title: Ceramide induces a loss in cytosolic peroxide levels in mononuclear cells.
abstract: Ceramide (a sphingolipid) and reactive oxygen species are each partly responsible for intracellular signal transduction in response to a variety of agents. It has been reported that ceramide and reactive oxygen species are intimately linked and show reciprocal regulation [Liu, Andreieu-Abadie, Levade, Zhang, Obeid and Hannun (1998) J. Biol. Chem. 273, 11313-11320]. Utilizing synthetic, short-chain ceramide to mimic the cellular responses to fluctuations in natural endogenous ceramide formation or using stimulation of CD95 to induce ceramide formation, we found that the principal redox-altering property of ceramide is to lower the [peroxide](cyt) (cytosolic peroxide concentration). Apoptosis of Jurkat T-cells, primary resting and phytohaemagglutinin-activated human peripheral blood T-lymphocytes was preceded by a loss in [peroxide](cyt), as measured by the peroxide-sensitive probe 2',7'-dichlorofluorescein diacetate (also reflected in a lower rate of superoxide dismutase-inhibitable cytochrome c reduction), and this was not associated with a loss of membrane integrity. Where growth arrest of U937 monocytes was observed without a loss of membrane integrity, the decrease in [peroxide](cyt) was of a lower magnitude when compared with that preceding the onset of apoptosis in T-cells. Furthermore, decreasing the cytosolic peroxide level in U937 monocytes before the application of synthetic ceramide by pretreatment with either of the antioxidants N -acetyl cysteine or glutathione conferred apoptosis. However, N -acetyl cysteine or glutathione did not affect the kinetics or magnitude of ceramide-induced apoptosis of Jurkat T-cells. Therefore the primary redox effect of cellular ceramide accumulation is to lower the [peroxide](cyt) of both primary and immortalized cells, the magnitude of which dictates the cellular response.
authors.affiliation: Molecular Biosciences Group, School of Life and Health Sciences, Aston University, Aston Triangle, Birmingham B4 7ET, UK.
authors.forename: Darren C
authors.initials: DC
authors.lastname: Phillips
authors.forename: Helen R
authors.initials: HR
authors.lastname: Griffiths
mesh_terms: D017209:Apoptosis / Q000187:drug effects; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002518:Ceramides / Q000378:metabolism / Q000494:pharmacology*; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D053938:DNA Fragmentation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D053222:Fas Ligand Protein; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D016130:Immunophenotyping; D019169:Jurkat Cells; D007963:Leukocytes, Mononuclear / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000494:pharmacology; D010545:Peroxides / Q000378:metabolism*; D020298:U937 Cells
chemical_list: D002518:Ceramides; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D010545:Peroxides; D005978:Glutathione