Efficacy of resveratrol metabolites on colon cancer cell growth

Polycarpou, Elena (2013) Efficacy of resveratrol metabolites on colon cancer cell growth. (PhD thesis), Kingston University, .


Resveratrol is a natural polyphenol present in many plant species and derived foods including grapes and red wine. It has been shown to possess chemotherapeutic properties in animal cancer models as well as many biological effects in vitro. In this study, the effects of three resveratrol metabolites including resveratrol-3-0-D-glucuronide, resveratrol-4'-O-D-glucuronide and resveratrol-3-0-D-sulphate on the growth of colon cancer cell lines have been investigated. The growth inhibitory effects of resveratrol, piceatannol, pterostilbene and the glucuronide and sulphate metabolites on Caco-2, CCL-228 and HCT-116 cells were assessed using the neutral red and MTT assays. Resveratrol and its metabolites inhibited the growth of cells with ICso values in the range of 9.8-31 f.lM whilst piceatannol and pterostilbene in the range of 21.7-29.6f.lM and 5-34.5f.lM respectively. Apoptotic assessment by DAPI staining, Z-VAD-FMK pre-treatment and percentage of cells in sub-Gl by FACS all revealed the absence of apoptosis. Treatment of differentiated Caco-2 mono layers showed that resveratrol was capable of inhibiting the growth of cells following treatment on the apical but not basolateral membrane and the effects were less profound with the metabolites. Only high concentrations (500f.lM) of metabolites (not used in any ofthe growth studies) appeared to be toxic to normal cells as measured by a haemolysis assay. Resveratrol was capable of causing S phase arrest in all 3 cell lines at 30f.lM whilst the two glucuronides caused GO/GI arrest in Caco-2 and CCL-228 cells only. Resveratrol-3-0-D-sulphate had no effect on the cell cycle. Growth inhibition caused by resveratrol and its two glucuronides was reversed by the addition of an AMP kinase inhibitor (compound C) or an adenosine A3 receptor antagonist (MRS-119l). Treatment with the highly selective A3 receptor agonist, 2CI-m-MECA caused growth inhibition and the A3 receptor was 'detected in all 3 cell lines. Levels of eyelin D 1 as measured by western blot were significantly reduced at higher concentrations (lOOf.lM) but p-AMPK was not reliably increased in all cases. Resveratrol glucuronides were shown to inhibit the growth of three colon cancer cell lines by GO/G 1 arrest and depletion of eyelin D 1. These findings strongly suggest a role of the adenosine A3 receptor in the observed inhibition therefore, providing a novel target for resveratrol and its metabolites.

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