The Study Of Cellular Cytotoxicity Of L-DOPA And Its Mechanism

AIM:To study the influence of L-DOPA (Levodopa, L-DOPA) on dopaminergic cells proliferation and survival, and the protective effect of Quinone oxidoreductase (Quinone oxidoreductase, NQO1) on L-DOPA cytotoxicity. To further investigate the pathogenesis of Parkinson’s disease and explore the potential therapeutic prevention methods of Levodopa failure syndrome.METHODS:The MTT assay (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, MTT) was used to detect the proliferation injury affection of levodopa (L-DOPA) on neuroblastoma cells (SH-SY5Y), to detect the combined cytotoxic effects of reserpine (Reserpine, RE) and different concentrations of L-DOPA on SH-SY5Y and the protective effect of sulforaphane (Sulforaphane, SF) on cytotoxicity induced by different concentrations of L-DOPA. The quinone protein was detected by the NBT/glycinate assay. Intracellular expression of NQO1 induced by sulforaphane (SF) was detected by Western Blots. Microplate Reader was used to detect the changes of lactate dehydrogenase activity.RESULTS:With the increasing concentration of L-DOPA, the cytotoxic effects on SH-SY5Y cells were enhanced. SH-SY5Y cells showed significant damage when the concentration of L-DOPA is 400μmol/L, the content of intracellular quinone protein increased with L-DOPA concentration increasing. The cytotoxicity effects on SHSY5Y much more increased when L-DOPA induced with reserpine, the same as quinone protein content increased; intracellular lactate dehydrogenase activity was decreased after L-DOPA effects on SH-SY5Y; sulforaphane increases the expression of NQO1 and alleviates L-DOPA-induced cytotoxicity in SH-SY5Y cells; there are no significant differences in content of quinone protein between L-DOPA plus SF and L-DOPA alone group.CONCLUSIONS:L-DOPA cytotoxic effects on SH-SY5Y cells in a dose-dependent manner, with increasing concentration of L-DOPA, the content of quinone protein increased, which can be speculated that there are a degree of correlation between L-DOPA cytotoxic effects and the formation of quinone protein. Reserpine can increases the cells cytotoxicity of L-DOPA, which also increases the content of quinone protein. L-DOPA can effectively reduce the Intracellular lactate dehydrogenase activity, suggesting that L-DOPA could convert lactate dehydrogenase to quinone protein, then affecting cellular oxidative metabolism and leading to cell injury.90μmol/L HT can’t protect the cytotoxicity effects of L-DOPA; SF can induce NQO1, which may play a protective effect on the cytotoxicity of L-DOPA; there are no significant effects on SF’s preventing the increasing of quinone protein induced by L-DOPA.