Inhibitory effect of siRNA-YAP1 on transforming growth factor β2-induced epithelial-mesenchymal transition in human lens epithelial cells

Authors: Zheng Liu,  Hu Chao,  Yang Binbin,  Yang Xinggang,  Ding Zhixiang

DOI: 10.3760/cma.j.cn115989-20191016-00447
Published 2021-04-10
Cite asChin J Exp Ophthalmol, 2021, 39(4): 289-296.

Abstract

Objective

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).

Methods

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.

Results

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).

Conclusions

YAP1 participates in the TGF-β2 induced EMT in human LECs, and siRNA-YAP1 can suppress the EMT process.

Key words:

Lens epithelial cells; Epithelial-mesenchymal transition; Yes-associated protein 1; Small interfering RNA; Inhibitory effect

Contributor Information

Zheng Liu
Department of Ophthalmology, Affiliated Hospital of Guilin Medical University, Guilin 541001, China
Hu Chao
Department of Ophthalmology, Affiliated Hospital of Guilin Medical University, Guilin 541001, China
Yang Binbin
Department of Ophthalmology, Affiliated Hospital of Guilin Medical University, Guilin 541001, China
Yang Xinggang
Department of Ophthalmology, Affiliated Hospital of Guilin Medical University, Guilin 541001, China
Ding Zhixiang
Department of Ophthalmology, Affiliated Hospital of Guilin Medical University, Guilin 541001, China
(Read 3 times, 1 visits today)
Updated: April 14, 2021 — 9:10 am