Synthesis of the oxysterol, 24(S), 25-epoxycholesterol, parallels cholesterol production and may protect against cellular accumulation of newly-synthesized cholesterol

doi:10.1186/1476-511X-6-10

Lipids in Health and Disease

Volume 6

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Synthesis of the oxysterol, 24(S), 25-epoxycholesterol, parallels cholesterol production and may protect against cellular accumulation of newly-synthesized cholesterol

Jenny Wong¹ , Carmel M Quinn² and Andrew J Brown¹

¹School of Biotechnology and Biomolecular Sciences, The University of New South Wales, Sydney, Australia

²Centre for Vascular Research at The University of New South Wales and Department of Haematology, Prince of Wales Hospital, Sydney, Australia

author email corresponding author email

Lipids in Health and Disease 2007, 6:10doi:10.1186/1476-511X-6-10


Published:	5 April 2007

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Additional file 1:

Validation of TLC/phosphorimaging approach for determining cholesterol and 24,25EC synthesis. (C) CHO-7 cells were incubated with [1-¹⁴C]-acetate for 24 h under conditions of partial OSC inhibition to yield comparable levels of cholesterol and 24,25EC. Samples were pooled and neutral lipids extracted. Varying amounts of the extracts were applied to a thin-layer chromatography plate and the greatest amount was assigned an arbitrary relative concentration of 1. After development, bands corresponding to authentic cholesterol and 24,25EC were visualized by phosphorimager and quantified by densitometry (A, B). Alternatively, bands were cut, eluted and quantified by scintillation counting (D, E). Equations for the lines of best fit (A) y = 1.01x; R²= 0.99; (B) y = 1.00x; R²= 0.99; (D) y = 1.00x; R²= 0.99; (E) y = 1.00x; R²= 0.99.

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24(S),25-epoxycholesterol synthesis is increased in HepG2 cells treated with GW534511X. HepG2 cells were incubated with increasing concentrations of GW534511X in the presence of [1-¹⁴C]-acetate for 24 h. Neutral lipid extracts were separated by thin-layer chromatography and bands corresponding to authentic cholesterol and 24,25EC were visualized by phosphorimager.

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