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Anuprita Ghosh, Swaminathan Sethu, Ganesh Ram Sahu, santosh gopi krishna gadde, Naresh Kumar Yadav, Rupesh Agrawal, Debasish Sinha, Arkasubhra Ghosh; A negative feedback loop between Angiogenin and Telomerase in AMD pathogenesis. Invest. Ophthalmol. Vis. Sci. 2018;59(9):358.
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© ARVO (1962-2015); The Authors (2016-present)
Age related macular degeneration(AMD) is characterised by dysfunction of retinal pigmented epithelium as well as neovascularisation. Telomerase(TERT) affects age associated tissue health by maintaining telomere length and controlling key intracellular signalling pathways. Angiogenin(ANG) regulates proliferation, angiogenesis and extracellular matrix modulation. Hence, we investigated molecular inter-regulation of these factors in AMD.
Patient samples were obtained after written patient consent and approval by the Institutional Ethics Committee. Aqueous humor (AH) samples from 87 controls and 78 AMD subjects were analysed for levels of ANG, VEGF and an additional 12 soluble factors by flow cytometry. ARPE19 cells were used to analyse secretion of ANG and VEGF under hypoxic and inflammatory stress. Modulation of TERT and ANG was done by overexpression or shRNA constructs. Telomere status and TERT activity were measured by TRAP (telomere repeat amplification protocol) and RTL (relative telomere length) assays. Gene expression in cells and PBMCs from patients was done by QPCR.
AMD patient AH had significantly higher levels of ANG (3951+385pg/ml) and VEGF (116+29pg/ml) compared to controls (2852+245pg/ml and 22+2.5pg/ml respectively). Hypoxic stress induced ANG, VEGF and TERT by 3.9; 15.3 and 4.0 folds, respectively in ARPE19. Additionally, inflammatory stimulus induced ANG and VEGF by 2.9 and 3.9 folds, respectively. ANG stimulation reduced TERT expression (0.23+0.09) but induced Akt activation and VEGF secretion(3.7 fold) in cells. However, overexpression of TERT reduced ANG levels to 0.5 fold. rhANG and TERT inhibitor (MST-312) reduced TERT activity (0.36 fold in normoxia; 0.58 fold in hypoxia). TRAP or RTL assays from AMD patients and age matched controls did not show significant differences. However, there was an inverse correlation of ANG and TERT levels in PBMCs of AMD patients, but not in controls.
Significantly high ANG levels in patients and its inverse regulation by TERT indicates a negative transcriptional feedback loop. Lack of telomere length differences in AMD patients suggests a non-canonical transcriptional role of TERT in AMD. Stress induced elevation of ANG levels via Akt phosphorylation and TERT reduction could be a novel mechanism in AMD pathology.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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