Effects of enhancing mitochondrial oxidative phosphorylation with reducing equivalents and ubiquinone on 1-methyl-4-phenylpyridinium toxicity and complex I-IV damage in neuroblastoma cells.
The effects of increasing mitochondrial oxidative phosphorylation (OXPHOS), by enhancing electron transport chain components, were evaluated on 1-methyl-4-phenylpyridinium (MPP+) toxicity in brain neuroblastoma cells. Although glucose is a direct energy source, ultimately nicotinamide and flavin reducing equivalents fuel ATP produced through OXPHOS. The findings indicate that cell respiration/mitochondrial O(2) consumption (MOC) (in cells not treated with MPP+) is not controlled by the supply of glucose, coenzyme Q(10) (Co-Q(10)), NADH+, NAD or nicotinic acid. In contrast, MOC in whole cells is highly regulated by the supply of flavins: riboflavin, flavin adenine dinucleotide (FAD) and flavin mononucleotide (FMN), where cell respiration reached up to 410% of controls. In isolated mitochondria, FAD and FMN drastically increased complex I rate of reaction (1300%) and (450%), respectively, having no effects on complex II or III. MPP+ reduced MOC in whole cells in a dose-dependent manner. In isolated mitochondria, MPP+ exerted mild inhibition at complex I, negligible effects on complexes II-III, and extensive inhibition of complex IV. Kinetic analysis of complex I revealed that MPP+ was competitive with NADH, and partially reversible by FAD and FMN. Co-Q(10) potentiated complex II ( approximately 200%), but not complex I or III. Despite positive influence of flavins and Co-Q(10) on complexes I-II function, neither protected against MPP+ toxicity, indicating inhibition of complex IV as the predominant target. The nicotinamides and glucose prevented MPP+ toxicity by fueling anaerobic glycolysis, evident by accumulation of lactate in the absence of MOC. The data also define a clear anomaly of neuroblastoma, indicating a preference for anaerobic conditions, and an adverse response to aerobic. An increase in CO(2), CO(2)/O(2) ratio, mitochondrial inhibition or O(2) deprivation was not directly toxic, but activated metabolism through glycolysis prompting depletion of glucose and starvation. In conclusion, the results of this study indicate that the mechanism of action for MPP+, involves the inhibition of complex I and and more specifically complex IV, leading to impaired OXPHOS and MOC. Moreover, flavin dervatives control the rate of complex I/cellular respiration and Co-Q10 augments complex II [corrected].
Mazzio
Elizabeth A
EA
College of Pharmacy and Pharmaceutical Sciences, Florida A&M University, Tallahassee, FL 32307, USA.
Soliman
Karam F A
KF
D015655:1-Methyl-4-phenylpyridinium / Q000633:toxicity*; D000332:Aerobiosis; D000818:Animals; D014450:Electron Transport Complex III / Q000378:metabolism; D006019:Glycolysis / Q000187:drug effects; D051379:Mice; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism; D009447:Neuroblastoma / Q000473:pathology; D010084:Oxidation-Reduction; D010085:Oxidative Phosphorylation / Q000187:drug effects; D010101:Oxygen Consumption / Q000187:drug effects; D014407:Tumor Cells, Cultured; D014451:Ubiquinone / Q000494:pharmacology*
D014451:Ubiquinone; D014450:Electron Transport Complex III; D015655:1-Methyl-4-phenylpyridinium