Establishment of intestinal organoid cultures modeling injury-associated epithelial regeneration

Abstract

The capacity of 3D organoids to mimic physiological tissue organization and functionality has provided an invaluable tool to model development and disease in vitro. However, conventional organoid cultures primarily represent the homeostasis of self-organizing stem cells and their derivatives. Here, we established a novel intestinal organoid culture system composed of 8 components, mainly including VPA, EPZ6438, LDN193189, and R-Spondin 1 conditioned medium, which mimics the gut epithelium regeneration that produces hyperplastic crypts following injury; therefore, these organoids were designated hyperplastic intestinal organoids (Hyper-organoids). Single-cell RNA sequencing identified different regenerative stem cell populations in our Hyper-organoids that shared molecular features with in vivo injury-responsive Lgr5+ stem cells or Clu+ revival stem cells. Further analysis revealed that VPA and EPZ6438 were indispensable for epigenome reprogramming and regeneration in Hyper-organoids, which functioned through epigenetically regulating YAP signaling. Furthermore, VPA and EPZ6438 synergistically promoted regenerative response in gut upon damage in vivo. In summary, our results demonstrated a new in vitro organoid model to study epithelial regeneration, highlighting the importance of epigenetic reprogramming that pioneers tissue repair.

MTG16 (CBFA2T3) regulates colonic epithelial differentiation, colitis, and tumorigenesis by repressing E protein transcription factors

“…Interestingly, RNA-seq and ASCL2 ChIP-seq showed that these cells were not enriched for Clu or fetal epithelial genes previously thought to enhance intestinal epithelial response to injury 120,121 .…”

Characterization of an in vitro 3D intestinal organoid model by using massive RNAseq-based transcriptome profiling

“…In their research, they also found markers of differentiated epithelial cells (Lyz1, Muc2, ChgA, ALP) expressed less in organoids than raw crypts which is consistent with our findings. Qu et al 33 established a novel hyperplastic intestinal organoids (Hyper-organoids) by applying an intestinal organoid culture system composed of 8 components mainly including VPA, EPZ6438, LDN193189, and R-Spondin 1 conditioned medium. VPA and EPZ6438 was revealed to be indispensable for epigenome reprogramming and regeneration in Hyper-organoids by functioning through epigenetically regulating YAP signaling.…”

Developments and Opportunities for 3D Bioprinted Organoids

“…Deng et al . constructed a new intestinal organ culture system that can simulate the regeneration process of proliferative crypts after intestinal epithelial injury and revealed the key role and mechanism of two epigenetic regulators (VPA and EPZ6438) in regulating regeneration after intestinal injury ( Figure 5A and B )[ 57 ]. Meanwhile, a novel engineered plant-based nanocellulose hydrogel was recently reported as a culture medium for small intestine organoids, which has the advantages of clear composition and low cost compared with the currently used organoid culture medium, Matrigel[ 58 ] ( Figure 5C ).…”

“…(from ref. [ 57 ] licensed under Creative Commons Attribution 4.0 license) and (from ref. [ 58 ] licensed under Creative Commons Attribution 4.0 license).…”