Effect of Rosuvastatin on MicroRNAs expression in human atherosclerotic plaques: results from Quasar Study

Background: Statins are competitive inhibitors of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase and are a leading therapy for the prevention of ischemic events. Although lowering of plasma low-density lipoproteins (LDL) is the most relevant effect, numerous studies suggest many other pleiotropic effects of statins, which involve improving of endothelial function, enhancing atherosclerotic plaque stability and decreasing oxidative stress and inflammation. MicroRNAs are small non-coding RNAs which act as post-transcriptional gene regulators. Growing evidences support the microRNAs involvement in cardiovascular disease and there are tantalizing hints of the effect of statin on microRNAs. The aim of this study is to investigate whether a short-time treatment with low or high dose of rosuvastatin may affect microRNAs expression in human atherosclerotic plaques. Material and Methods: In the “QUalitative Analysis of plaque Stability After Rosuvastatin therapy in asymptomatic patients enlisted to undergo carotid endoarterectomy” (QUASAR) 70 patients with severe stenosis of the internal carotid artery were randomized to receive a 12 week low (10 mg/day) or high (40 mg/day) doses of rosuvastatin before the elective endoarterectomy. Total RNA was extracted from plaques using Trizol reagent. Pools creation for MiRNome qPCR analysis was carried out using total RNA extracted from respectively 11 selected plaques of rosuvastatin 10 mg and 40 mg groups and from 11 plaques of naive hypercholesterolemic patients (control group). MiRNome qPCR analysis was performed by using miRCURY LNA Universal RT microRNA PCR system. MicroRNAs validation study was performed on all plaques from the pooled samples by qPCR. Results: MiRNome qPCR analysis of 742 microRNAs on pooled hypercholesterolemic samples versus respectively pooled rosuvastatin 10mg and rosuvastatin 40mg samples showed several microRNAs dysregulated in rosuvastatin groups versus hypercholesterolemic one. We have paid attention on nine microRNAs on the strength of their predicted target genes involved in atherosclerosis. Real-time PCR validation studies on all plaques from the pooled samples showed that both rosuvastatin doses significantly up-regulated mir-9 (p<0.004), mir-20b (p<0.001), mir-133a/b (p=0.001), mir-144 (p<0.001), mir-301a (p=0.01), and mir-377 (p=0.002), respect to hypercholesterolemic patients. Mir-150 and mir-155 were not found significantly down-regulated in the qPCR validation analysis. Conclusions: These data showed that short-term rosuvastatin treatment may affect microRNAs expression profile in human atherosclerotic plaques. Further studies on their predicted target gene are required to better understand microRNAs involvement in the beneficial effects observed with statin-based lipid lowering therapies.