Inhibition of vascular smooth muscle cell proliferation by sodium salicylate mediated by upregulation of p21Waf1 and p27Kip1

BACKGROUND: Salicylates may have direct vascular effects by mechanisms that are independent of platelet inhibition. METHODS AND RESULTS: We investigated the effect of salicylates on vascular smooth muscle cell (SMC) proliferation in response to platelet-derived growth factor (PDGF) in vitro. Salicylate concentrations of 5 and 10 mmol/L inhibited serum- or PDGF-induced SMC cell count and [(3)H]thymidine incorporation by 62% and 81%, respectively. There was no evidence of cellular toxicity or apoptosis as determined by trypan blue exclusion and FACS analyses. Because cell cycle progression is regulated by hyperphosphorylation of the retinoblastoma (Rb) protein, we examined the effects of salicylate on Rb hyperphosphorylation. Treatment with salicylate, but not indomethacin, inhibited nuclear factor-kappaB activation and completely abolished Rb hyperphosphorylation in PDGF-treated SMCs. This effect was associated with a decrease in cyclin-dependent kinase (Cdk)-2 and, to a lesser extent, Cdk-6, but not Cdk-4 activity, without changes in Cdk-2, -4, and -6 and cyclin D and E protein levels. Because Cdk-2 activity is regulated by the Cdk inhibitors p21(Waf1) and p27(Kip1), we studied the effects of salicylate on p21(Waf1) and p27(Kip1) expression. Treatment with salicylate prevented PDGF-induced downregulation of p21(Waf1) and p27(Kip1) but not of the Cdk-4/-6 inhibitor p16(Ink4). CONCLUSIONS: These findings indicate that high doses of salicylates inhibit SMC proliferation by cell cycle arrest at the G(1)-S phase and suggest a beneficial role for high-dose salicylates in the treatment of vascular proliferative disorder