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Arpita R. Dave, Eunice Sze Yin Ng, Zhichun Jiang, Jane G Hu, Michael B Gorin, Anna Matynia, Linsey Stiles, Roxana A Radu; MeCP2-mediated mitochondrial impairment evidenced in Stargardt disease. Invest. Ophthalmol. Vis. Sci. 2021;62(8):269.
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© ARVO (1962-2015); The Authors (2016-present)
Clinical and basic studies implicate the retinal pigment epithelium (RPE) as the primary site of two related central blinding maculopathies such as AMD and Stargardt disease (STGD1). Links of epigenetics to aging have been proposed but a specific epigenetic-mediated mechanism responsible for the RPE cellular death is yet to be identified. Preliminary data suggest a role of Methyl-CpG-binding protein 2 (MeCP2), an epigenetic factor, in mitochondria dynamics of aged and diseased RPE cells. Using Abca4-/- mice and human STGD1 iPSC-derived RPE cells, we correlated the MeCP2 protein profile and mitochondrial functions. We employed proteomics analysis to identify MeCP2-regulated molecular targets responsible for age-dependent mitochondrial changes.
1-, 3-, and 6-mo old background-matched wild-type (WT) and Abca4-/- mice and human normal and STGD1 iPSC-derived RPE cells homogenates were used to measure the levels of MeCP2 by quantitative immunoblotting (normalized to beta-actin; n=3). By LC-MS proteomics analysis, we determined oxidative phosphorylation (OXPHOS) and glycolysis proteins using 4 mouse RPE sheets (n=3). Mouse and human primary RPE cultures (n=4) were used to evaluate mitochondria functions by Seahorse respirometry. Statistical analysis was done using t-test or One-way ANOVA.
By immunoblotting, we found a ~2- and ~4-fold increased MeCP2 level in 3-mo- and in 6-mo-old respectively when compared to 1-mo-old Abca4-/- RPE cells. Importantly, RPE cell homogenate from 6-mo-old Abca4-/- mice showed ~2-fold higher MeCP2 level vs age-matched WT mice. Analysis of human iPSC-derived RPE grown for 2-mo in culture indicated increased MeCP2 level in STGD1 vs normal cells. Proteomics analysis revealed significantly (p<0.05 to p<0.001) reduced levels of the OXPHOS complexes (I, II, III, and V) and elevated levels of the glycolysis proteins. Consistently, Seahorse respirometry analysis indicated increased glycolytic activity in Abca4-/- vs WT RPE cells.
Our data evidenced an age-dependent MeCP2 protein changes that correlated with altered mitochondrial functions in the RPE cells of the STGD1 models. These studies inform on a specific regulatory role of MeCP2 and its targeted molecular players in the RPE cells that are involved in disease progression. Importantly, modulation of MeCP2 in the RPE cells may potentially rescue the mitochondria deficiency to mitigate RPE functions and prevent blindness.
This is a 2021 ARVO Annual Meeting abstract.
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