DNA methylation defines regional identity of human intestinal epithelial organoids and undergoes dynamic changes during development

Abstract

ObjectiveHuman intestinal epithelial organoids (IEOs) are increasingly being recognised as a highly promising translational research tool. However, our understanding of their epigenetic molecular characteristics and behaviour in culture remains limited.DesignWe performed genome-wide DNA methylation and transcriptomic profiling of human IEOs derived from paediatric/adult and fetal small and large bowel as well as matching purified human gut epithelium. Furthermore, organoids were subjected to in vitro differentiation and genome editing using CRISPR/Cas9 technology.ResultsWe discovered stable epigenetic signatures which define regional differences in gut epithelial function, including induction of segment-specific genes during cellular differentiation. Established DNA methylation profiles were independent of cellular environment since organoids retained their regional DNA methylation over prolonged culture periods. In contrast to paediatric and adult organoids, fetal gut-derived organoids showed distinct dynamic changes of DNA methylation and gene expression in culture, indicative of an in vitro maturation. By applying CRISPR/Cas9 genome editing to fetal organoids, we demonstrate that this process is partly regulated by TET1, an enzyme involved in the DNA demethylation process. Lastly, generating IEOs from a child diagnosed with gastric heterotopia revealed persistent and distinct disease-associated DNA methylation differences, highlighting the use of organoids as disease-specific research models.ConclusionsOur study demonstrates striking similarities of epigenetic signatures in mucosa-derived IEOs with matching primary epithelium. Moreover, these results suggest that intestinal stem cell-intrinsic DNA methylation patterns establish and maintain regional gut specification and are involved in early epithelial development and disease.

Post‐natal co‐development of the microbiota and gut barrier function follows different paths in the small and large intestine in piglets

“…Pearson correlations were performed between jejunal mucosa data (morphology, barrier, and PRR expression) of all post-natal ages except PND0 and the relative abundance of the main jejunal families (with a median abundance greater than 0.1%). 26 Interestingly, colonic mucosa barrier and defense parameters do not display major changes with post-natal age. respectively, 6,26 suggesting that adult intestinal epithelial stem cells are programmed to engage toward small intestine or colon-type epithelia.…”

“…respectively, 6,26 suggesting that adult intestinal epithelial stem cells are programmed to engage toward small intestine or colon-type epithelia. Intestinal organoids derived from small intestine or colon of adult mice recapitulate markers of small intestine and colon epithelia, F I G U R E 8 Correlation matrix of microbiota (at the family level) and mucosal parameters in the jejunum.…”

Single-cell sequencing of developing human gut reveals transcriptional links to childhood Crohn’s disease

“…The ability to generate self-organising, intestinal epithelial organoids from fetal gut epithelium as early as 8-10 PCW implies the presence of epithelial cells that are capable of giving rise to such three-dimensional structures (Fordham et al, 2013). Indeed, the use of organoid models as tools to investigate early fetal intestinal development has been demonstrated previously (Kraiczy et al, 2019). Nevertheless, the cross-talk between epithelial cell subsets and other mucosal cell types as well as cell lineage trajectories remains unknown.…”

Non-coding DNA in IBD: from sequence variation in DNA regulatory elements to novel therapeutic potential

“…Surprisingly, only a minority of these locations (approximately 5000) overlapped between the two cell types examined, showing that enhancers exhibit cell type-specific patterns. The contribution of epigenetic signatures on development is further established by the finding that fetal gut shows distinct DNA methylation patterns and dynamics from paediatric and adult intestinal tissue 44. By identifying the divergent patterns of enhancer activity in different cell types within one organism, enhancer activity was shown to play an important role during differentiation and development 45 46…”