Zn<sup>2+</sup>-dependent increase in cells with phosphatidylserine-exposed membranes after treatment with submicromolar concentrations of 2-<i>n</i>-octyl-4-isothiazolin-3-one in rat thymocytes
Some household products have high levels of the antimicrobial 2-<i>n</i>-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<sup>2+</sup> levels in rat thymocytes. However, because Ca<sup>2+</sup> is considered the essential cation that causes cell injury and death, we examined whether Ca<sup>2+</sup> and Zn<sup>2+</sup> 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<sup>2+</sup> levels but not intracellular Ca<sup>2+</sup> levels. Therefore, the involvement of Zn<sup>2+</sup> was studied further. The simultaneous application of 0.3 µM OIT and 3 µM ZnCl<sub>2</sub> significantly increased cells with phosphatidylserine-exposed membranes without changing the dead cells. In contrast, applications of 0.3 µM OIT or 3 µM ZnCl<sub>2</sub> alone had no effects. The combination of OIT (0.1-1 µM) and ZnCl<sub>2</sub> (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<sup>2+</sup> chelator. These results suggested that submicromolar concentrations of OIT induced Zn<sup>2+</sup>-dependent cytotoxicity in the presence of micromolar concentrations of external Zn<sup>2+</sup>. Because the threshold of OIT levels that affected cellular parameters in the presence of micromolar concentrations of Zn<sup>2+</sup> are much lower than the OIT contents in some household products, the adverse effects of OIT are of great concern.
- Fundamental Toxicological Sciences
Fundamental Toxicological Sciences 2(5), 209-216, 2015