Some household products have high levels of the antimicrobial 2-
n-octyl-4-isothiazolin-3-one (OIT). Although the diverse effects of OIT are of concern, information regarding its cellular actions is limited. In a previous study, we found that OIT increased intracellular Zn
2+ levels in rat thymocytes. However, because Ca
2+ is considered the essential cation that causes cell injury and death, we examined whether Ca
2+ and Zn
2+ were involved in OIT-induced cytotoxicity and proposed the mechanisms underlying these results. The effects of OIT on the membrane and cellular parameters of rat thymocytes were examined with a flow cytometer and appropriate fluorescent probes. OIT (0.3-3 µM) increased intracellular Zn
2+ levels but not intracellular Ca
2+ levels. Therefore, the involvement of Zn
2+ was studied further. The simultaneous application of 0.3 µM OIT and 3 µM ZnCl
2 significantly increased cells with phosphatidylserine-exposed membranes without changing the dead cells. In contrast, applications of 0.3 µM OIT or 3 µM ZnCl
2 alone had no effects. The combination of OIT (0.1-1 µM) and ZnCl
2 (1-3 µM) significantly decreased the cellular non-protein thiol contents. These changes that were induced by their combination were completely suppressed by adding an intracellular Zn
2+ chelator. These results suggested that submicromolar concentrations of OIT induced Zn
2+-dependent cytotoxicity in the presence of micromolar concentrations of external Zn
2+. Because the threshold of OIT levels that affected cellular parameters in the presence of micromolar concentrations of Zn
2+ are much lower than the OIT contents in some household products, the adverse effects of OIT are of great concern.
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