Tissue architecture affects cell state. (a) An H&E-stained histological slice from three separate EwS tissue samples, evaluated by a pathologist. Pseudorosettes structures outlined with dashed line. Note both elongated and rounded presentations of pseudorosettes. (b) XY maximum intensity projections of TC71 cells expressing nuclear marker H2B-mcherry in the HBV, PVS, or CHT. Numbered dashed lines indicate the corresponding optical XZ projections depicted below. Note that only the HBV but not the other regions contains pseudorosette-like structures (indicated by arrowheads) throughout the cell cluster. (c) Proportion of contact with other TC71 cells is plotted against volume (b) and sphericity (c). While no clear relationship is observed for volume, a few embedded (E) cells but not other groups (NN, no neighbor; PE, partially exposed) show cells with distinctly low sphericity (<0.8). Arrows indicate cells from all three regions with distinctive cell morphological states, rendered to the right and color-coded by k-means clustering in Fig. 2 e. Scale bars, 20 µm.