Ceramide mimics tumour necrosis factor-alpha in the induction of cell cycle arrest in endothelial cells - Induction of the tumour suppressor p53 withdecrease in retinoblastoma/protein levels

Tumour necrosis factor (TNF)-alpha induces a transient increase in N-octano ylsphingosine (C8-ceramide) which has been postulated as an intracellular m ediator in TNF-alpha signalling. We tested the ability of C8-ceramide to re produce the TNF-alpha-mediated interference with endothelial cell prolifera tion and DNA synthesis. TNF-alpha (10 ng.mL(-1)) and C8-ceramide (20 mu m) inhibited the incorporation of [H-3]thymidine into DNA and led to an accumu lation of cells in the G(1) phase of the cell cycle. When the responses of the tumour suppressors p53 and RB were analysed, it was found that TNF-alph a and C8-ceramide induced increased expression of p53. Treatment with TNF-a lpha or C8-ceramide lead to a significant decrease in total retinoblastoma protein (RB) content that correlated with high levels of p53. These results suggest that p53 and RB may complement each other in their contribution to cell cycle arrest. TNF-alpha prevented RB phosphorylation whereas C8-ceram ide did not interfere with this process, suggesting that it follows a ceram ide-independent pathway.