Publisher Correction: Three-dimensional mapping of intact ovaries reveals the aging dynamics of the ovarian reserve.
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Published Erratum↗
Biomedical subjects
James Sharpe
Publications and source records attributed to James Sharpe.
Tomographic molecular imaging and 3D quantification within adult mouse organs.
A convenient technology to quantify three-dimensional (3D) morphological features would have widespread applications in biomedical research. Based on combined improvements in sample preparation, tomographic imaging and computational processing, we present a procedure for high-resolution 3D quantification of structures within intact adult mouse organs. Using the nonobese diabetic (NOD) mouse model, we demonstrate a correlation between total islet beta-cell volume and the onset of type-1 diabetes.
Animals↗
Spleen versus pancreas: strict control of organ interrelationship revealed by analyses of Bapx1-/- mice.
During early stages of pancreatic development, the mesenchyme that contributes to the spleen overlies the dorsal pancreatic endoderm. Here, we show that interactions between splenic mesenchyme and pancreas proceed via a highly orchestrated morphogenetic program. Disruption of morphogenesis, as occurs in the Bapx1(Nkx3.2)(-/-) embryo, results in transformation of these tissues into well-organized, ectopic gut-like structures. Bapx1 plays a crucial organizing role effecting position and separation of the spleen and pancreas to prevent this metaplastic transformation. Similar transformations occur in organ cultures employing wild-type pancreatic endoderm and spleen mesenchyme, revealing the developmental plasticity of the pancreas and that precise spatial and temporal control of tissue interactions are required for development of both organs.
Animals↗
Visualizing plant development and gene expression in three dimensions using optical projection tomography.
A deeper understanding of the mechanisms that underlie plant growth and development requires quantitative data on three-dimensional (3D) morphology and gene activity at a variety of stages and scales. To address this, we have explored the use of optical projection tomography (OPT) as a method for capturing 3D data from plant specimens. We show that OPT can be conveniently applied to a wide variety of plant material at a range of scales, including seedlings, leaves, flowers, roots, seeds, embryos, and meristems. At the highest resolution, large individual cells can be seen in the context of the surrounding plant structure. For naturally semitransparent structures, such as roots, live 3D imaging using OPT is also possible. 3D domains of gene expression can be visualized using either marker genes, such as beta-glucuronidase, or more directly by whole-mount in situ hybridization. We also describe tools and software that allow the 3D data to be readily quantified and visualized interactively in different ways.
Flowers↗
Correction of artefacts in optical projection tomography.
A new imaging technique called optical projection tomography (OPT), essentially an optical version of x-ray computed tomography (CT), provides molecular specificity, cellular resolution and larger specimen coverage ( approximately 1 cubic centimetre) than was previously possible with other imaging techniques. It is ideally suited to gene expression studies in small animals. Reconstructed OPT images demonstrate several artefacts which reduce the overall image quality. In this paper, we describe methods to prevent smear artefacts due to illumination intensity fluctuation, ring artefacts due to CCD pixel sensitivity variation and a new 'detector edge' artefact caused by non-zero background signal. We also present an automated method to align the position of the rotational axis during image reconstruction. Finally, we propose a method to eliminate bowl artefacts due to projection truncation using a lower resolution OPT scan of the same specimen. This solution also provides OPT with the ability to obtain a high-resolution reconstruction from a region of interest of a specimen that is larger than the field of view. Implementation of these corrections and modifications increases the accuracy of the OPT imaging technique and extends its capabilities to obtain higher resolution data from within a whole specimen.
Algorithms↗
3D modelling, gene expression mapping and post-mapping image analysis in the developing human brain.
As human brain development proceeds, there are complex changes in size and shape, most notably in the developing forebrain. Molecular technologies enable us to characterise the gene expression patterns that underlie these changes. To interpret these patterns the location of expression must be identified and, often, gene expression patterns compared for several genes or across several developmental stages. To facilitate interpretation we have generated a set of three-dimensional models using a recently developed technique, optical projection tomography. The models act as a framework onto which gene expression patterns are mapped and anatomical domains identified using custom-designed software, MAPaint. Here, we demonstrate their use to compare forebrain development at two embryonic stages (Carnegie stages 18 and 21; 44 and 52 days post conception, respectively) and as a means of recording, storing and visualising gene expression data for three example genes EMX1, EMX2 and OTX2. Anatomical domains were also mapped to the models and the comparison of gene expression and anatomical data is demonstrated at Carnegie stage 21. The three-dimensional models and sophisticated software facilitate the analysis and visualisation of morphological changes and gene expression patterns during early brain development and can be applied to the development of other complex structures.
Brain↗
3 dimensional modelling of early human brain development using optical projection tomography.
BACKGROUND: As development proceeds the human embryo attains an ever more complex three dimensional (3D) structure. Analyzing the gene expression patterns that underlie these changes and interpreting their significance depends on identifying the anatomical structures to which they map and following these patterns in developing 3D structures over time. The difficulty of this task greatly increases as more gene expression patterns are added, particularly in organs with complex 3D structures such as the brain. Optical Projection Tomography (OPT) is a new technology which has been developed for rapidly generating digital 3D models of intact specimens. We have assessed the resolution of unstained neuronal structures within a Carnegie Stage (CS)17 OPT model and tested its use as a framework onto which anatomical structures can be defined and gene expression data mapped. RESULTS: Resolution of the OPT models was assessed by comparison of digital sections with physical sections stained, either with haematoxylin and eosin (H&E) or by immunocytochemistry for GAP43 or PAX6, to identify specific anatomical features. Despite the 3D models being of unstained tissue, peripheral nervous system structures from the trigeminal ganglion (approximately 300 microm by approximately 150 microm) to the rootlets of cranial nerve XII (approximately 20 microm in diameter) were clearly identifiable, as were structures in the developing neural tube such as the zona limitans intrathalamica (core is approximately 30 microm thick). Fourteen anatomical domains have been identified and visualised within the CS17 model. Two 3D gene expression domains, known to be defined by Pax6 expression in the mouse, were clearly visible when PAX6 data from 2D sections were mapped to the CS17 model. The feasibility of applying the OPT technology to all stages from CS12 to CS23, which encompasses the major period of organogenesis for the human developing central nervous system, was successfully demonstrated. CONCLUSION: In the CS17 model considerable detail is visible within the developing nervous system at a minimum resolution of approximately 20 microm and 3D anatomical and gene expression domains can be defined and visualised successfully. The OPT models and accompanying technologies for manipulating them provide a powerful approach to visualising and analysing gene expression and morphology during early human brain development.
Brain↗
Optical projection tomography.
Optical projection tomography is a new approach for three-dimensional (3-D) imaging of small biological specimens. It fills an imaging gap between MRI and confocal microscopy, being most suited to specimens that are from 1 to 10 mm across. The tomographic principles of optical projection tomography (OPT) are explained, its most important applications in biomedical research explored, and comparisons drawn of its pros and cons compared to a number of alternative imaging technologies.
Animals↗
Optical projection tomography as a new tool for studying embryo anatomy.
Optical projection tomography (OPT) is a new technique for three-dimensional (3D) imaging of small biological tissues. It is particularly useful for reconstructing vertebrate embryos and for examining the 3D anatomy of developing organs. The advantages of this technique over previous methods will be explained: in particular, its ability to image at a higher resolution than magnetic resonance imaging (MRI), while at the same time being able to image specimens much larger than those possible using confocal laser-scanning microscopy. Being an optical technique, OPT is also able to take advantage of the many coloured and fluorescent dyes which have been developed for tissue-specific or gene-specific staining. This becomes particularly important for the visualization of the 3D shapes of specific organs and tissues as it allows the computer to automatically determine the outline of the desired structure.
Animals↗
EMAP and EMAGE: a framework for understanding spatially organized data.
The Edinburgh MouseAtlas Project (EMAP) is a time-series of mouse-embryo volumetric models. The models provide a context-free spatial framework onto which structural interpretations and experimental data can be mapped. This enables collation, comparison, and query of complex spatial patterns with respect to each other and with respect to known or hypothesized structure. The atlas also includes a time-dependent anatomical ontology and mapping between the ontology and the spatial models in the form of delineated anatomical regions or tissues. The models provide a natural, graphical context for browsing and visualizing complex data. The Edinburgh Mouse Atlas Gene-Expression Database (EMAGE) is one of the first applications of the EMAP framework and provides a spatially mapped gene-expression database with associated tools for data mapping, submission, and query. In this article, we describe the underlying principles of the Atlas and the gene-expression database, and provide a practical introduction to the use of the EMAP and EMAGE tools, including use of new techniques for whole body gene-expression data capture and mapping.
Animals↗
Optical projection tomography as a tool for 3D microscopy and gene expression studies.
Current techniques for three-dimensional (3D) optical microscopy (deconvolution, confocal microscopy, and optical coherence tomography) generate 3D data by "optically sectioning" the specimen. This places severe constraints on the maximum thickness of a specimen that can be imaged. We have developed a microscopy technique that uses optical projection tomography (OPT) to produce high-resolution 3D images of both fluorescent and nonfluorescent biological specimens with a thickness of up to 15 millimeters. OPT microscopy allows the rapid mapping of the tissue distribution of RNA and protein expression in intact embryos or organ systems and can therefore be instrumental in studies of developmental biology or gene function.
Anatomy, Cross-Sectional↗