Fig 1.
Overview of combined experimental and computational pipeline to compare hemodynamic environments and shape variation across a cohort of embryonic chick hearts.
Table 1.
Reported HH18 Embryonic Chick Morphological and Hemodynamic Properties.
Fig 2.
Light sheet fluorescence microscopy optically sections whole embryos for generating anatomy reconstructions.
(A) The LCA 649 channel distinguished endothelial cells lining the lumen and was primarily used to inform segmentations. (B) The autofluorescence channel distinguished soft tissue and was used to verify segmentations. (C) Together these channels were used to construct segmentations of the developing heart, AAs, and dorsal aorta. (D-F) The imaging captured anatomical features that are often difficult to obtain, due to their size and position. In the segmentation panel, the segmented portions of interest are highlighted as solid yellow within the entire segmented anatomy (blue). (D) AAs, which are notoriously difficult to image due to their small size and position, were segmented using the LCA 649 channel. (E) The segmented heart and outflow tract (red). The LCA channel distinguished the lumen from other soft tissue, such as the cardiac jelly, which can be seen in the autofluorescence channel. (F) The bifurcating dorsal aorta segmented using the LCA 649 channel.
Fig 3.
Geometric Morphometric Methods can be applied to segmented embryo cohorts and used to identify outliers and variation across individuals.
(A) A template embryo was selected and 275 pseudolandmarks were placed along the contours of geometric features, such as the AAs, for the analysis. (B) Originally, two embryos that were missing multiple AAs were included in the GPA. The PCA scatter plot of the Procrustes residuals separates these two embryos from the rest of the cohort, with an inlay plot showing the detail. X-axis is shown in log scale. (C) The variance of each pseudolandmark is plotted by representing each as an ellipsoid. The radius length of each ellipsoid corresponds to the variance in that dimension of the pseudoLM. (D)The GPA was repeated on only embryos containing all six AAs. The PCA plot displays how the embryos varied in PC 1 and PC 2. The mean shape (grey), and the mean shape warped by PC 1 (blue) and PC 2 (red) is displayed. (E) Detailed views of the different anatomical regions of the mean shape, pseudolandmarks and warped mean shapes, and warped pseudoLMs. (F) View comparing the mean shape and PC 1-warped mean shape and PC 2-warped mean shape, pseudoLMs, and warped pseudoLMs.
Fig 4.
Wall shear stress is elevated in locally stenosed regions such as the AAs.
(A) (A) WSS was observed during peak flow and (B) accelerating flow. As expected, peak WSS occurred during peak flow. Peak WSS locations varied by individual anatomical features within embryos. (C) In the case 1 embryo with less stenosis in the outflow tract, peak WSS occurred in the AAs. (C) Conversely, in the case 2 embryo with a highly stenosed outflow tract, peak WSS occurred in the outflow tract. (E) An embryo with a moderately stenosed outflow tract experienced elevated WSS throughout locally stenosed regions including the outflow tract and AAs. WSS spatial patterning is similar during peak flow compared to accelerating flow. Regions that experience the highest WSS during peak flow experience high WSS during accelerating flow, indicating that this flow regime is not heavily driven by convective effects. (F) The pressure difference between the outflow tract and dorsal aorta was larger during peak flow compared to accelerating flow, as expected. (G) Average WSS was highest in the AA III pair, which is the centrally located of the three pairs present during this stage of development, despite all pairs of the AAs experiencing consistent pressure and the higher degree of stenosis in flanking arches of the AA III pair.
Fig 5.
Blood flow velocity streamlines are consistent with WSS patterns.
(A) Blood flow velocity streamlines are observed during peak and accelerating flow, exhibiting similar temporal and spatial patterns compared to WSS, consistent with the finding that there is low convective flow. As expected, blood flow velocity streamlines indicate elevated flow in the same regions as elevated WSS across variations in embryonic anatomies. (B) An embryo with little stenosis in the outflow tract experiences elevated velocity primarily in the AAs, consistent with WSS patterns. (C) An embryo with highly stenosed regions in the outflow tract experiences elevated flow velocity in the outflow tract, consistent with the elevated WSS. (D) An embryo with moderate outflow tract stenosis experiences elevated WSS throughout locally stenosed regions in the outflow tract and in the AAs.
Fig 6.
Streamline tracing confirms that there is low convective flow.
Streamlines identified through each aortic arch. Paths were traced from the outflow tract to the dorsal aorta to understand streamline spatial configuration in relation to before and after arch flow. Streamlines through the left AAs (A) and right AAs (B) of an embryo show that there is little mixing of flow streams, even after exiting the AAs. [C-D] Two views of the dorsal aorta demonstrate that the streamlines through AA IV pair are split by the other arch pair streamlines while traveling down the dorsal aorta. (E) We examined streamlines in the cross sections in the outflow tract, upstream of the AAs, (F) and in the cross section of the dorsal aorta, downstream of the AAs. The order of the streamlines in the outflow tract was maintained in the dorsal aorta, and confirms that the AA IV pair separates after flowing though the aortic arches. There is minimal mixing of streamlines, consistent with the finding that there is little convective influence in this flow regime. (G) A second embryo illustrates the spatial order of streamlines is largely conserved across anatomical variations. (H) Examined streamlines through an embryo where AA III is smaller than AA II, showing that the distribution but not order of streamlines is altered. Detailed views of the cross sections are shown to highlight all the AA streamlines.