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Long-term antiglaucoma eye drop therapy prior to trabeculectomy is a risk factor for surgical failure resulting from Tenon’s capsule fibrosis at the fistula site. The study tested the hypothesis that secretion of the proinflammatory cytokine interleukin (IL)-6 by wounded Tenon’s capsule-fibroblasts is elevated by prior long-term antiglaucoma eye drop treatment.Fibroblast cultures were established from Tenon’s capsule biopsies during trabeculectomy. Twice daily and for four and a half days, confluent secondary (2–4 passages) cultures were treated (30 minutes at 37°C) with the following drugs (diluted at 1:400–1:100): 0.2% brimonidine-tartrate (Alphagan), 2.0% dorzolamide-HCl (Trusopt), 0.5% timolol-maleate (Timoptic), 2.0% dorzolamide-HCl/0.5% timolol-maleate (Cosopt), 2 and 4% pilocarpine-HCl (Akarpine and Pilocar), 0.005% latanoprost (Xalatan), placebos for Trusopt and Timoptic, and 0.01% benzalkonium chloride. Subsequently, cultures were wounded by removing cells grown on half of each culture dishes along with the medium and conditioned for 20 hours in serum-free growth medium, which was then collected for ELISA for IL-6 (and TNF-α and IL-1β). Cultures were grown for four additional days to show the maintenance of culture sterility.Latanoprost, pilocarpine-HCl, and timolol-maleate increased IL-6 levels in the conditioned medium in a dilution factor-dependent manner (P < 0.05, ANOVA). IL-6 concentrations were increased most significantly by latanoprost and were (pg/ml; mean ± SEM; N = 3 cultures) 186 ± 37, 187 ± 33, 295 ± 46 and 336 ± 76 in cultures treated at 1:400, 1:250, 1:150, and 1:100 dilutions, respectively, whereas those of six control cultures averaged 80 ± 9. Benzalkonium chloride, brimonidine-tartrate, dorzolamide-HCl, Cosopt placebo, Timoptic placebo, and dorzolamide-HCl/timolol-maleate did not significantly elevate IL-6 concentrations. IL-1β and TNF-α were not detected in the medium of control cultures and those treated with pilocarpine (1:200).The present study demonstrated for the first time that the level of IL-6 secretion by wounded Tenon’s capsule fibroblast cell cultures was increased by repeat pulsing of these cultures with some, but not all, antiglaucoma eye drops prior to wounding.