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Journal of Lipid Research, Vol. 49, 2124-2134, October 2008 Compartmentalization of stearoyl-coenzyme A desaturase 1 activity in HepG2 cells*
* Department of Pediatrics, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA * This project was supported by Grant T32 DK-007571 from the National Institute of Diabetes and Digestive and Kidney Diseases and by Grant 2 K12 HD-034610 from the National Institutes of Health/National Institute of Child Health and Human Development (to J.K.Y.). The GC-MS facility is supported by Public Health Grant P01 CA-042710 to the UCLA Clinical Nutrition Research Unit and Stable Isotope Core, Public Health Grant 1P01 AT-003960 to the Metabolomics Core of the UCLA Center for Excellence in Pancreatic Diseases (to W-N.P.L.), and Public Health Grant M01 RR-00425 to the General Clinical Research Center. Published, JLR Papers in Press, July 3, 2008. 2 The cell assay was performed by incubating HepG2 cells with inhibitor and [14C]palmitate. After saponification, [14C]palmitate, [14C]stearate, and the desaturated species [14C]palmitoleate and [14C]oleate were measured by radioactivity in the respective peaks by HPLC (flow counter). Activity was calculated as the ratio of desaturated to saturated 14C fatty acid. The IC50 was the concentration calculated to give 50% of the control value.
1 To whom correspondence should be addressed. e-mail: lee{at}labiomed.org
Stearoyl-coenzyme A desaturase 1 (SCD1) catalyzes the conversion of stearate (18:0) to oleate (18:1n-9) and of palmitate (16:0) to palmitoleate (16:1), which are key steps in triglyceride synthesis in the fatty acid metabolic network. This study investigated the role of SCD1 in fatty acid metabolism in HepG2 cells using SCD1 inhibitors and stable isotope tracers. HepG2 cells were cultured with [U-13C]stearate, [U-13C]palmitate, or [1,2-13C]acetate and (1) DMSO, (2) compound CGX0168 or CGX0290, or (3) trans-10,cis-12 conjugated linoleic acid (CLA). 13C incorporation into fatty acids was determined by GC-MS and desaturation indices calculated from the respective ion chromatograms. FAS, SCD1, peroxisome proliferator-activated receptor
Supplementary key words desaturation index fatty acid metabolism HepG2 cells stable isotope
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