Both estrogen and raloxifene cause G1 arrest of vascular smooth muscle cells

Abstract

The proliferation of vascular smooth muscle cells (VSMC) is a crucial pathophysiological process in the development of atherosclerosis. Although estrogen is known to inhibit the proliferation of VSMC, the mechanism responsible for this effect remains to be elucidated. In addition, the effect of raloxifene on VSMC remains unknown. We have shown here that 17 -estradiol (E 2 ) and raloxifene significantly inhibited the platelet-derived growth factor (PDGF)-stimulated proliferation of cultured human VSMC. Flow cytometry demonstrated that PDGFstimulated S-phase progression of the cell cycle in VSMC was also suppressed by E 2 or raloxifene. We found that PDGF-induced phosphorylation of retinoblastoma protein (pRb), whose hyperphosphorylation is a hallmark of the G1-S transition in the cell cycle, was significantly inhibited by E 2 and raloxifene. These effects were associated with a decrease in cyclin D1 expression, without a change in cyclin-dependent kinase 4 or cyclin-dependent kinase inhibitor, p27 kip1 expression. ICI 182,780 abolished the inhibitory effects of E 2 and raloxifene on PDGFinduced pRb phosphorylation. Next, we examined which estrogen receptor (ER) is necessary for these effects of E 2 and raloxifene. Since VSMC express both ER and ER , A10, a rat aortic smooth muscle cell line that expresses ER but not ER , was used. The dose-dependent stimulation of A10 cell proliferation by PDGF was not inhibited by E 2 or raloxifene in contrast to the results obtained in VSMC. Moreover, E 2 and raloxifene significantly inhibited the PDGF-induced cyclin D1 promoter activity in A10 cells transfected with cDNA for ER but not in the parental cells. These results suggested that E 2 and raloxifene exert an antiproliferative effect in VSMC treated with PDGF, at least in part through inhibition of pRb phosphorylation, and that the inhibitory effects of E 2 and raloxifene may be mainly mediated by ER .

Hormonal effects on blood vessels

“…[39][40][41] It has also been reported that stimulation of GPR30 blocks VSMC proliferation. 37,42 The vasodilator action of G-1 may be mediated by NO-independent 40 and NO-dependent 37,39,40 pathways; the latter involves GPR30-induced endothelial NO synthase (eNOS) phosphorylation. 43 Also, G-1 decreases nicotinamide adenine dinucleotide phosphate-stimulated superoxide production by the carotid and intracranial arteries, indicating the scavenging effects of GPR30 on superoxide anions.…”

Raloxifene Elicits Combined Rapid Vasorelaxation and Long-Term Anti-Inflammatory Actions in Rat Aorta

“…Once 5 ϫ 10 3 or 8 ϫ 10 3 cells/well were plated in a 96-well plate, they were first incubated for 24 h in M199 with 10% FCS and then switched to phenol red-free M199 with 10% FCS for 2 days and phenol red-free M199 with 0.4% FCS for 24 h. Cell viability was assessed after a 24-h incubation with the LPS/cytokine mixture in the presence or absence of 1 M raloxifene. The number of surviving cells was assessed as described elsewhere (Takahashi et al, 2003) by recording the absorbance at 490 nm generated by the bioreduction of the MTS tetrazolium compound into a colored soluble formazan product (Cell Titer 96 AQueous One Solution Cell Proliferation Assay; Promega, Madison, WI).…”

Molecular Mechanism of Action of Selective Estrogen Receptor Modulator in Target Tissues

“…2), in part through direct inhibition of gene expression of cyclin D1 followed by dephosphorylation of Rb and suppression of E2F activity mediated by ERa in a transcription-dependent manner, a so-called genomic mechanism [38] as reported the effect of estrogen on Rb phosphorylation [39,40], and in part due to induction of apoptosis through a p38 cascade whose activation is mediated by ERa in a transcriptionindependent manner, a so-called nongenomic mechanism [41] (Fig. 3).…”