The aim of this study was to evaluate the effects of the HMG-CoA reductase inhibitor fluvastatin on expression of the extracellular matrix protein thrombospondin-1 (TSP-1) in human aortic (HASMC) and human coronary artery smooth muscle cells (HCASMC). TSP-1 stimulates proliferation and migration of vascular smooth muscle cells in culture, is up-regulated following vascular injury induced by angioplasty and stenting and is thought to play a major role in the pathogenesis of in-stent restenosis (Roth et al., 1998).

Cultured HASMC and HCASMC were incubated with increasing concentrations of fluvastatin (0.1-10 µM) for up to 72h. TSP-1 protein expression was assessed by Western blotting analysis. Gels (7%) were run under non-reduced conditions where the anti-TSP-1 antibody recognises the trimeric (450kDa) version of the protein. TSP-1 mRNA levels were also assessed by RT-PCR analysis. HASMC proliferation was assessed using a BrDU incorporation ELISA while HCASMC proliferation was assessed using an Alamar Blue assay. Cell viability was also assessed using the methylthiazol tetrazolium (MTT) assay. Cell morphology was examined by phase-contrast microscopy. Data were analysed by ANOVA using the statistical package PRISM: a p-value <0.05 was taken to indicate significance.

Treatment of HASMC and HCASMC with 0.5 µM fluvastatin reduced TSP-1 mRNA levels to 60.1 ± 4.5 and 35.2 ± 4.1 % of control levels respectively (p<0.01, n=4). Likewise, treatment of HASMC and HCASMC with 1 µM fluvastatin significantly reduced TSP-1 protein expression to 69.7 ± 3.4 and 36.4 ± 1.3 % of control levels respectively (p<0.01, n=5). A significant reduction in both HASMC and HCASMC proliferation to 11.8 ± 0.4 and 30.6 ± 1 % of control levels was observed after treatment with 0.5 µM fluvastatin (p<0.01, n=6). Fluvastatin did not reduce viability of HCASMC at concentrations below 10 µM .

In conclusion, this study has shown that fluvastatin decreases both mRNA and protein expression of the matricellular glycoprotein TSP-1 in human vascular smooth muscle cells. Fluvastatin is well known to inhibit vascular smooth muscle cell proliferation and one mechanism through which it may achieve this is by suppressing TSP-1 expression within this cell type. Targeted delivery of fluvastatin may therefore be a useful therapeutic objective for prevention of the intimal hyperplasia associated with in-stent restenosis.

Liao J.K. (2002) Int J Cardiol., 86, 5-18
Roth, J.J et al., (1998) J Surg Res., 74, 11-16

Supported by the Irish Health Research Board