Influence of Cryopreservation on Structural, Chemical, and Immunoenzymatic Properties of Aortic Valve Allografts

Abstract

Objectives: The problems in preparing (including cryopreservation) and implanting aortic valve allografts (AVAs) is widely elaborated, but some issues need explanation.

Material and methods: Twenty AVAs cryopreserved in dimethylsulphoxide/RPMI solution under -160°C for 1-15 years and 3 controls stored at +4°C up to 2 weeks, from 19 male and 4 female donors, aged 20-51, ±30.8 years, were examined using light (LM), digital (DM), and scanning electron microscopy (SEM), energy dispersion X-ray spectroscopy (EDS), and enzyme-linked immunosorbent assay immunoenzymatic tests (PECAM1, CD34).

Results: All AVAs were macroscopically correct. LM revealed normal structure of leaflets but massive endothelial decellularization (±59 cells remained on the surface of 5 mm scraps). DM and SEM demonstrated generally normal collagen structures, but local alterations, probably influenced by freezing-thawing (gaps, separated plates) or being initial phase of native degeneration (grains). EDS detected a little elevated calcium amount in 1 specimen only. The mean PECAM1 and CD34 concentrations were at similar low level in all probes.

Conclusions: Fresh and cryopreservation technologies did not significantly influence the basic properties of AVA leaflets; however, massive endothelial decellularization was present in both groups. Therefore, no endocardial cell activity nor signs of inflammation were observed. These results were independent of donors' age and sex, processing technology, and time of storage of cryopreserved AVAs.