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Terry Gaasterland, Amy Dubinsky, Douglas E Gaasterland, Karl Willert; Study of diseased and normal optic nerve and retina to identify microRNAs involved in glaucomatous optic neuropathy. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):4384.
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We studied microRNA activity in optic nerve in glaucoma, a leading cause of irreversible blindness. Primary open angle glaucoma (POAG) is the most common subtype of glaucoma with visual field loss due to apoptosis of retinal ganglion cells and consequent thinning of the retinal nerve fiber layer and associated optic nerve axon loss. While a number of candidate genes have been connected to glaucoma through genetic studies, none yet explain glaucoma molecular mechanisms. In 2012, the CDKN2B-AS1 region, which harbors CDKN2B (p15) in its first intron, was strongly associated with POAG in a genome-wide association study. Our lab determined that a microRNA binding site in the 3’ untranslated region of CDKN2B is disrupted by a risk SNP. These findings suggested that control of cell cycle progression through pathways that activate CDK4 may have a powerful role in onset and severity of disease.
We surveyed microRNA expression in optic nerve and retina from 2 glaucomatous and 3 normal human donor eyes. Total RNA was extracted from optic nerve, retina, and additional tissues and subjected to high throughput sequencing for small RNA and messenger RNA (mRNA). Reads from small RNA were matched against 2565 known human microRNA sequences, counted, and normalized. We stratified microRNAs by high to low expression and grouped them by tissue expression patterns to compare glaucoma vs. normal and retina vs. optic nerve. Seed sites for binding of microRNA to mRNA were identified computationally.
Several microRNAs were absent in diseased but present in normal optic nerve. One microRNA was absent in normal optic nerve but detected as significantly expressed in diseased tissue. The microRNA targeting CDKN2B was among 43 differentially expressed microRNAs as were several microRNAs targeting POAG genes established through GWAS and exome sequencing. Computational analysis of mRNA seed sites for the 43 microRNAs indicated a regulatory network that ties cell cycle regulation to optic neuropathy.
Forty-three microRNAs were differentially expressed in optic nerve and retina from disease and normal human eyes. Several of their target genes were previously associated with optic neuropathy. The differentially expressed microRNAs and their mRNA targets provide a foundation to define a regulatory network for microRNA modulation of neuron death through apoptosis.
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