Development of Supercooling Preservation Method for Adherently Cultured Endothelial Cells and Its Application to...

Area of Science:

• Biotechnology and Bioengineering
• Cell Biology
• Drug Discovery Technologies

Background:

• Microphysiological systems (MPS) are valuable tools for drug discovery and toxicity testing, mimicking human organ functions.
• Preserving MPS with adherent cells presents a significant challenge, limiting their practical application and storage.
• Current preservation methods often lead to cell damage or death, necessitating improved techniques.

Purpose of the Study:

• To develop and optimize a supercooling preservation method for human-derived adherent cells in MPS.
• To evaluate the efficacy of supercooling for preserving cell viability, morphology, and function in both 2D and 3D MPS models.
• To establish practical storage conditions for MPS to facilitate their use in various research applications.

Main Methods:

• Human hepatic sinusoidal endothelial cells (TMNK-1) were used to optimize supercooling conditions, including temperature, cooling/rewarming rates, and preservation solutions.
• Preservation was performed at -4 °C with a cooling rate of -0.028 °C/min and a rewarming rate of +1.0 °C/min.
• The optimized method was applied to both 2D and 3D MPS models containing TMNK-1 or HepG2 cells.

Main Results:

• Optimized supercooling preservation at -4 °C maintained high cell viability and preserved morphology for up to 7 days.
• The method successfully preserved cell viability in both 2D and 3D MPS models containing different human cell types.
• No significant cell death or cytoskeletal disruption was observed post-preservation, indicating the method's gentleness.

Conclusions:

• Supercooling preservation offers a viable, non-freezing method for the short-term storage of MPS.
• This technique addresses a critical challenge in MPS research, enhancing their utility in drug discovery and toxicity assessments.
• The developed method provides a practical strategy for maintaining the integrity of MPS for extended periods.