Résumé
We aimed at understanding the differentiation processes that drive airway epithelial cell phenotype as it may offer therapeutic avenues in chronic airway diseases.NHBE (Normal Human Bronchial Epithelial) were cultured at ALI (Air Liquid Interface) for 7 days with or without Y27632, DapT and LDN as these molecules are known to affect Rho, Notch and BMP pathways respectively. The four fold increase in CCSP (Club Cell Secretory Protein) mRNA, positive CCSP cells (from 0 to 1000/mm²) and CCSP concentration (from 0 to 30 ng/ml) were observed from day 0 to day 7 when NHBE were cultured with traditional medium. Moreover this was extended by another four to five fold increase mRNA with the RhoA/ROCK inhibitor Y27632. Cell density increased up to 6000/mm² (p<0.01) and CCSP concentration increased up to 45 ng/ml (p<0.01). The BMP inhibitor, LDN, produced the same effect but was not additive to Y27632. Interestingly, CCSP upregulation by Y27632 and LDN was accompanied by an increase in MUC5AC (Mucin 5AC) positive cells mRNA, and this was more specifically achieved with LDN. However, inhibiting the Notch γ secretase with DapT specifically increased the ciliated cells marker, FoxJ1 (Forkhead Box J1), while hampering CCSP and MUC5AC mRNA, proteins and density of positive cells. Main airway epithelial cell fate drivers were identified, Y27632 and LDN promoting differentiation toward club and goblet cells whereas DapT promoted ciliated cells.