Near-infrared II fluorescence for imaging hindlimb vessel regeneration with dynamic tissue perfusion measurement

Background-Real-time vascular imaging that provides both anatomic and hemodynamic information could greatly facilitate the diagnosis of vascular diseases and provide accurate assessment of therapeutic effects. Here, we have developed a novel fuorescence-based all-optical method, named near-infrared II (NIR-II) fuorescence imaging, to image murine hindlimb vasculature and blood fow in an experimental model of peripheral arterial disease, by exploiting fuorescence in the NIR-II region (1000-1400 nm) of photon wavelengths. Methods and Results-Because of the reduced photon scattering of NIR-II fuorescence compared with traditional NIR fuorescence imaging and thus much deeper penetration depth into the body, we demonstrated that the mouse hindlimb vasculature could be imaged with higher spatial resolution than in vivo microscopic computed tomography. Furthermore, imaging during 26 days revealed a signifcant increase in hindlimb microvascular density in response to experimentally induced ischemia within the frst 8 days of the surgery (P<0.005), which was confrmed by histological analysis of microvascular density. Moreover, the tissue perfusion in the ischemic hindlimb could be quantitatively measured by the dynamic NIR-II method, revealing the temporal kinetics of blood fow recovery that resembled microbead-based blood fowmetry and laser Doppler blood spectroscopy. Conclusions-The penetration depth of millimeters, high spatial resolution, and fast acquisition rate of NIR-II imaging make it a useful imaging tool for murine models of vascular disease.