470-P: Mitochondrial Long Noncoding RNA and the Compromised Electron Transport Chain Machinery in Diabetic Retinopathy

Abstract

Purpose: Diabetic retinopathy has a complex pathogenesis, and mitochondrial damage is considered to play a central role. Diabetes dysfunctions the mitochondria and damages their DNA (mtDNA) , decreasing the transcription of mtDNA-encoded genes including Cytochrome B (CYTB) , which is important for the functioning of complex III of the electron transport chain system (ETC) . This propagate the vicious cycle of free radicals. Diabetes also decreases mtDNA-encoded long non-coding RNA Cytochrome B (LncCytB) . Although long noncoding RNAs have no open reading frame for translation, they can bind to the DNA or RNA in a sequence specific manner and alter the gene expression. Our aim was to investigate the role of LncCytB in regulating CYTB-ETC system in diabetic retinopathy. Methods: Human retinal endothelial cells, incubated in 5mM (normal) or 20mM (high) D-glucose, were used to quantify the expressions of LncCytB (by strand-specific PCR) and CYTB (qRT-PCR) . The binding of LncCytB at CYTB promoter was determined by Chromatin isolation by RNA Purification method. Role of LncCytB in regulating CYTB expression and mitochondrial damage was performed in the cells overexpressing LncCytB. Results: Compared to cells in normal glucose, LncCytB and CYTB levels were decreased by over 50% in the cells exposed to high glucose, the binding of LncCytB at CYTB promoter was reduced by ∼40% and the activity of the complex III was inhibited. Overexpression of LncCytB ameliorated glucose-induced decrease in CYTB gene transcripts, and increase in mtDNA damage and in mitochondrial reactive oxygen species. Conclusions: Due to decreased interactions between LncCytB-CYTB in hyperglycemic milieu, CYTB transcripts are decreased, and this compromises the ETC system, resulting in increased mitochondrial ROS. Thus, regulating LncCytB levels in diabetes could halt the self-propagating cycle of free radicals, and prevent the development of diabetic retinopathy. Funding EY014370, EY017313, and EY022230