Date Published: June 12, 2019
Publisher: Public Library of Science
Author(s): Michel Desjarlais, Jose Carlos Rivera, Isabelle Lahaie, Gaël Cagnone, Maëlle Wirt, Samy Omri, Sylvain Chemtob, Jing Chen.
Ischemic retinopathies (IRs) are leading causes of visual impairment. They are characterized by an initial phase of microvascular degeneration and a second phase of aberrant pre-retinal neovascularization (NV). microRNAs (miRNAs) regulate gene expression, and a number play a role in normal and pathological NV. But, post-transcriptional modulation of miRNAs in the eye during the development of IRs has not been systematically evaluated.
Using Next Generation Sequencing (NGS) we profiled miRNA expression in the retina and choroid during vasodegenerative and NV phases of oxygen-induced retinopathy (OIR).
Approximately 20% of total miRNAs exhibited altered expression (up- or down-regulation); 6% of miRNA were found highly expressed in retina and choroid of rats subjected to OIR. During OIR-induced vessel degeneration phase, miR-199a-3p, -199a-5p, -1b, -126a-3p displayed a robust decreased expression (> 85%) in the retina. While in the choroid, miR-152-3p, -142-3p, -148a-3p, -532-3p were upregulated (>200%) and miR-96-5p, -124-3p, -9a-3p, -190b-5p, -181a-1-3p, -9a-5p, -183-5p were downregulated (>70%) compared to controls. During peak pathological NV, miR-30a-5p, -30e-5p and 190b-5p were markedly reduced (>70%), and miR-30e-3p, miR-335, -30b-5p strongly augmented (by up to 300%) in the retina. Whereas in choroid, miR-let-7f-5p, miR-126a-5p and miR-101a-3p were downregulated by (>81%), and miR-125a-5p, let-7e-5p and let-7g-5p were upregulated by (>570%) during NV. Changes in miRNA observed using NGS were validated using qRT-PCR for the 24 most modulated miRNAs. In silico approach to predict miRNA target genes (using algorithms of miRSystem database) identified potential new target genes with pro-inflammatory, apoptotic and angiogenic properties.
The present study is the first comprehensive description of retinal/choroidal miRNAs profiling in OIR (using NGS technology). Our results provide a valuable framework for the characterization and possible therapeutic potential of specific miRNAs involved in ocular IR-triggered inflammation, angiogenesis and degeneration.
Ischemic retinopathies (IRs) such as retinopathy of prematurity (ROP) and diabetic retinopathy (DR) remain the leading causes of visual impairment and blindness worldwide1. IRs are characterized by an initial phase of microvascular degeneration followed by ischemia and a subsequent phase of pathological pre-retinal neovascularization (NV). Several mechanisms that control microvascular development and vascular repair are altered during IRs [1,2] including those related to the expression of microRNAs (miRNAs), a family of non-coding RNAs (20–25 nucleotides) involved in post-transcriptional regulation of genes [3,4]. miRNAs regulate a wide range of physiological and pathological process [5,6]. miRNAs can repress their targets by inhibiting protein translation or by degrading specific mRNA with a perfectly complementary target binding sequence (miRNA/mRNA) . It is estimated that miRNAs negatively regulate the expression of more than 60% of genes involved in various cellular processes such as oxidative stress, apoptosis/survival, autophagy, inflammation, cell migration, proliferation and growth . As anticipated, the level of miRNA expression differs between cells types and pathological states [5–7], suggesting a critical role of miRNAs in the progression of various diseases including cancer , cardiovascular diseases , degenerative diseases , and retinopathies [3,10].
Although miRNAs are recognized as critical factors in the regulation of numerous biological and cellular processes such as growth, apoptosis, inflammation, metabolism and angiogenesis [3–6], their specific participation in IRs remains to be defined. With the intention of providing a broad scope miRNA profile in different phases of OIR using a sensitive and accurate approach, we conducted the present study using NGS [14,15]. We identified a number of miRNAs in retina and choroid with altered expression during the different phases of OIR (relative to control). We found around 1000 miRNAs expressed in retinal and choroidal tissues, but we focused our attention on the most abundant miRNAs (≥1000 RPM) that were modulated by more than 20% in OIR. Our results showed around 6% (60–69 miRNA) of the total of miRNAs highly expressed in retina and choroid. This range was very similar to a previous study that evaluated the modulation pattern of miRNAs in plasma of human patients with stage 3 ROP , and in retina of OIR mice model . RT-PCR evaluation of the 24 most modulated miRNAs validated the changes observed by NGS.