Biology subjects

Amack, J. D.

Publications and source records attributed to Amack, J. D..

7 recordsLinked to original sources

bioRxiv2025

Size-Modulated Mesoderm-Endoderm Divergence and Myocardial Cavitation in Micropatterned Cardioids

The human heart, originating from the splanchnic mesoderm, is the first functional organ to develop, co-evolving with the foregut endoderm through reciprocal signaling. Previously, cardioid models offered new insights on cardiovascular cell lineages and tissue morphogenesis during heart development, while mesoderm-endoderm crosstalk remain incompletely understood. Here, we integrated micropatterned cardioids, CRISPR-engineered reporter hiPSCs, deep-tissue imaging, and single-cell RNA sequencing (scRNA-seq) to explore synergistic mesoderm-endoderm co-development. scRNA-seq with PHATE trajectory mapping reconstructed lineage bifurcations of mesoderm-heart and endoderm-foregut lineages, identifying key cell types in cardiac and hepatic development. Ligand-receptor interaction analysis highlighted mesodermal cells enriched in non-canonical WNT, NRG, and TGF-{beta} signaling, while endodermal cells exhibited VEGF and Hedgehog activity. We found that micropattern sizes influenced cellular composition, cardioid cavitation, contractile functions, and mesoderm-endoderm signaling crosstalk. The cardioids generated from 600 {micro}m diameter circle patterns showed larger cavity formation resembling early heart chamber formation. Our findings establish micropatterned cardioids as a model for mesoderm-endoderm co-development, enhancing our understanding of heart-foregut synergy during early embryogenesis.

bioengineering↗

bioRxiv2025

Fhod3 in zebrafish supports myofibril stability during growth of embryonic skeletal muscle

BackgroundActin filament organization in cardiomyocytes critically depends on the formin Fhod3, but a role for Fhod3 in skeletal muscle development has not yet been described. ResultsWe demonstrate here that in zebrafish mutated for one of two fhod3 paralog genes, fhod3a, skeletal muscle of the trunk appears normal through 2 days post-fertilization, but afterward exhibits myofibril damage, including gaps between myofibrils and myofibril fragmentation. Despite the apparent progressive nature of myofibril damage, fhod3a mutant embryos differ from muscular dystrophy models in that damage is exacerbated by inhibition of muscle activity, and fhod3a mutants show no evidence of sarcolemma disruption. Rather, our results suggest myofibril damage coincides with growth of the skeletal muscle fiber contractile apparatus. We find that neither the second fhod3 paralog, fhod3b, nor the related fhod1 contribute to embryonic skeletal muscle development, but simultaneous mutation of fhod3a and fhod3b was associated with pericardial edema suggestive of cardiac dysfunction. ConclusionsTaken together, these results indicate fhod3-encoded formins are dispensable for initial myofibril assembly in skeletal muscle, but promote myofibril stability during muscle fiber growth. This is the first demonstration that Fhod3 contributes to skeletal muscle development in a vertebrate.

developmental biology↗

bioRxiv2024

Dynamical forces drive organ morphology changes during embryonic development

Cells, tissues, and organs must change shape in precise ways during embryonic development to execute their functions. Multiple mechanisms including biochemical signaling pathways and biophysical forces help drive these morphology changes, but it has been difficult to tease apart their contributions, especially from tissue-scale dynamic forces that are typically ignored. We use a combination of mathematical models and in vivo experiments to study a simple organ in the zebrafish embryo called Kupffers vesicle. Modeling indicates that dynamic forces generated by tissue movements in the embryo produce shape changes in Kupffers vesicle that are observed during development. Laser ablations in the zebrafish embryo that alter these forces result in altered organ shapes matching model predictions. These results demonstrate that dynamic forces sculpt cell and organ shape during embryo development. Significance StatementWe aim to understand the mechanisms that control precise cell and tissue shape changes required for organ function. Many studies focused on cell shapes have ignored the role of dynamic forces self-generated by slow tissue flows, but recent work showing tissues are near a jamming transition with diverging relaxation timescales suggests slow motion could give rise to large forces. Through a combination of mathematical modeling, imaging, and mechanical perturbations to in vivo experiments, our work demonstrates that tissue-scale dynamic forces are sculpting the shape of an epithelial organ in the zebrafish embryo called Kupffers vesicle (KV). Because there are many processes during development that occur at similarly slow rates, this suggests that self-generated dynamic forces should be investigated more broadly.

developmental biology↗

bioRxiv2021

3D viscoelastic drag forces drive changes to cell shapes during organogenesis in the zebrafish embryo

The left-right organizer in zebrafish embryos, Kupffers Vesicle (KV), is a simple organ that undergoes programmed asymmetric cell shape changes that are necessary to establish the left-right axis of the embryo. We use simulations and experiments to investigate whether 3D mechanical drag forces generated by the posteriorly-directed motion of the KV through the tailbud tissue are sufficient to drive such shape changes. We develop a fully 3D vertex-like (Voronoi) model for the tissue architecture, and demonstrate that the tissue can generate drag forces and drive cell shape changes. Furthermore, we find that tailbud tissue presents a shear-thinning, viscoelastic behavior consistent with those observed in published experiments. We then perform live imaging experiments and particle image velocimetry analysis to quantify the precise tissue velocity gradients around KV as a function of developmental time. We observe robust velocity gradients around the KV, indicating that mechanical drag forces must be exerted on the KV by the tailbud tissue. We demonstrate that experimentally observed velocity fields are consistent with the viscoelastic response seen in simulations. This work also suggests that 3D viscoelastic drag forces could be a generic mechanism for cell shape change in other biological processes. HighlightsO_LInew physics-based simulation method allows study of dynamic tissue structures in 3D C_LIO_LImovement of an organ through tissue generates viscoelastic drag forces on the organ C_LIO_LIthese drag forces can generate precisely the cell shape changes seen in experiment C_LIO_LIPIV analysis of experimental data matches simulations and probes tissue mechanics C_LI Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

developmental biology↗

bioRxiv2020

PLK1- and PLK4-mediated asymmetric mitotic centrosome size and positioning in the early zebrafish embryo.

Factors that regulate mitotic spindle positioning have been elucidated in vitro, however it remains unclear how a spindle is placed within the confines of extremely large cells. Our studies identified a uniquely large centrosome structure in the early zebrafish embryo (246.44{+/-}11.93m2 mitotic centrosome in a 126.86{+/-}0.35m diameter cell), whereas C. elegans centrosomes are notably smaller (6.75{+/-}0.28m2 mitotic centrosome in a 55.83{+/-}1.04m diameter cell). During early embryonic cell divisions, cell size changes rapidly in C. elegans and zebrafish embryos. Notably, mitotic centrosome area scales closely with changing cell size compared to changes in spindle length for both organisms. One interesting difference between the two is that mitotic centrosomes are asymmetric in size across embryonic zebrafish spindles, with the larger mitotic centrosome being 2.14{+/-}0.13-fold larger in size than the smaller. The largest mitotic centrosome is placed towards the embryo center in a Polo-Like Kinase (PLK) 1 and PLK4 dependent manner 87.14{+/-}4.16% of the time. We propose a model in which uniquely large centrosomes direct spindle placement within the disproportionately large zebrafish embryo cells to orchestrate cell divisions during early embryogenesis.

cell biology↗

bioRxiv2018

Tissue flow induces cell shape changes during organogenesis

In embryonic development, cell shape changes are essential for building functional organs, but in many cases the mechanisms that precisely regulate these changes remain unknown. We propose that fluid-like drag forces generated by the motion of an organ through surrounding tissue could generate changes to its structure that are important for its function. To test this hypothesis, we study the zebrafish left-right organizer, Kupffers vesicle (KV), using experiments and mathematical modeling. During development, monociliated cells that comprise the KV undergo region-specific shape changes along the anterior-posterior axis that are critical for KV function: anterior cells become long and thin, while posterior cells become short and squat. Here, we develop a mathematical vertex-like model for cell shapes, which incorporates both tissue rheology and cell motility, and constrain the model parameters using previously published rheological data for the zebrafish tailbud [Serwane et al.] as well as our own measurements of the KV speed. We find that drag forces due to dynamics of cells surrounding the KV could be sufficient to drive KV cell shape changes during KV development. More broadly, these results suggest that cell shape changes could be driven by dynamic forces not typically considered in models or experiments.

biophysics↗

bioRxiv2017

Asymmetric cell volume changes regulate epithelial morphogenesis in zebrafish Kupffers vesicle

How epithelial cell behaviors are coordinately regulated to sculpt tissue architecture is a fundamental question in biology. Kupffer's vesicle (KV), a transient organ with a fluid - filled lumen, provides a simple system to investigate the interplay between intrinsic cellular mechanisms and external forces during epithelial morphogenesis. Using 3 - dimensional (3D) analyses of single cells we identify asymmetric cell volume changes along the anteroposterior axis of KV that coincide with asymmetric cell shape changes. Blocking ion flux prevents these cell volume changes and cell shape changes. Vertex simulations suggest cell shape changes do not depend on lumen expansion. Consistent with this prediction, asymmetric changes in KV cell volume and shape occur normally when KV lumen growth fails due to leaky cell adhesions. These results indicate ion flux mediates asymmetric cell volume changes that contribute to asymmetric cell shape changes in KV, and that these changes in epithelial morphology are separable from lumen - generated forces.

cell biology↗