Authors: Zheng Liu, Hu Chao, Yang Binbin, Yang Xinggang, Ding Zhixiang
To investigate the inhibitory effect of small interfering RNA-Yes-associated protein 1 (siRNA-YAP1) on epithelial-mesenchymal transition (EMT) in human lens epithelial cells (LECs) induced by transforming growth factor-β2 (TGF-β2).
Human LECs line (HLEB-3) was cultured and divided into normal control group and TGF-β2 induced group.The cells in the normal control group were treated with serum-free low-glucose medium for 24 hours, and the cells in the TGF-β2 induced group were treated with additional 10 ng/ml TGF-β2 for 24 hours.The cultured HLEB-3 cells were divided into siRNA empty vector group, siRNA-YAP1 transfection group, siRNA empty vector+ TGF-β2 group and siRNA-YAP1+ TGF-β2 group, and the cells were transfected with plasmid including siRNA empty vector or siRNA-YAP1 sequence according to grouping.The relative expression levels of YAP1 mRNA and protein in various groups were detected and compared by quantitative real-time polymerase chain reaction (PCR), immunofluorescence and Western blot assay, respectively.The relative expression levels of EMT marker proteins (E-cadherin and Vimentin proteins) in various groups were detected by immunofluorescence and Western blot assay.
Compared with the normal control group, the expression level of E-cadherin protein was decreased (1.180±0.118 vs.0.830±0.104) and the Vimentin protein was increased (0.797±0.110 vs.1.240±0.110) in the TGF-β2 induced group, with significant differences between the two groups (t=3.857, P=0.018; t=-4.933, P=0.008).The relative expression levels of YAP1 mRNA and protein in the TGF-β2 induced group were significantly increased in comparison with the normal control group (2.200±0.193 vs.1.136±0.123; 1.203±0.121 vs.0.967±0.025), with significant differences between the two groups (t=-9.288, P<0.01; t=-3.329, P=0.029).Compared with the siRNA empty vector group, the expression levels of YAP1 mRNA and protein in the siRNA-YAP1 transfection group were significantly reduced (both at P<0.01).Compared with the siRNA empty vector+ TGF-β2 group, the relative expression level of E-cadherin protein was significantly enhanced and the expression level of Vimentin protein was significantly reduced in the siRNA-YAP1+ TGF-β2 group (both at P<0.01).
YAP1 participates in the TGF-β2 induced EMT in human LECs, and siRNA-YAP1 can suppress the EMT process.