Quantitative Geometric Modeling of Blood Cells from X-ray Histotomograms of Whole Zebrafish Larvae

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Abstract

Tissue phenotyping is foundational to understanding and assessing the cellular aspects of disease in organismal context and an important adjunct to molecular studies in the dissection of gene function, chemical effects, and disease. As a first step toward computational tissue phenotyping, we explore the potential of cellular phenotyping from 3-Dimensional (3D), 0.74 µm isotropic voxel resolution, whole zebrafish larval images derived from X-ray histotomography, a form of micro-CT customized for histopathology. As proof of principle towards computational tissue phenotyping of cells, we created a semi-automated mechanism for the segmentation of blood cells in the vascular spaces of zebrafish larvae, followed by modeling and extraction of quantitative geometric parameters. Manually segmented cells were used to train a random forest classifier for blood cells, enabling the use of a generalized cellular segmentation algorithm for the accurate segmentation of blood cells. These models were used to create an automated data segmentation and analysis pipeline to guide the steps in a 3D workflow including blood cell region prediction, cell boundary extraction, and statistical characterization of 3D geometric and cytological features. We were able to distinguish blood cells at two stages in development (4- and 5-days-post-fertilization) and wild-type vs.polA2 huli hutu(hht) mutants. The application of geometric modeling across cell types to and across organisms and sample types may comprise a valuable foundation for computational phenotyping that is more open, informative, rapid, objective, and reproducible.

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