Sterol carrier protein-2 (SCP-2) involvement in cholesterol hydroperoxide cytotoxicity as revealed by SCP-2 inhibitor effects

• Published in: Journal of lipid research 2010-11, Vol.51 (11), p.3174-3184
• Main Authors: Kriska, Tamas, Pilat, Anna, Schmitt, Jared C., Girotti, Albert W.
• Format: Article
• Language: English
• Subjects: Acetanilides - metabolism; Acetanilides - pharmacology; Animals; Apoptosis - drug effects; Benzoxazines - chemistry; Benzoxazines - metabolism; Benzoxazines - pharmacology; Biological Transport - drug effects; Carrier Proteins - antagonists & inhibitors; Carrier Proteins - metabolism; Cell Line; Cholesterol - analogs & derivatives; Cholesterol - metabolism; Cholesterol - toxicity; Humans; Hydrogen Peroxide - metabolism; Hydrogen Peroxide - toxicity; lipid peroxidation; lipid peroxide translocation; Membrane Potential, Mitochondrial - drug effects; Mice; oxidative stress; Rats; Reactive Oxygen Species - metabolism; Thiazoles - metabolism; Thiazoles - pharmacology
• ISSN: 0022-2275; 1539-7262
• DOI: 10.1194/jlr.M008342

Sterol carrier protein-2 (SCP-2) plays an important role in cholesterol trafficking and metabolism in mammalian cells. The purpose of this study was to determine whether SCP-2, under oxidative stress conditions, might also traffic hydroperoxides of cholesterol, thereby disseminating their cytotoxic effects. Two inhibitors, SCPI-1 and SCPI-3, known to block cholesterol binding by an insect SCP-2, were used to investigate this. A mouse fibroblast transfectant clone (SC2F) overexpressing SCP-2 was found to be substantially more sensitive to apoptotic killing induced by liposomal 7α-hydroperoxycholesterol (7α-OOH) than a wild-type control. 7α-OOH uptake by SC2F cells and resulting apoptosis were both inhibited by SCPI-1 or SCPI-3 at a subtoxic concentration. Preceding cell death, reactive oxidant accumulation and loss of mitochondrial membrane potential were also strongly inhibited. Similar SCPI protection against 7α-OOH was observed with two other types of SCP-2-expressing mammalian cells. In striking contrast, neither inhibitor had any effect on H2O2-induced cell killing. To learn whether 7α-OOH cytotoxicity is due to uptake/transport by SCP-2, we used a fluorescence-based competitive binding assay involving recombinant SCP-2, NBD-cholesterol, and SCPI-1/SCPI-3 or 7α-OOH. The results clearly showed that 7α-OOH binds to SCP-2 in SCPI-inhibitable fashion. Our findings suggest that cellular SCP-2 not only binds and translocates cholesterol but also cholesterol hydroperoxides, thus expanding their redox toxicity and signaling ranges under oxidative stress conditions.