Glaucocalyxin A and B-induced Cell Death is Related to GSH Perturbation in Human Leukemia HL-60 Cells

Glaucocalyxin (Gla) A-C are major ent-kauranoid diterpenoids isolated from Rabdosia japonica var. glaucocalyx, a plant used in Chinese traditional medicine as an antitumor and anti-inflammatory agent. The present investigation was carried out to observe whether cellular reduced glutathione (GSH) plays important roles in Gla -induced cytotoxicity. Among major ent-kauranoid diterpenoids isolated, Gla A and B dose-dependently decreased the growth of HL-60 cells with an IC50 of approximately 6.15 and 5.86 µM at 24 h, respectively. Both Gla A and B could induce apoptosis, G2/M-phase cycle arrest, DNA damage and the accumulation of reactive oxygen species (ROS) in HL-60 cells. Moreover, Gla A, B caused rapid decrease of the intracellular GSH content, while inhibition of cellular GSH synthesis by buthionine sulfoximine (BSO) augmented the induced cytotoxicity and apoptosis in HL-60 cells. On the other hand, the administration of GSH or GSH precursor N-acetyl-cysteine (NAC) could rescue Gla A, B-depleted cellular GSH, and abrogate the induced cytotoxicity, G2/M-phase cycle arrest, DNA damage and ROS accumulation in HL-60 cells. Furthermore, Gla A, B decreased the activity of the GSH-related enzymes including glutathione reductase (GR) and glutathione peroxidase (GPX). These data suggest that the intracellular GSH redox system plays important roles in regulating the Gla A, B-induced cytotoxicity on HL-60 cells.