Mechanisms underlying early-stage changes in cellular function of optic nerve head astrocytes and retinal ganglion cells during oxidative stress as novel targets for neuro- and glioprotection

Abstract

Purpose : Exposure of optic nerve head astrocytes (ONHAs) and retinal ganglion cells (RGCs) to oxidative stress can contribute to axon damage, glaucoma-mediated tissue remodeling and additional hallmarks of glaucoma. Insights in the mechanisms underlying oxidative stress-mediated cell damage to both ONHAs and RGCs can form the basis for strategies to protect both neurons and glia, thereby preventing the progression of glaucoma and related disorders. The study was designed to test the hypothesis that both ONHAs and RGCs respond to oxidative stress with altered proteolytic cleavage of tau protein contributing to cellular degeneration, which can be prevented by treatment with 17β-estradiol.

Methods : Rat ONHAs and RGCs were isolated and cultured. Cells were exposed to oxidative stress using tert-butyl hydroperoxide (tBHP) modeling extracellular oxidative stress as it occurs in the glaucomatous retina and ONH due to inflammation. We combined this model with an intervention approach, pre-treatment of cells with 17β-estradiol. Dephosphorylation of tau protein, tau protein cleavage and estrogen-dependent neuro- and glioprotective signaling proteins were measured using immunoblotting and immunocytochemistry. Changes in caspase activity, cellular viability and estrogen-dependent intracellular signaling were measured using fluorimetric assays.

Results : Exposure of cells to tBHP-mediated oxidative stress generated significantly elevated activation of caspase-3, dephosphorylation of tau at Ser⁴²², and tau protein cleavage associated with the formation of neurofibrillary tangles. 17β-estradiol treatment prevented these degenerative changes and oxidative stress-mediated increases in ONHA and RGC degeneration and apoptosis.

Conclusions : Oxidative stress triggers distinct pathways of cellular damage, which potentially result in decreased cell function and can ultimately activate apoptosis in ONHAs and RGCs. In addition, we provide experimental evidence for effective prevention of the activation of these degenerative signaling pathways by 17β-estradiol not only as a potentially therapeutically relevant compound, but also as a means to accelerate the discovery of biologically relevant target engagement relevant for the protection of ONHA function and for the prevention of neurodegeneration in glaucoma and related disorders.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.