Intracellular glutathione depletion and reactive oxygen species generation are important in -hederin-induced apoptosis of P388 cells

Authors: Huat B.T.K.¹; Swamy S.M.K.²

Source: Molecular and Cellular Biochemistry, Volume 245, Numbers 1-2, March 2003 , pp. 127-139(13)

Publisher: Springer

Abstract:

-Hederin, a pentacyclic triterpene saponin isolated from the seeds of Nigella sativa, was recently reported to have potent in vivo antitumor activity against LL/2 (Lewis Lung carcinoma) in BDF1 mice. In this study we observed that -hederin caused a dose- and time-dependent increase in apoptosis of murine leukemia P388 cells. In order to evaluate the possible mechanisms for apoptosis, the effects of -hederin on intracellular thiol concentration, including reduced glutathione (GSH), and protein thiols, and the effects of pretreatment with N-acetlycysteine (NAC), a precursor of intracellular GSH synthesis, or buthionine sulfoxime (BSO), a specific inhibitor of intracellular GSH synthesis, on -hederin-induced apoptosis were investigated. It was found that -hederin rapidly depleted intracellular GSH and protein thiols prior to the occurrence of apoptosis. NAC significantly alleviated -hederin-induced apoptosis, while BSO augmented -hederin-induced apoptosis significantly. The depletion of cellular thiols observed after -hederin treatment caused disruption of mitochondrial membrane potential (_m) and subsequently increased the production of reactive oxygen species (ROS) in P388 cells at an early time point. Bongkrekic acid (BA), a ligand of the mitochondrial adenine nucleotide translocator, and cyclosporin (CsA) attenuated the -hederin-induced loss of _m, and ROS production. Thus, oxidative stress after -hederin treatment is an important event in -hederin-induced apoptosis. As observed in this study, permeability transition of mitochondrial membrane occurs after depletion of GSH and precedes a state of reactive oxygen species (ROS) generation. Further, we observed that -hederin caused the release of cytochrome c from the mitochondria to cytosol, leading to caspase-3 activation. Our findings thus demonstrate that changes in intracellular thiols and redox status leading to perturbance of mitochondrial functions are important components in the mechanism of -hederin-induced cell death.

Keywords: Nigella sativa; alpha-hederin; triterpene; reduced glutathione; ROS; apoptosis

Language: English

Document Type: Research article

Affiliations: 1: Department of Pharmacology, Faculty of Medicine, National University of Singapore, Singapore 119260 E-mail: phctankh@nus.edu.sg 2: Department of Pharmacology, Faculty of Medicine, National University of Singapore, Singapore 119260

Publication date: 2003-03-01

• In this: publication
• By this: publisher
• In this Subject: Anatomy & Physiology ,  Biology ,  Biochemistry
• By this author: Huat B.T.K. ;  Swamy S.M.K.

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