Ultrasound molecular imaging of angiogenesis induced by mutant forms of hypoxia-inducible factor-1α

Aims Targeted point mutants of hypoxia-inducible factor-1 alpha (HIF-1 alpha) are potential optimal agents for angiogenesis therapy. Data are limited regarding the angiogenic response of HIF-1 alpha mutants. We aimed to compare the angiogenic effect of wild-type and mutant HIF-1 alpha by contrast ultrasound molecular imaging (UMI) of alpha(v)-integrin expression. Methods and results The wild-type gene of human HIF-1 alpha, a gene with double mutations (HIF-1 alpha(564/803)), a gene with triple mutations (HIF-1 alpha(564/803/402)), or the LacZ gene (control) was transfected into the ischaemic hind limbs of C57BL/6 mice using an adenovirus vector. The video intensity of microbubbles targeted to alpha(v)-integrins in the ischaemic limbs increased along with the number of point mutations of HIF-1 alpha. Immunohistochemical expression of endothelial alpha(v)-integrins was higher in the mutant HIF-1 alpha(564/803/402) group than the other groups as was the density of both capillaries and arterioles in ischaemic muscle. Expression of both the mRNA and protein for HIF-1 alpha and VEGF was significantly higher in the mutant HIF-1 alpha(564/803/402) group than in the other groups. The half-life of HIF-1 alpha and VEGF mRNA was longer in HIF-1 alpha mutant-transfected cells than in wild-type HIF-1 alpha or LacZ-transfected cells. Conclusion HIF-1 alpha mutants were more effective for enhancing angiogenesis in ischaemic muscle tissue than wild-type HIF-1 alpha, and the response could be qualitatively evaluated by UMI of alpha(v)-integrins expression.