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. Author manuscript; available in PMC: 2011 May 3.

Published in final edited form as: Biochem J. 2010 Mar 29;427(2):265–274. doi: 10.1042/BJ20091589

SMS-KCNR cells were treated with indicated concentrations of hydrogen peroxide, menadione and tert-butylhydroperoxide for 1h. Levels of endogenous ceramide and dihydroceramide (A,B) as well as dihydroceramide desaturase activity (C) were measured by LC/MS. Average of 2 independent experiments performed in duplicates. ***p<0.001. (D) In situ assay with addition of substrate (C12-dhCCPS) together with inhibitors (left panel) or for 1h after pretreatment with inhibitors (right panel). (E) After treatment with menadione and tert-butylhydroperoxide levels of endogenous DEGS-1 were determined by Western blot. Blots were stripped and re-probed for β-actin to normalize for loading. Insets are representative of two independent experiments.

SMS-KCNR cells were treated with indicated concentrations of hydrogen peroxide, menadione and tert-butylhydroperoxide for 1h. Levels of endogenous ceramide and dihydroceramide (A,B) as well as dihydroceramide desaturase activity (C) were measured by LC/MS. Average of 2 independent experiments performed in duplicates. ***p<0.001. (D) In situ assay with addition of substrate (C12-dhCCPS) together with inhibitors (left panel) or for 1h after pretreatment with inhibitors (right panel). (E) After treatment with menadione and tert-butylhydroperoxide levels of endogenous DEGS-1 were determined by Western blot. Blots were stripped and re-probed for β-actin to normalize for loading. Insets are representative of two independent experiments.