Distal nerve degeneration in the cornea is initiated by oxidative stress.

Abstract

Purpose : Apart from its function in perception of stimuli, the sensory innervation of the cornea is important for the maintenance of hydration and homeosthasis of the ocular surface. Corneal nerve degeneration is progressive during normal aging. Aging reduces the ability of mitochondria to get rid of free radicals and oxidative stress. The aim of this work is to study the effect of induced oxidative stress on the corneal sensitive innervation.

Methods : Oxidative stress was induced by intense light exposure to trigeminal ganglion cell cultures from adutl Wistar rats (blue light: 470 nm and 500 lux, 4 hours a day for 10 days). Also, rats were exposed to a source of blue light under the same conditions and corneas and trigeminal ganglia were collected. An equivalent group of animals and cultures were maintained in the dark during light exposure cicles and were used as controls. A group of rats and cultures were treated with antioxidants before light exposure.
Immunofluorescence analysis and qPCR for β tubulin III, tau-p, CaMKII, Hemoxigenase-1, Tomm 20 and Cytochrome C Oxidase, were performed to evaluate oxidative stress and axonal cytoskeleton degeneration. Alexa Fluor 594 and 488 secondary antibodies were used to visualize the results. WST-1 assay was performed to evaluate cell viability.
Whole mounted cornea preparations were immunolabeled with β tubulin III and imaged under a Leica SP8 confocal microscopy. FIJI software (ImageJ 1.49d, NIH) was used to analyze nerve loss on the subbasal plexus.
Rats were handled and cared following ARVO statements for the use of animals.

Results : Exposure to intense light caused expression of HE-1 (oxidative stress inducible) and cleavage of Cit C from the mitochondrion membrane. Activated αCaMKII participated in tau phosphorylation and axonal microtubule disassembly. This cascade of events caused the rupture of neurites in cultured trigeminal neurons without affecting severely to neuronal survival. Distal degeneration was also observed as a reduction of corneal subbasal nerve plexus density.

Conclusions : Axonal distal degeneration of sensory nerve terminals in the cornea may be initiated by oxidative insults, such as intense light exposure or normal aging. Reduction of corneal nerve density is asociated with ocular diseases such as age-related dry eye. Reducing oxidative stress may prevent axonal loss and functionality.

This is a 2021 ARVO Annual Meeting abstract.