Fig. 8: Biomechanical phase diagram of gut spheroid fission.

a Phase diagram of different spheroid morphogenic states. (Inset) Simulation results are representative of spheroid morphogenesis with impaired surface tension (surface tension inhibited), or normal tension (normal), respectively. b (upper) Representative confocal micrographs showing X-Y sections of staining of F-actin, phosphorylated myosin light chain (pMLC), and the nuclei (DAPI) in protruding, pre-fission tissue columns formed in micropatterned gut spheroid generator with Y27632 treatment. (upper right) Representative tissue buds (prospective spheroid-forming regions) were selected from pre-fission tissue columns and shown in zoom-in images. (lower right) Fluorescence intensities of F-actin and pMLC along the diameter of selected tissue buds (white lines in zoom-in images). Black arrows indicate locations of outer tissue surface, and black arrowheads show the location of inner / lumenal tissue surface, of selected tissue buds. n = 3 independent experiments. Scale bar: 100 μm. c Violin plots showing average fluorescence intensities of F-actin and pMLC at the lumenal and outer surfaces of pre-fission tissue columns under indicated conditions on day 7. The lines in the violin plots represent median values. n_{pre-fission tissue columns} > 30 and n_micropattern > 5 for each experiment. n = 3 independent experiments. d (left) Representative phase images showing posterior foregut tissues that remained attached to micropatterned gut spheroid generator after removing all readily-formed spheroids from the supernatant under indicated conditions on day 7. Scale bar: 400 μm. (right) Red dashed squares mark colonies for zoom-in. e Percentage of colonies with remnant pearling under the indicated conditions. n = 4 independent experiments. f Representative phase images showing readily-formed spheroids collected from μGSG under indicated conditions. Scale bar: 500 μm. g Normalized number of spheroids generated from indicated conditions. Data were plotted as mean ± s.d. n = 4 independent experiments. h Representative phase images showing readily-formed spheroids collected from μGSG under indicated conditions. Scale bar: 500 μm. i Normalized the number of spheroids generated from indicated conditions. Data were plotted as mean ± s.d. n = 3 independent experiments. P-values were calculated using an unpaired, two-sided Student’s t-test. Source data are provided as a Source Data file.