Evaluation of a Room-Temperature Preservation Method Maintaining Viability and Function in Human Cardiac Organoids

Three-dimensional (3D) cardiac models, including spheroids, organoids, and organ-on-chips, are advanced systems for studying human physiology, disease, and drug responses with greater biological relevance than 2D models. As their use expands in biomedical research, tissue engineering, and regenerative medicine, reliable preservation methods are needed. However, cryopreservation often fails to protect 3D systems due to limited cryoprotectant penetration, ice formation, and mechanical stress during freezing and thawing. Room-temperature (RT) preservation has emerged as a promising alternative for short-term transport. This study evaluated a RT-based transport medium (CellShip^®) for preserving cardiac organoids for up to seven days, compared with conventional cryopreservation using slow-freezing in Cryostor^®CS10. Viability and functionality were assessed using apoptosis, ATP levels, beating activity, proliferation, and size. During maturation, organoids showed increased size, ATP levels, and beating capacity. Cryopreservation reduced size, proliferation, ATP levels, and altered beating, while increasing apoptosis. In contrast, RT preservation maintained stable viability and functionality after recovery. These findings demonstrate that RT preservation effectively maintains cardiac organoid integrity and function, offering a promising alternative for short-term storage and transport, with potential terrestrial and nonterrestrial applications.