The Effects Of Abnormal Expression Of YAP In Nasal Polyps-derived Fibroblasts On Nasal Epithelial Barrier Function

Objective: Previous studies suggested that Yes-associated protein(YAP)could affect the proliferation and remodeling of nasal mucosal epithelial cells from chronic sinusitis with nasal polyps.However,the expression and potential mechanism of YAP in human nasal fibroblasts remain unclear.In this study,the indirect co-culture model of primary nasal mucosal epithelial cells and fibroblasts derived from nasal polyps was established to clearly regulate the effect of YAP in nasal polyp-derived fibroblasts on the nasal mucosal epithelial barrier function,and to explore and improve the potential mechanism of action of YAP in chronic sinusitis with nasal polyps.It provides a new intervention index for optimizing the treatment of chronic sinusitis with nasal polyps.Method: Nasal polyp tissues and inferior turbinate mucosa tissues of patients undergoing functional nasal endoscopic surgery in the Department of Otolaryngology and Head and Neck Surgery,Affiliated Hospital of Qingdao University were collected.We used q-PCR and western blotting methods to detect the expressions of Hippo pathway downstream genes such as YAP,SAV1,TEAD and MST1,and candidate differential genes were screened.Bioinformatics was used to analyze the protein interaction network,possible cell functions and signaling pathways of differential genes,which were verified by subsequent experiments.Nasal epithelial cells and fibroblasts were extracted from nasal polyp tissue,and the indirect co-culture model of nasal epithelial cells and nasal polyp derived fibroblasts was established.On this basis,lentivirus coated with overexpressed target gene plasmids was used to infect the nasal fibroblasts in the lower layer of the model,thus the expression of target gene was up-regulated in the fibroblasts.After the differentiation of the upper nasal mucosal epithelial cells in the co-cultured model was finished,q-PCR,western blotting and immunofluorescence methods were used to detect the tight junction of nasal epithelial cells,epithelial-mesenchymal transformation,and epithelial differentiation.Results: The expression of YAP,the key factor of Hippo pathway,was increased in nasal polyps compared with normal inferior turbinate mucosa.In addition,the results further suggested that YAP expression was up-regulated in nasal polyps-derived fibroblasts compared with controls.In the indirect co-culture model of nasal epithelial cells and nasal fibroblasts,overexpression of YAP in the lower layer of nasal fibroblasts showed that(1)the expression of tight junction proteins ZO-1,Occludin,and Claudin was down-regulated;(2)The expression of E-cadherin was down-regulated,and the expression of epithelial to mesenchymal transition related genes such as Fibronectin,Snail1,β-catenin,EGFR,MMP1 and MMP9 was up-regulated.(3)The expression of Muc5 AC,a marker of goblet cells,was up-regulated,while β-Ⅳ tubulin,a marker of ciliated columnar epithelial cells,was downregulated.Conclusion: Compared with the control group,the expression of YAP is up-regulated in human nasal polyps and their derived nasal fibroblasts.Overexpression of YAP gene in nasal polyp-derived fibroblasts can reduce the expression of epithelial junctional proteins in nasal mucosa,increase the number of goblet cells,decrease the number of ciliated cells,and affect the epithelial to mesenchymal transition of nasal mucosa,thereby affecting the epithelial barrier function.Therefore,this study explores the interaction between nasal polyp-derived fibroblasts and nasal mucosal epithelial cells,and YAP can change the characteristics of nasal mucosal epithelial cells by acting on nasal polyp-derived fibroblasts.This study is conducive to further exploring the theoretical basis of submucosal involvement in the pathogenesis of nasal polyps,and provides a new target for optimizing the diagnosis and treatment of chronic sinusitis complicated with nasal polyps.