title: Cyclosporine A prevents ex vivo PCO formation through induction of autophagy-mediated cell death.
abstract: The purpose of this study was to determine the Cyclosporine A (CsA) dose and minimum drug delivery time needed to prevent posterior capsule opacification (PCO) in an ex vivo canine model and evaluate the mechanism of CsA-induced cell death. Canine lens epithelial cells (LEC) were treated with CsA and changes in cell migration, proliferation, and density were monitored over time. CsA-treated LEC underwent transmission electron microscopy (TEM), immunofluorescence, and immunoblotting in the presence or absence of autophagy inhibitors to evaluate the mechanism of cell death. Lens capsules were harvested from canine cadaver eyes for an ex vivo model of PCO. Lens capsules were treated with CsA for 1, 2, 3, 4, 5, 6, or 7 days, and subsequently maintained in culture for a total of 28 days in the absence of drug. CsA reduced LEC viability in a dose dependent manner. 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. A small non-significant induction of cleaved caspase-3 was observed in CsA-treated LEC. Lens capsules treated with 5, 6, or 7 days of 10 μg/mL CsA showed a significant decrease in ex vivo PCO formation; 6 days of drug delivery prevented PCO. This study finds that morphologic changes, formation of acidic vesicles, and increased expression of LC3-II supports the hypothesis that CsA mediates LEC death via autophagy; this is a novel finding in the lens. Induction of CsA-induced apoptosis was minimal. Six days of intracapsular CsA drug delivery prevented ex vivo PCO formation.
authors.affiliation: The Ohio State University, College of Optometry, 320 West 10th Avenue, Columbus, OH 43210, USA; The Ohio State University, College of Veterinary Medicine, 601 Vernon Tharp Street, Columbus, OH 43210, USA. Electronic address: chandler.111@osu.edu.
authors.forename: Heather L
authors.initials: HL
authors.lastname: Chandler
authors.affiliation: The Ohio State University, College of Optometry, 320 West 10th Avenue, Columbus, OH 43210, USA.
authors.forename: Kristen J
authors.initials: KJ
authors.lastname: Gervais
authors.affiliation: The Ohio State University, College of Veterinary Medicine, 601 Vernon Tharp Street, Columbus, OH 43210, USA.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Lutz
authors.affiliation: The Ohio State University, College of Optometry, 320 West 10th Avenue, Columbus, OH 43210, USA.
authors.forename: Elizabeth M
authors.initials: EM
authors.lastname: Curto
authors.affiliation: The Ohio State University, College of Optometry, 320 West 10th Avenue, Columbus, OH 43210, USA.
authors.forename: Rachel B
authors.initials: RB
authors.lastname: Matusow
authors.affiliation: The Ohio State University, College of Veterinary Medicine, 601 Vernon Tharp Street, Columbus, OH 43210, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Wilkie
authors.affiliation: The Ohio State University, College of Veterinary Medicine, 601 Vernon Tharp Street, Columbus, OH 43210, USA.
authors.forename: Anne J
authors.initials: AJ
authors.lastname: Gemensky-Metzler
mesh_terms: D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D058442:Capsule Opacification / Q000378:metabolism / Q000473:pathology / Q000517:prevention & control*; D002452:Cell Count; D016923:Cell Death / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D016572:Cyclosporine / Q000008:administration & dosage / Q000494:pharmacology*; D004285:Dogs; D004591:Electrophoresis, Polyacrylamide Gel; D004847:Epithelial Cells / Q000187:drug effects / Q000473:pathology; D015151:Immunoblotting; D007166:Immunosuppressive Agents / Q000008:administration & dosage / Q000494:pharmacology*; D046529:Microscopy, Electron, Transmission; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D009924:Organ Culture Techniques; D057852:Posterior Capsule of the Lens / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D007166:Immunosuppressive Agents; D008869:Microtubule-Associated Proteins; D016572:Cyclosporine
keywords: Autophagy; Canine; Cyclosporine A; Lens epithelial cell; Posterior capsule opacification