Role of ceramide in cellular senescence.
Recently the sphingomyelin cycle, involving the hydrolysis of membrane sphingomyelin by an activated sphingomyelinase to generate ceramide, has emerged as a key pathway in cell differentiation and apoptosis in leukemic and other cell types. Here we investigate a role for this pathway in the senescence of WI-38 human diploid fibroblasts (HDF). We found that endogenous levels of ceramide increased considerably (4-fold) and specifically (compared with other lipids) as cells entered the senescent phase. Investigation of the mechanism of increased ceramide led to the discovery that neutral sphingomyelinase activity is elevated 8-10 fold in senescent cells. There were no changes in sphingomyelinase activity or ceramide levels as HDF entered quiescence following serum withdrawal or contact inhibition. Thus, the activation of the sphingomyelinase/ceramide pathway in HDF is due to senescence and supports the hypotheses that senescence represents a distinct program of cell development that can be differentiated from quiescence. Additional studies disclosed the ability of ceramide to induce a senescent phenotype. Thus, when exogenous ceramide (15 microM) was administered to young WI-38 HDF, it produced endogenous levels comparable to those observed in senescent cells (as determined by metabolic labeling studies). Ceramide concentrations of 10-15 microM inhibited the growth of young HDF and induced a senescent phenotype by its ability to inhibit DNA synthesis and mitogenesis. These concentrations of ceramide also induced retinoblastoma dephosphorylation and inhibited serum-induced AP-1 activation in young HDF, thus recapitulating basic biochemical and molecular changes of senescence. Sphingomyelinase and ceramide may thus be implicated as mediators of cellular senescence.
Venable
M E
ME
Department of Medicine, Duke University Medical Center, Durham, NC 27710, USA.
Lee
J Y
JY
Smyth
M J
MJ
Bielawska
A
A
Obeid
L M
LM
D001483:Base Sequence; D001665:Binding Sites; D016923:Cell Death; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D016922:Cellular Senescence / Q000187:drug effects / Q000502:physiology*; D002518:Ceramides / Q000378:metabolism / Q000494:pharmacology* / Q000502:physiology*; D003260:Contact Inhibition; D016895:Culture Media, Serum-Free; D004247:DNA / Q000096:biosynthesis; D006801:Humans; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D013108:Sphingomyelin Phosphodiesterase / Q000378:metabolism*; D013936:Thymidine / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism
D002518:Ceramides; D016895:Culture Media, Serum-Free; D009838:Oligodeoxyribonucleotides; D018808:Transcription Factor AP-1; D004247:DNA; D013108:Sphingomyelin Phosphodiesterase; D013936:Thymidine