Protective Effects of Organic Acids against Xanthine/Xanthine Oxidase-Induced Cell Death by Reducing the Intracellular Level of Hydrogen Peroxide

Abstract

Enhanced production of superoxide is considered to play a pivotal role in the pathogenesis of various chronic diseases. In the present study, we examined the toxic effects of superoxide and hydrogen peroxide (H₂O₂) produced by xanthine (XA) plus xanthine oxidase (XO), and the protective effects of various organic acids against them by use of a cellular model of COS7 cells, an African green monkey cell line. Here, we report that superoxide and H₂O₂ generated by XA/XO triggered cell death associated with the increase in the intracellular level of H₂O₂. The reactive oxygen species (ROS) levels were measured by use of 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) and a multi-well fluorescence spectrophotometer. XA/XO induced an ROS burst before initiating the loss of cell viability. Catalase and N-acetylcysteine protected the cells from the XA/XO-induced cell death, indicating that the effector of the cell death was indeed H₂O₂. Further, we found that organic acids involved in aerobic energy metabolism, such as pyruvate, oxaloacetate, and α-ketoglutarate, had significant protective effects against the cells death by reducing the levels of H₂O₂. These organic acids all shared a common chemical structure, i.e., that of α-keto acid, which directly reacted with H₂O₂. In contrast, other organic acids, such as lactate, succinate, fumarate, and malate, which do not have the α-keto acid structure, but may produce it by dehydrogenase systems, did not efficiently protect the cells, suggesting that this structure was essential for the protective action of organic acids against oxidative stress.