Cilostazol, a quinolinone derivative, is approved by the Food and Drug Administration for the treatment of intermittent claudication.1) It has been shown that Cilostazol has an inhibitory effect on platelet aggregation and thronbosis in feline cerebral ischemia.2) Cilostazol has also been shown to significantly reduce the risk of recurrent strokes without affecting the occurrence of intracranial hemorrhage. Recently, the Cilostazol Stroke Prevention Study revealed that Cilostazol reduced the risk of secondary stroke by 41.7% as compared with a placebo 3) suggesting that Cilostazol has a specific effect on small-vessel disease, although the mechanism for this is not well known. It has been shown that Cilostazol and its metabolites reduced the intracellular reactive oxygen species (ROS) induced by LPS in human umbilical vein endothelial cells by directly scavenging hydroxyl radicals.
4)However, the anti-stroke effect of Cilostazol in vivo seems not to be due to its antioxidative effect, because its blood clearance is so rapid, and its concentration in the brain would be too low to directly scavenge ROS. Thus, Cilostazol may cause an anti-stroke effect via the induction of a putative anti-oxidative molecule in the brain.Metallothionein (MT) is a small sulfhydryl-rich protein, the level of which is elevated by various inducers of metals, hormones and cytokines. The induction of MT synthesis has been reported in various tissues, but rarely in the brain. MT is thought to be a multifunctional protein detoxifying heavy metals such as cadmium and inorganic mercury, maintaining zinc and copper homeostasis, and regulating the biosynthesis and activity of zinc-binding proteins such as zinc-dependent transcription factors. MT also has the ability to scavenge ROS. 5,6) Concerning the anti-oxidative activities of MT, a role of MT in neurodegenerative diseases has been proposed.7) Thus, in this experiment, we examined the induction of MT by Cilostazol in the mouse brain and liver, and also in human cultured neuronal cells in order to confirm the possibility that Cilostazol directly induces MT in the human brain as an anti-oxidative effecter molecule. Animals and Treatments Male C57BL/6J mice were purchased from Clea Japan Inc. (Tokyo Japan). They were housed in plastic cages at 23°C with a 12 h light/dark cycle and were given lab food and tap water ad libitum. Mice (7 weeks old) were randomly assigned to treatment groups and were injected i.p. with 0.5% Cilostazol suspended with 0.5% carboxylmethyl cellulose solution or vehicle alone. After various time intervals, the liver and brain were removed from each mouse under anesthetization with sodium pentobarbital and were then stored at Ϫ80°C for the determination of MT level. The mice received humane care throughout the experiments according to the guidelines of Tokushima Bunri University.
MATERIALS AND METHODS
MaterialsCell Cultures IMR-32 cells, a human neuroblastoma cell line, were obtained from the American Type Culture Collection (ATCC, Rockville, MD, U.S.A.), and maintained ...
