PROJECT SUMMARY The clinical translation of stem cell-based therapeutics for regenerative medicine is currently hindered by the lack of suitable imaging approaches that allow long-term monitoring of transplanted cells. Nuclear imaging techniques together with reporter gene transgenic expression provide a highly sensitive, non-invasive tool to monitor the fate of viable transplanted cells in vivo. The objective of this application is to develop a clinical- applicable, non-invasive imaging technology that enables the longitudinal monitoring of patient-specific stem cells and their effects on host tissues for peripheral artery disease (PAD). To that end, patient-specific vascular endothelial cells (ECs) have been generated by us using induced pluripotent stem cell (iPSC) technology from multiple PAD patients. In addition, ectopic expression of human sodium iodide symporter (hNIS) been shown to enable live cell tracking via Single Photon Emission Computed Tomography /Computed Tomography (SPECT/CT) without affecting EC functions. The hypothesis of the project is that by engineering patient-specific iPSC-ECs with reporter gene hNIS, the long-term survival, engraftment and biodistribution of those cells in vivo can be tracked non-invasively using SPECT/CT, without negatively impacting cell or host tissue functions. This will be achieved by firstly optimizing hNIS transgenic expression in patient-specific iPSC-ECs in vitro, followed by evaluating cell tracking and vascular regeneration in a hindlimb ischemia model in vivo. Through this work, it is expected that a clinical applicable imaging method will be established that can track the survival, engraftment, and distribution of patient-specific ECs. This study will also provide critical information on the feasibility of using autologous iPSC-ECs from PAD patients for vascular regeneration, which may significantly expand the therapeutic options for various cardiovascular and ischemic diseases. This is a feasibility test to establish an optimized cell modification and imaging protocol both in vitro and in vivo, before future studies in larger animal models and patients.