Abstract
Abstract: :
Purpose: In a previous study we showed that the latanoprost, in its commercial presentation, appeared, in vitro, less toxic than the benzalkonium chloride it contained. Therefore we investigate with microplate cytometry if the three antiglaucoma prostanoids, commercially available, could protect in vitro, conjunctiva–derived cells toward benzalkonium chloride (BAC) toxicity and if an antioxidative mechanism could be involved in prostaglandin effects. Methods: Human conjunctiva–derived epithelial cells from Chang cell line were exposed to latanoprost, travoprost and bimatoprost, and to three formulations of benzalkonium chloride (2x10–2 %, 1.5x10–2 % and 0.5x10–2 %) corresponding to the same concentrations contained in the three prostanoid eye drops. Each solution was diluted to 1/10 and applied for 30 minutes. Cellular membrane integrity, cytosolic H2O2, cytosolic O2.– and apoptosis were evaluated using, respectively, Neutral Red, H2DCF–DA, Hydroethidine and Yopro–1 probes. Results: Cellular viability decreased as benzalkonium chloride concentration increased with a concentration–dependent toxicity. Toxicity of latanoprost and travoprost were statistically significantly lower than their respective BAC concentration (p<0.01) while bimatoprost toxicity was not different compared with control. H2O2 detection statistically decreased with cells exposed to latanoprost (p<0.01) and travoprost (p<0.01) and O2.– detection decreased with cells exposed to latanoprost (p<0.01) compared with their corresponding BAC concentration alone. The Yopro–1 test showed a BAC–induced apoptotic effect, which increased with its concentration. Latanoprost and travoprost were responsible for a pro–apoptotic effect compared with control (p<0.01) but lower than their respective preservative concentration with a significant statistical difference (p<0.01). Conclusions: Latanoprost and travoprost are responsible for a protective effect toward benzalkonium chloride toxicity on conjunctiva–derived epithelial cells in vitro, probably related to antioxidative properties. The low toxicity of bimatoprost could not make it possible to reveal a hypothetic antioxidative effect. The reduction of the reactive oxygen species could be related to a diminution of the BAC pro–apoptotic effect.
Keywords: antioxidants • conjunctiva • cell death/apoptosis