Simvastatin inhibits thromboxane-induced contraction in porcine coronary artery: role of extracellular calcium

M. N. Saarti¹, M. D. Randall², R. E. Roberts². ¹school of life science, University of nottingham, Nottingham, United Kingdom, ²School of Life Sciences, University of Nottingham, Nottingham, United Kingdom.

Introduction: Calcium plays an important role in thromboxane-induced contraction in smooth muscle (1). Previous studies have indicated that simvastatin may regulate vascular tone through effects on calcium influx and/or inhibition of mitochondrial function (2). The aim of this study was to determine if inhibition of extracellular calcium-dependent contraction by simvastatin is associated with inhibition of mitochondrial complexes.

Method: Coronary arteries from pigs (large white hybrid pigs, 4-6 months old, male and female) were mounted in isolated tissue baths. Tissues were incubated with simvastatin (1, 3, or 10 μM) for 2 hrs in presence or absence of calcium, with or without the mitochondrial complex inhibitors rotenone and myxothiazol (10 μM). After the incubation period, thromboxane mimetic U46619 was added cumulatively (1nM-300nM). In other experiments, tissues were incubated with a single concentration simvastatin (10 μM) with and without mitochondrial complexes inhibitors rotenone and myxothiazol (10 μM) in calcium-free buffer. After the incubation period, tissues were exposed to a maximum concentration of U46619 (300nM). CaCl₂ was then added cumulatively (10μM-3mM) to induce a contraction.

Results: Simvastatin inhibited thromboxane-induced contraction in porcine coronary artery (PCA) in the presence of calcium (Rmax 88±5 % v control 111 ±7.1%, n=14, P<0.05), which was prevented by rotenone and myxothiazol (inhibitors of complex I and III respectively). The U46619-induced contraction was reduced in the absence of calcium and under these conditions simvastatin had no inhibitory effect. Simvastatin inhibited the CaCl₂-induced contraction (Rmax 44±5 % v control 62 ±6.2%, n=9, P<0.05). In this case rotenone and myxothiazol did not prevent the inhibitory effect of simvastatin.

Conclusion: This study showed that simvastatin inhibited thromboxane-induced contraction only in the presence of calcium and inhibited the contraction in response to re-addition of extracellular calcium, suggesting that simvastatin inhibits the contraction caused by influx of extracellular calcium. Inhibition of mitochondrial complexes I (Rotenone) and III (Myxothiazol) prevented the inhibitory effect of simvastatin in the presence of calcium, suggesting that the inhibitory effect of simvastatin on the U46619-induced contraction may be related to mitochondrial regulation of calcium influx. On the other hand, inhibition of complexes I and III had no effect on the inhibitory effect of simvastatin on calcium-induced contraction. This could be explained by differences in the calcium channels involved in initiation of the thromboxane contraction and those involved in maintenance of the contraction. Further studies are required to determine if this is the case.