Introduction: Clinical solutions for peripheral artery disease (PAD) remain elusive due to the many challenges of surgical intervention. Stem cell therapy offer a promising option, but barriers exist to their clinical translatability. Among these include lifestyle choices like tobacco use that may influence the efficacy of stem cell therapy. The objective of this study aims to examine the potential effects of nicotine on human induced pluripotent stem cell derived endothelial cells (iPSC-ECs) therapy in the context of PAD. Hypothesis: Chronic nicotine exposure negatively affects the therapeutic potential of implanted iPSC-ECs in a murine model of PAD. Methods: Mice were implanted with an osmotic pump for controlled exposed to nicotine or saline (control) for 28 days. Plasma cotinine was measured to confirm nicotine exposure. After 28 days of nicotine or saline exposure, the mice underwent induction of hindlimb ischemia and before receiving an injection of 10 6 iPSC-ECs. A second dose of cells were injected to the ischemic limb on day 7 post-surgery. All mice were monitored for blood perfusion recovery using laser Doppler spectroscopy imaging. Survival of the transplanted cells in vivo were tracked by bioluminescence imaging. The mice receiving iPSC-ECs were also imaged for cell survival using bioluminescence imaging. At day 14 post-surgery, mice were euthanized, and the gastrocnemius muscle and thigh muscle were harvested for histological analysis. Results: The average cotinine concentration in sera was ~200 ng/mL, which is within physiological range for a tobacco user. Blood perfusion recovery at day 14 showed trends of nicotine treated groups have hindered perfusion recovery, compared to those treated with saline. Importantly, nicotine exposure trended with poorer perfusion recovery after cell therapy. Bioluminescent imaging showed iPSC-ECs persisted in vivo for all groups without significant difference. Conclusions: Nicotine exposure was associated with poorer blood perfusion recovery after iPSC-EC therapy, compared to control saline exposure, suggesting that nicotine may have adverse effects on iPSC-EC therapy for treatment of PAD.