Effects of different freezing methods on morphology and viability of rabbit carotids

Objective: To assess the effects of vitrification and slow freezing method on the morphology and viability of rabbit carotids. Methods: Rabbit carotids were harvested and cryopreserved by vitrification and slow freezing method for 2 weeks, and then thawed slowly in water bath at 37°C. The fresh carotids were taken as controls. The morphological changes of all carotids were observed with the naked eye, optical microscope and electron microscope. The fresh and frozen-thawed arteries were cultured in vitro. Viability, regeneration and proliferation of the cells were observed and described by cellular growth curve. Results: The morphological studies demonstrated that some of endothelium cells (ECs) fell off, some of the smooth muscle cells (SMCs) and fibroblast cells swelled or denaturalized after freeze-thawing procedure. Vitrification, however, was less detrimental to vessel structure than slow freezing method. The cells of vitrified carotids and frozen carotids grew out from tissues on the 6th and 7th day. The biological characteristics were similar to those of the fresh carotids. The growth curve showed that cell viability and proliferation were preserved perfectly in vitrification group compared to slow freezing group, but inferior to the fresh carotids. The cells cultured from all carotids were identified through immunohistochemical staining and proved to be SMCs. Conclusion: Vitrification does less damage to the morphology and viability of vessels, so rabbit carotids preserved by vitrification are better than those preserved by slow freezing method.