Dual effects of estrogen on vascular smooth muscle cells: Receptor-mediated proliferative vs. metabolite-induced pro-senescent actions

Abstract Objective To investigate the mechanism for the dual effects of estrogen on vascular smooth muscle cells (VSMCs). Methods Cultured rat VSMCs were exposed to gradient concentrations (10 −9 –10 −5  M) of 17β-estradiol (E 2 ) with or without pre-administration of a broad-spectrum CYP450 inhibitor 1-aminobenzotriazole (ABT) (10 × 10 −6  M) and an estrogen receptor (ER) antagonist ICI 182,780 (10 −6  M), respectively. The growth, cell cycle progression, premature senescence, estrogen metabolites, reactive oxygen species (ROS) and DNA damage of the cells were analyzed with cell counting assay, flow cytometry, Western blot, liquid chromatography–mass spectrometry and comet assay, respectively. Results E 2 in its physiological levels from 10 −9  M to 10 −8  M had a concentration-dependent promoting effect on growth of VSMCs. However, when the concentration increased over 10 −8  M, the growth-promoting effect gradually reversed to a growth-inhibiting action. When the activity of CYP450s was blocked by ABT, the growth-promoting effect of E 2 increased and did not reverse at high concentrations. Whereas when the ERs were blocked by ICI 182,780, E 2 showed a pure growth-inhibiting effect. The E 2 metabolites 2- and 4-hydroxyestradiols accumulated with the increase of E 2 over 10 −8  M, which accompanied by increased ROS, DNA damage and cellular senescence. All of these changes were eliminated by block of CYP450s, indicating that the VSMC growth inhibition by E 2 is due to an increased production of ROS from accumulated E 2 metabolites which induces DNA damage, leading to VSMC premature senescence. Conclusion The complex effect of E 2 is due to two opposite actions: one ER-mediated and proliferative, and the other estrogen metabolite-induced and pro-senescent.