Culture-Associated DNA Methylation Changes Impact on Cellular Function of Human Intestinal Organoids

Area of Science:

• Gastroenterology
• Epigenetics
• Cell Biology

Background:

• Human intestinal epithelial organoids (IEOs) model intestinal development and disease.
• In vitro culturing requires passaging, which may alter cell characteristics.

Purpose of the Study:

• To investigate the impact of culture duration on DNA methylation (DNAm) in human IEOs.
• To assess how DNAm changes affect gene expression and cellular function.

Main Methods:

• Analyzed 173 human IEO lines from small and large bowels.
• Examined DNA methylation changes over culture duration.
• Correlated DNAm changes with gene expression and cellular function.

Main Results:

• Culture duration reproducibly altered DNAm in IEOs across multiple cohorts.
• While global function remained stable, a subset of DNAm changes correlated with altered gene expression.
• Epigenetic changes were enriched in cancer-associated genomic regions.

Conclusions:

• Culture-associated epigenetic changes occur in human IEOs.
• These epigenetic alterations can impact gene expression and cellular function.
• Passage number is a critical factor to consider in IEO research.