422 A DORMANT BLOOD MICROBIOME MAY UNDERLIE THE PERSISTENT AND RELAPSING INFLAMMATION OF CROHN'S DISEASE

Background: The development of the esophagus results from the balanced coordination of esophageal progenitor cell (EPC) proliferation and differentiation.Much remains unknown about the molecular mechanisms related to the development of the esophagus.YAP is an important regulator of proliferation in several endodermal derived organs; however, the role of YAP in esophageal development remains to be elucidated.We hypothesize that YAP is required for the proliferation and stratification of EPCs in the developing esophagus.To address this we used genetic mouse models and 3D human pluripotent stem cell (hPSC)derived esophageal organoids to evaluate the role of YAP in esophageal development.Methods: We deleted Yap using Shh-Cre; Yap loxp/loxp mutants to examine the role of Yap deletion in the developing mouse esophagus.To investigate the effect of Yap overexpression we generated Shh-Cre;Yap loxp/loxp ;R26 Yap5SA mutants in which endogenous Yap is replaced by constitutively nuclear Yap.We utilized our previously established protocol to generate EPCs from human pluripotent stem cells (hPSCs) that are able to self-renew and differentiate into a stratified squamous epithelium in 3D organoid culture.We treated hPSCs-derived EPCs with the YAP inhibitor Verteporfin to examine its effects on proliferation and differentiation of EPCs.Results: Deletion of Yap in the mouse esophagus reduces the number of epithelial layers and lumen size.Reduction of the numbers of p63 + , pHH3 + and Edu + EPCs suggests that EPC proliferation is affected by Yap loss.Deletion of Yap affects stratification, as seen in the reduction in the thickness of the Krt13 + epithelium and reduction of Krt8 + suprabasal cells.Yap overexpression causes a hyperplastic phenotype of the epithelium in which the thickness of basal cell layers (p63 + Krt5 + ) and differentiated cell layers (Krt13 + ) are increased.Additionally, the number of p63 + basal cells and pHH3 + proliferating epithelial cells are increased.YAP inhibition with Verteporfin reduces the number and size of esophageal organoids.Consistently, YAP knockdown by siRNA reduces the numbers proliferation (Ki67 + ) of EPCs and stratification of hPSC-derived esophageal organoids.Conclusions: Our results show that YAP plays a conserved role in the development of the mouse and human esophageal epithelium by regulating EPC proliferation.Notably, YAP has been shown to be increased in dysplastic esophageal epithelium and esophageal squamous cell carcinoma.Future directions include evaluating whether YAP can act as an oncogene to transform these EPCs at early stages of these malignancies.