Effect of MiRNA 204-5P Mimics and Lipopolysaccharide-Induced Inflammation on Transcription Factor Levels, Cell Maintenance, and Retinoic Acid Signaling in Primary Limbal Epithelial Cells

Abstract

MicroRNA-204-5p (miR-204-5p) is a critical regulator of differentiation, structural maintenance, and inflammation in limbal epithelial cells (LECs). This study examined the role of miR-204-5p in modulating the gene expression related to transcription factors, cell structure, extracellular matrix remodeling, and retinoic acid signaling under normal and lipopolysaccharide (LPS)-induced inflammatory conditions. Using qPCR, we analyzed the mRNA levels of FOSL2, FOXC1, Meis2, PPARγ, ABCG2, PTGES2, IL-1β, IL-6, KRT3, KRT12, MMP2, MMP9, RARA, RARB, RXRA, RXRB, CRABP2, RBP1, RDH10, ADH7, ADH1A1, FABP5, CYP1B1, and CYP26A1, while changes in protein levels were assessed via Western blot or ELISA. Our data revealed that the overexpression of miR-204-5p reduced the mRNA levels of FOXC1, KRT12, and RDH10 under normal and inflammatory conditions (p ≤ 0.039). Additionally, it decreased FOSL2 and RXRA mRNA under normal conditions (p = 0.006, p = 0.011) and KRT3 and FABP5 mRNA under inflammatory conditions (p = 0.010, p = 0.001). The IL-6 mRNA expression was significantly increased following the LPS treatment in cells overexpressing miR-204-5p (p = 0.029). A protein analysis revealed significant reductions in FOXC1 and KRT3 in the miR-204-5p-transfected cells during LPS-induced inflammation (p = 0.020, p = 0.030). These findings suggest that miR-204-5p modulates genes critical to the differentiation, migration, and inflammatory response of LECs. The modulation of FOXC1 and KRT3 by miR-204-5p highlights these proteins as novel targets under inflammatory conditions.