Mir-16 Modulates Endothelial Dysfunction After Balloon Injury in a Rat Model of Chronic Peripheral Ischemia

Background: Peripheral ischemia is associated with endothelial dysfunction in the coronary district and represents an important predictor of future cardiovascular events such as myocardial infarction and stroke. MicroRNAs (miRs) play an important regulatory role in cells of the vascular wall. Thus, the aim of the present study was to evaluate the role of miRs in mediating remote endothelial dysfunction in a model of peripheral ischemia.Methods and Results: Wistar male rats, were divided into the following groups: no treatment (sham); vascular injury on the carotid artery by balloon catheter (BI); femoral artery ligation (AL); ligation of the femoral artery followed, after 21 days, by balloon injury (AL+BI). At 21-days-evaluation, real time RT-PCR showed a profound up-regulation of miR-16 levels within ECs extracted by injured carotid artery of AL+BI group compared to the BI group.A bioinformatics approach revealed highly conserved binding sites for miR-16 in the 3’UTR of ARHGDIA (Rho-GDP dissociation inhibitor 1), an inhibitor of Rho signaling. Interestingly up-regulation of miR-16 was associated with a reduction of the ARHGDIA-transcript levels and a higher activation of the RhoA pathway, with consequent reduction of nitric oxide bioavailability. Accordingly, inhibition of miR-16 in ECs resulted in increased mRNA and protein levels of eNOS, with increased NO production, while up-regulation of miR-16 resulted in increased NF-kB activation in ECs.Conclusions: The expression levels of miR-16 are increased in the vascular endothelium of injured arteries in rats with a model of chronic peripheral ischemia, over rats without peripheral ischemia. Such modulation of miR-16 levels may play a key role in modulating endothelial dysfunction through modulation of the RhoA signaling pathway and the transcription factor NF-kB.