Fig. 9.

Altered expression level of GRHL2 in human IPF in vitro manipulation of Grhl2 in mouse epithelial cells lines highlights the functional role of GRHL2 in IPF. A: GRHL2 expression level in IPF patients showed substantial decrease compared with non-IPF donors. GAPDH was used as endogenous control for normalization. B: epithelial and mesenchymal expression level in Grhl2 knockdown studies in MLE15 cells showed decrease in epithelial genes such as Cdh1 and Cldn4 but showed increased expression of Vim and Zeb1. C: cell morphology of MLE15 cells transduced with nonsilencing (Nonsilence) and Grhl2 shRNA. Grhl2 knockdown in MLE15 shows cell flattening of MLE15 after 92 h of incubation. D: Grhl2 staining in E10 cells transduced with CMV-GRHL2-UBC-GFP to show that stable transfections and expression of Grhl2. E: overexpression of Grhl2 in E10 cells showed upregulation of Grhl2 and Cdh1 with modest increase in mesenchymal genes. E10 cells were either transduced with lentivirus containing CMV-Grhl2-UBC-GFP (E10-Grhl2) or with empty vector (CMV-dsred-UBC-GFP). GRHL2 expression studies were performed with 11 IPF patients and 8 non-IPF donors. Grhl2 knockdown and overexpression studies were performed in triplicate.