New Reproducible Porcine Aortic Root Calcification Model: An Ex Vivo Study Under Dynamic Conditions

Abstract

Background: Calcific aortic valve disease results in severe aortic valve stenosis, the most frequent valvular disorder. Ex vivo animal models provide an essential resource to understand the mechanism of valvular calcification. Therefore, we aimed to develop a new ex vivo porcine aortic root calcification model.

Methods: Porcine aortic roots were subjected to a procalcific treatment with a buffer solution containing a defined calcium concentration (CaCl₂ = 2.2 mmol) in a durability tester (Hi-Cycle tester) with over 53.2 million cycles. The control group consisted of native porcine aortic roots that were not treated. Cusps of porcine aortic valves of both groups (total n = 10) were compared through macroscopic evaluation, analysis of tissue thickness, scanning and transmission electron microscopy, histological examination and calcium determination.

Results: After durability testing, macroscopic examination demonstrated pronounced calcification at regions with high mechanical stress—the commissures and the nadirs of the cusps. Calcific nodules cause tissue thickness after Hi-Cycle testing. Hydroxyapatite crystals were found by scanning electron microscopy, and calcium deposits were noticed by transmission electron microscopy within calcified cusps in the calcified group. The proof of cusp calcification was seen histologically in the calcified group. Calcium content of the aortic cusps differed significantly after treatment with calcification buffer vs control group (7.240 [6.383–9.494] vs. 3.178 [3.140–3.701] μg/cm² cusp area, p = 0.008).

Conclusion: We established a new reproducible and dynamic porcine aortic root calcification model. This ex vivo model may be a helpful alternative for investigating treatment modalities of calcification and functional analysis of heart valves instead of a complex animal model.