ToxSci Advance Access published online on December 4, 2007
Toxicological Sciences, doi:10.1093/toxsci/kfm292
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Apoptosis of Cultured Astrocytes Induced by the Copper and Neocuproine Complex Through Oxidative Stress and JNK Activation
a Institutes of Toxicology b Pharmacology c Biochemistry College of Medicine, National Taiwan University, Taipei, Taiwan
* Corresponding author. Institute of Pharmacology, College of Medicine, National Taiwan University, Section 1, Jen-Ai Road, No. 1, Taipei, 10043, Taiwan. Fax. No.: +886-2-23915297, Email: syl{at}ha.mc.ntu.edu.tw, Tel. No.: +886-2-23123456-ext 8313
Received September 8, 2007; revision received November 29, 2007; accepted November 29, 2007
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Abstract |
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Astrocytes play a critical neurotrophic and neuroprotective role in the brain, and improper function of these cells may contribute to the onset of neurodegenerative diseases. Since astrocytes are known to be enriched with Cu chaperone proteins, it is important to understand the factors that may lead to cytotoxic effects of Cu on astrocytes. In this report, we demonstrated a dramatic potentiating effect of neocuproine (NCP), a membrane permeable metal chelator, on Cu, but not Fe or Pb, in inducing apoptosis of cultured astrocytes. It was estimated that individually, CuCl2 and NCP only weakly exhibited cytotoxic effects on astrocytes, with EC50 of 180 µM and 600 µM respectively. However, NCP at a nontoxic concentration of 10 µM markedly reduced EC50 of Cu to 0.35 µM (physiological concentration) and Cu (10 µM) reduced EC50 of NCP down to 0.06 µM. The mechanisms underlying these dramatic potentiation effects are elucidated. NCP increased the intracellular concentration of Cu in astrocytes and a non-permeable Cu chelator, bathocuproine disulfonate (BCPS) was able to abolish all of the apoptotic signaling. Cell death was determined to be via apoptosis due to increased ROS production, mitochondrial dysfunction, depletion of GSH and ATP, cytochrome c release, JNK and caspase-3 activation, and PARP degradation. This finding, coupled with our previous reports, suggests that metal chelators (NCP, dithiocarbamate and disulfiram) should be cautiously used as they may potentiate a cytotoxic effect of endogenous Cu on astrocytes. Their clinical implications in the etiology of neurodegenerative diseases deserve further investigation.
Key Words: neocuproine; Cu; apoptosis; astrocytes; oxidative stress; JNK; caspase-3.