Mariana Aparecida Rosales, Magali Saint-Geniez; Loss of PGC-1α function in RPE induces EMT and promotes retinal degeneration. Invest. Ophthalmol. Vis. Sci. 2018;59(9):1185.
Purpose : Epithelial-to-mesenchymal transition (EMT) of retinal pigment epithelial (RPE) cells is central to numerous degenerative ocular diseases including proliferative vitreoretinopathy and AMD. We have previously shown that PGC-1α, a core regulator of oxidative metabolism, is involved in RPE differentiation and that its silencing triggers rapid mitochondrial and autophagy dysfunctions. As metabolic defects are known to promote EMT, we examined the consequences of PGC-1α deletion on RPE transdifferentiation in vivo and in vitro.
Methods : PGC1α-deficient RPE cells were generated by stable expression of shPGC-1α in ARPE-19. shPGC-1α and shControl ARPE-19 cells were maturated and maintained in serum free condition for up to 21 days. Change in EMT-associated genes was analyzed by qPCR and immunohistochemistry. RPE-specific PGC-1s KO mice were generated by subretinal injection of AAV2/1-Cre in 8 weeks old C57Bl/6J;α,β double floxed mice (n=10; 6 females, 4 males). AAV2/1-GFP injected mice were used as controls (n= 9; 4 females, 5 males). Specificity and efficiency of Cre expression was confirmed by histology and qPCR. Two and four months after AAV-delivery, morphological and functional changes were evaluated by fundus, OCT, ERG and electron microscopy.
Results : EMT in PGC-1α silenced cells was first evidenced at day 7 by the reorganization of the fibrillar intermediate filaments into dense prenuclear coils. These morphological changes were associated with progressive loss of tight-junctions and cell-contact inhibition at day 14. Robust upregulation of the EMT-associated genes ZEB1 (p≤0.05), VIM (p≤0.01) and TWIST1 (p≤0.01) was detected at day 21. Similar RPE dedifferentiation was observed 2 and 4 months following RPE-specific deletion of the PGC-1 isoforms in adult mice. RPE anomalies were characterized by mitochondrial defects, pigmentary changes, cellular dissociation and migration. RPE dysfunction was associated with significant retinal degeneration demonstrated by reduced a- (p≤0.001), b- (p≤0.001), and c-waves (p≤0.01) amplitudes from ERG recordings and decreased total retinal thickness (p≤0.0001) measured by OCT.
Conclusions : These results demonstrate that sustained PGC-1α silencing in adult RPE cells induces EMT and subsequent retinal degeneration. Our findings support a role for PGC-1α and metabolic dysfunction in ocular pathologies associated with RPE EMT.