Decellularization of human donor aortic and pulmonary valved conduits using low concentration sodium dodecyl sulfate

Abstract

The clinical use of decellularized cardiac valve allografts is increasing. Long‐term data will be required to determine whether they outperform conventional cryopreserved allografts. Valves decellularized using different processes may show varied long‐term outcomes. It is therefore important to understand the effects of specific decellularization technologies on the characteristics of donor heart valves. Human cryopreserved aortic and pulmonary valved conduits were decellularized using hypotonic buffer, 0.1% (w/v) sodium dodecyl sulfate and nuclease digestion. The decellularized tissues were compared to cellular cryopreserved valve tissues using histology, immunohistochemistry, quantitation of total deoxyribose nucleic acid, collagen and glycosaminoglycan content, in vitro cytotoxicity assays, uniaxial tensile testing and subcutaneous implantation in mice. The decellularized tissues showed no histological evidence of cells or cell remnants and >97% deoxyribose nucleic acid removal in all regions (arterial wall, muscle, leaflet and junction). The decellularized tissues retained collagen IV and von Willebrand factor staining with some loss of fibronectin, laminin and chondroitin sulfate staining. There was an absence of major histocompatibility complex Class I staining in decellularized pulmonary valve tissues, with only residual staining in isolated areas of decellularized aortic valve tissues. The collagen content of the tissues was not decreased following decellularization however the glycosaminoglycan content was reduced. Only moderate changes in the maximum load to failure of the tissues were recorded postdecellularization. The decellularized tissues were noncytotoxic in vitro, and were biocompatible in vivo in a mouse subcutaneous implant model. The decellularization process will now be translated into a good manufacturing practices‐compatible process for donor cryopreserved valves with a view to future clinical use. Copyright © 2016 The Authors Tissue Engineering and Regenerative Medicine published by John Wiley & Sons, Ltd.

In vitro biomechanical and hydrodynamic characterisation of decellularised human pulmonary and aortic roots

“…During the decellularisation treatment, complete cell removal was demonstrated histologically and biochemically (Vafaee et al, 2016) however, it is crucial to retain the tissue function and biomechanical properties whilst completely removing all cellular components.…”

“…In Phase I, the effect of decellularisation on the biomechanical and hydrodynamic properties of human aortic and pulmonary roots was investigated by comparison to cellular valve allografts (unpaired controls). The decellularised tissue was prepared as reported by Vafaee et al which included the removal of excess fat and connective tissue, and scraping of the adventitial layer of the valve root using a scalpel blade (referred to throughout as 'scraping'), to allow diffusion of decellularisation solutions to achieve complete decellularisation (Vafaee et al, 2016). During this phase, there was a concern that the scraping process to disrupt the outer layer of the adventitia may have compromised the structure of the vessel.…”

“…Four aortic roots (mean diameter 21.3 ± 2.7 mm) and four pulmonary roots (mean diameter 24.3 ± 2.4 mm) were used as the cellular tissue controls. Four aortic (mean diameter 23.3 ± 2.7 mm) and four pulmonary roots (mean diameter 24.8 ± 3.0 mm) were decellularised as previously described by Vafaee et al Vafaee et al (2016). Each root underwent sequential in vitro hydrodynamic and biomechanical test protocols carried out over two consecutive days [ Fig.…”

“…The decellularised pulmonary leaflets were significantly stiffer in the collagen dominant phase and failed at significantly higher UTS than the cellular leaflets in the circumferential direction. Such changes may be due to the significant reduction of glycosaminoglycan (GAGs) and removal of cells reported by Vafaee et al (2016). The loss of GAGs in the decellularised leaflets, may have led to loosening of the fibrous structure, and uncrimping of the circumferentially aligned collagen fibers.…”

“…A fully comprehensive description of the process and biological characteristics of the decellularised roots has been reported previously (Vafaee et al, 2016). The aim of this part of the study was to investigate the influence of decellularisation on the hydrodynamic performance and biomechanical properties of the low concentration SDS decellularised human aortic and pulmonary roots.…”