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1From the Department of Ophthalmology, University Hospital of Tours, Tours, France; the 2Department of Toxicology, Faculty of Biological and Pharmacological Sciences, University of Paris 5, Paris, France; the 3Department of Ophthalmology, Quinze-Vingts National Ophthalmology Hospital and INSERM U598, Paris, France; 4Ambroise Paré Hospital, APHP, Paris Ouest School of Medicine, Paris, France; and the 5Department of Ophthalmology, University Hospital of Dijon, Dijon, France.
PURPOSE. To compare the toxicity of latanoprost and preserved and unpreserved timolol on conjunctival cells. Expression of inflammatory markers and MUC5AC-related mucin production were evaluated by impression cytology in a case–control ex vivo study. The proapoptotic effect of the same drugs was also evaluated in vitro in a conjunctival cell line and compared with that of benzalkonium chloride (BAC).
METHODS. Impression cytology (IC) specimens were obtained from a series of normal subjects and from patients with glaucoma treated for at least 1 year with latanoprost eye drops or preserved or unpreserved timolol. All groups were comparable in age and duration of treatment. Expression of HLA-DR, intercellular adhesion molecule (ICAM)-1, and mucin was evaluated in a masked manner by flow cytometry. For the in vitro study, a human conjunctiva–derived cell line was treated with 0.02% BAC-containing latanoprost or timolol, unpreserved timolol, or 0.02% BAC alone for 15 minutes, followed or not by 4 or 24 hours of cell recovery in normal medium. Cell viability and chromatin condensation were evaluated using microplate cold light cytofluorometry with the neutral red and the Hoechst 33342 tests, respectively. The Hoechst–neutral red ratio was defined for the apoptosis assay, and cytoskeleton changes were assessed by confocal microscopy.
RESULTS. No difference was found between normal eyes and those receiving unpreserved timolol. Preserved latanoprost and timolol significantly increased the inflammatory marker expression and decreased MUC5AC expression, but to a significantly higher extent in the preserved timolol group compared with latanoprost. In vitro, 0.02% BAC-containing timolol and latanoprost triggered conjunctival cell apoptosis—however, to a significantly lesser extent than did 0.02% BAC alone. Unpreserved timolol did not cause any cell toxicity.
CONCLUSIONS. These ex vivo and in vitro studies demonstrate that BAC-containing latanoprost and timolol exhibit higher proinflammatory and proapoptotic effects on conjunctival cells than does unpreserved timolol. Latanoprost caused less toxicity, however, than preserved timolol, and both drugs were less toxic than BAC alone. These results suggest a potential protective effect of the prostaglandin analogue and to a lesser extent of timolol against the toxicity of BAC in conjunctival cells.
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