Estimating Cell Count and Distribution in Labeled Histological Samples Using Incremental Cell Search

Cell proliferation is critical to the outgrowth of biological structures including the face and limbs. This cellular process has traditionally been studied via sequential histological sampling of these tissues. The length and tedium of traditional sampling is a major impediment to analyzing the large datasets required to accurately model cellular processes. Computerized cell localization and quantification is critical for high-throughput morphometric analysis of developing embryonic tissues. We have developed the Incremental Cell Search (ICS), a novel software tool that expedites the analysis of relationships between morphological outgrowth and cell proliferation in embryonic tissues. Based on an estimated average cell size and stain color, ICS rapidly indicates the approximate location and amount of cells in histological images of labeled embryonic tissue and provides estimates of cell counts in regions with saturated fluorescence and blurred cell boundaries. This capacity opens the door to high-throughput 3D and 4D quantitative analyses of developmental patterns

Estimating Cell Count and Distribution in Labeled Histological Samples Using Incremental Cell Search

Cell proliferation is critical to the outgrowth of biological structures including the face and limbs. This cellular process has traditionally been studied via sequential histological sampling of these tissues. The length and tedium of traditional sampling is a major impediment to analyzing the large datasets required to accurately model cellular processes. Computerized cell localization and quantification is critical for high-throughput morphometric analysis of developing embryonic tissues. We have developed the Incremental Cell Search (ICS), a novel software tool that expedites the analysis of relationships between morphological outgrowth and cell proliferation in embryonic tissues. Based on an estimated average cell size and stain color, ICS rapidly indicates the approximate location and amount of cells in histological images of labeled embryonic tissue and provides estimates of cell counts in regions with saturated fluorescence and blurred cell boundaries. This capacity opens the door to high-throughput 3D and 4D quantitative analyses of developmental patterns

Speciation through the lens of biomechanics: locomotion, prey capture and reproductive isolation

Speciation is a multifaceted process that involves numerous aspects of the biological sciences and occurs for multiple reasons. Ecology plays a major role, including both abiotic and biotic factors. Whether populations experience similar or divergent ecological environments, they often adapt to local conditions through divergence in biomechanical traits. We investigate the role of biomechanics in speciation using fish predator–prey interactions, a primary driver of fitness for both predators and prey. We highlight specific groups of fishes, or specific species, that have been particularly valuable for understanding these dynamic interactions and offer the best opportunities for future studies that link genetic architecture to biomechanics and reproductive isolation (RI). In addition to emphasizing the key biomechanical techniques that will be instrumental, we also propose that the movement towards linking biomechanics and speciation will include (i) establishing the genetic basis of biomechanical traits, (ii) testing whether similar and divergent selection lead to biomechanical divergence, and (iii) testing whether/how biomechanical traits affect RI. Future investigations that examine speciation through the lens of biomechanics will propel our understanding of this key process

Micro-computed tomography-based phenotypic approaches in embryology: procedural artifacts on assessments of embryonic craniofacial growth and development

<p>Abstract</p> <p>Background</p> <p>Growing demand for three dimensional (3D) digital images of embryos for purposes of phenotypic assessment drives implementation of new histological and imaging techniques. Among these micro-computed tomography (μCT) has recently been utilized as an effective and practical method for generating images at resolutions permitting 3D quantitative analysis of gross morphological attributes of developing tissues and organs in embryonic mice. However, histological processing in preparation for μCT scanning induces changes in organ size and shape. Establishing normative expectations for experimentally induced changes in size and shape will be an important feature of 3D μCT-based phenotypic assessments, especially if quantifying differences in the values of those parameters between comparison sets of developing embryos is a primary aim. Toward that end, we assessed the nature and degree of morphological artifacts attending μCT scanning following use of common fixatives, using a two dimensional (2D) landmark geometric morphometric approach to track the accumulation of distortions affecting the embryonic head from the native, uterine state through to fixation and subsequent scanning.</p> <p>Results</p> <p>Bouin's fixation reduced average centroid sizes of embryonic mouse crania by approximately 30% and substantially altered the morphometric shape, as measured by the shift in Procrustes distance, from the unfixed state, after the data were normalized for naturally occurring shape variation. Subsequent μCT scanning produced negligible changes in size but did appear to reduce or even reverse fixation-induced random shape changes. Mixtures of paraformaldehyde + glutaraldehyde reduced average centroid sizes by 2-3%. Changes in craniofacial shape progressively increased post-fixation.</p> <p>Conclusions</p> <p>The degree to which artifacts are introduced in the generation of random craniofacial shape variation relates to the degree of specimen dehydration during the initial fixation. Fixation methods that better maintain original craniofacial dimensions at reduced levels of dehydration and tissue shrinkage lead to the progressive accumulation of random shape variation during handling and data acquisition. In general, to the degree that embryonic organ size and shape factor into μCT-based phenotypic assessments, procedurally induced artifacts associated with fixation and scanning will influence results. Experimental designs will need to address these significant effects, either by employing alternative methods that minimize artifacts in the region of focus or in the interpretation of statistical patterns.</p

Conditional Creation and Rescue of Nipbl-Deficiency in Mice Reveals Multiple Determinants of Risk for Congenital Heart Defects

Elucidating the causes of congenital heart defects is made difficult by the complex morphogenesis of the mammalian heart, which takes place early in development, involves contributions from multiple germ layers, and is controlled by many genes. Here, we use a conditional/invertible genetic strategy to identify the cell lineage(s) responsible for the development of heart defects in a Nipbl-deficient mouse model of Cornelia de Lange Syndrome, in which global yet subtle transcriptional dysregulation leads to development of atrial septal defects (ASDs) at high frequency. Using an approach that allows for recombinase-mediated creation or rescue of Nipbl deficiency in different lineages, we uncover complex interactions between the cardiac mesoderm, endoderm, and the rest of the embryo, whereby the risk conferred by genetic abnormality in any one lineage is modified, in a surprisingly non-additive way, by the status of others. We argue that these results are best understood in the context of a model in which the risk of heart defects is associated with the adequacy of early progenitor cell populations relative to the sizes of the structures they must eventually form

Zoological Endeavors Inspired by A. Richard Palmer: Introduction, Biography, and Bibliography

The Professor stood in front of the lecture room, basking in the applause of his colleagues and former students, smiles all around. Underneath his dark jacket was a t-shirt with pictures of curious and puzzling Ediacaran organisms; in his hand, a beer mug engraved with an acorn barnacle reaching out with its feeding legs and penis (Fig. 1), with a quote from the letter written by Charles Darwin to Charles Lyell on 9 August 1838:The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Freshwater influence is associated with differences in bone mineral density and armour configuration in threespine stickleback (Gasterosteus aculeatus)

Threespine stickleback (Gasterosteus aculeatus Linnaeus, 1758) exhibit a well-documented reduction in plate number associated with adaptation to freshwater environments. We tested the hypothesis that changes in plate number are accompanied by changes in plate bone mineral density and plate shape, reflecting the presence of a complex plate “armour” phenotype and a complex adaptive response to different selective pressures in changing habitats. We used traditional and novel morphometric techniques to characterize armour traits from stickleback occupying three marine habitats and one tidally influenced freshwater stream in southwestern British Columbia. Stickleback inhabiting marine environments share a conserved plate phenotype that includes a full complement of highly mineralized plates that exhibit a characteristic density profile along the plate. Stickleback inhabiting tidally influenced fresh water display an average reduction in plate number along with increased variation in number and reduced total mineralization despite maintenance of a marine-like density profile. Further, we found that although mineralization and armour shape are correlated with size, after accounting for size variation in both traits remains attributable to habitat. Our results hint at an important role for development in structuring phenotypic variation during the process of adaptive change in stickleback