Topical administration of a Rock/Net inhibitor promotes retinal ganglion cell survival and axon regeneration after optic nerve injury

Abstract

Intraocular pressure (IOP)-lowering ophthalmic solutions that inhibit Rho-associated protein kinases (Rock) and norepinephrine transporters (Net) are currently under clinical evaluation. Here we evaluate topical application of one such drug for its effects on retinal ganglion cell (RGC) survival and axon regeneration after optic nerve crush injury. We performed unilateral optic nerve crush on young rats (P18) and topically applied Rock/Net inhibitor AR-13324 or placebo 3 times a day for 14 days. IOP was measured starting 3 days before and up to 9 days after injury. On day 12, cholera toxin B (CTB) was injected intravitreally to trace optic nerve regeneration. On day 14, retinas and optic nerves were collected. The retinas were flat-mounted and stained with RBPMS to quantify RGC survival and the optic nerves were sectioned for optic nerve axon quantification using fluorescent and confocal microscopy. Rock phosphorylation targets implicated in axon growth including cofilin and LIMK were examined by fluorescence microscopy and quantitative western blotting. AR-13324 lowered IOP as expected. RGC survival and optic nerve axon regeneration were significantly higher with Rock/Net inhibitor treatment compared with placebo. Furthermore, topical therapy decreased Rock target protein phosphorylation in the retinas and proximal optic nerves. These data suggest that topical administration of a Rock/Net inhibitor promotes RGC survival and regeneration after optic nerve injury, with associated molecular changes indicative of posterior drug activity. Coordinated IOP lowering and neuroprotective or regenerative effects may be advantageous in the treatment of patients with glaucoma.

Rho-Kinase Inhibition Reduces Myofibroblast Differentiation and Proliferation of Scleral Fibroblasts Induced by Transforming Growth Factor β and Experimental Glaucoma

“… 38 Interestingly, topical application of the ROCK/Net inhibitor netarsudil reduced RGC loss following rat optic nerve crush. 39 Topical application of netarsudil inhibited phosphorylation of the downstream ROCK targets cofillin and LIM kinase in isolated retina—suggesting ROCK inhibition in the posterior segment of the eye.…”

The Role of Endogenous Neuroprotective Mechanisms in the Prevention of Retinal Ganglion Cells Degeneration

“…It is worth remembering that RhoA/ROCK signaling modulates AH outflow; ROCK inhibitors lower intraocular pressure (IOP) via a direct effect on TM and Schlemm’s canal, and they are currently under clinical evaluation in glaucoma ( Wang and Chang, 2014 ). Interestingly, it has been reported that topical administration of a ROCK inhibitor promotes RGC survival and axon regeneration after ON injury in vivo ( Shaw et al, 2017 ), suggesting that the beneficial effects of this class of inhibitors may go beyond their action on IOP.…”

IGF-1, Inflammation and Retinal Degeneration: A Close Network

“…Retinal degenerative diseases are heterogeneous pathologies; among them, age-related macular degeneration (AMD), retinitis pigmentosa (RP), and diabetic retinopathy (DR) have a high incidence and prevalence in humans (Hernandez et al, 2016 ; Narayan et al, 2016 ; Shaw et al, 2017 ). In fact, a plethora of factors including genetic alterations, aging, vascular defects, chemical insults, oxidative stress, or light-induced damage are responsible of the development of retinal degeneration (Semeraro et al, 2015 ; van Norren and Vos, 2016 ).…”