MicroRNA-145 regulates human corneal epithelial differentiation

Area of Science:

• Ophthalmology
• Molecular Biology
• Stem Cell Biology

Background:

• Epigenetic factors, including microRNAs (miRNAs), are key regulators of stem cell self-renewal and differentiation.
• Human limbal-peripheral corneal (LPC) epithelia harbor corneal epithelial progenitor cells (CEPCs) and transit amplifying cells crucial for corneal maintenance.

Purpose of the Study:

• To investigate miRNAs expressed in LPC epithelia and their function in corneal epithelium.
• To identify the role of specific miRNAs in the regulation of corneal epithelial progenitor cells and corneal integrity.

Main Methods:

• Small RNA sequencing and microarray analysis of human limbal-peripheral corneal (LPC) and central corneal (CC) epithelia.
• Quantitative polymerase chain reaction (qPCR) and locked nucleic acid-based in situ hybridization for miRNA validation.
• Lentiviral transfection of CEPCs with miR-145 and subsequent organotypic culture and gene expression analysis.
• Luciferase reporter assays to confirm miRNA-target interactions.

Main Results:

• Differential expression of 18 miRNAs between LPC and CC epithelia was identified, with the miR-143/145 cluster strongly expressed in LPC.
• miR-145 transfection in CEPCs led to defective epithelium, altered cytokeratin and connexin expression, and changes in stem cell markers.
• miR-145 modulated global gene expression, up-regulating immune response genes and down-regulating epithelial development and stem cell maintenance genes.
• miR-145 was confirmed to suppress integrin β8 (ITGB8) expression in corneal epithelial cells and CEPCs.

Conclusions:

• The miR-143/145 cluster is expressed in human corneal epithelium.
• miR-145 is a critical regulator of corneal epithelium formation and maintenance of epithelial integrity.
• Targeting of ITGB8 by miR-145 is a key mechanism in its role in corneal homeostasis.