Disruption of Pyridine Nucleotide Redox Status During Oxidative Challenge at Normal and Low-Glucose States: Implications for Cellular Adenosine Triphosphate,Mitochondrial Respiratory Activity,and Reducing Capacity in Colon Epithelial Cells

Abstract

We recently demonstrated that menadione (MQ), a redox cycling quinone, mediated the loss of mitochondrial glutathione/glutathione disulfide redox balance. In this study, we showed that MQ significantly disrupted cellular pyridine nucleotide (NAD⁺/NADH, NADP⁺/NADPH) redox balance that compromised cellular ATP, mitochondrial respiratory activity, and NADPH-dependent reducing capacity in colonic epithelial cells, a scenario that was exaggerated by low glucose. In the cytosol, MQ induced NAD⁺ loss concurrent with increased NADP⁺ and NAD kinase activity, but decreased NADPH. In the mitochondria, NADH loss occurred in conjunction with increased nicotinamide nucleotide transhydrogenase activity and NADP⁺, and decreased NADPH. These results are consistent with cytosolic NAD⁺-to-NADP⁺ and mitochondrial NADH-to-NADPH shifts, but compromised NADPH availability. Thus, despite the sacrifice of NAD⁺/NADH in favor of NADPH generation, steady-state NADPH levels were not maintained during MQ challenge. Impairments of cellular bioenergetics were evidenced by ATP losses and increased mitochondrial O₂ dependence of pyridine nucleotide oxidation–reduction; half-maximal oxidation (P₅₀) was 10-fold higher in low glucose, which was lowered by glutamate or succinate supplementation. This exaggerated O₂ dependence is consistent with increased O₂ diversion to nonmitochondrial O₂ consumption by MQ-semiquinone redox cycling secondary to decreased NADPH-dependent MQ detoxication at low glucose, a situation that was corrected by glucose-sparing mitochondrial substrates. Antioxid. Redox Signal. 14, 2151–2162.