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

Resveratrol is a naturally occurring polyphenol with cancer chemopreventive properties. The objective of the current study was to investigate the effe ct 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 mu mol/L) as assessed by crystal violet assay, [H-3]thymidine and [C-14] leucine incorporation. Furthermore, apoptosis was determined by measuring caspase-3 activity, which increased significantly after 24 and 48 h of treatment with 200 mu mol/L resveratrol. Perturbed cel l cycle progression from the S to G2 phase was observed for concentrations up to 50 mu mol/L, whereas higher concentrations led to reversal of the S p hase arrest. These effects were specific for resveratrol; they were not obs erved after incubation with the stilbene analogs stilbenemethanol and rhapo ntin. Levels of cyclin DI and cyclin-dependent kinase (cdk) 4 proteins were decreased, as revealed by immunoblotting. In addition, resveratrol enhance d the expression of cyclin E and cyclin A. The protein levels of cdk2, cdk6 and proliferating cell nuclear antigen were unaffected. Similar results we re obtained for the colon carcinoma cell line HCT-116, indicating that cell cycle inhibition by resveratrol is independent of cyclooxygenase inhibitio n. The phosphorylation state of the retinoblastoma protein in Caco-2 cells was shifted from hyperphosphorylated to hypophosphorylated at 200 mu mol/L, which may account for reversal of the S phase block at concentrations exce eding 50 mu mol/L. These findings suggest that resveratrol exerts chemoprev entive effects on colonic cancer cells by inhibition of the cell cycle.