Single-Cell Sequencing Unveils Distal and Proximal Features of Induced Pluripotent Stem Cell-Derived Intestinal Organoids and cAMP’s role in enhancing multicellularity intestinal lineage specification.

This study aims to advance intestinal organoid models by characterizing mesenchyme-free regional-specific induced pluripotent stem cell (iPSC)-derived human intestinal organoids (HIOs) at single-cell resolution and investigating the impact of cyclic AMP (cAMP) on intestinal differentiation lineage specification. We evaluated various culture conditions' effects on iPSC-derived HIO differentiation by performing single-cell RNA sequencing (scRNA-seq) on proximal HIOs (n=2), distal HIOs cultured with (n=2), and without cAMP (n=2). Canonical marker analysis and validation against published scRNA-seq datasets were conducted. Immunohistochemistry and immunofluorescence assays supported our findings.