Noninvasive Evaluation of Angiogenesis and Therapeutic Response after Hindlimb Ischemia with an Integrin-Targeted Tracer by PET

Background: Peripheral arterial disease (PAD) can severely compromise limb vitality and function. Angiogenesis plays an important role in healing of ischemic lesions. Radiolabeled RGD (Arg-Gly-Asp) peptides specifically targeting a(v)beta(3) integrin are promising tracers for imaging angiogenesis. In this study, we investigated the application of a one-step labeled RGD in evaluation of angiogenesis and therapy response in a mouse model of hindlimb ischemia (HI) by positron emission tomography (PET). Methods: HI was induced by ablation of the femoral artery in mice. PET imaging using F-18-AlF-NOTA-PRGD2 (F-18-PRGD2) tracer was performed at day 0 (presurgery) and days 3, 7, 14, and 21 after surgery to evaluate hindlimb angiogenesis longitudinally and noninvasively. The control peptide RAD (Arg-Ala-Asp) labeled with a similar procedure and a block agent were used to confirm the specific binding of F-18-PRGD2 to a(v)beta(3) integrin. Ex vivo CD31 staining was performed to detect angiogenesis. In addition, the angiogenic therapy response was monitored with F-18-PRGD2 tracer and immunofluorescence staining to confirm the imaging data. Results: The successful establishment of HI model was confirmed by ultrasound imaging and laser doppler perfusion imaging (LDPI). Specific binding of F-18-PRGD2 to a(v)beta(3) integrin was validated by minimal tracer uptake of the control peptide RAD and significant decrease of tracer accumulation when a block agent was added. Local accumulation of F-18-RRGD2 in ischemic hindlimb was detected as early as 3 days and reached a peak at 7 days after surgery. The temporal change of focal tracer uptake was positively correlated with the pattern of vascular density. Moreover, vascular endothelial growth factor (VEGF) treatment increased the tracer uptake and enhanced angiogenesis, which is consistent with integrin beta 3 expression. Conclusions: PET imaging of a one-step labeled tracer F-18-PRGD2 targeted to a(v)beta(3)integrin allows longitudinal monitoring of ischemia-induced angiogenesis and noninvasive assessment of VEGF treatment response in a mouse model of hindlimb ischemia. The simple synthesis procedure and in vivo performance of this PET tracer enables the feasibility of future clinical translation in ischemic cardiovascular diseases.