Lotta Hof1, Till Moreth1, Michael Koch1, Tim Liebisch2, Marina Kurtz3, Julia Tarnick4, Susanna M Lissek5, Monique M A Verstegen6, Luc J W van der Laan6, Meritxell Huch7,8, Franziska Matthäus2, Ernst H K Stelzer1, Francesco Pampaloni9. 1. Physical Biology Group, Buchmann Institute for Molecular Life Sciences (BMLS), Goethe-Universität Frankfurt am Main, Frankfurt am Main, Germany. 2. Frankfurt Institute for Advanced Studies and Faculty of Biological Sciences, Goethe-Universität Frankfurt am Main, Frankfurt am Main, Germany. 3. Department of Physics, Goethe Universität Frankfurt am Main, Frankfurt am Main, Germany. 4. Deanery of Biomedical Science, University of Edinburgh, Edinburgh, UK. 5. Experimental Medicine and Therapy Research, University of Regensburg, Regensburg, Germany. 6. Department of Surgery, Erasmus MC - University Medical Center, Rotterdam, The Netherlands. 7. The Wellcome Trust/CRUK Gurdon Institute, University of Cambridge, Cambridge, UK. 8. Present address: Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany. 9. Physical Biology Group, Buchmann Institute for Molecular Life Sciences (BMLS), Goethe-Universität Frankfurt am Main, Frankfurt am Main, Germany. fpampalo@bio.uni-frankfurt.de.
Abstract
BACKGROUND: Organoids are morphologically heterogeneous three-dimensional cell culture systems and serve as an ideal model for understanding the principles of collective cell behaviour in mammalian organs during development, homeostasis, regeneration, and pathogenesis. To investigate the underlying cell organisation principles of organoids, we imaged hundreds of pancreas and cholangiocarcinoma organoids in parallel using light sheet and bright-field microscopy for up to 7 days. RESULTS: We quantified organoid behaviour at single-cell (microscale), individual-organoid (mesoscale), and entire-culture (macroscale) levels. At single-cell resolution, we monitored formation, monolayer polarisation, and degeneration and identified diverse behaviours, including lumen expansion and decline (size oscillation), migration, rotation, and multi-organoid fusion. Detailed individual organoid quantifications lead to a mechanical 3D agent-based model. A derived scaling law and simulations support the hypotheses that size oscillations depend on organoid properties and cell division dynamics, which is confirmed by bright-field microscopy analysis of entire cultures. CONCLUSION: Our multiscale analysis provides a systematic picture of the diversity of cell organisation in organoids by identifying and quantifying the core regulatory principles of organoid morphogenesis.
BACKGROUND: Organoids are morphologically heterogeneous three-dimensional cell culture systems and serve as an ideal model for understanding the principles of collective cell behaviour in mammalian organs during development, homeostasis, regeneration, and pathogenesis. To investigate the underlying cell organisation principles of organoids, we imaged hundreds of pancreas and cholangiocarcinoma organoids in parallel using light sheet and bright-field microscopy for up to 7 days. RESULTS: We quantified organoid behaviour at single-cell (microscale), individual-organoid (mesoscale), and entire-culture (macroscale) levels. At single-cell resolution, we monitored formation, monolayer polarisation, and degeneration and identified diverse behaviours, including lumen expansion and decline (size oscillation), migration, rotation, and multi-organoid fusion. Detailed individual organoid quantifications lead to a mechanical 3D agent-based model. A derived scaling law and simulations support the hypotheses that size oscillations depend on organoid properties and cell division dynamics, which is confirmed by bright-field microscopy analysis of entire cultures. CONCLUSION: Our multiscale analysis provides a systematic picture of the diversity of cell organisation in organoids by identifying and quantifying the core regulatory principles of organoid morphogenesis.
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