Quaternary ammonium cytotoxicity in a human conjunctival cell line

Purpose: Ophthalmic preparations can induce conjunctival toxicity, often ca used by preservatives. The aim of this study was to evaluate in vitro cytot oxicity of cytotoxicity ammoniums. Methods : Cytotoxicity tests were done on a continuous human conjunctival c ell line using microplate cold light cytofluorimetry. Membrane integrity (n eutral red test), DNA condensation (Hoechst 33342 test) and reactive oxygen species (ROS) production (dichiorofluoresceine diacetate and hydroethidine tests) were evaluated on living cells treated with different concentration s of benzalkonium chloride, benzododecinium bromide and cetrimide (0.00001 to 0.01 %) after minutes of treatment or 15 minutes and 24 hours of cell re covery. Results : All the compounds tested showed similar in vitro effects. Using: the neutral red test, we observed a decrease in membrane integrity even at 0.005 % and 0.01 % (p < 0.001) and after a short time (15 minutes). A stimu lation of ROS production (H2O2 and O-2(-)) was observed at 0.00001 % and ab ove (p < 0.001), associated with a chromatine condensation due to an apopto tic phenomenon. Conclusion: A necrotic phenomenon is suggested at high concentrations-of qu aternary ammonium preservatives whereas an apoptotic mechanism exists for l ower concentrations. This toxicity observed in vitro can explain some of th e ocular surface damage caused by long-term use of preserved eye-drops.