GCLC nanomedicine for mitigation of Acrolein toxicity in the cornea

Abstract

Purpose : Acrolein vapor to eye causes severe corneal damage, eyelid swelling, and vision loss. How acrolein damages cornea and other ocular tissues still largely unknown. This study aimed to (1) define the severity (length and extent) of damage to the cornea, eyelid, and conjunctiva in vivo using rabbit model of acrolein-vapor exposure recently developed in our lab, and (2) characterize molecular mechanisms by which acrolein damages corneal tissues using human in vitro and ex vivo models of corneal wound healing and fibrosis

Methods : New Zealand White rabbits were used for in vivo studies. Eyes were exposed to acrolein vapor via a 7mm disc soaked in acrolein within a doggle to create mild (10ml acrolein=0.25 mg/ml) and severe wound (20ml acrolein=0.8 mg/ml). Clinical eye exam, slit-lamp biomicroscopy, pachymetry, and tonometry in live rabbits done before and after acrolein exposure (4h, 24h, 3-day, 7-day, 10-day and 14-day). Cornea, eyelid and sclera tissues were harvested and cryo-preserved. Human corneal stromal fibroblasts (hCSF) and ex vivo human cornea organ culture model were used for mechanistic studies. Ocular irritation, H&E, western blotting, immunofluorescence, qRT-PCR and commercial kit quantified damage to cornea and other ocular tissues.

Results : Acrolein caused morbid levels of inflammation severe inflammation within 15 min, which was continued for 24h, then gradually declined, severe haze appeared at 7-day, and grade-3 clinical damage to eye (haze+neovascularization) observed at 14-day. Also, acrolein exposure caused remarkable decrease (~50%) in glutathione (GSH) content and increase in thiobarbituric acid reactive substances (TBARS; >4-fold) in the rabbit cornea in vivo. Acrolein-treated human cornea showed significant compromised epithelium, reduced keratocyte density and increased aSMA (haze marker), CD31 (neovascularization marker), TUNEL+ cells (apoptosis marker) ex vivo. In vitro studies using nanoparticles-assisted gene delivery of Glutamate Cysteine Ligase Catalytic subunit (GCLC) into hCSFs restored GSH, ROS and LPO levels to the control non-treated group after acrolein exposure.

Conclusions : Acrolein causes severe acute toxicity to the eye but significantly more damage to the cornea in vivo. GCLC gene therapy has potential to counteract acrolein toxicity in the cornea.

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