MIR182 and Ocular Hypertension.

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Abstract Despite the significant research effort of microRNAs (miRNAs) in the pathogenesis of glaucoma, their specific roles in the aqueous outflow pathway remain unclear. Lowering intraocular pressure (IOP) is the only clinical treatment for primary open-angle glaucoma (POAG), no matter whether the patients have high (IOP > 21 mmHg) or normal-tension glaucoma (IOP ≤ 21 mmHg). Significant achievements have been made in understanding the regulation of IOP and identifying therapeutic targets to lower IOP. We and others have identified the association of rs76481776 in the MIR182 gene with POAG. However, it remains unknown how miR-182-5p contributes to POAG risk. Without such knowledge, the development of miRNA-based glaucoma therapy will likely remain difficult. Our overall objective is to determine how elevated expression of miR-182-5p contributes to POAG with high IOP. The risk allele A of POAG-associated rs76481776 with POAG has been reported to increase mature miR-182-5p expression in vitro, suggesting that elevated miR-182-5p expression may contribute to POAG. We identified 2-fold higher miR-182-5p expression in glaucomatous aqueous humor compared to controls. Our central hypothesis is that elevated expression of miR-182-5p affects IOP by regulating the cellular functions of human trabecular meshwork (TM) and Schlemm's canal (SC) endothelial cells. Human TM cells with induced premature senescence have 7-9 fold higher miR-182-5p expression, and overexpression of miR-182-5p in these HTM cells leads to partial cellular senescence. miR-182-5p has been shown to target genes in many pathways, such as CHEK2, FOXO1, MTSS1, and CYLD. The expression of miR-182-5p could be induced by TGF-β treatment in cancer cells, leading to prolonged NF-κB activation, while TGF-β related pathways have been shown to play essential roles in POAG and fibrosis. The rationale for this proposal is that, once we determine how elevated miR-182-5p expression affects the cellular functions of human TM and SC, certain genes or miRNAs could be manipulated either up or down pharmacologically, resulting in new and innovative approaches to lower IOP and to delay/prevent the progression of glaucoma. In Aim 1, we will identify molecular targets of miR-182-5p in primary human TM cells in vitro. Human TM cells will be subject to cyclic mechanical stretch and tissue train 3D cell culture with uniaxial tissue stretch. We will perform whole proteomics profiling and strand-specific RNA-Seq analysis to identify miR-182-5p target genes. In Aim 2, we will determine the impact of miR-182-5p overexpression in the TM and SC on outflow facility and IOP levels in vivo using transgenic mice. Upon successful completion, we expect to identify the specific molecular targets of miR-182- 5p in human TM and SC to lower IOP more effectively in glaucoma patients.