(A) Sketch showing approximate position of the SBF-SEM imaged planes (x-y planes), and image stacks (spanning the z-direction depth) with respect to the heart morphology. For the imaged transmural planes, the y-direction approximately corresponds to the radial direction in the left ventricle (LV), and the x-direction is approximately parallel to the LV wall boundary (circumferential direction). The stack depth direction (z-direction) corresponds to the heart longitudinal direction. Myofibril orientation angles were quantified for the x-y and x-z planes: the transmural angle θ is the myofibril angle in the x-y plane, with respect to the circumferential (x) direction (wall direction); the elliptical angle Φ is the orientation angle in the x-z plane with respect to the circumferential (x) direction. Segmentations of organelles on the x-y and x-z planes, together with frequency versus angle histograms, are shown in (B) for the control heart, ROI A (CON A); and (C) for the TOF heart, ROI A (TOF A). Each bar in the histogram represents a 10° range, hence there are 18 bars to represent 0° to 180° angles. Organelle segmentation is color-coded. Red: nuclei; Blue: mitochondria; Green: myofibrils. The extracellular matrix is shown in yellow. Scale bars: 10 µm. The details of the angle calculations are included as supplementary files (Figure 10—source datas 1–4).