Investigation of the effects of hydroxytyrosol on cytotoxicity, apoptosis, Pi3K/Akt and ERK 1/2 pathways in ovarian cancer cell cultures

It has been shown that hydroxytyrosol (HT) has antitumoral, anti-proliferative and apoptotic effects on various cancer cell types. It was also studied in several tumor cell lines in terms of its action mechanisms. In this study, we aimed to evaluate the cytotoxic, apoptotic effects of hydroxytyrosol and whether its cytotoxic effect on through phosphatidylinositol 3- kinase / Akt (PI3K/Akt) and extracellular signal-regulated kinase 1/2 (ERK 1/2) signal transduction pathways in human ovarian cancer cell cultures OVCAR-3 and MDAH-2774. To investigate the cytotoxic effect of increasing concentrations of HT (50-400 µM) XTT assay was used in OVCAR-3 and MDAH-2774 cell lines at 24., 48.,72. and 96. hours. To determine the apoptotic effect of HT, DNA fragmentation assay and protein antibody array methods were used. XTT assay was used to determine whether hydroxytyrosol is effective on PI3K/Akt and ERK 1/2 signal tranduction pathways, specific inhibitors of these pathways were used. The findings were evaluated by ANOVA followed by Dunnett's statistical analyzes. IC50 value of HT in MDAH-2774 cell cultures was calculated as 102.3 µM at 72 h and 51.5 µM in OVCAR-3 cell cultures at 96 h. Apoptotic effect of HT in MDAH-2774 cells at 72 h was the highest at 100 µM, and kept lowering at 150 µM concentrations and was not seen at 200 µM and higher concentrations. Highest apoptotic effect was seen at 100 µM concentration in OVCAR-3 cells at 96 h, however apoptotic effect was decreased over 100 µM concentrations. In our study, HT increased the levels of pro-apoptotic molecules Bad, Bax, active caspase-3, Htra2/Omi and SMAC/DIABLO and also increased the levels of pro-apoptotic death receptors TRAIL R1/DR4, TRAIL R2/DR5 and FAS/TNFRSF6, however, it decreased the levels of Survivin which is one of the inhibitor of apoptosis protein (IAP) family in MDAH-2774 cells. In OVCAR-3 cells, HT decreased the levels of anti-apoptotic proteins Bcl-, pro-caspase 3 and IAP family proteins CIAP-1, CIAP-2, XIAP, Livin and Survivin and increased the levels of cytochrome c. We have shown that cytotoxic effect of HT is not through PI3K/Akt signaling pathway however, it may effect through ERK1/2 signaling pathway in both OVCAR-3 and MDAH-2774 cells. These findings show that HT has cytotoxic and apoptotic effects on ovarian cancer cell lines and it may show its cytotoxic effect through ERK 1/2 signalling pathway.