Abstract
Purpose:
To determine the appropriate Cyclosporine A (CsA) dose and minimum drug delivery time needed to prevent posterior capsule opacification (PCO) in an ex vivo model and evaluate the mechanism of CsA-induced cell death.
Methods:
Lens capsules were harvested from canine cadaver eyes using an established ex vivo model of PCO. Lens capsules were treated with 0, 5, or 10 µg/mL CsA for 0, 2, 3, 4, 5, 6, or 7 days, and then maintained in culture for a total of 28 days in the absence of drug. CsA-treated lens epithelial cells (LEC) underwent routine transmission electron microscopy (TEM), western blotting, and fluorescent staining to evaluate the mechanism of cell death.
Results:
Lens capsules treated with 5, 6, or 7 days of 10 µg/mL CsA showed a significant decrease in ex vivo PCO formation; 7 days of drug delivery was sufficient to prevent PCO. Morphologically, CsA-treated LEC were swollen, had intact nuclei, lacked peripheral chromatin condensation, and demonstrated prominent vacuolization; TEM revealed autophagosomes. LC3-II protein expression and acridine orange fluorescence increased in CsA-treated cells. Dose dependent changes were observed in all experiments.
Conclusions:
Seven days of intracapsular CsA drug delivery prevented ex vivo PCO formation. Morphologic changes and TEM suggest that CsA is able to induce LEC death via autophagy; this is a novel finding in the lens. Acridine orange is a marker for acidic vesicles and LC3-II is a protein involved in mediating autophagic death. Expression of these markers in CsA-treated LEC further supports this new mechanism of drug-induced LEC death.
Keywords: 652 posterior capsular opacification (PCO)