Targeting miR-146a to Treat Delayed Wound Healing in Human Diabetic Organ-Cultured Corneas

Limbal epithelial stem cells (LESC) residing at the corneal periphery are largely responsible for maintaining corneal optical transparency by continuously supplying new corneal epithelial cells, which mature during their radial migration to the central cornea. Diabetes mellitus (DM) affects all the structures of the eye including the cornea. Frequent epithelial erosions, delayed wound healing, and microbial infections are common alterations of the diabetic eye that can result in vision loss. MicroRNAs (miRNAs) are short non-coding oligonucleotides that regulate gene expression by repressing translation. Our purpose was to understand the role of miR-146a in the human limbal versus central corneal epithelial compartment in normal and pathological conditions such as diabetes mellitus. Using quantitative real-time PCR (QPCR) we found miR-146a enrichment in the limbal corneal compartment. This miRNA was also expressed at higher levels in the diabetic vs. normal limbus. Cell migration and wound closure were significantly delayed in normal and diabetic primary limbal epithelial cells (LEC) transfected with miR-146a. Cells treated with miR-146a had decreased levels of phosphorylated (activated) p38 and EGFR, mediators of epithelial wound healing. Conversely, inhibition of miR-146a significantly enhanced cell migration in both normal and diabetic primary LEC and in diabetic organ-cultured corneas by nearly 40% vs. scrambled miRNA control, accompanied by increased phosphorylated signaling intermediates. Transfection of miR-146a in cultured LEC resulted in an increased immunoreactivity for putative LEC markers Frizzled-7 and K15, whereas inhibition of miR-146a decreased their expressions. These data suggest that miR-146a plays a role in LEC maintenance at the corneal periphery, and its expression is downregulated during their migration towards the central cornea and accompanying terminal differentiation. Furthermore, abnormal miR-146a upregulation may be an important mechanism of delayed wound healing in the diabetic cornea.