Topical Benzalkonium Chloride Treatment Causes Corneal Neurotoxicity

Abstract

Purpose: : Although topical application of benzalkonium chloride (BAK), a common bactericidal preservative in ophthalmic preparations produces chemical denervation in non-ocular tissues, its neurotoxic effect on corneal nerves remains unclear. We hypothesize that topical application of benzalkonium chloride (BAK) is accompanied by corneal inflammation and neurotoxicity.

Methods: : 6-8 week old thy1-YFP neurofluorescent mice eyes were treated topically either with vehicle or BAK-containing eyedrops once a day for 1 week. In vivo stereofluorescence imaging was performed to sequentially image and monitor changes in stromal nerve fiber density (NFD) in the treated corneas every week upto 4 weeks. The extent of DED was evaluated by aqueous tear production measurement (modified Schirmer test) and corneal fluorescein staining. Whole-mount immunofluorescence staining of corneas was performed with antibodies to regeneration-associated and nerve structural markers. Immunoblot analyses were performed on trigeminal ganglion and corneal lysates to determine expression levels of proteins associated with neurotoxicity and regeneration. In vitro Campenot compartmental culture of trigeminal ganglion neurons was performed in specialized devices to determine the effect of BAK on neurite outgrowth.

Results: : Aqueous tear production and stromal NFD were found to be significantly reduced (p<0.05) along with increased fluorescein staining after 1 week of 0.1% BAK treatment. Sequential in vivo imaging of corneas showed two forms of BAK-induced neurotoxicity; (i) Reversible Neurotoxicity characterized by axonopathy and recovery of lost nerves and (ii) Irreversible Neurotoxicity characterized by nerve degeneration and regeneration. Elevated protein levels of regeneration-associated and nerve structural proteins in corneal lysates confirmed regeneration. BAK addition (0.0001% and 0.001%) to in vitro compartmental cultures of dissociated trigeminal ganglion cells caused a significant dose-related neurite reduction and an exacerbated effect with the higher dose (p<0.001).

Conclusions: : Topical BAK application to the eye causes corneal neurotoxicity accompanied by inflammation and reduced aqueous tear production.

Keywords: cornea: basic science