Caspase-dependent apoptotic death by gadolinium chloride (GdCl 3 ) via reactive oxygen species production and MAPK signaling in rat C6 glioma cells

Abstract

Gadolinium (Gd) compounds serve as magnetic resonance imaging contrast agents and exert certain anticancer activities. Yet, the molecular signaling underlying the antitumor effect of Gd chloride (GdCl 3 ) on glioma remains unclear. In the present study, we aimed to ascertain the apoptotic mechanisms of GdCl 3 on rat glioma C6 cells. Our results demonstrated that GdCl 3 significantly reduced cell viability and shrunk cell morphology of C6 cells in a concentration-dependent manner. GdCl 3 led to apoptotic C6 cell death as detected by TUNEL staining. An increase in cleaved caspase-3, cleaved caspase-8 and cleaved caspase-9 occurred in GdCl 3 -treated C6 cells as detected by immunoblotting analysis. The activities of caspase-3, caspase-8 and caspase-9 were increased, and the specific inhibitors of caspase-3/-8/-9 individually reversed cell viability, which caused apoptotic death in C6 cells prior to GdCl 3 exposure. GdCl 3 also caused an elevation in the cytoplasmic Ca 2+ level and reactive oxygen species (ROS) production, as well as the loss of mitochondrial membrane potential (ΔΨm) as shown by flow cytometric analysis in C6 cells. The results from the immunoblotting analysis demonstrated that there were upregulated protein levels of cytochrome c and Bax but a downregulated protein level of Bcl-2 in C6 cells after GdCl 3 treatment. Additionally, GdCl 3 decreased the protein levels of phosphorylated-extracellular signal-regulated kinases, phosphorylated-c-Jun N-terminal kinase and phosphorylated-p38 mitogen-activated protein kinases in C6 cells. In conclusion, ROS production and MAPKs signaling pathways contribute to GdCl 3 -induced caspase cascade-mediated apoptosis in C6 cells. Our findings provide a better understanding of the molecular mechanisms underlying the role of GdCl 3 in rat glioma C6 cells.