Role of ceramide in lipopolysaccharide (LPS)-induced signaling - LPS increases ceramide rather than acting as a structural homolog

Ceramide and ceramide activated enzymes have been implicated in responses to bacterial lipopolysaccharide (LPS) and the proinflammatory cytokines tumor necrosis factor-alpha (TNF) and interleukin-1 beta (IL-1). Although TNF and IL-1 cause elevation of cellular ceramide, which is thought to act as a second messenger, LPS has been proposed to signal by virtue of structural similarity to ceramide. We have investigated the relationship between ceramide and LPS by comparing the effects of a cell-permeable ceramide analog (C-2-ceramide) and LPS on murine macrophage cell lines and by measuring ceramide levels in macrophages exposed to LPS, We found that while both C-2-ceramide and LPS activated c-Jun N-terminal kinase (JNK), only LPS also activated extracellular signal-regulated kinases (ERKs). C-2-ceramide was also unable to activate NF-kappa B, a transcription factor important for LPS-induced gene expression. Upon measurement of cellular ceramide in macrophage lines, we observed a small but rapid rise in ceramide, similar to that seen upon IL-1 or TNF treatment, suggesting LPS induces an increase in ceramide rather than interacting directly with ceramide-responsive enzymes. We found that C-2-ceramide activated JNK and induced growth arrest in macrophages cell lines from both normal mice (Lps(n)) and mice genetically unresponsive to LPS (Lps(d)), whereas only Lps(n) macrophages made these responses to LPS. Surprisingly, LPS treatment of Lps(d) macrophages induced a rise in ceramide similar to that observed in LPS-responsive cells. These results indicate that the wild type Lps allele is not required for LPS-induced ceramide generation and suggest that ceramide elevation alone is insufficent stimulus for most responses to LPS.