Intestinal epithelial organoids established from gut tissue have become a widely used research tool. However, it remains unclear how environmental cues, divergent microbiota composition and other sources of variation before, during and after establishment confound organoid properties, and how these properties relate to the original tissue. While environmental influences cannot be easily addressed in human organoids, mice offer a controlled assay‐system. Here, we probed the effect of donor microbiota differences, previously identified as a confounding factor in murine in vivo studies, on organoids. We analyzed the proteomes and transcriptomes of primary organoid cultures established from two colonized and one germ‐free mouse colony of C57BL/6J genetic background, and compared them to their tissue of origin and commonly used cell lines. While an imprint of microbiota‐exposure was observed on the proteome of epithelial samples, the long‐term global impact of donor microbiota on organoid expression patterns was negligible. Instead, stochastic culture‐to‐culture differences accounted for a moderate variability between independently established organoids. Integration of transcriptome and proteome datasets revealed an organoid‐typic expression signature comprising 14 transcripts and 10 proteins that distinguished organoids across all donors from murine epithelial cell lines and fibroblasts and closely mimicked expression patterns in the gut epithelium.
Read more: Wiley Online Library