Computational methods to create and analyze a digital gene expression atlas of embryo development from microscopy images

Abstract The creation of atlases, or digital models where information from different subjects can be combined, is a field of increasing interest in biomedical imaging. When a single image does not contain enough information to appropriately describe the organism under study, it is then necessary to acquire images of several individuals, each of them containing complementary data with respect to the rest of the components in the cohort. This approach allows creating digital prototypes, ranging from anatomical atlases of human patients and organs, obtained for instance from Magnetic Resonance Imaging, to gene expression cartographies of embryo development, typically achieved from Light Microscopy. Within such context, in this PhD Thesis we propose, develop and validate new dedicated image processing methodologies that, based on image registration techniques, bring information from multiple individuals into alignment within a single digital atlas model. We also elaborate a dedicated software visualization platform to explore the resulting wealth of multi-dimensional data and novel analysis algo-rithms to automatically mine the generated resource in search of bio¬logical insights. In particular, this work focuses on gene expression data from developing zebrafish embryos imaged at the cellular resolution level with Two-Photon Laser Scanning Microscopy. Disposing of quantitative measurements relating multiple gene expressions to cell position and their evolution in time is a fundamental prerequisite to understand embryogenesis multi-scale processes. However, the number of gene expressions that can be simultaneously stained in one acquisition is limited due to optical and labeling constraints. These limitations motivate the implementation of atlasing strategies that can recreate a virtual gene expression multiplex. The developed computational tools have been tested in two different scenarios. The first one is the early zebrafish embryogenesis where the resulting atlas constitutes a link between the phenotype and the genotype at the cellular level. The second one is the late zebrafish brain where the resulting atlas allows studies relating gene expression to brain regionalization and neurogenesis. The proposed computational frameworks have been adapted to the requirements of both scenarios, such as the integration of partial views of the embryo into a whole embryo model with cellular resolution or the registration of anatom¬ical traits with deformable transformation models non-dependent on any specific labeling. The software implementation of the atlas generation tool (Match-IT) and the visualization platform (Atlas-IT) together with the gene expression atlas resources developed in this Thesis are to be made freely available to the scientific community. Lastly, a novel proof-of-concept experiment integrates for the first time 3D gene expression atlas resources with cell lineages extracted from live embryos, opening up the door to correlate genetic and cellular spatio-temporal dynamics. La creación de atlas, o modelos digitales, donde la información de distintos sujetos puede ser combinada, es un campo de creciente interés en imagen biomédica. Cuando una sola imagen no contiene suficientes datos como para describir apropiadamente el organismo objeto de estudio, se hace necesario adquirir imágenes de varios individuos, cada una de las cuales contiene información complementaria respecto al resto de componentes del grupo. De este modo, es posible crear prototipos digitales, que pueden ir desde atlas anatómicos de órganos y pacientes humanos, adquiridos por ejemplo mediante Resonancia Magnética, hasta cartografías de la expresión genética del desarrollo de embrionario, típicamente adquiridas mediante Microscopía Optica. Dentro de este contexto, en esta Tesis Doctoral se introducen, desarrollan y validan nuevos métodos de procesado de imagen que, basándose en técnicas de registro de imagen, son capaces de alinear imágenes y datos provenientes de múltiples individuos en un solo atlas digital. Además, se ha elaborado una plataforma de visualization específicamente diseñada para explorar la gran cantidad de datos, caracterizados por su multi-dimensionalidad, que resulta de estos métodos. Asimismo, se han propuesto novedosos algoritmos de análisis y minería de datos que permiten inspeccionar automáticamente los atlas generados en busca de conclusiones biológicas significativas. En particular, este trabajo se centra en datos de expresión genética del desarrollo embrionario del pez cebra, adquiridos mediante Microscopía dos fotones con resolución celular. Disponer de medidas cuantitativas que relacionen estas expresiones genéticas con las posiciones celulares y su evolución en el tiempo es un prerrequisito fundamental para comprender los procesos multi-escala característicos de la morfogénesis. Sin embargo, el número de expresiones genéticos que pueden ser simultáneamente etiquetados en una sola adquisición es reducido debido a limitaciones tanto ópticas como del etiquetado. Estas limitaciones requieren la implementación de estrategias de creación de atlas que puedan recrear un multiplexado virtual de expresiones genéticas. Las herramientas computacionales desarrolladas han sido validadas en dos escenarios distintos. El primer escenario es el desarrollo embrionario temprano del pez cebra, donde el atlas resultante permite constituir un vínculo, a nivel celular, entre el fenotipo y el genotipo de este organismo modelo. El segundo escenario corresponde a estadios tardíos del desarrollo del cerebro del pez cebra, donde el atlas resultante permite relacionar expresiones genéticas con la regionalización del cerebro y la formación de neuronas. La plataforma computacional desarrollada ha sido adaptada a los requisitos y retos planteados en ambos escenarios, como la integración, a resolución celular, de vistas parciales dentro de un modelo consistente en un embrión completo, o el alineamiento entre estructuras de referencia anatómica equivalentes, logrado mediante el uso de modelos de transformación deformables que no requieren ningún marcador específico. Está previsto poner a disposición de la comunidad científica tanto la herramienta de generación de atlas (Match-IT), como su plataforma de visualización (Atlas-IT), así como las bases de datos de expresión genética creadas a partir de estas herramientas. Por último, dentro de la presente Tesis Doctoral, se ha incluido una prueba conceptual innovadora que permite integrar los mencionados atlas de expresión genética tridimensionales dentro del linaje celular extraído de una adquisición in vivo de un embrión. Esta prueba conceptual abre la puerta a la posibilidad de correlar, por primera vez, las dinámicas espacio-temporales de genes y células.

Infrared thermography to quantify the risk of breast cancer.

From the last decades, infrared thermography is quite often associated with things other than clinical medicine. For example, the chemical, automobile, aeronautic industries and civil engineering. However, thermography is where infrared images of the breast are analyzed by board certified thermographers and an abnormal thermogram is reported as the significant risk for the existence of breast tumor (Ng, 2009). Thermography is a painless, noninvasive, no radiation, as well as being cheaper and faster, easier access. The aim of this review was to identify the views of clinicians on the use of thermography for quantifying the risk of breast cancer. We used articles published recently in a reliable database. Thermography has been convicted over the years; it has been labeled by subjective interpretation. Most of the reviewed articles agree that mammography is currently the main examination chosen by doctors for the screening of breast cancer (Acharya et al., 2010; Kennedy et al., 2009). However, several studies have reported promising results for the technique (Wang et al., 2010). Additionally, some authors suggest that thermography is complementary to other diagnostic methods, and that the best strategy for the early detection of breast cancer would be to use them together (Kennedy et al., 2009; Hersh, 2004). The combination of thermal imaging with other tests would increase accuracy, sensitivity and specificity of the evaluation and allow a better quantification of the risk of breast cancer.

Overexpression of transforming growth factor β1 in arterial endothelium causes hyperplasia, apoptosis, and cartilaginous metaplasia

Uninjured rat arteries transduced with an adenoviral vector expressing an active form of transforming growth factor β1 (TGF-β1) developed a cellular and matrix-rich neointima, with cartilaginous metaplasia of the vascular media. Explant cultures of transduced arteries showed that secretion of active TGF-β1 ceased by 4 weeks, the time of maximal intimal thickening. Between 4 and 8 weeks, the cartilaginous metaplasia resolved and the intimal lesions regressed almost completely, in large part because of massive apoptosis. Thus, locally expressed TGF-β1 promotes intimal growth and appears to cause transdifferentiation of vascular smooth muscle cells into chondrocytes. Moreover, TGF-β1 withdrawal is associated with regression of vascular lesions. These data suggest an unexpected plasticity of the adult vascular smooth muscle cell phenotype and provide an etiology for cartilaginous metaplasia of the arterial wall. Our observations may help to reconcile divergent views of the role of TGF-β1 in vascular disease.

Variations in frequencies of drug resistance in Plasmodium falciparum

Continual exposure of malarial parasite populations to different drugs may have selected not only for resistance to individual drugs but also for genetic traits that favor initiation of resistance to novel unrelated antimalarials. To test this hypothesis, different Plasmodium falciparum clones having varying numbers of preexisting resistance mechanisms were treated with two new antimalarial agents: 5-fluoroorotate and atovaquone. All parasite populations were equally susceptible in small numbers. However, when large populations of these clones were challenged with either of the two compounds, significant variations in frequencies of resistance became apparent. On one extreme, clone D6 from West Africa, which was sensitive to all traditional antimalarial agents, failed to develop resistance under simple nonmutagenic conditions in vitro. In sharp contrast, the Indochina clone W2, which was known to be resistant to all traditional antimalarial drugs, independently acquired resistance to both new compounds as much as a 1,000 times more frequently than D6. Additional clones that were resistant to some (but not all) traditional antimalarial agents acquired resistance to atovaquone at high frequency, but not to 5-fluoroorotate. These findings were unexpected and surprising based on current views of the evolution of drug resistance in P. falciparum populations. Such new phenotypes, named accelerated resistance to multiple drugs (ARMD), raise important questions about the genetic and biochemical mechanisms related to the initiation of drug resistance in malarial parasites. Some potential mechanisms underlying ARMD phenotypes have public health implications that are ominous.

Reactions of participants to the results of a randomised controlled trial: exploratory study

Objectives: To assess views of parents of babies who participated in a neonatal trial, about feedback of trial results.

Comparative analysis of the gene-dense ACHE/TFR2 region on human chromosome 7q22 with the orthologous region on mouse chromosome 5

Chromosome 7q22 has been the focus of many cytogenetic and molecular studies aimed at delineating regions commonly deleted in myeloid leukemias and myelodysplastic syndromes. We have compared a gene-dense, GC-rich sub-region of 7q22 with the orthologous region on mouse chromosome 5. A physical map of 640 kb of genomic DNA from mouse chromosome 5 was derived from a series of overlapping bacterial artificial chromosomes. A 296 kb segment from the physical map, spanning Ache to Tfr2, was compared with 267 kb of human sequence. We identified a conserved linkage of 12 genes including an open reading frame flanked by Ache and Asr2, a novel cation-chloride cotransporter interacting protein Cip1, Ephb4, Zan and Perq1. While some of these genes have been previously described, in each case we present new data derived from our comparative sequence analysis. Adjacent unfinished sequence data from the mouse contains an orthologous block of 10 additional genes including three novel cDNA sequences that we subsequently mapped to human 7q22. Methods for displaying comparative genomic information, including unfinished sequence data, are becoming increasingly important. We supplement our printed comparative analysis with a new, Web-based program called Laj (local alignments with java). Laj provides interactive access to archived pairwise sequence alignments via the WWW. It displays synchronized views of a dot-plot, a percent identity plot, a nucleotide-level local alignment and a variety of relevant annotations. Our mouse–human comparison can be viewed at http://web.uvic.ca/~bioweb/laj.html. Laj is available at http://bio.cse.psu.edu/, along with online documentation and additional examples of annotated genomic regions.

Effects of temporal association on recognition memory

The influence of temporal association on the representation and recognition of objects was investigated. Observers were shown sequences of novel faces in which the identity of the face changed as the head rotated. As a result, observers showed a tendency to treat the views as if they were of the same person. Additional experiments revealed that this was only true if the training sequences depicted head rotations rather than jumbled views; in other words, the sequence had to be spatially as well as temporally smooth. Results suggest that we are continuously associating views of objects to support later recognition, and that we do so not only on the basis of the physical similarity, but also the correlated appearance in time of the objects.

Mechanical bases of frequency tuning and neural excitation at the base of the cochlea: Comparison of basilar-membrane vibrations and auditory-nerve-fiber responses in chinchilla

We review the mechanical origin of auditory-nerve excitation, focusing on comparisons of the magnitudes and phases of basilar-membrane (BM) vibrations and auditory-nerve fiber responses to tones at a basal site of the chinchilla cochlea with characteristic frequency ≈ 9 kHz located 3.5 mm from the oval window. At this location, characteristic frequency thresholds of fibers with high spontaneous activity correspond to magnitudes of BM displacement or velocity in the order of 1 nm or 50 μm/s. Over a wide range of stimulus frequencies, neural thresholds are not determined solely by BM displacement but rather by a function of both displacement and velocity. Near-threshold, auditory-nerve responses to low-frequency tones are synchronous with peak BM velocity toward scala tympani but at 80–90 dB sound pressure level (in decibels relative to 20 microPascals) and at 100–110 dB sound pressure level responses undergo two large phase shifts approaching 180°. These drastic phase changes have no counterparts in BM vibrations. Thus, although at threshold levels the encoding of BM vibrations into spike trains appears to involve only relatively minor signal transformations, the polarity of auditory-nerve responses does not conform with traditional views of how BM vibrations are transmitted to the inner hair cells. The response polarity at threshold levels, as well as the intensity-dependent phase changes, apparently reflect micromechanical interactions between the organ of Corti, the tectorial membrane and the subtectorial fluid, and/or electrical and synaptic processes at the inner hair cells.

Processos e significados da experiência de pais, mães ou cuidador primário de crianças abusadas sexualmente

Introdução - O abuso sexual de crianças constitui um problema de saúde pública, com aproximadamente 73 milhões de casos de meninos e 150 milhões de meninas registrados anualmente no mundo. O abuso sexual gera consequências negativas e condutas de risco que contribuem com algumas das principais causas de morte, doença e deficiência nas vítimas do abuso. Pais ou cuidadores primários são fundamentais no processo de orientação e de cuidado das crianças abusadas, no sentido de prevenir as consequências cognitivas, comportamentais e emocionais evidenciadas no futuro dessas crianças. Entretanto, as habilidades das famílias de cuidadores para lidar com a problemática ainda são insuficientes. Objetivo - Descrever os processos e significados da experiência vivida pelos pais ou cuidadores primários de crianças abusadas sexualmente. Método - Os dados empíricos foram tratados utilizando-se o Discurso do Sujeito Coletivo (DSC), fundamentado na Teoria das Representações Sociais, que viabiliza a emergência das representações sociais por meio da construção dos discursos coletivos obtidos de depoimentos de um grupo específico. Foram avaliados 60 pais ou cuidadores primários não estupradores, que responderam à cinco situações-problema, cada uma com questões correspondentes, residentes nos municípios de Cajicá e Tabio de Bogotá, Colômbia. O processamento e a análise dos dados foram realizados no sofware Qualiquantisoft, associado à metodologia do DSC. Resultados - Na primeira situação-problema, que aborda o porquê do silêncio do filho sobre o abuso, os entrevistados enfatizaram que é fundamental o relacionamento pais e filhos (45,7 por cento , n = 43), bem como melhorar o papel de pais por meio da escuta, do diálogo e do confiar, dedicando mais tempo às crianças; também acham que o silêncio se deve ao medo por parte das crianças e a ameaças e intimidação por parte do abusador, Na situação-problema 2, relativa à identificação do abuso sexual como problema real, o significado atribuído configura cadeias que se repetem por transmissão intergeracional (26,9 por cento , n = 21). Na situação-problema 3, o que fazer no futuro, 53,3 por cento dos entrevistados (n = 32) acham que a criança está comprometida comportamentalmente e enfatizam a homossexualidade com perda da identidade como consequência da violência sexual. Na situação-problema 4, que enfatiza o papel da rede social quanto ao cuidado da criança, os entrevistados acreditam que a solução é dar proteção (29,1 por cento ; n = 32), com ações que visem a afastar a criança do ambiente agressor, dar orientação, apoio e segurança à criança e à família. A quinta situação-problema que diz respeito ao cuidado das crianças abusadas; 34,26 por cento dos entrevistados (n = 37) enfatizam o apoio e a ajuda com a interveniência da rede de apoio social e afetivo. Conclusão - Para os pais ou cuidadores primários de crianças abusadas sexualmente, os significados se expressam como afetivo, coragem, superação, não ter medo e saber reconhecer as falhas dos pais. Em função dos resultados, que identificam posturas tradicionais dos respondentes, recomenda-se programas inovadores com um alto componente educativo, onde se contextualize o abuso sexual por meio de situações reais em escolas, delegacias, nas famílias e na comunidade, com interveniência das redes de apoio social; enfatiza-se igualmente a necessidade de formação mais humanizada dos profissionais de apoio social.

Família à mesa: com a palavra, mães e filhos

Introdução O comportamento alimentar do indivíduo resulta de decisões, conscientes ou não, relacionadas à cultura alimentar de sua região, à tradição alimentar de seu convívio social e às transformações decorridas do acesso à informação científica e popular. Objetivo Analisar a alimentação de adolescentes, residentes na região Norte do município de São José dos Campos (SP), na perspectiva da família. Materiais e métodos Estudo qualitativo, realizado em 2014 e 2015, com mães e adolescentes do 6o ano do ensino fundamental municipal. Dados sobre percepção, conhecimento, comportamento e barreiras para uma alimentação saudável dos adolescentes foram produzidos por meio de grupos focais. Entrevistas semiestruturadas foram realizadas com as mães dos estudantes para investigar aspectos sobre alimentação da família e relação com formação de práticas e comportamentos alimentares dos filhos. Os discursos foram registrados com auxílio de um gravador de voz, e transcritos posteriormente para análise, por meio da técnica de análise de conteúdo temática. Foram realizados 4 grupos focais com 6 estudantes cada, e 15 entrevistas com mães. Resultados e discussão Os discursos foram classificados em temas: comportamento alimentar, preferências alimentares e conhecimentos e crenças. Verificou-se que a alimentação dos adolescentes pode ser influenciada pelo comportamento alimentar dos pais. Mães e adolescentes percebemse conscientes e julgam sobre o que comem. Entretanto, a fala muitas vezes está distante da prática, seja por negligencia, dificuldade em perceber possibilidades de ação, questões emotivas ou pelo hábito formado, considerado a maior dificuldade para mudanças relacionadas à alimentação, tanto das mães quanto adolescentes. Possibilidades de soluções, quando referidas, eram por vezes permeadas por conflitos, causando insatisfação em mães e filhos. O contexto no qual esses individuos estão inseridos deve ser considerado, como o bairro e a escola, os quais contribuem para a determinação das escolhas alimentares. Conclusão a família exerce importante papel no comportamento alimentar de adolescentes, entretanto, a compreensão sobre o tema deve considerar, sobretudo, a cultura e o meio social em que a dinâmica familiar está contextualizada.

Psychological Perspectives on the Civil Liberties of Transgendered Inmates in a Prison Setting

As society becomes increasingly less binary, and moves towards a more spectrum based approach to mental illness, medical illness, and personality, it becomes necessary to address this shift within formerly rigid institutions. This paper explores this shift as it is occurring within correctional settings around the United States concerning the medical care, housing, and safety of transgendered inmates. As there is no legal standard for the housing or access to gender-affirming medical care (i.e., hormone therapy, sexual reassignment surgery), these issues are addressed on an institutional level, with very little consistency throughout the country. Currently, most institutions follow a genitalia-based system of classification. Within the system, core beliefs are held, some adaptive and some no longer adaptive, that drive the system's behavior and outcomes. With regard to transgendered inmates, several underlying beliefs within the system serve to maintain the status quo; however, the most basic underpinning is the system's reliance on a binary gender system. As views of humane treatment of the incarcerated expand and modernize, the role of mental health within corrections has also expanded. Psychologists, social workers, counselors, and psychiatrists are found in almost all correctional facilities, and have become a voice of advocacy for an often underserved population.

Examining the impact of pre-k and kindergarten classroom alignment on school readiness outcomes

This study explored children’s experiences of instructional alignment from prekindergarten to kindergarten and analyzed the impact of those alignment experiences on children’s school readiness outcomes. The study answered the following overarching research question: Does the alignment of children’s learning experiences between prekindergarten and kindergarten impact school readiness outcomes? Three sub-questions drove the research design: (1) How do children’s prekindergarten and kindergarten learning experiences align; (2) To what extent does the alignment of early learning experiences predict children’s school readiness outcomes; and (3) Does the quality of prekindergarten classroom teacher interactions moderate the impact of any PK-K alignment effects? Using cluster analysis and hierarchical linear modeling (HLM) to analyze data from over 1,300 children in the 2009 Head Start Family and Child Experiences Survey (FACES), the study found that children have distinct and definable experiences of PK-K alignment. Results also indicated a disparity in children’s PK-K alignment experiences, with Hispanic/Latino children more likely to attend Head Start programs with poor systems transition practices followed by kindergartens with poor classroom structures. The study found that growth in the use of instructional activity centers from prekindergarten to kindergarten is predictive of better literacy and math outcomes. Findings further suggested that boys, minority students, and children from lower income households are predicted to score lower than girls, white classmates, and higher-income peers across school readiness measures. Findings support the need for equitable transition and alignment practices for children from all racial and ethnic groups. They also argue for an increase in child-directed activity centers in kindergarten. With one exception, the current findings did not support the hypothesis that prekindergarten teacher quality is a moderator of alignment effects on children’s school readiness outcomes. The study presents suggestions for further research.

Factors Affecting Women Leaders in the Denver Group of the Colorado Mountain Club

A study of women leaders in the Colorado Mountain Club (CMC) demonstrated that this group perceived pace as an impediment to leadership growth. This study is an exploratory-quantitative inquiry that assessed the views of 20 of the active women hike leaders in the Denver group. The author designed a survey of factors women hike leaders would rate according to their CMC experiences. Although there are more women members of the Denver group, women leaders comprise only 30% of the leadership group The results from this first ever survey of CMC's women leaders provides a knowledge base for CMC and other interested parties. This study clearly demonstrated the need for more research into its topic of women in leadership positions.

A metasynthesis of qualitative studies regarding opinions and perceptions about barriers and determinants of health services’ accessibility in economic migrants

Background: Access to health services is an important health determinant. New research in health equity is required, especially amongst economic migrants from developing countries. Studies conducted on the use of health services by migrant populations highlight existing gaps in understanding which factors affect access to these services from a qualitative perspective. We aim to describe the views of the migrants regarding barriers and determinants of access to health services in the international literature (1997–2011). Methods: A systematic review was conducted for Qualitative research papers (English/Spanish) published in 13 electronic databases. A selection of articles that accomplished the inclusion criteria and a quality evaluation of the studies were carried out. The findings of the selected studies were synthesised by means of metasynthesis using different analysis categories according to Andersen’s conceptual framework of access and use of health services and by incorporating other emergent categories. Results: We located 3,025 titles, 36 studies achieved the inclusion criteria. After quality evaluation, 28 articles were definitively synthesised. 12 studies (46.2%) were carried out in the U.S and 11 studies (42.3%) dealt with primary care services. The participating population varied depending mainly on type of host country. Barriers were described, such as the lack of communication between health services providers and migrants, due to idiomatic difficulties and cultural differences. Other barriers were linked to the economic system, the health service characteristics and the legislation in each country. This situation has consequences for the lack of health control by migrants and their social vulnerability. Conclusions: Economic migrants faced individual and structural barriers to the health services in host countries, especially those with undocumented situation and those experimented idiomatic difficulties. Strategies to improve the structures of health systems and social policies are needed.

Experiences on using Arduino for laboratory experiments of Automatic Control and Robotics

The free hardware platforms have become very important in engineering education in recent years. Among these platforms, Arduino highlights, characterized by its versatility, popularity and low price. This paper describes the implementation of four laboratory experiments for Automatic Control and Robotics courses at the University of Alicante, which have been developed based on Arduino and other existing equipment. Results were evaluated taking into account the views of students, concluding that the proposed experiments have been attractive to them, and they have acquired the knowledge about hardware configuration and programming that was intended.