Minor polar components of extra-virgin olive oil: inhibition of NF-kB activity at gastric level

The transcription factor modulates the expression of several genes involved in the inflammatory process and several studies demonstrate its involvement in gastritis and peptic ulcer, in the presence or not of the bacterium H. pylori infection. In vivo studies show that the minor polar components of extra-virgin olive oil possess anti-inflammatory properties. The aim of this work was to investigate the mechanism of action of the anti-inflammatory effect, by evaluating whether the phenolic extract from extra-virgin olive oil inhibited the NF-kB driven transcription in gastric adenocarcinoma cells (AGS), and assessing the contribution of individual phenols. Phenolic extracts were obtained from two commercial extra-virgin olive oils, respectively from Italy and Spain, by extraction with MeOH:H2O, 80:20 v:v, and quantified with Folin-Ciocalteu assay. Secoiridoids (oleuropein aglycone and ligustroside aglycone) were analyzed by LC-MS, and phenolic alcohols (tyrosol and hydroxytyrosol) by GC-MS. AGS cells were transfected with luciferase reporter plasmid and stimulated with TNF as a pro-inflammatory agent. Both phenolic extracts inhibited NF-kB activity in a concentration-dependent manner (IC50 of 0.86 ± 0.12 μg/ml, and 1.28 ± 0.13 μg/ml for the Italian and Spanish oil respectively; mean + sd). Secoiridoids represented over the 70% of the phenolic extracts while phenolic alcohols corresponded to less than 7%. All individual compounds inhibited the activity of NF-kB in a statistically significant manner (-50% at 10 μM). Since hydroxytyrosol is catabolized to homovanillic alcohol by catechol-o-methyltransferase in humans, the effect of the later was also evaluated even if not present in the oil. The inhibitory effect reached statistically significant values at 1 μM. The effect of the two phenolic extracts was then compared to the reconstituted mixtures of the four constituents. Ligustroside aglycone and oleuropein aglycone, not commercially available, were prepared from their glucosides by hydrolysis with β-glucosidase. Reconstituted mixtures had an inhibitory effect comparable to that of the total phenolic extracts, indicating that the compounds are the main responsible of the activity of the extract. In conclusion, the results of this study suggest that the anti-inflammatory effect of the polar fraction of extra-virgin olive oil may be mediated by the inhibition of NF-kB activity. The concentration at which the phenolic extracts are active are compatible with a consumption of 30-50 g/die of oil, as estimated in the Mediterranean dietary regimen. Secoiridoids and phenolic alcohols, as well as homovanillic alcohol, contribute to anti-inflammatory action in gastric cells.