Genome-wide Expression Profiling and Pathway Analysis in Cyclic Stretched Human Trabecular Meshwork Cells

Abstract Purpose Regulation of intraocular pressure is dependent upon homeostatic responses of trabecular meshwork (TM) cells to mechanical stretch. Despite the important roles of miRNAs in regulating TM function and aqueous outflow, it remains unclear how miRNA and their target genes interact in response to physiological cyclic mechanical stretch. We aimed to identify differentially expressed miRNAs and their potential targets in human TM cells in response to cyclic mechanical stress. Methods Monolayers of TM cells from non-glaucomatous donors (n=3-6) were cultured in the presence or absence of 15% mechanical stretch, 1 cycle/s, for 6 or 24-hours using computer-controlled Flexcell Unit. We profiled the expression of 800 miRNAs using NanoString Human miRNA assays and identified differentially expressed miRNAs using the Bioconductor Limma package. We identified differentially expressed genes using Operon Human Oligo Arrays with GeneSpring software. Pathway analysis with WebGestalt identified stretch-related pathways. We used Integrative miRNA Target Finder from Ingenuity Pathway Analysis to identify potential miRNA-mRNA regulations. Results We identified 540 unique genes and 74 miRNAs with differential expression in TM cells upon cyclic mechanical stretch. Pathway analysis indicated the significant enrichment of genes involved in Wnt-signaling, receptor protein serine/threonine kinase signaling, TGF-β pathway, and response to unfolded protein. We also identified several miRNA master regulators, including miR-19b-3p and miR-93-5p, which may act as switches to control several mechano-responsive genes. Conclusions This study suggests that cyclic mechanical stress of TM cells triggers alterations in the expression of both mRNAs and miRNAs implicated in glaucoma-associated pathways.