Increased transcription of IL-8 in endothelial cells is differentially regulated by TNF-alpha and oxidized phospholipids

Oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (Ox-PAPC ) upregulates a spectrum of inflammatory cytokines and adhesion molecules d ifferent from those induced by classic inflammatory mediators such as tumor necrosis factor-alpha (TNF-alpha) or lipopolysaccharide. Interestingly, Ox -PAPC also induces the expression of a set of proteins similar to those ind uced by TNF-alpha or lipopolysaccharide, which include the chemokines monoc yte chemotactic protein-1 (MCP-1) and interleukin (IL)-8. To elucidate the molecular mechanisms of Ox-PAPC-induced gene expression and to determine wh ether Ox-PAPC and other inflammatory mediators such as TNF-alpha utilize co mmon signaling pathways, we examined the transcriptional regulation of IL-8 by Ox-PAPC and TNF-alpha in human aortic endothelial cells. Both Ox-PAPC a nd TNF-alpha induced the expression of IL-8 mRNA in a dose-dependent fashio n; however, the kinetics of IL-8 mRNA accumulation between the 2 ligands di ffered. Ox-PAPC-induced IL-8 mRNA was seen as early as 30 minutes, peaked b etween 4 and 8 hours, and decreased substantially by 24 hours. In contrast, TNF-alpha -induced IL-8 mRNA synthesis was elevated at 30 minutes, peaked at 2 hours, and reached basal/undetectable levels by 6 hours. Actinomycin D experiments suggested that both Ox-PAPC and TNF-alpha regulate the express ion of IL-8 at the transcriptional level. Furthermore, the half-life of IL- 8 mRNA for both ligands was similar (<30 minutes), suggesting that mRNA sta bility was not responsible for the differences in the kinetics of IL-8 accu mulation between the 2 ligands. Transient transfection studies with reporte r constructs containing 1.48 kb of the IL-8 promoter identified an Ox-PAPC- specific response region between -133 and -1481 bp of the IL-8 promoter. In contrast, TNF-<alpha> activation of the IL-8 promoter was mediated almost entirely through the nuclear factor-kappaB and activation protein-1 respons e elements present between -70 and -133 bp of the IL-8 promoter. Thus, alth ough Ox-PAPC and TNF-a both induced IL-8 synthesis, our data suggest that t he 2 ligands utilize different mechanisms in the regulation of IL-8 transcr iption.