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Investigative ophthalmology & visual science

MicroRNA Expression Profile and the Role of miR-204 in Corneal Wound Healing.


PMID 26047168

Abstract

MicroRNAs (miRNAs) are endogenous short chain (∼ 22-nucleotide) noncoding RNAs that inhibit protein translation through binding to target mRNAs. Recent studies have implied that miRNAs play a regulatory role in corneal development. Here we profile their involvement in corneal epithelial renewal, develop an miRNA-target network that affects wound healing outcome, and investigate the function of miR-204 in this response. NanoString nCounter technology and bioinformatics analyzed miRNA expression levels and their targets during mouse corneal epithelial wound healing. Real-time RT-PCR was performed to detect miR-204 expression in mouse corneal epithelium. Human corneal epithelial cells (HCECs) were transfected with miR-204 using transfection reagent. MTS (3-[4,5-dimethylthiazol-2-yl]-5-[3-carboxymethoxyphenyl]-2-[4-sulfophenyl]-2H-tetrazolium, inner salt) and a scratch wound-healing assay evaluated the effects of miR-204 expression on HCEC proliferation and migration, respectively. Cell cycle analysis was performed by flow cytometry. Expression of sirtuin 1 (SIRT1) was determined by Western blot analysis. Fifteen miRNAs were dramatically downregulated, whereas 14 other miRNAs were markedly upregulated during corneal wound healing. Expression of miR-204 fell the most during this process. Transfection of miR-204 into HCECs led to a significant decline in cell proliferation and induced cell cycle G1-arrest. Furthermore, in these cells, miR-204 also inhibited migration. Sirtuin 1 was confirmed as a target of miR-204. During mouse corneal epithelial wound healing, a complex miRNA-gene network was resolved that is modulated by changes in miR-204 expression. Downregulation of this miRNA appears to be an essential response to injury since its decline promotes human corneal epithelial cell proliferation and migration. Therefore, miR-204 could be a biomarker of this process.