This study aimed to compare the effects of an acute Pilates program under hypoxic vs. normoxic conditions on the metabolic, cardiac, and vascular functions of the participants. Ten healthy female Pilates experts completed a 50-min tubing Pilates program under normoxic conditions (N trial) and under 3000 m (inspired oxygen fraction = 14.5%) hypobaric hypoxia conditions (H trial) after a 30-min exposure in the respective environments on different days. Blood pressure, branchial ankle pulse wave velocity, and flow-mediated dilation (FMD) in the branchial artery were measured before and after the exercise. Metabolic parameters and cardiac function were assessed every minute during the exercise. Both trials showed a significant increase in FMD; however, the increase in FMD was significantly higher after the H trial than that after the N trial. Furthermore, FMD before exercise was significantly higher in the H trial than in the N trial. In terms of metabolic parameters, minute ventilation, carbon dioxide excretion, respiratory exchange ratio, and carbohydrate oxidation were significantly higher but fat oxidation was lower during the H trial than during the N trial. In terms of cardiac function, heart rate was significantly increased during the H trial than during the N trial. Our results suggested that, compared to that under normoxic conditions, Pilates exercise under hypoxic conditions led to greater metabolic and cardiac responses and also elicited an additive effect on vascular endothelial function.