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Amany M Tawfik, Khaled Elmasry, Yutao Liu, Mohamed Al-Sayed Al-Shabrawey; Epigenetic modifications associated with Hyperhomocysteinemia; potential role in Blood Retinal Barrier dysfunction. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3566. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To study Homocysteine (Hcy) induced epigenetic modifications as potential mechanisms of blood retinal barrier (BRB) dysfunction.
Retinal tissues isolated from 3 weeks old mice with elevated levels of Hcy due to lack of the enzyme cystathionine-β-synthase (cbs+/- and cbs-/-) and wild type (cbs+/+) mice as well as Human Retinal Endothelial cells (HRECS) and Human retinal pigmented epithelial cell line (ARPE-19) treated with or without Hcy (20,50 and 100mM) for 24 hours were subjected to evaluation of 1)- Histone deacetylases (HDAC) using HDAC Activity Colorimetric Assay Kit 2)- DNA methylation using EpiQuik DNA Methyltransferase (DNMT) Activity/Inhibition Assay Kit . 3) - miRNA analysis from retinas of mice with elevated Hcy level (cbs mice) was done, briefly, 12 mouse retinal RNA samples (4 cbs+/+, 4 cbs+/- and 4 cbs-/-) were assayed using Affymetrix GeneChip miRNA 3.0 array. Expression data was analyzed using Partek Genomics Suite. List of differentially expressed miRNAs were generated and imported into Ingenuity Pathway Analysis (Qiagen) for detection of predicted target genes and associated pathways.
Hcy induced significant increase in HDAC and DNMT activity in HRECs, ARPE-19, cbs+/- and cbs-/- mice retina. In addition miRNA profiling of cbs+/- and cbs-/- detected 127 miRNAs in cbs+/- (49 downregulated and 78 upregulated) and 39 miRNAs in cbs-/- (20 miRNAs downregulated and 19 upregulated) were statistically significant and differentially expressed in comparison to cbs+/+. miRNAs pathway analysis showed their involvement in ER stress, oxidative stress, inflammation, Hypoxia and angiogenesis pathways.
Hcy induced Epigenetic modifications could represent novel biomarkers or therapeutic targets for retinal diseases associated with elevated Hcy such as age related macular degeneration and diabetic retinopathy.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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