This study investigates the effects of space microgravity on human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs), revealing significant metabolic and physiological changes. Under microgravity, hPSC-CMs exhibited reduced beating rates and abnormal calcium cycling. Metabolomic and transcriptomic analyses showed impaired thiamine metabolism, with microgravity blocking thiamine intake and utilization. This disruption affected the tricarboxylic acid (TCA) cycle, reducing ATP production and leading to cytoskeletal remodeling and calcium homeostasis imbalance. Thiamine supplementation reversed these effects in vitro and in vivo, suggesting it could be a viable countermeasure against microgravity-induced cardiac changes. The study, conducted on the China Space Station (CSS), is the first astrobiological experiment in space, providing insights into the mechanisms of microgravity-induced cardiac adaptation. Thiamine, a critical coenzyme in TCA cycle metabolism, was found to be underutilized in microgravity, impairing energy production and cardiac function. Thiamine antagonists like amprolium exacerbated these effects, while thiamine supplementation restored normal metabolic function and cardiac performance. The study also demonstrated that thiamine treatment improved cardiac function in mice subjected to simulated microgravity, reducing cardiac atrophy and enhancing systolic and diastolic function. These findings suggest that thiamine supplementation could be a potential strategy to mitigate microgravity-induced cardiac adaptations during long-duration spaceflight. The research highlights the importance of understanding metabolic reprogramming in space environments and underscores the potential of thiamine as a therapeutic intervention for space-related cardiovascular issues. The study provides a foundation for further research into space medicine and the development of countermeasures for long-term space travel.This study investigates the effects of space microgravity on human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs), revealing significant metabolic and physiological changes. Under microgravity, hPSC-CMs exhibited reduced beating rates and abnormal calcium cycling. Metabolomic and transcriptomic analyses showed impaired thiamine metabolism, with microgravity blocking thiamine intake and utilization. This disruption affected the tricarboxylic acid (TCA) cycle, reducing ATP production and leading to cytoskeletal remodeling and calcium homeostasis imbalance. Thiamine supplementation reversed these effects in vitro and in vivo, suggesting it could be a viable countermeasure against microgravity-induced cardiac changes. The study, conducted on the China Space Station (CSS), is the first astrobiological experiment in space, providing insights into the mechanisms of microgravity-induced cardiac adaptation. Thiamine, a critical coenzyme in TCA cycle metabolism, was found to be underutilized in microgravity, impairing energy production and cardiac function. Thiamine antagonists like amprolium exacerbated these effects, while thiamine supplementation restored normal metabolic function and cardiac performance. The study also demonstrated that thiamine treatment improved cardiac function in mice subjected to simulated microgravity, reducing cardiac atrophy and enhancing systolic and diastolic function. These findings suggest that thiamine supplementation could be a potential strategy to mitigate microgravity-induced cardiac adaptations during long-duration spaceflight. The research highlights the importance of understanding metabolic reprogramming in space environments and underscores the potential of thiamine as a therapeutic intervention for space-related cardiovascular issues. The study provides a foundation for further research into space medicine and the development of countermeasures for long-term space travel.