The pattern of blood flow has long been thought to play a significant role in vascular morphogenesis, yet the flow-sensing mechanism that is involved at early embryonic stages, when flow forces are low, remains unclear. It has been proposed that endothelial cells use primary cilia to sense flow, but this has never been tested in vivo. Here we show, by noninvasive, high-resolution imaging of live zebrafish embryos, that endothelial cilia progressively deflect at the onset of blood flow and that the deflection angle correlates with calcium levels in endothelial cells. We demonstrate that alterations in shear stress, ciliogenesis, or expression of the calcium channel PKD2 impair the endothelial calcium level and both increase and perturb vascular morphogenesis. Altogether, these results demonstrate that endothelial cilia constitute a highly sensitive structure that permits the detection of low shear forces during vascular morphogenesis.

Highlights d Oscillatory flow amplitude scales with klf2a expression and calcium levels d TRPP2 and TRPV4 are mechanosensitive channels in the endocardium d TRPV4 and TRPP2 control valve development d TRPV4 and TRPP2 control klf2a expression and intracellular calcium

Hemodynamic forces are fundamental to development. Indeed, much of cardiovascular morphogenesis reflects a two-way interaction between mechanical forces and the gene network activated in endothelial cells via mechanotransduction feedback loops. As these interactions are becoming better understood in different model organisms, it is possible to identify common mechanogenetic rules, which are strikingly conserved and shared in many tissues and species. Here, we discuss recent findings showing how hemodynamic forces potentially modulate cardiovascular development as well as the underlying fluid and tissue mechanics, with special attention given to the flow characteristics that are unique to the small scales of embryos.

Mechanical forces are instrumental to cardiovascular development and physiology. The heart beats approximately 2.6 billion times in a human lifetime and heart valves ensure that these contractions result in an efficient, unidirectional flow of the blood. Composed of endocardial cells (EdCs) and extracellular matrix (ECM), cardiac valves are among the most mechanically challenged structures of the body both during and after their development. Understanding how hemodynamic forces modulate cardiovascular function and morphogenesis is key to unraveling the relationship between normal and pathological cardiovascular development and physiology. Most valve diseases have their origins in embryogenesis, either as signs of abnormal developmental processes or the aberrant re-expression of fetal gene programs normally quiescent in adulthood. Here we review recent discoveries in the mechanobiology of cardiac valve development and introduce the latest technologies being developed in the zebrafish, including live cell imaging and optical technologies, as well as modeling approaches that are currently transforming this field. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.

Myocardial contractility and blood flow provide essential mechanical cues for the morphogenesis of the heart. In general, endothelial cells change their migratory behavior in response to shear stress patterns, according to flow directionality. Here, we assessed the impact of shear stress patterns and flow directionality on the behavior of endocardial cells, the specialized endothelial cells of the heart. At the early stages of zebrafish heart valve formation, we show that endocardial cells are converging to the valve-forming area and that this behavior depends upon mechanical forces. Quantitative live imaging and mathematical modeling allow us to correlate this tissue convergence with the underlying flow forces. We predict that tissue convergence is associated with the direction of the mean wall shear stress and of the gradient of harmonic phase-averaged shear stresses, which surprisingly do not match the overall direction of the flow. This contrasts with the usual role of flow directionality in vascular development and suggests that the full spatial and temporal complexity of the wall shear stress should be taken into account when studying endothelial cell responses to flow in vivo.

Hemodynamic shear stress is sensed by the endocardial cells composing the inner cell layer of the heart, and plays a major role in cardiac morphogenesis. Yet, the underlying hemodynamics and the associated mechanical stimuli experienced by endocardial cells remains poorly understood. Progress in the field has been hampered by the need for high temporal resolution imaging allowing the flow profiles generated in the beating heart to be resolved. To fill this gap, we propose a method to analyze the wall dynamics, the flow field, and the wall shear stress of the developing zebrafish heart. This method combines live confocal imaging and computational fluid dynamics to overcome difficulties related to live imaging of blood flow in the developing heart. To provide an example of the applicability of the method, we discuss the hemodynamic frequency content sensed by endocardial cells at the onset of valve formation, and how the fundamental frequency of the wall shear stress represents a unique mechanical cue to endocardial, heart-valve precursors.

Desminopathies belong to a family of muscle disorders called myofibrillar myopathies that are caused by Desmin mutations and lead to protein aggregates in muscle fibers. To date, the initial pathological steps of desminopathies and the impact of desmin aggregates in the genesis of the disease are unclear. Using live, high-resolution microscopy, we show that Desmin loss of function and Desmin aggregates promote skeletal muscle defects and alter heart biomechanics. In addition, we show that the calcium dynamics associated with heart contraction are impaired and are associated with sarcoplasmic reticulum dilatation as well as abnormal subcellular distribution of Ryanodine receptors. Our results demonstrate that desminopathies are associated with perturbed excitation-contraction coupling machinery and that aggregates are more detrimental than Desmin loss of function. Additionally, we show that pharmacological inhibition of aggregate formation and Desmin knockdown revert these phenotypes. Our data suggest alternative therapeutic approaches and further our understanding of the molecular determinants modulating Desmin aggregate formation.

Highlights d Endothelial cell (EC) nuclei are enriched in the ventral dorsal aorta (DA) during EC movement d DA wall deforms asymmetrically in response to pulsatile blood flow d Low blood flow or abnormal mechanosensing results in increased cell extrusion