Hydroxytyrosol, an olive oil phenolic compound, inhibits cyclooxygenase-2 transcription in phorbol--ester-treated human mocytes

Introduction: Inflammation plays a critical role in atherosclerotic plaque development and instability. Cyclooxygenase(COX)-2 is a key mediator of inflammation. It is up-regulated in activated monocytes and macrophages of human atherosclerotic lesions. The extra-virgin olive oil is a source of bioactive compounds which can be responsible of Mediterranean diet athero-protective effect. Aim of the study was to examine the effects of hydroxytyrosol (HT), a phenolic compound from olives and extra-virgin olive oil, on COX-2 expression in phorbol-ester stimulated monocytic-macrophagic cells and to explore the mechanisms involved. Methods: U937 monocytoid cells were pre-treated with HT (0-10 micromol/L) for 60 min before stimulation with 20 nmol/L phorbol myristate acetate (PMA) in RPMI 1640 medium with 5%SFB for 20 h. Cell supernatants were then tested for the release of PGE2 by EIA kit and cell extracts were analysed for COX-2 expression by Western blot analysis. COX-2 mRNA was investigated by semi-quantitative RT-PCR and COX-2 promoter activity was assessed by transient transfection of full length and partially deleted or mutagenized COX-2 promoter constructs. Results: Stimulation of U937 cells with 20 nmol/L PMA for 20 h caused a marked increase in the production of prostaglandin(PG) E2. This effect was inhibited by 1 micromol/L HT both in PMA-treated U937 (by about 25%) and even more in U937 derived-macrophages (> 40%). Pre-treatment of U937 cells with 0.1-10 micromol/L HT reduced PMA-induced COX-2 protein and mRNA in a concentration dependent manner with a 30% reduction already at 1 mmol/L (p