Regulatory role of ceramide in interleukin (IL)-1 beta-induced E-selectin expression in human umbilical vein endothelial cells - Ceramide enhances IL-1 beta action, but is not sufficient for E-selectin expression

Recent studies indicate that sphingolipids mediate several cellular processes, We assessed roles of sphingolipids in the regulation of E-selectin expression in human umbilical vein endothelial cells, All exogenously-added sphingolipids (sphingosine, C-2-ceramide, sphingosine 1-phosphate, and N,N-dimethylsphingosine) failed to induce E-selectin expression by themselves. C-2-ceramide at 5 mu M enhanced interleukin-1 beta (IL-1 beta)-induced E-selectin expression 2.7-fold, whereas other sphingolipids tested had no effects on this process, Sphingomyelinase, but not phospholipases A(2), C, or D, mimicked the enhancing effect of C-2-ceramide, Northern blot analyses revealed that C-2-ceramide and sphingomyelinase increased interleukin-1 beta-induced E-selectin gene transcription levels, C-2-ceramide and sphingomyelinase induced NF-kappa B activation by themselves and enhanced activation by IL-1 beta, which is essential for E-selectin expression, Immunological analyses with anti-NF-kappa B antibodies showed that subunit composition of NF-kappa B activated by IL-1 beta differs from that activated by C-2-ceramide, suggesting that signaling pathways utilized by these stimuli may be different. Treatment with C-2-ceramide or sphingomyelinase did not alter NF-ELAM1 specific binding activity, IL-1 beta induced sphingomyelin hydrolysis to ceramide; intracellular ceramide level increased to 182% of control value at 30 min. Taken together, these findings suggest that (i) sphingomyelin hydrolysis to ceramide does not trigger, but rather enhances cytokine-induced E-selectin expression, ill part through NF-kappa B; (ii) sphingomyelin hydrolysis to ceramide does not mediate all the effects of IL-1 beta, although it may play important roles in IL-1 beta signal transduction in human umbilical vein endothelial cells.