Detergent-based decellularization strategy preserves macro- and microstructure of heart valves

Abstract

Objectives: Biological tissue has great potential to function as bioprostheses in patients for heart valve replacement. As these matrices are mainly xenogenic, the immunogenicity needs to be reduced by decellularization steps. Reseeding of bioscaffolds has tremendous potential to prevent calcification upon implantation, so intact microstructure of the material is mandatory. An optimal decellularization protocol of heart valves resulting in adequate preservation of the extracellular architecture has still not been developed. Biological scaffolds must be decellularized to remove the antigenic potential while preserving the complex mixture of structural and functional proteins that constitute the extracellular matrix.

Methods: Here, we compared 3 different decellularization strategies for their efficiency to remove cells completely while preserving the porcine heart valve ultrastructure. Porcine pulmonary heart valves were treated either with trypsin-ethylenediaminetetraacetic acid (TRP), a protocol using detergents in combination with nucleases (DET + ENZ), or with Accutase® solution followed by nuclease treatment (ACC + ENZ). The treated heart valves then were subjected to histological, DNA and scanning electron microscopic analyses.

Results: All DNA fragments were removed after ACC + ENZ treatment, whereas cellular removal was incomplete in the TRP group. TRP and ACC + ENZ-treated valves were enlarged and showed a disrupted architecture and degraded ultrastructure. In contrast, fully acellular heart valves with intact architecture, layer composition and surface topography were achieved with DET + ENZ treatment. DET + ENZ treatment yielded excellent results in terms of preservation of material architecture and removal of DNA content.

Conclusions: Compared to TRP and ACC + ENZ procedures, DET + ENZ-treated porcine pulmonary heart valves demonstrated well-preserved macroscopic structures and microscopic matrix components and represent an excellent scaffold for further application in tissue engineering.

Keywords: Decellularization; Detergent; Enzyme; Pulmonary heart valve; Tissue engineering.