Find in Library

In this paper, we report the metabolic characterization of two <i>foci</i>, F1 and F3, obtained at the end of Cell Transformation Assay (CTA), performed by treating C3H10T1/2Cl8 mouse embryo fibroblasts with 1 μM CdCl₂ for 24 h. The elucidation of the cadmium action mechanism can be useful both to improve the in vitro CTA and to yield insights into carcinogenesis. The metabolism of the two <i>foci</i> was investigated through Seahorse and enzyme activity assays; mitochondria were studied in confocal microscopy and reactive oxygen species were detected by flow cytometry. The results showed that F1 <i>focus</i> has higher glycolytic and TCA fluxes compared to F3 <i>focus</i>, and a more negative mitochondrial membrane potential, so that most ATP synthesis is performed through oxidative phosphorylation. Confocal microscopy showed mitochondria crowded in the perinuclear region. On the other hand, F3 <i>focus</i> showed lower metabolic rates, with ATP mainly produced by glycolysis and damaged mitochondria. Overall, our results showed that cadmium treatment induced lasting metabolic alterations in both <i>foci</i>. Triggered by the loss of the Pasteur effect in F1 <i>focus</i> and by mitochondrial impairment in F3 <i>focus</i>, these alterations lead to a loss of coordination among glycolysis, TCA and oxidative phosphorylation, which leads to malignant transformation.