Aggregation of cryopreserved mid-hindgut endoderm for more reliable and reproducible hPSC-derived small intestinal organoid generation

Suspension culture in a rotating bioreactor for efficient generation of human intestinal organoids

“…This was also confirmed with mRNA expression analysis of cultured organoids. s-HIOs cultured in E(N)RA maintained comparable CDX2 and SOX2 expression to s-Spheroids but had greater expressions of vimentin and forkhead box F1 ( FOXF1 ), well-known mesenchymal markers in HIO, 8 , 17 , 25 suggesting that the suspension culture in the rotating bioreactor promotes not only the efficient growth of s-Spheroids but also their maturation to s-HIOs, likely through the improved exchange of culture factors and gases ( Figure 3 G). The fact that s-HIOs were surrounded by vimentin (VIM)-expressing mesenchyme suggests that the rotational floating culture condition is suitable for the maturation of HIOs to the same extent as t-HIOs in three-dimensional culture.…”

“…One induces hindgut spheroids from dissociated single cells in suspension, similar to our method, but requires three-dimensional culture for subsequent maturation. 17 The other reports suspension culture of HIOs; however, the induction of hindgut spheroids, which is the most complicated and variable part, follows the conventional method. 30 Our study demonstrates effective suspension culture throughout the entire culture process from hindgut spheroid induction to HIO maturation.…”

“…Furthermore, the amount and size of generated spheroids differ greatly between iPSC lines. 17 Therefore, the development of a simpler and more robust protocol for obtaining homogeneous spheroids and mature HIOs would provide a solid foundational basis for future clinical applications.…”

Host-microbiota interaction in intestinal stem cell homeostasis

“… 206 To address this challenge, Mayhew et al introduced a straightforward, reproducible, and scalable approach for generating HIOs through aggregating cryopreservable hPSC-derived mid-hindgut endoderm (MHE) monolayers, significantly enhancing HIO production by approximately tenfold. 214 Given the high incidence of diseases that impact the large intestine such as colitis and colon cancer, Wells et al detailed the differentiation of human colonic organoids (HCOs) from hPSCs through transient activation of BMP signaling. This innovative approach further expands the utility of HCO technology in studying colonic pathologies.…”

Scalable production of homogeneous cardiac organoids derived from human pluripotent stem cells

“…One notable technique within the latter category involves the utilization of bioreactors and non-adhesive-surface-treated microwells. 8 , 24 , 37 , 38 , 39 , 40 , 41 Bioreactors offer a practical approach for generating a substantial quantity of spheroids, characterized by a relatively uniform structure. These spheroids can be effectively guided to undergo differentiation into CMs originating from hiPSC spheroids.…”