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ARS Home » Plains Area » Grand Forks, North Dakota » Grand Forks Human Nutrition Research Center » Healthy Body Weight Research » Research » Publications at this Location » Publication #153839
Title: SUPPLEMENTAL ASCORBATE OR ALPHA-TOCOPHEROL INDUCES CELL DEATH IN CU-DEFICIENT HL-60 CELLS

Author
item RAYMOND, LAURA - ENERGY & ENVIRON RES CTR
item Johnson, William

Submitted to: Experimental Biology and Medicine
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/8/2004
Publication Date: 10/1/2004
Citation: Raymond, L.J., Johnson, W.T. 2004. Supplemental ascorbate or alpha-tocopherol induces cell death in Cu-deficient HL-60 cells. Experimental Biology and Medicine. 229:885-894.

Interpretive Summary: Mitochondria are cellular components that provide energy for a number of biological processes. In order to provide this energy, mitochondria convert the oxygen we breathe into high energy chemical compounds. However, instead of being converted into high energy compounds, some of the oxygen utilized by mitochondria is instead converted to chemical forms, called reactive oxygen intermediates (ROS), that can damage cellular proteins, lipids, and DNA. An important component of mitochondrial oxygen metabolism is a copper-containing enzyme called cytochrome c oxidase. When the activity of cytochrome c oxidase is lowered, as is the case during copper deficiency, the production of ROS by mitochondria and the potential for oxidative damage are both increased. ROS produced by mitochondria can lead to oxidative stress which can trigger a program for cell death known as apoptosis. It was found that mitochondria obtained from copper-deprived human leukemia cells had lowered cytochrome c oxidase activity and elevated manganese superoxide dismutase (MnSOD) content. MnSOD helps protect mitochondria from oxidative damage and its content is dependent on mitochondrial ROS production. Thus, the increase in MnSOD content indicates that copper deprivation elevated mitochondrial ROS production. Copper deprivation also disrupted the voltage across the mitochondrial inner membrane indicating that mitochondrial damage occurred as a result of the oxidative stress resulting from increased mitochondrial ROS production. However, the damage was not sufficient to trigger apoptosis. Vitamins C and E, which normally function to protect cells against oxidative damage, stimulated apoptosis in cells deprived of copper. This suggests that in addition to protecting cells from ROS, vitamin C and vitamin E may also facilitate the elimination of oxidatively stressed and damaged cells by stimulating apoptosis.

Technical Abstract: Cytochrome c oxidase (CCO) is the Cu-dependent, terminal respiratory complex of the mitochondrial electron transport chain. Inhibition of CCO can promote oxidative stress by increasing mitochondrial production of reactive oxygen species (ROS). As mitochondria have an important role in apoptosis as both a target and source for ROS, enhanced ROS production resulting from inhibition of CCO by Cu deficiency may trigger apoptosis. The present study focuses on the mitochondrial effects of a compound, TET, which inhibits CCO by chelating Cu and causing cellular Cu deficiency, and the antioxidants, ascorbate and alpha-tocopherol, in a human promyelocytic leukemia cell line (HL-60). The following effects were observed: (1) TET reduced both cell growth and viability only in the presence of ascorbate or alpha-tocopherol; (2) TET reduced CCO activity and increased mitochondrial ROS production as indicated by increased expression of Mn superoxide dismutase, but the induction of Mn superoxide dismutase was not affected by ascorbate or alpha-tocopherol; (3) TET acted independently of ascorbate or alpha-tocopherol in disrupting mitochondrial membrane potential; (4) TET did not increase caspase 8 activity in the absence of ascorbate or alpha-tocopherol; and (5) TET did not increase transfer of cytochrome c from mitochondria to cytosol unless alpha-tocopherol was present. These findings indicate that reduction in CCO activity by TET-induced Cu deficiency increased oxidative stress in HL-60 cells sufficiently to disrupt the electrochemical gradient of the inner mitochondrial membrane but did not trigger apoptosis. Also, ascorbate and alpha-tocopherol did not alleviate oxidative stress but may have become pro-oxidants, adding to the oxidant burden sufficiently to trigger apoptosis in TET-treated cells.