Extended Data Fig. 4: CDH1-tagged and TUBB-tagged human hepatocyte organoids as a tool for tracing cell movement and mitotic spindle dynamics.
a, Tagging of CDH1 reveals that hepatocytes tend to form rosette structures around lumina (indicated by the white arrows) (left). Representative example of analysis of cell movement in CDH1::mNEON human hepatocyte organoids (right). Individual cell movements were traced based on changes of cell centroid positioning over time. The green dots represent the initial position of the cell centroid and length and orientation of each green arrow represent individual cell centroid movement from begin to end of the experiment. The red line indicates lumen movement. (t = 9 hours, 45 min intervals). The experiment was repeated five times independently with similar results. b, Representative examples of tracing of cell movement based on changes of cell centroid positioning over time in CDH1::mNEON (left) and CDH1::tdTomato (right) human hepatocyte organoids. Initial and final positioning and cell shape outline are represented in blue and pink, respectively, and cell movements are indicated by arrows. Similarly, the initial and final position of the lumen is outlined and its movement is visualized. Note that hepatocytes tend to rotate around the lumen. c, Bar plot showing the average cell rotation for individual organoids with a lumen (light green) or without a lumen (dark green). Note that in organoids in which there is no lumen cells tend to rotate less. as previously mentioned, organoids with a lumen tend to rotate. n=9 clonal organoids with a lumen and n=7 clonal organoids without a lumen. Two-sided unpaired t-test; *p<0.05. d, Example of the determination of mitotic spindle dynamics in TUBB::GFP human hepatocyte organoids. The mitotic spindle orientation was traced over time by marking the initial (gray) and final (red) spindle orientation. The thin line indicates the average position of the spindle poles at every time point between the two. Scale bars, a: left: 75 µm, right: 100 µm; d: 100 µm. Statistics source data are provided in Statistical Source Data Extended Data Fig. 4.
