Inhibition of the pesticide rotenone-induced Ca2+ signaling, cytotoxicity and oxidative stress in HCN-2 neuronal cells by the phenolic compound hydroxytyrosol

Abstract

Rotenone, a plant-derived pesticide belonging to genera Derris and Lonchorcarpus, is an inhibitor of NADH dehydrogenase complex. Studies have shown that rotenone was applied as a neurotoxic agent in various neuronal models. Hydroxytyrosol [2-(3,4-dihydroxyphenyl)-ethanol] is a natural phenolic compound found in the olive (Olea europaea L.). Studies of hydroxytyrosol have dramatically increased because this compound may contribute to the prevention of neurodegenerative diseases. Although hydroxytyrosol has received increasing attention due to its multiple pharmacological activities, it is not explored whether hydroxytyrosol inhibited rotenone-induced cytotoxicity in the neuronal cell model. The aim of this study was to explore whether hydroxytyrosol prevented rotenone-induced Ca2+ signaling, cytotoxicity and oxidative stress in HCN-2 neuronal cell line. In HCN-2 cells, rotenone (5–30 μM) concentration-dependently induced cytosolic Ca2+ concentrations ([Ca2+]i) rises and cytotoxicity. Treatment with hydroxytyrosol (30 μM) reversed rotenone (20 μM)-induced cytotoxic responses. In Ca2+-containing medium, rotenone-induced Ca2+ entry was inhibited by 2-APB (a store-operated Ca2+ channel modulator) or hydroxytyrosol. In Ca2+-free medium, treatment with thapsigargin (an endoplasmic reticulum Ca2+ pump inhibitor) or hydroxytyrosol significantly inhibited rotenone-induced [Ca2+]i rises. Furthermore, treatment with hydroxytyrosol reversed ROS levels, cytotoxic responses, and antioxidant enzyme activities (SOD, GPX and CAT) in rotenone-treated cells. Together, in HCN-2 cells, rotenone induced Ca2+ influx via store-operated Ca2+ entry and Ca2+ release from the endoplasmic reticulum and caused oxidative stress. Moreover, hydroxytyrosol ameliorated Ca2+ or ROS-associated cytotoxicity. It suggests that hydroxytyrosol might have a protective effect on rotenone-induced neurotoxicity in human neuronal cells.