Downregulation of the cyclin D1/Cdk4 complex occurs during resveratrol-induced cell cycle arrest in colon cancer cell lines

Abstract

Resveratrol is a naturally occurring polyphenol with cancer chemopreventive properties. The objective of the current study was to investigate the effect of resveratrol on the human colonic adenocarcinoma cell line Caco-2. The compound inhibited cell growth and proliferation of Caco-2 cells in a dose-dependent manner (12.5-200 μmol/L) as assessed by crystal violet assay, [3H]thymidine and [14C]leucine incorporation. Furthermore, apoptosis was determined by measuring caspase-3 activity, which increased significantly after 24 and 48 h of treatment with 200 μmol/L, resveratrol. Perturbed cell cycle progression from the S to G2 phase was observed for concentrations up to 50 μmol/L, whereas higher concentrations led to reversal of the S phase arrest. These effects were specific for resveratrol; they were not observed after incubation with the stilbene analogs stilbenemethanol and rhapontin. Levels of cyclin D1 and cyclin-dependent kinase (cdk) 4 proteins were decreased, as revealed by immunoblotting. In addition, resveratrol enhanced the expression of cyclin E and cyclin A. The protein levels of cdk2, cdk6 and proliferating cell nuclear antigen were unaffected. Similar results were obtained for the colon carcinoma cell line HCT-116, indicating that cell cycle inhibition by resveratrol is independent of cyclooxygenase inhibition. The phosphorylation state of the retinoblastoma protein in Caco-2 cells was shifted from hyperphosphorylated to hypophosphorylated at 200 μmol/L, which may account for reversal of the S phase block at concentrations exceeding 50 μmol/L. These findings suggest that resveratrol exerts chemopreventive effects on colonic cancer cells by inhibition of the cell cycle.