimental data. CONCLUSIONS: These data address a fundamental deficiency in our understanding of the model eukaryote Saccharomyces cerevisiae and its response to the most basic of environments. While choice of carbon source has the greatest impact on cell growth, specific effects due to nitrogen source and interactions between the nutrients are frequent. We demonstrate utility in characterizing genes of unknown function and illustrate how these data can be integrated with other whole-genome screens to interpret similarities between seemingly diverse perturbation types.
Assuntos
Deleção de Genes , Genoma Fúngico , Metaboloma , Saccharomyces cerevisiae/genética , Carbono/metabolismo , Respiração Celular , Fermentação , Nitrogênio/metabolismo , Saccharomyces cerevisiae/metabolismoRESUMO
Scaffolds for tissue engineering and regenerative medicine applications are commonly manufactured from synthetic materials, intact or isolated components of extracellular matrix (ECM), or a combination of such materials. After surgical implantation, the metabolic requirements of cells that populate the scaffold depend upon adequate gas and nutrient exchange with the surrounding microenvironment. The present study measured the oxygen transfer through three biologic scaffold materials composed of ECM including small intestinal submucosa (SIS), urinary bladder submucosa (UBS), and urinary bladder matrix (UBM), and one synthetic biomaterial, Dacron. The oxygen diffusivity was calculated from Fick's first law of diffusion. Each material permitted measurable oxygen diffusion. The diffusivity of SIS was found to be dependent on the direction of oxygen transfer; the oxygen transfer in the abluminal-to-luminal direction was significantly greater than the luminal-to-abluminal direction. The oxygen diffusivity of UBM and UBS were similar despite the presence of an intact basement membrane on the luminal surface of UBM. Dacron showed oxygen diffusivity values seven times greater than the ECM biomaterials. The current study showed that each material has unique oxygen diffusivity values, and these values may be dependent on the scaffold's ultrastructure.