Purpose:
DNA damage response (DDR) plays crucial roles in the fates of cells and the mechanism of DDR in neuro-degenerative diseases are focused. But little is known of the role of DDR in the loss of retinal ganglion cells (RGCs) in glaucoma. Here, we investigate the role DDRs play in RGC death under normoxic or hypoxic conditions in rat RGCs in vitro and in vivo.
Methods:
Using primary cultured rat RGCs under normoxic or hypoxic conditions, the number of 53BP1 nuclear foci, a marker for DNA double strand breaks (DSB) and DNA repair, and the number of apoptotic cells were counted. Ataxia telangiectasia mutated (ATM) protein is known to play a pivotal role in DDR by phosphorylating apoptosis-related proteins at cell cycle checkpoints. Therefore, RGC-5 cells were treated with the ATM inhibitor, KU55933 and caffeine, and the numbers of 53BP1 foci and annexin V-positive cells, an apoptosis marker, were counted. We also counted 53BP1 foci in RGCs in a rat optic nerve (ON) crush model.
Results:
Hypoxia significantly suppressed 53BP1 focus formation in cultured rat RGCs. 53BP1 positive cells were decreased from 17.9 % to 3.7 % (p < 0.05). Correspondingly, RGCs with more than five foci were decreased from 23.8 % to 10.9 % (p < 0.01) in a rat ON crush model. The ATM inhibitors significantly decreased the number of 53BP1 foci and increased apoptosis in hypoxic as well as normoxic culture conditions (p < 0.05).
Conclusions:
These findings support that both ATM inhibition and hypoxia down-regulates 53BP1 focus formation and facilitates apoptosis of RGCs. This is the first demonstration of the possible roles for ATM-dependent DDR in hypoxia-induced RGC death.
Keywords: apoptosis/cell death • ganglion cells • hypoxia