Our current scope of research is to investigate the roles of circadian rhythms in intestinal epithelial cell proliferation and differentiation. We are trying to determine whether the circadian clock regulates cell proliferation and differentiation from intestinal stem cells. This protocol presents a real-time monitoring method for circadian studies in three-dimensional organoids using a bioluminescence assay.
Unlike performing bioluminescence assays in a conventional two-dimensional cell culture, this approach enables us to investigate circadian rhythms and their functions in a more physiologically relevant organoid model. Our findings show disrupted circadian rhythms in stem cell-enriched conditions while demonstrating robust circadian oscillations in differentiated human intestinal organoids, suggesting a remodeling of circadian clock machinery from stem cells to differentiated cells. Usages of patient-derived organoids with circadian bioluminescent reporters will help us advance our understanding of circadian rhythms in peripheral organs and different disease states.
