@article{4b142667e7cf4c408c73729e62b40f3c,
title = "Hyperglycemia-induced miR182-5p drives glycolytic and angiogenic response in Proliferative Diabetic Retinopathy and RPE cells via depleting FoxO1",
abstract = "Purpose: Diabetic Retinopathy (DR) is associated with metabolic dysfunction in cells such as retinal pigmented epithelium (RPE). Small molecular weight microRNAs can simultaneously regulate multiple gene products thus having pivotal roles in disease pathogenesis. Since miR182-5p is involved in regulating glycolysis and angiogenesis, two pathologic processes of DR, we investigated its status in DR eyes and in high glucose model in vitro. Method: ology: Total RNA was extracted from vitreous humor of PDR (n = 48) and macular hole (n = 22) subjects followed by quantification of miR182-5p and its target genes. ARPE-19 cells, cultured in DMEM under differential glucose conditions (5 mM and 25 mM) were used for metabolic and biochemical assays. Cells were transfected with miRNA182 mimic or antagomir to evaluate the gain and loss of function effects. Results: PDR patient eyes had high levels of miR182-5p levels (p < 0.05). RPE cells under high glucose stress elevated miR182-5p expression with altered glycolytic pathway drivers such as HK2, PFKP and PKM2 over extended durations. Additionally, RPE cells under high glucose conditions exhibited reduced FoxO1 and enhanced Akt activation. RPE cells transfected with miR182-5p mimic phenocopied the enhanced basal and compensatory glycolytic rates observed under high glucose conditions with increased VEGF secretion. Conversely, inhibiting miR182-5p reduced Akt activation, glycolytic pathway proteins, and VEGF while stabilizing FoxO1. Conclusion: Glycolysis-associated proteins downstream of the FoxO1-Akt axis were regulated by miR182-5p. Further, miR182-5p increased expression of VEGFR2 and VEGF levels, likely via inhibition of ZNF24. Thus, the FoxO1-Akt-glycolysis/VEGF pathway driving metabolic dysfunction with concurrent angiogenic signaling in PDR may be potentially targeted for treatment via miR182-5p modulation.",
keywords = "Angiogenesis, Diabetic retinopathy, Glycolysis, microRNA, miR182-5p",
author = "Shaika Shanbagh and Gadde, {Santosh Gopikrishna} and Rohit Shetty and Stephane Heymans and Abilash, {V. G.} and Chaurasia, {Shyam S.} and Arkasubhra Ghosh",
note = "Funding Information: This work was supported by NNF (Narayana Nethralaya Foundation) and funding by National Eye Institute, NIH, Bethesda , MD grant numbers RO1EY029795 to SSC. The funders had no role in the study design, data collection, and analysis, decision to publish, or preparation of the manuscript. Funding Information: Therefore, we investigated the downstream impact of miR182-5p mediated regulation of FoxO1, Akt, and VEGF. Under normoglycemic conditions, augmenting miR182-5p by transfecting a mimic led to the inhibition of FoxO1, activation of Akt and increased VEGFR2 levels compared to control, thereby phenocopying the effects of high glucose in RPE cells and patient vitreous humor. Furthermore, we also observed elevated expression of angiogenic and glycolytic factors, including GLUT1, HK2, PKM2, PFKP, LDHA, PDHA, VEGF, ZnF24, and VEGFR2 in miR182-5p mimic transfected cells, supporting the results obtained in the high glucose model. Previous studies suggest that GLUT1 overexpression leads to a higher level of VEGF in retinal endothelial cells as well as diabetic kidney disease (Cai and Boulton, 2002; Pf{\"a}fflin et al., 2006). Conversely, inhibiting miR182-5p under high glucose conditions attenuated Akt phosphorylation, VEGF and glycolytic gene levels while rescuing FoxO1 and ZnF24 expression. Importantly, miR182-5p inhibition significantly reduced VEGF secretion from the RPE cells under hyperglycaemia. Thus, ablation of miR182-5p may have a potential inhibitory role in controlling the pathologic molecular changes in DR.This work was supported by NNF (Narayana Nethralaya Foundation) and funding by National Eye Institute, NIH, Bethesda, MD grant numbers RO1EY029795 to SSC. The funders had no role in the study design, data collection, and analysis, decision to publish, or preparation of the manuscript. Publisher Copyright: {\textcopyright} 2023 Elsevier Ltd",
year = "2024",
month = jan,
day = "1",
doi = "10.1016/j.exer.2023.109713",
language = "English",
volume = "238",
journal = "Experimental Eye Research",
issn = "0014-4835",
publisher = "Academic Press",
}