Development of an Ex Vivo Porcine Aortic Valve Calcification Model for Hydrodynamic Testing

Background: Aortic valve calcification at some stage results in aortic valve stenosis. The underlying mechanisms of calcification are not fully understood so far. Currently, no ex vivo disease model of aortic stenosis exists that allows for the testing of treatment modalities. Therefore, the aim of this study was to establish an ex vivo model of leaflet calcification for functional analyses.

Method: Porcine aortic roots (aortic valve with left ventricular outflow tract and aorta ascendens) were perfused with double-distilled water and physiological buffer (pH = 7.4; NaCl = 135 mmol, CaCl2 = 2.2 mmol, KH2PO4 = 1.2 mmol, MOPS(3-(N-morpholinopropan)sulfonsäure) = 0.5 mmol) at 37°C in a Hi-Cycler with over 53.2 million cycles which corresponds to ~20 months in a patient (treatment group). Native porcine aortic roots without treatment served as negative controls. Cusps of porcine aortic valves of both groups (total n = 10) were compared by histological examination (Hematoxylin eosin, elastica van Gieson, Kossa, Alizarin), electron microscopy (EM), transmission electron microscopy (TEM), transmission light absorption method (TLAM) and calcium titration.

Results: Calcium absorption in the aortic cusps differed regarding different swine aortic cusps with the highest calcium value 20 µg/cm² and the lowest calcium value 6 µg/cm² cusp-area after Hi-Cycler testing. Calcium titration showed significantly higher calcium value in the treatment group after Hi-Cycler testing versus the control group (10.8 ± 5.6 vs. 3.5 ± 0.5 µg/cm² cusp-area, p < 0.001). Histological examination demonstrated calcification of the cusps after Hi-Cycle testing while no calcification was found in the control group. TLAM showed pronounced calcification at the commissures and the nadirs of the cusps. Hydroxyapatite crystals were found in the calcified cusps in EM.

Conclusion: In this study, we established a reproducible dynamic ex vivo calcification model. This calcified porcine conduit model may be useful for the analysis of technical aspects of different biological prostheses and for the investigation of therapeutic options without the use of a complex animal model.

Publication History

Article published online:
03 February 2022