Cardiomyocyte segmentation and cross-sectional area calculation

The high resolution achieved by PB X-PCI allowed identifying and analyzing individual cardiomyocytes in the HR datasets, that were segmented and their CSA was quantified using open-source software (Fiji-Image J and 3D Slicer)^48,49.

In our cardiomyocyte analysis, we included two post-MI and the control hearts. Cardiomyocytes were analyzed in sub-volumes sized 551.2 µm × 551.2 µm × 260 µm (length × width × height) of the HR datasets. In post-MI hearts, cardiomyocytes in the area of infarction (including preserved cells adjoining the fibrotic post-MI tissue), as well as in the non-affected myocardium were analyzed. The infarcted area was part of the anterior wall, while the non-affected area analyzed was located in the septum. The same corresponding areas were also quantified in the control rat heart. Two sub-volumes were selected per area, one closer to the endocardium, and the other closer to the epicardium. Individual cardiomyocytes were segmented by manual delineation. Then CSA calculation was performed in Fiji-ImageJ and expressed as the median value with interquartile ranges of measurements of 10 cells per sub-volume.