Impaired mitochondrial functions contribute to 3-bromopyruvate toxicity in primary rat and mouse hepatocytes.

Abstract

A compound with promising anticancer properties, 3-bromopyruvate (3-BP) is a synthetic derivative of a pyruvate molecule; however, its toxicity in non-malignant cells has not yet been fully elucidated. Therefore, we elected to study the effects of 3-BP on primary hepatocytes in monolayer cultures, permeabilized hepatocytes and isolated mitochondria. After a 1-h treatment with 100μM 3-BP cell viability of rat hepatocytes was decreased by 30% as measured by the WST-1 test (p<0.001); after 3-h exposure to ≥200μM 3-BP lactate dehydrogenase leakage was increased (p<0.001). Reactive oxygen species production was increased in the cell cultures after a 1-h treatment at concentrations ≥100μmol/l (p<0.01), and caspase 3 activity was increased after a 20-h incubation with 150μM and 200μM 3-BP (p<0.001). This toxic effect of 3-BP was also proved using primary mouse hepatocytes. In isolated mitochondria, 3-BP induced a dose- and time-dependent decrease of mitochondrial membrane potential during a 10-min incubation both with Complex I substrates glutamate + malate or Complex II substrate succinate, although this decrease was more pronounced with the latter. We also measured the effect of 3-BP on respiration of isolated mitochondria. ADP-activated respiration was inhibited by 20μM 3-BP within 10min. Similar effects were also found in permeabilized hepatocytes of both species.