Publications
77 results found
Yaganoglu S, Kalyviotis K, Vagena-Pantoula C, et al., 2023, Author Correction: Highly specific and non-invasive imaging of Piezo1-dependent activity across scales using GenEPi, Nature Communications, Vol: 14, ISSN: 2041-1723
Steib E, Vagena-Pantoula C, Vermot J, 2023, TissUExM protocol for ultrastructure expansion microscopy of zebrafish larvae and mouse embryos, STAR Protocols, Vol: 4, ISSN: 2666-1667
Expansion microscopy of millimeter-large mechanically heterogeneous tissues, such as whole vertebrate embryos, has been limited, particularly when combined with post-expansion immunofluorescence. Here, we present a protocol to perform ultrastructure expansion microscopy of whole vertebrate embryos, optimized to perform post-expansion labeling. We describe steps for embedding and denaturing zebrafish larvae or mouse embryos. We then detail procedures for hydrogel handling and mounting. This protocol is particularly well suited for super-resolution imaging of macromolecular protein complexes in situ but does not preserve lipids. For complete details on the use and execution of this protocol, please refer to Steib et al.1.
Steib E, Tetley R, Laine RF, et al., 2022, TissUExM enables quantitative ultrastructural analysis in whole vertebrate embryos by expansion microscopy, Cell Reports: Methods, Vol: 2, ISSN: 2667-2375
Super-resolution microscopy reveals the molecular organization of biological structures down to the nanoscale. While it allows the study of protein complexes in single cells, small organisms, or thin tissue sections, there is currently no versatile approach for ultrastructural analysis compatible with whole vertebrate embryos. Here, we present tissue ultrastructure expansion microscopy (TissUExM), a method to expand millimeter-scale and mechanically heterogeneous whole embryonic tissues, including Drosophila wing discs, whole zebrafish, and mouse embryos. TissUExM is designed for the observation of endogenous proteins. It permits quantitative characterization of protein complexes in various organelles at super-resolution in a range of ∼3 mm-sized tissues using conventional microscopes. We demonstrate its strength by investigating tissue-specific ciliary architecture heterogeneity and ultrastructural defects observed upon ciliary protein overexpression. Overall, TissUExM is ideal for performing ultrastructural studies and molecular mapping in situ in whole embryos.
Vignes H, Vagena-Pantoula C, Vermot J, 2022, Mechanical control of tissue shape: Cell-extrinsic and -intrinsic mechanisms join forces to regulate morphogenesis, SEMINARS IN CELL & DEVELOPMENTAL BIOLOGY, Vol: 130, Pages: 45-55, ISSN: 1084-9521
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- Citations: 4
Cairelli AG, Chow RW-Y, Vermot J, et al., 2022, Fluid mechanics of the zebrafish embryonic heart trabeculation, PLoS Computational Biology, Vol: 18, ISSN: 1553-734X
Embryonic heart development is a mechanosensitive process, where specific fluid forces are needed for the correct development, and abnormal mechanical stimuli can lead to malformations. It is thus important to understand the nature of embryonic heart fluid forces. However, the fluid dynamical behaviour close to the embryonic endocardial surface is very sensitive to the geometry and motion dynamics of fine-scale cardiac trabecular surface structures. Here, we conducted image-based computational fluid dynamics (CFD) simulations to quantify the fluid mechanics associated with the zebrafish embryonic heart trabeculae. To capture trabecular geometric and motion details, we used a fish line that expresses fluorescence at the endocardial cell membrane, and high resolution 3D confocal microscopy. Our endocardial wall shear stress (WSS) results were found to exceed those reported in existing literature, which were estimated using myocardial rather than endocardial boundaries. By conducting simulations of single intra-trabecular spaces under varied scenarios, where the translational or deformational motions (caused by contraction) were removed, we found that a squeeze flow effect was responsible for most of the WSS magnitude in the intra-trabecular spaces, rather than the shear interaction with the flow in the main ventricular chamber. We found that trabecular structures were responsible for the high spatial variability of the magnitude and oscillatory nature of WSS, and for reducing the endocardial deformational burden. We further found cells attached to the endocardium within the intra-trabecular spaces, which were likely embryonic hemogenic cells, whose presence increased endocardial WSS. Overall, our results suggested that a complex multi-component consideration of both anatomic features and motion dynamics were needed to quantify the trabeculated embryonic heart fluid mechanics.
Vignes H, Vagena-Pantoula C, Prakash M, et al., 2022, Extracellular mechanical forces drive endocardial cell volume decrease during zebrafish cardiac valve morphogenesis, DEVELOPMENTAL CELL, Vol: 57, Pages: 598-+, ISSN: 1534-5807
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- Citations: 12
Chow RW-Y, Fukui H, Chan WX, et al., 2022, Cardiac forces regulate zebrafish heart valve delamination by modulating Nfat signaling, PLoS Biology, Vol: 20, ISSN: 1544-9173
In the clinic, most cases of congenital heart valve defects are thought to arise through errors that occur after the endothelial–mesenchymal transition (EndoMT) stage of valve development. Although mechanical forces caused by heartbeat are essential modulators of cardiovascular development, their role in these later developmental events is poorly understood. To address this question, we used the zebrafish superior atrioventricular valve (AV) as a model. We found that cellularized cushions of the superior atrioventricular canal (AVC) morph into valve leaflets via mesenchymal–endothelial transition (MEndoT) and tissue sheet delamination. Defects in delamination result in thickened, hyperplastic valves, and reduced heart function. Mechanical, chemical, and genetic perturbation of cardiac forces showed that mechanical stimuli are important regulators of valve delamination. Mechanistically, we show that forces modulate Nfatc activity to control delamination. Together, our results establish the cellular and molecular signature of cardiac valve delamination in vivo and demonstrate the continuous regulatory role of mechanical forces and blood flow during valve formation.
Fukui H, Chow RW-Y, Xie J, et al., 2021, Bioelectric signaling and the control of cardiac cell identity in response to mechanical forces, Science, Vol: 374, Pages: 351-354, ISSN: 0036-8075
Developing cardiovascular systems use mechanical forces to take shape, but how ubiquitous blood flow forces instruct local cardiac cell identity is still unclear. By manipulating mechanical forces in vivo, we show here that shear stress is necessary and sufficient to promote valvulogenesis. We found that valve formation is associated with the activation of an extracellular adenosine triphosphate (ATP)–dependent purinergic receptor pathway, specifically triggering calcium ion (Ca2+) pulses and nuclear factor of activated T cells 1 (Nfatc1) activation. Thus, mechanical forces are converted into discrete bioelectric signals by an ATP-Ca2+-Nfatc1–mechanosensitive pathway to generate positional information and control valve formation.
Peralta M, Lopez LO, Jerabkova K, et al., 2020, Intraflagellar Transport Complex B Proteins Regulate the Hippo Effector Yap1 during Cardiogenesis, CELL REPORTS, Vol: 32, ISSN: 2211-1247
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- Citations: 12
Campinho P, Vilfan A, Vermot J, 2020, Blood flow forces in shaping the vascular system: a focus on endothelial cell behavior, Frontiers in Physiology, Vol: 11, ISSN: 1664-042X
The endothelium is the cell monolayer that lines the interior of the blood vessels separating the vessel lumen where blood circulates, from the surrounding tissues. During embryonic development, endothelial cells (ECs) must ensure that a tight barrier function is maintained whilst dynamically adapting to the growing vascular tree that is being formed and remodeled. Blood circulation generates mechanical forces, such as shear stress and circumferential stretch that are directly acting on the endothelium. ECs actively respond to flow-derived mechanical cues by becoming polarized, migrating and changing neighbors, undergoing shape changes, proliferating or even leaving the tissue and changing identity. It is now accepted that coordinated changes at the single cell level drive fundamental processes governing vascular network morphogenesis such as angiogenic sprouting, network pruning, lumen formation, regulation of vessel caliber and stability or cell fate transitions. Here we summarize the cell biology and mechanics of ECs in response to flow-derived forces, discuss the latest advances made at the single cell level with particular emphasis on in vivo studies and highlight potential implications for vascular pathologies.
Campinho P, Lamperti P, Boselli F, et al., 2020, Blood Flow Limits Endothelial Cell Extrusion in the Zebrafish Dorsal Aorta, CELL REPORTS, Vol: 31, ISSN: 2211-1247
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- Citations: 14
Duchemin A-L, Vignes H, Vermot J, 2019, Mechanically activated piezo channels modulate outflow tract valve development through the Yap1 and Klf2-Notch signaling axis, eLife, Vol: 8, Pages: 1-27, ISSN: 2050-084X
Mechanical forces are well known for modulating heart valve developmental programs. Yet, it is still unclear how genetic programs and mechanosensation interact during heart valve development. Here, we assessed the mechanosensitive pathways involved during zebrafish outflow tract (OFT) valve development in vivo. Our results show that the hippo effector Yap1, Klf2, and the Notch signaling pathway are all essential for OFT valve morphogenesis in response to mechanical forces, albeit active in different cell layers. Furthermore, we show that Piezo and TRP mechanosensitive channels are important factors modulating these pathways. In addition, live reporters reveal that Piezo controls Klf2 and Notch activity in the endothelium and Yap1 localization in the smooth muscle progenitors to coordinate OFT valve morphogenesis. Together, this work identifies a unique morphogenetic program during OFT valve formation and places Piezo as a central modulator of the cell response to forces in this process.
Duchemin A-L, Vignes H, Vermot J, et al., 2019, Mechanotransduction in cardiovascular morphogenesis and tissue engineering, CURRENT OPINION IN GENETICS & DEVELOPMENT, Vol: 57, Pages: 106-116, ISSN: 0959-437X
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- Citations: 25
Ferreira RR, Fukui H, Chow R, et al., 2019, The cilium as a force sensor-myth versus reality, JOURNAL OF CELL SCIENCE, Vol: 132, ISSN: 0021-9533
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- Citations: 49
Andres-Delgado L, Ernst A, Galardi-Castilla M, et al., 2019, Actin dynamics and the Bmp pathway drive apical extrusion of proepicardial cells, DEVELOPMENT, Vol: 146, ISSN: 0950-1991
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- Citations: 12
Galvez-Santisteban M, Chen D, Zhang R, et al., 2019, Hemodynamic-mediated endocardial signaling controls in vivo myocardial reprogramming, ELIFE, Vol: 8, ISSN: 2050-084X
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- Citations: 20
Ferreira RR, Pakula G, Klaeyle L, et al., 2018, Chiral Cilia orientation in the left-right organizer, Cell Reports, Vol: 25, Pages: 2008-2016.e4, ISSN: 2211-1247
Chirality is a property of asymmetry between an object and its mirror image. Most biomolecules and many cell types are chiral. In the left-right organizer (LRO), cilia-driven flows transfer such chirality to the body scale. However, the existence of cellular chirality within tissues remains unknown. Here, we investigate this question in Kupffer’s vesicle (KV), the zebrafish LRO. Quantitative live imaging reveals that cilia populating the KV display asymmetric orientation between the right and left sides, resulting in a chiral structure, which is different from the chiral cilia rotation. This KV chirality establishment is dynamic and depends on planar cell polarity. While its impact on left-right (LR) symmetry breaking remains unclear, we show that this asymmetry does not depend on the LR signaling pathway or flow. This work identifies a different type of tissue asymmetry and sheds light on chirality genesis in developing tissues.
Campinho P, Lamperti P, Boselli F, et al., 2018, Three-dimensional microscopy and image analysis methodology for mapping and quantification of nuclear positions in tissues with approximate cylindrical geometry, PHILOSOPHICAL TRANSACTIONS OF THE ROYAL SOCIETY B-BIOLOGICAL SCIENCES, Vol: 373, ISSN: 0962-8436
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- Citations: 6
Fukui H, Miyazaki T, Chow RW-Y, et al., 2018, Hippo signaling determines the number of venous pole cells that originate from the anterior lateral plate mesoderm in zebrafish, ELIFE, Vol: 7, ISSN: 2050-084X
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- Citations: 16
Chow RW-Y, Lamperti P, Steed E, et al., 2018, Following Endocardial Tissue Movements via Cell Photoconversion in the Zebrafish Embryo, JOVE-JOURNAL OF VISUALIZED EXPERIMENTS, ISSN: 1940-087X
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- Citations: 5
Boselli F, Steed E, Freund JB, et al., 2017, Anisotropic shear stress patterns predict the orientation of convergent tissue movements in the embryonic heart, DEVELOPMENT, Vol: 144, Pages: 4322-4327, ISSN: 0950-1991
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- Citations: 37
Goddard LM, Duchemin A-L, Ramalingan H, et al., 2017, Hemodynamic Forces Sculpt Developing Heart Valves through a KLF2-WNT9B Paracrine Signaling Axis, DEVELOPMENTAL CELL, Vol: 43, Pages: 274-+, ISSN: 1534-5807
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- Citations: 81
Bouchaala R, Anton N, Anton H, et al., 2017, Light-triggered release from dye-loaded fluorescent lipid nanocarriers <i>in vitro</i> and <i>in vivo</i>, COLLOIDS AND SURFACES B-BIOINTERFACES, Vol: 156, Pages: 414-421, ISSN: 0927-7765
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- Citations: 16
Ferreira RR, Vilfan A, Juelicher F, et al., 2017, Physical limits of flow sensing in the left right organizer, ELIFE, Vol: 6, ISSN: 2050-084X
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- Citations: 32
Ferreira RR, Vermot J, 2017, The balancing roles of mechanical forces during left-right patterning and asymmetric morphogenesis, MECHANISMS OF DEVELOPMENT, Vol: 144, Pages: 71-80, ISSN: 0925-4773
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- Citations: 8
Chow RW-Y, Vermot J, 2017, The rise of photoresponsive protein technologies applications in vivo: a spotlight on zebrafish developmental and cell biology., F1000Res, Vol: 6, ISSN: 2046-1402
The zebrafish ( Danio rerio) is a powerful vertebrate model to study cellular and developmental processes in vivo. The optical clarity and their amenability to genetic manipulation make zebrafish a model of choice when it comes to applying optical techniques involving genetically encoded photoresponsive protein technologies. In recent years, a number of fluorescent protein and optogenetic technologies have emerged that allow new ways to visualize, quantify, and perturb developmental dynamics. Here, we explain the principles of these new tools and describe some of their representative applications in zebrafish.
Jacob L, Sawma P, Garnier N, et al., 2016, Inhibition of PlexA1-mediated brain tumor growth and tumor-associated angiogenesis using a transmembrane domain targeting peptide, ONCOTARGET, Vol: 7, Pages: 57851-57865
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- Citations: 23
Steed E, Boselli F, Vermot J, 2016, Hemodynamics driven cardiac valve morphogenesis, BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH, Vol: 1863, Pages: 1760-1766, ISSN: 0167-4889
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- Citations: 41
Steed E, Faggianelli N, Roth S, et al., 2016, klf2a couples mechanotransduction and zebrafish valve morphogenesis through fibronectin synthesis, Nature Communications, Vol: 7, ISSN: 2041-1723
The heartbeat and blood flow signal to endocardial cell progenitors through mechanosensitive proteins that modulate the genetic program controlling heart valve morphogenesis. To date, the mechanism by which mechanical forces coordinate tissue morphogenesis is poorly understood. Here we use high-resolution imaging to uncover the coordinated cell behaviours leading to heart valve formation. We find that heart valves originate from progenitors located in the ventricle and atrium that generate the valve leaflets through a coordinated set of endocardial tissue movements. Gene profiling analyses and live imaging reveal that this reorganization is dependent on extracellular matrix proteins, in particular on the expression of fibronectin1b. We show that blood flow and klf2a, a major endocardial flow-responsive gene, control these cell behaviours and fibronectin1b synthesis. Our results uncover a unique multicellular layering process leading to leaflet formation and demonstrate that endocardial mechanotransduction and valve morphogenesis are coupled via cellular rearrangements mediated by fibronectin synthesis.
Boselli F, Vermot J, 2016, Live imaging and modeling for shear stress quantification in the embryonic zebrafish heart, METHODS, Vol: 94, Pages: 129-134, ISSN: 1046-2023
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- Citations: 29
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