Ellagic acid, a polyphenolic compound found in berries, fruits and nuts, has been shown to possess growth-inhibiting and apoptosis promoting activities in cancer cell lines in vitro. The objective of this study was to investigate the effect of ellagic acid in human neuroblastoma SH-SY5Y cells. In cultures of SH-SY5Y cells incubated with ellagic acid, time- and concentration-dependent inhibitory effects on cell number were demonstrated.Ellagic acid induced cell detachment, decreased cell viability and induced apoptosis as measured by DNA strand breaks. Ellagic acid-induced alterations in cell cycle were also observed. Simultaneous treatment with all-trans retinoic acid did not rescue the cells from ellagic acid effects. Furthermore, the results suggested that pre-treatment with all-trans retinoic acid to induce differentiation and cell cycle arrest did not rescue the cells from ellagic acid-induced cell death.

The polyphenol ellagic acid is found in many natural food sources and has been proposed as a candidate compound for clinical applications due to its anti-oxidative capacity and as a potential anti-tumorigenic compound. The objective of the present study was to evaluate the sensitivity to and possible apoptosis mechanism induced by ellagic acid in neuronal tumor cells. As a model the human neuroblastoma SH-SY5Y cell line was used. The methods applied were bright field and phase contrast microscopy, XTT- and LDH-assays, western blot, and flow cytometric analysis of DNA degradation and mitochondrial membrane potential. Ellagic acid treatment was found to induce a reduction in cell number preceded by alterations of the mitochondrial membrane potential and activation of caspase-9 and -3, DNA-fragmentation and cell death by apoptosis. The apoptotic cell death studied was not due to anoikis since it was significant in the adherent fraction of the cells. We conclude that ellagic acid induces dose- and time-dependent apoptosis, at least partly by the mitochondrial pathway, in an embryonal neuronal tumor cell system. This finding is in agreement with previously reported data on adult carcinoma cells thus suggesting a more general effect of ellagic acid on tumor cells.

Ellagic acid has previously been reported to induce reduced proliferation and activation of apoptosis in several tumor cell lines including our own previous data from non-differentiated human neuroblastoma SH-SY5Y cells. The aim of this study was now to investigate if in vitro differentiation with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate or the vitamin A derivative all-trans retinoic acid altered the sensitivity to ellagic acid in SH-SY5Y cells. The methods used were cell counting and LDH-assay for evaluation of cell number and cell death, flow cytometric analysis of SubG(1)-and TUNEL-analysis for apoptosis and western blot for expression of apoptosis-associated proteins. In vitro differentiation was shown to reduce the sensitivity to ellagic acid with respect to cell detachment, loss of viability and activation of apoptosis. The protective effect was phenotype-specific and most prominent in all-trans retinoic acid-differentiated cultures. Differentiation-dependent up-regulation of Bcl-2 and integrin expression is introduced as possible protective mechanisms. The presented data also point to a positive correlation between proliferative activity and sensitivity to ellagic-acid-induced cell detachment. In conclusion, the presented data emphasize the need to consider degree of neuronal differentiation and phenotype of neuroblastoma cells when discussing a potential pharmaceutical application of ellagic acid in tumor treatment.