De novo ceramide biosynthesis is associated with resveratrol-induced inhibition of ornithine decarboxylase activity.
Previous studies could demonstrate, that the naturally occuring polyphenol resveratrol inhibits cell growth of colon carcinoma cells at least in part by inhibition of protooncogene ornithine decarboxylase (ODC). The objective of this study was to provide several lines of evidence suggesting that the induction of ceramide synthesis is involved in this regulatory mechanisms. Cell growth was determined by BrdU incorporation and crystal violet staining. Ceramide concentrations were detected by HPLC-coupled mass-spectrometry. Protein levels were examined by Western blot analysis. ODC activity was assayed radiometrically measuring [(14)CO(2)]-liberation. A dominant-negative PPARgamma mutant was transfected in Caco-2 cells to suppress PPARgamma-mediated functions. Antiproliferative effects of resveratrol closely correlate with a dose-dependent increase of endogenous ceramides (p<0.001). Compared to controls the cell-permeable ceramide analogues C2- and C6-ceramide significantly inhibit ODC-activity (p<0.001) in colorectal cancer cells. C6-ceramide further diminished protein levels of protooncogenes c-myc (p<0.05) and ODC (p<0.01), which is strictly related to the ability of ceramides to inhibit cell growth in a time- and dose-dependent manner. These results were further confirmed using inhibitors of sphingolipid metabolism, where only co-incubation with a serine palmitoyltransferase (SPT) inhibitor could significantly counteract resveratrol-mediated actions. These data suggest that the induction of ceramide de novo biosynthesis but not hydrolysis of sphingomyelin is involved in resveratrol-mediated inhibition of ODC. In contrast to the regulation of catabolic spermidine/spermine acetyltransferase by resveratrol, inhibitory effects on ODC occur PPARgamma-independently, indicating independent pathways of resveratrol-action. Due to our findings resveratrol could show great chemopreventive and therapeutic potential in the treatment of colorectal cancers.
Ulrich
Sandra
S
First Department of Internal Medicine-ZAFES, Johann Wolfgang Goethe University, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany.
Huwiler
Andrea
A
Loitsch
Stefan
S
Schmidt
Helmut
H
Stein
Jürgen M
JM
D018938:Caco-2 Cells; D049109:Cell Proliferation / Q000187:drug effects; D002518:Ceramides / Q000096:biosynthesis*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D009955:Ornithine Decarboxylase / Q000378:metabolism; D065108:Ornithine Decarboxylase Inhibitors; D047495:PPAR gamma / Q000502:physiology; D016271:Proto-Oncogene Proteins c-myc / Q000032:analysis; D000077185:Resveratrol; D013096:Spermine / Q000494:pharmacology; D013267:Stilbenes / Q000494:pharmacology*
D002518:Ceramides; D004791:Enzyme Inhibitors; D065108:Ornithine Decarboxylase Inhibitors; D047495:PPAR gamma; D016271:Proto-Oncogene Proteins c-myc; D013267:Stilbenes; D013096:Spermine; D009955:Ornithine Decarboxylase; D000077185:Resveratrol