Silencing of int6 gene restores function of the ischaemic hindlimb in a rat model of peripheral arterial disease

Aims Intermittent claudication (IC) is one of the serious symptoms of peripheral arterial disease (PAD) and is characterized by pain in the legs or buttocks that worsens with exercise and subsides with rest. The concept of 'therapeutic angiogenesis' for PAD has been widely proposed; however, the methodology, including cell transplantation, is still unclear. In this study, we examined the clinical efficacy of silencing the int6 gene, which encodes a protein that stabilizes hypoxia-inducible factor (HIF)-2 alpha, on angiogenesis in PAD. Methods and results An animal model for IC was established in Sprague-Dawley rats by external iliac artery ligation and evaluated by quantitative analysis of gait disturbance. Next, we explored the therapeutic effects of int6 siRNA injected into the adductor magnus muscle on IC. Recovery of hindlimb function occurred in the early stages after int6 siRNA injection. The number of blood vessels showed an obvious increase in the int6 siRNA-treated muscles. Angiography revealed the recovery of peripheral circulation at the affected sites. Early up-regulation of HIF-2 alpha and other angiogenic factors, including basic fibroblast growth factor and hepatocyte growth factor, was also apparent in the int6 siRNA-treated sites. We also confirmed the up-regulation of HIF-2 alpha and its translocation to the nucleus in the int6 siRNA-injected muscle. Conclusion A single injection of int6 siRNA promoted angiogenesis via up-regulation of HIF-2 alpha-related angiogenic factors in the muscles of the affected hindlimb and reduced gait disturbance. The int6 gene may be a novel therapeutic target for the treatment of IC in patients with PAD.