Simvastatin Induces Delayed Apoptosis Through Disruption of Glycolysis and Mitochondrial Impairment in Neuroblastoma Cells.

Simvastatin, a commonly used cholesterol-lowering drug, inhibits the mevalonate pathway involved in the synthesis of the mitochondrial electron carrier coenzyme Q10 (CoQ10), as well as other bioenergetics substrates. The purpose of this study was to investigate simvastatin exposure on mitochondrial respiration, metabolic fuel preferences, and glucose utilization. We hypothesized that simvastatin at a noncytotoxic dose will impair energy metabolism in human neuroblastoma cells. SK-N-AS cells were exposed at acute and chronic time points and evaluated in a Seahorse XF analyzer, revealing decreased mitochondrial and glycolytic parameters. Flow cytometry showed a significant induction of apoptosis in simvastatin-treated cells at 48 hours. Finally, multiple techniques were used to show that simvastatin-mediated impairment of bioenergetics is more complex than CoQ10 depletion or hampered glucose uptake. Therefore, the data reported here represent a biphasic hit to mitochondria followed by reduction in glucose and glutamine metabolism in neuroblastoma; adding mechanism to potential pleotropic effects of statins.

Acute Maneb Exposure Significantly Alters Both Glycolysis and Mitochondrial Function in Neuroblastoma Cells.

The pesticides paraquat (PQ) and maneb (MB) have been described as environmental risk factors for Parkinson's disease (PD), with mechanisms associated with mitochondrial dysfunction and reactive oxygen species generation. A combined exposure of PQ and MB in murine models and neuroblastoma cells has been utilized to further advance understanding of the PD phenotype. MB acts as a redox modulator through alkylation of protein thiols and has been previously characterized to inhibit complex III of the electron transport chain and uncouple the mitochondrial proton gradient. The purpose of this study was to analyze ATP-linked respiration and glycolysis in human neuroblastoma cells utilizing the Seahorse extracellular flux platform. Employing an acute, subtoxic exposure of MB, this investigation revealed a MB-mediated decrease in mitochondrial oxygen consumption at baseline and maximal respiration, with inhibition of ATP synthesis and coupling efficiency. Additionally, MB-treated cells showed an increase in nonmitochondrial respiration and proton leak. Further investigation into mitochondrial fuel flex revealed an elimination of fuel flexibility across all 3 major substrates, with a decrease in pyruvate capacity as well as glutamine dependency. Analyses of glycolytic function showed a substantial decrease in glycolytic acidification caused by lactic acid export. This inhibition of glycolytic parameters was also observed after titrating the MB dose as low as 6 µM, and appears to be dependent on the dithiocarbamate functional group, with manganese possibly potentiating the effect. Further studies into cellular ATP and NAD levels revealed a drastic decrease in cells treated with MB. In summary, MB significantly impacted both aerobic and anaerobic energy production; therefore, further characterization of MB's effect on cellular energetics may provide insight into the specificity of PD to dopaminergic neurons.