Retinoic acid receptor γ activation promotes differentiation of human induced pluripotent stem cells into esophageal epithelium

Background The esophagus is known to be derived from the foregut. However, the mechanisms regulating this process remain unclear. In particular, the details of the human esophagus itself have been poorly researched. In this decade, studies using human induced pluripotent stem cells (hiPSCs) have proven powerful tools for clarifying the developmental biology of various human organs. Several studies using hiPSCs have demonstrated that retinoic acid (RA) signaling promotes the differentiation of foregut into tissues such as lung and pancreas. However, the effect of RA signaling on the differentiation of foregut into esophagus remains unclear. Methods We established a novel stepwise protocol with transwell culture and an air-liquid interface system for esophageal epithelial cell (EEC) differentiation from hiPSCs. We then evaluated the effect of all-trans retinoic acid (ATRA), which is a retinoic acid receptor (RAR)alpha, RAR beta and RAR gamma agonist, on the differentiation from the hiPSC-derived foregut. Finally, to identify which RAR subtype was involved in the differentiation, we used synthetic agonists and antagonists of RAR alpha and RAR gamma, which are known to be expressed in esophagus. Results We successfully generated stratified layers of cells expressing EEC marker genes that were positive for lugol staining. The enhancing effect of ATRA on EEC differentiation was clearly demonstrated with quantitative reverse transcription polymerase chain reaction, immunohistology, lugol-staining and RNA sequencing analyses. RAR gamma agonist and antagonist enhanced and suppressed EEC differentiation, respectively. RAR alpha agonist had no effect on the differentiation. Conclusion We revealed that RAR gamma activation promotes the differentiation of hiPSCs-derived foregut into EECs.