Aim of the study: Danxiongfang (DF) is a new Chinese medicine formula used to treat atherosclerosis and vascular restenosis. The active ingredients in DF are danshensu (DSS), tanshinones (cryptotanshinone, CT) and ferulic acid (FA). The aim of present study was to evaluate the inhibitory effects of DF and its active ingredients on cell proliferation and protection against hydrogen peroxide (H(2)O(2))-induced injury in rat vascular smooth muscle cells (VSMC) in vitro.
Methods: VSMC proliferation was assayed by cell counting and measurement of cell viability using the 3-(4, 5-dimethylthiazol -2yl)-2, 5-diphenyltetrazolium bromide (MTT) method and protein content was measured by the Bradford method. The nitric oxide (NO) level was detected by an assay kit. The endothelin-1 (ET-1) level was measured by ELISA. The protective effects of DF and its active ingredients on H(2)O(2)-induced cell injury was evaluated in terms of cell viability (MTT assay), superoxide dismutase (SOD) activity and malondialdehyde (MDA) levels. Hydroxyl free radicals generated by the Fenton reaction was detected with the spin-trapping technique on an electron spin resonance spectrometer.
Results: The results suggest that DSS, CT, FA and DF inhibited VSMC proliferation by increasing the NO level and decreasing the ET-1 content. In rat VSMCs exposed to H(2)O(2), FA, DSS, CT and the six formulations of DF increased cell viability and SOD activity, and reduced the levels of MDA and hydroxyl free radicals. These effects of FA, DSS and CT occurred in a dose-dependent manner. Of the six formulas, DF 4 and DF 5 had the more significant activities. The effects of DF were much greater than those of the individual ingredients, even though the concentrations of these ingredients in the DF formulas were much lower than the doses of the individual ingredients used in each study, indicating markedly synergistic effects of DSS, CT and FA in DF on rat VSMCs.
Conclusions: these findings provide a pharmacological foundation for the clinical use of DF in the prevention and treatment of hyperlipidemia and atherosclerosis relevant to endothelial cell proliferation and damage.