Optimization of hypothermosol for the hypothermic storage of cardiac cells - Addition of EDTA

Hypothermic solutions have been developed for shipping and storing organs, tissues, and cells. Cells are immersed in these solutions and kept at 4°C where they remain in a state of near suspended animation for long periods of time. A number of modifications to current cold-storage solutions have been suggested that improve the ability of these media to protect cells during hypothermic storage. We show that the Ca²⁺/Mg²⁺ chelator, ethylenediaminetetraacetic acid (EDTA), can confer added protection to a mixed chick cardiac cell population stored in the cold-storage solution, HypoThermosol. Both the base HypoThermosol and the EDTA-supplemented HypoThermosol are better at protecting the cardiac cells from cold-induced damage than is ViaSpan [University of Wisconsin (UW) solution]. To further understand why EDTA confers a protective effect, cardiac cells were cold-stored in HypoThermosol containing 5 mM Mg²⁺ and 0 mM Ca²⁺ in the presence or absence of EDTA. HypoThermosol containing 0 mM Ca²⁺, 5 mM Mg²⁺, and no EDTA was less effective as a cold-storage solution than the base HypoThermosol solution that contained 0.05 mM Ca²⁺ and 5 mM Mg²⁺; whereas the HypoThermosol containing 0 mM Ca²⁺, 5 mM Mg²⁺, and EDTA was the best at preserving the cardiac cells. When cardiac cells were stored in the base HypoThermosol solution containing 0.05 mM Ca²⁺ and 5 mM Mg²⁺, there was a time dependent leakage of Ca²⁺ from the cells that peaked at 7 days of cold storage. Therefore, we conclude that the positive effect conferred by EDTA on the ability of HypoThermosol to act as a cold-storage solution might be due to its ability to sequester intracellular calcium (Ca²⁺_i) that leaks from the cells and accumulates as Ca²⁺ during cold storage. The fact that a similar EDTA enhancing effect is seen in the cold storage of other cells suggests that the leakage of Ca²⁺_i should be seriously considered in the design of future cold-storage solutions.