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miR-365 promotes diabetic retinopathy through inhibiting Timp3 and increasing oxidative stress
Juan Wang1, Jieping Zhang1, Xin Chen1
1Department of Ophthalmology of Shanghai Tenth People's Hospital, Tongji Eye Institute, Tongji University School of Medicine, Shanghai, China; Laboratory of Clinical Visual Science, Department of Regenerative Medicine, Stem Cell Research Center, Tongji University School of Medicine, Shanghai, China.
Abstract:
miRs play critical roles in oxidative stress-related retinopathy pathogenesis. miR-365 was identified in a previously constructed library from glyoxal-treated rat Müller cell. This report explores epigenetic alterations in Müller cells under oxidative stress to develop a novel therapeutic strategy. To examine the miR-365 expression pattern, in situ hybridization and quantitative RT-PCR were performed. Bioinformatical analysis and dual luciferase report assay were applied to identify and confirm target genes. Streptozotocin (STZ)-treated rats were used as the diabetic retinopathy (DR) model. Lentivirus-mediated anti-miR-365 was delivered subretinally and intravitreally into the rats' eyes. The functional and structural changes were evaluated by electroretinogram (ERG), histologically, and through examination of expression levels of metallopeptidase inhibitor 3 (Timp3), glial fibrillary acidic protein (Gfap), recoverin (Rcvrn) and vascular endothelia growth factor A (Vegfa). Oxidative stress factors and pro-inflammatory cytokines were analyzed. miR-365 expression was confirmed in the glyoxal-treated rat Müller cell line (glyoxal-treated rMC-1). In the retina, miR-365 mainly localized in the inner nuclear layer (INL). The increased miR-365 participated in Müller cell gliosis through oxidative stress aggravation, as observed in glyoxal-treated rMC-1 and DR rats before 6 weeks. Timp3 was a target and negatively regulated by miR-365. When miR-365 was inhibited, Timp3 expression was upregulated, Müller cell gliosis was alleviated, and retinal oxidative stress was attenuated. Visual function was also partially rescued as detected by ERG. miR-365 was found to be highly expressed in the retina and the abnormality of miR-365/Timp3 pathway is closely related to the pathology, like Müller gliosis, and the visual injury in DR. The mechanism might be through oxidative stress, and miR-365/Timp3 could be a potential therapeutic target for treating DR.
Insights
In diabetic retinopathy, inhibiting miR-365 reduces oxidative stress and Müller cell gliosis by upregulating Timp3, partially restoring visual function. This suggests miR-365/Timp3 pathway as a therapeutic target.
Area of Science:
- Ophthalmology
- Molecular Biology
- Cell Biology
Background:
- MicroRNAs (miRs) are implicated in oxidative stress-related retinopathy.
- Müller cells play a critical role in retinal homeostasis and disease.
- Oxidative stress contributes to the pathogenesis of diabetic retinopathy (DR).
Purpose of the Study:
- To investigate the role of miR-365 in oxidative stress-induced Müller cell alterations.
- To explore the miR-365/Timp3 pathway as a potential therapeutic target for DR.
- To develop a novel therapeutic strategy targeting epigenetic modifications in Müller cells.
Main Methods:
- miR-365 expression analysis using in situ hybridization and quantitative RT-PCR.
- Target gene identification via bioinformatical analysis and dual luciferase reporter assay.
- Diabetic retinopathy model in Streptozotocin (STZ)-treated rats; lentivirus-mediated anti-miR-365 delivery; functional and structural evaluation using ERG, histology, and molecular markers (Timp3, Gfap, Rcvrn, Vegfa).
Main Results:
- miR-365 expression was confirmed in glyoxal-treated rat Müller cells and localized in the inner nuclear layer of the retina.
- Increased miR-365 aggravated Müller cell gliosis and oxidative stress in vitro and in vivo (DR model).
- Metallopeptidase inhibitor 3 (Timp3) was identified as a direct target negatively regulated by miR-365. Inhibition of miR-365 upregulated Timp3, alleviated Müller cell gliosis, attenuated retinal oxidative stress, and partially rescued visual function.
Conclusions:
- The miR-365/Timp3 pathway is closely linked to Müller cell gliosis and visual impairment in diabetic retinopathy.
- miR-365 contributes to DR pathology through oxidative stress and Müller cell gliosis.
- Inhibition of miR-365 represents a promising therapeutic strategy for treating diabetic retinopathy.
