A Semi-automated Organoid Screening Method Demonstrates Epigenetic Control of Intestinal Epithelial Differentiation

Abstract

Intestinal organoids are an excellent model to study epithelial biology. Yet, the selection of analytical tools to accurately quantify heterogeneous organoid cultures remains limited. Here, we developed a semi-automated organoid screening method, which we applied to a library of highly specific chemical probes to identify epigenetic regulators of intestinal epithelial biology. The role of epigenetic modifiers in adult stem cell systems, such as the intestinal epithelium, is still undefined. Based on this resource dataset, we identified several targets that affected epithelial cell differentiation, including HDACs, EP300/CREBBP, LSD1, and type I PRMTs, which were verified by complementary methods. For example, we show that inhibiting type I PRMTs, which leads enhanced epithelial differentiation, blocks the growth of adenoma but not normal organoid cultures. Thus, epigenetic probes are powerful tools to study intestinal epithelial biology and may have therapeutic potential.

BMP signaling in the intestinal epithelium drives a critical feedback loop to restrain IL-13–driven tuft cell hyperplasia

“…Lgr5+ stem cells and Paneth cells are found in the buds in budding organoids (mimicking crypts) while spheroids consist of less differentiated, proliferating cells ( 11 ). Building on our recent work on organoid segmentation ( 12 ), we here developed an automated image analysis pipeline that segments organoid objects from the background and subsequently classifies them into “spheroid” or “budding” categories based on a convolutional neural network ( Fig. 1A ).…”

An off-the-shelf multi-well scaffold-supported platform for tumour organoid-based tissues

“…Further, hydrogel plugs do not facilitate longitudinal in situ imaging of organoid phenotype due to the curved surface and high gel thickness associated with the hydrogel meniscus. The ability to perform longitudinal imaging of tumour cell cultures is important as we [35], [36] and others [25], [71], have shown the benefits of using this approach to provide a deeper understanding of organoid response to therapy. Here, we described the use of a thin paper scaffold to create supported hydrogel microtissues without a meniscus that enable the culture and long-term imaging of tumour organoids for analysis of drug response.…”

Single-cell technologies: a new lens into epigenetic regulation in development

“…Unlike comparatively inaccessible mouse embryos, these in vitro models enable streamlined application of chemical perturbation to affect lineage trajectories with greater experimental control, allowing researchers to direct these structures into different cell identities [61,64,66]. In parallel, chemical compound libraries to selectively inhibit epigenetic regulator function have been developed and used in high-throughput toxicology screens in 2D culture [68,69], 3D adult organoids [70], or in vivo [71].…”