Tormentic acid inhibits proliferation and induces apoptosis in vascular smooth muscle cells

Background

Disturbance in the balance between vascular smooth muscle cell (VSMC) proliferation and death is directly involved in diseases such as atherosclerosis and post-PTCA restenosis. Drugs that modulate such phenotypic alterations may be useful in the control of these and other clinical situations.

Aim

The present study analyzes the effect of the triterpenoid tormentic acid (TA, from the plant Rubus sieboldii), a DNA polymerase inhibitor with anti-inflammatory properties, on cultured VSMC.

Methods

Subconfluent cultures of embryonic rat aortic VSMC (A7r5) were exposed to increasing concentrations of TA or vehicle (DMSO) for 24 h, in the absence or presence of 10% fetal bovine serum (FBS), and apoptosis rates were evaluated through chromatin morphology analysis following DNA staining with the fluorescent dye HOE33342. In a separate series of experiment, A7r5 cells were cultured in 10% FBS continuously incubated with TA or vehicle. Cells were electronically counted (Coulter counter) at 2-day intervals.

Results

TA (30 μM) significantly increased apoptosis of A7r5 cells under serum deprivation conditions (DMSO: 9.6 ± 1.5% vs TA: 17.9 ± 1.5%, n = 9, P < 0.001), while cells cultured in FBS were not affected by the drug (DMSO: 2.0 ± 0.4% vs TA: 2.8 ± 0.4%, n = 9, P = 0.187). A7r5 cell proliferation was significantly inhibited by TA from the 6^th day of culture on (a 30% cell count reduction, n = 12, P < 0.001), with the relative difference in cell number remaining stable through the 8^th day (33% reduction, n = 6, P < 0.001).

Conclusions

Our data indicate that TA is a VSMC apoptosis inducer and proliferation inhibitor. The absence of a pro-apoptotic effect associated with an anti-mitotic action in the presence of serum suggests that TA may be useful in preventing proliferative vascular diseases without negatively affecting normal vasculature. In vivo models of vascular lesion are currently being employed in order to investigate this hypothesis.