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Arupratan Das, Claire Wenger, Cindy Berlinicke, Nicholas Marsh-Armstrong, Donald J Zack; Switching from the proteasomal to the lysosomal pathway for MQC during human RGC differentiation is essential for RGC survival. Invest. Ophthalmol. Vis. Sci. 2019;60(9):654.
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© ARVO (1962-2015); The Authors (2016-present)
Mitochondrial quality control (MQC), which helps to maintain healthy mitochondria, is essential for cell survival, and defects in the clearance of damaged mitochondria has been implicated in various neurodegeneration. To explore the possible role of MQC in contributing to retinal ganglion cell (RGC) degeneration in glaucoma and other optic neuropathies, we have been investigating the molecular pathways of MQC in human RGCs.
We have developed methods to differentiate and purify human RGCs from pluripotent stem cells, and the gene expression patterns and electrophysiology properties of the resulting cells match well with native RGCs. To study RGC degeneration, we have developed a fluorescence plate reader-based assay that measures both RGC death and cellular apoptosis. For mitochondrial degradation, we have developed methods to measure mitochondrial content using qPCR and flow-cytometry-based analysis. We used these techniques to measure relative contributions of the proteasomal and the lysosomal-autophagy pathways in degrading damaged mitochondria and their effect on hRGC viability.
Proteasomal and the lysosomal pathways are the two major pathways for degrading damaged organelles and proteins for maintaining healthy cellular homeostasis. We find that when mitochondria are damaged using the uncoupler carbonyl cyanide m-chlorophenyl hydrazone (CCCP), RGCs more efficiently clear damaged mitochondria than their precursor stem cells. To clear damaged mitochondria, stem cells use both the proteasomal and the lysosomal pathways while RGCs predominantly use the lysosomal pathway. Inhibiting proteasomal or lysosomal pathways acutely causes cell death through apoptosis in stem cells, while such an acute effect is only observed for lysosomal inhibition in RGCs.
Overall, our study suggests that the lysosomal pathway is critical for the clearance of damaged mitochondria and RGC survival. These results suggest that manipulating the lysosomal/autophagic pathways could be therapeutically relevant for enhancing RGC survival in glaucoma and in other forms of optic nerve disease.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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