[Cilostazol inhibits proliferation and induces apoptosis in rat vascular smooth muscle cells through Rb-p53-p21 pathways].

OBJECTIVE: To explore the effects and related mechanisms of cilostazol on rat vascular smooth muscle cells (VSMCs)proliferation. METHODS: VSMCs were treated with DMEM (control) and various doses of cilostazol (1.0×10(-7), 2.5×10(-7), 5.0×10(-7), 7.5×10(-7) and 1.0×10(-6) mol/L) for 13 d (cell counting) or 72 h. Proliferation of VSMCs was investigated by cell-counting, MTT and flow cytometry analysis. Cell apoptosis was determined by TUNEL staining. mRNA and protein expressions of cell cycle regulatory proteins, such as Rb, p53 and p21 were detected by RT-PCR and Western blot, respectively. RESULTS: Cilostazol inhibited VSMCs proliferation and induced VSMCs arrest at G1 phase in a dose-dependent manner. High dose of cilostazol (7.5×10(-7) and 1.0×10(-6) mol/L) induced VSMCs apoptosis. p53 mRNA expression in 2.5×10(-7) mol/L to 7.5×10(-7) mol/L groups as well as 1.0×10(-6) mol/L group (3.22 ± 0.45 vs. 1.75 ± 0.32) and p53 protein expression in 7.5×10(-7) mol/L group and 1.0×10(-6) mol/L group (0.53 ± 0.11 vs. 0.18 ± 0.06) were significantly upregulated after 72 h culture (all P < 0.05 vs. control). Low dose of cilostazol (1.0×10(-7), 2.5×10(-7) and 5.0×10(-7) mol/L) significantly upregulated p21 mRNA expression compared to control group (1.86 ± 0.19, 2.20 ± 0.24 and 2.10 ± 0.18 vs. 1.210 ± 0.18, all P < 0.05). Similarly, Rb mRNA expression was significantly upregulated in 1.0×10(-7), 2.5×10(-7) and 5.0×10(-7) mol/L groups (0.89 ± 0.07 vs. 0.38 ± 0.04)compared with control group (all P < 0.05). However, high dose cilostazol (7.5×10(-7) and 1.0×10(-6) mol/L) significantly downregulated p21 mRNA expression (0.81 ± 0.09 vs. 1.21 ± 0.18, 0.36 ± 0.10 vs. 1.21 ± 0.18, all P < 0.05 vs. control) and Rb mRNA expression (0.12 ± 0.02 and 0.11 ± 0.02 vs. 0.38 ± 0.04, all P < 0.05 vs. control). p21 and Rb protein expressions also upregulated at low concentrations of cilostazol and downregulated at high concentrations of cilostazol. CONCLUSION: Cilostazol could inhibit the proliferation of rat VSMCs through modulating Rb-p53-p21 pathway and induce VSMCs apoptosis through upregulating p53.