Antioxidation activity of tetrahydrobiopterin in pheochromocytoma PC 12 cells

Rat pheochromocytoma PC 12 cells are susceptible to the oxidative toxicity caused by H2O2, nitrofurantoin, dopamine, and xanthine/xanthine oxidase reaction. The cytotoxicities of these agents are greatly reduced by the simultaneous presence of 0.1 mM tetrahydrobiopterin (BH4), 3 units/ml horseradish peroxidase, 0.2 mM NADH, and 0.1 units/ml sheep liver dihydropteridine reductase (DHPR). Individually, BH4, NADH and DHPR have no protection against H2O2 toxicity in PC 12 cells. Peroxidase alone offers 58% of protection if cells are incubated in the medium but only 3% in Dulbecco's phosphate buffered saline. The efficiency of the BH4-mediated antioxidation system in PC 12 cells is equal to or better than ascorbic acid and catalase, depending on the source of the reactive O2 species (ROS). The reactions responsible for the BH4-antioxidation system may consist of the non-enzymatic and the peroxidase-catalyzed reduction of H2O2 to H2O by BH4 and the regeneration of BH4 by DHPR using NADH as the cofactor. The components of this defence mechanism against ROS are all normal cellular constituents and are ubiquitous in nature. This DHPR-catalyzed redox cycling of BH4 may constitute an as yet little-known antioxidation system in mammalian cells.