MicroRNAs as unique biomarkers and potential therapeutic targets for diabetic retinopathy

Abstract

Purpose : To investigate the circulatory microRNA(miRNA) profiles of aqueous, vitreous, and plasma in order to identify biomarkers in aqueous, vitreous and plasma, that could be used for targeted therapy.

Methods : Aqueous, vitreous and plasma samples were collected from a total of 27 patients - 11 controls and 16 patients with diabetes mellitus(DM) undergoing vitreoretinal surgery: DM-Type I with proliferative diabetic retinopathy (DMI-PDR), DMType II with PDR(DMII-PDR) and DMType II with nonproliferative DR(DMII-NPDR). Dysregulated miRNAs were identified by profiling on Affymetrix GeneChip miRNA 3.0 microarrays, followed by data analysis on Expression Console(EC), Transcriptome Analysis Console(TAC), and IPA software. Animal eye imaging was done using SLO/OCT and Fluorescein angiography using Micron IV. Retinal flatmount vasculature was stained with Isolectin-GS-B4-AlexaFluor 647, and analyzed by confocal imaging and Fiji.

Results : Our comparison of the circulatory miRNA population of ocular fluids and plasma showed that out of a total of 847 human miRNA probes on the Affymetrix GeneChip miRNA 3.0 we found common and unique miRNAs for both aqueous and vitreous, dependent on the DM type and presence of retinopathy. Dysregulation of miRNAs in aqueous generally do not appear to be a good representative of the miRNA abundance in vitreous, or plasma, although we did identify some common biomarkers: let-7b, miR-320b, miR-762 and miR-4488. Additionally, each of the DR subtypes showed a set of miRNA that is uniquely dysregulated in each fluid, for example in aqueous samples for DMII-NPDR it was miR-455-3p, for DMII-PDR was miR-296, and for DMI-PDR it was miR-3202. Pathway analysis identified TGF-beta and VEGF pathways as the common targets for miRNAs dysregulated in DR aqueous and vitreous. In order to identify future therapeutic targets, we have used an in vitro angiogenesis assay and in vivo animal model Akimba to test pro- or anti-angiogenic activity of candidate miRNA.

Conclusions : The comparative profiling of circulatory miRNAs in aqueous, vitreous, and plasma showed that a small number of circulatory miRNAs displayed differential presence in controls vs. diabetic retinopathy. A pattern is emerging of sets of miRNA that are common or uniquely dysregulated in the blood plasma or ocular fluids of DR subtypes, offering promise for the use of ocular fluids and plasma for identifying diagnostic and therapeutic targets.

This is a 2020 ARVO Annual Meeting abstract.