Inhibition of enhancer of zeste homolog 2 prevents corneal myofibroblast transformation in vitro

Kai Liao; Zekai Cui; Yong Zeng; Jian Liu; Yini Wang; Zhijie Wang; Shibo Tang; Jiansu Chen

doi:10.1016/j.exer.2021.108611

Inhibition of enhancer of zeste homolog 2 prevents corneal myofibroblast transformation in vitro

Exp Eye Res. 2021 Jul:208:108611. doi: 10.1016/j.exer.2021.108611. Epub 2021 May 13.

Authors

Kai Liao¹, Zekai Cui², Yong Zeng³, Jian Liu², Yini Wang², Zhijie Wang¹, Shibo Tang⁴, Jiansu Chen⁵

Affiliations

¹ Aier School of Ophthalmology, Central South University, Changsha, Hunan, China; Aier Eye Institute, Changsha, Hunan Province, China.
² Aier Eye Institute, Changsha, Hunan Province, China.
³ Department of Ophthalmology, Sichuan Academy of Medical Sciences and Sichuan Provincial People's Hospital, Chengdu, China.
⁴ Aier School of Ophthalmology, Central South University, Changsha, Hunan, China; Aier Eye Institute, Changsha, Hunan Province, China; CAS Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, Shanghai, China. Electronic address: tangshibo@vip.163.com.
⁵ Aier School of Ophthalmology, Central South University, Changsha, Hunan, China; Aier Eye Institute, Changsha, Hunan Province, China; Key Laboratory for Regenerative Medicine, Ministry of Education, Jinan University, Guangzhou, China. Electronic address: chenjiansu2000@163.com.

PMID: 33992624
DOI: 10.1016/j.exer.2021.108611

Abstract

Purpose: Corneal fibroblast can be transformed into corneal myofibroblasts by TGF-β1. Enhancer of zeste homolog 2 (EZH2) upregulation has been observed in the occurrence of other fibrotic disorders. We investigated the role of EZH2 in the progression of corneal fibrosis and the antifibrotic effect of EZH2 inhibition in corneal fibroblasts (CFs).

Methods: Primary CFs were isolated from corneal limbi and the CFs were treated with TGF-β1 to induce fibrosis. EPZ-6438 and EZH2 siRNA were used to inhibit EZH2 expression. Myofibroblast activation and extracellular matrix (ECM) protein synthesis was detected by quantitative real-time PCR, western blotting, and immunofluorescence staining assay. The functions of myofibroblast were evaluated by cell migration and collagen gel contraction assays. Molecular mechanisms involved in EZH2 inhibition were investigated by RNA sequencing.

Results: TGF-β1 activated EZH2 expression in CFs. Treatment with EPZ-6438 (5 μM) and EZH2 siRNA considerably suppressed corneal myofibroblast activation and ECM protein synthesis in CFs induced by TGF-β1 when compared to the control group. EPZ-6438 (5 μM) suppressed cell migration and gel contraction in CFs. RNA sequencing results revealed that antifibrotic genes were activated after EZH2 inhibition to suppress corneal myofibroblast activation.

Conclusion: Inhibition of EZH2 suppresses corneal myofibroblast activation and ECM protein synthesis, and could serve as a novel therapeutic target for preventing corneal scarring.

Keywords: Corneal fibroblast; EZH2; Fibrosis; Transcriptomics.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Cell Movement
Cells, Cultured
Cornea / metabolism*
Cornea / pathology
Corneal Diseases / genetics
Corneal Diseases / pathology
Corneal Diseases / therapy*
Disease Models, Animal
Enhancer of Zeste Homolog 2 Protein / antagonists & inhibitors*
Enhancer of Zeste Homolog 2 Protein / biosynthesis
Female
Gene Expression Regulation*
Humans
Male
Mice
Mice, Inbred C57BL
Myofibroblasts / metabolism*
Myofibroblasts / pathology
RNA / genetics*

Substances

RNA
Enhancer of Zeste Homolog 2 Protein