Decellularized cryopreserved human pericardium: a validation study towards tissue bank practice

Abstract

Pericardial patches are currently used as reconstructive material in cardiac surgery for surgical treatment of cardiac septal defects. Autologous pericardial patches, either treated with glutaraldehyde or not, can be used as an alternative to synthetic materials or xenograft in congenital septal defects repair. The availability of an allogenic decellularized pericardium could reduce complication during and after surgery and could be a valid alternative. Decellularization of allogenic tissues aims at reducing the immunogenic reaction that might trigger inflammation and tissue calcification over time. The ideal graft for congenital heart disease repair should be biocompatible, mechanically resistant, non-immunogenic, and should have the ability to growth with the patients. The aim of the present study is the evaluation of the efficacy of a new decellularization protocol of homologous pericardium, even after cryopreservation. The technique has proven to be suitable as a tissue bank procedure and highly successful in the removal of cells and nucleic acids content, but also in the preservation of collagen and biomechanical properties of the human pericardium.

Keywords: Allograft; Cardiac septal defects; Decellularization; Pericardium; Tissue bank.

Fig. 1

Cruciform specimen representation (a) and gripping (b)

Fig. 2

Haematoxylin and eosin staining of representative histological sections of non-decellularized pericardium (a) and decellularized pericardium (b), 40 × magnification Absence of nuclei is demonstrated in decellularized samples. Extracellular matrix is similar between non-decellularized pericardium (a) and decellularized pericardium (b)

Fig. 3

Nuclear fluorescence staining with Hoechst in representative samples of non-decellularized pericardium (a) and decellularized pericardium (b). Nuclei are absent in decellularized sample. 20 × magnification

Fig. 4

DNA quantity in non-decellularized samples (NDPCs) and decellularized ones (DPCs), obtained from three batches of pericardium (3 different donors). The reduction was statistically significant (p < 0.001) and the residual DNA in DPCs was below the limit of 50 ng/mg of dry tissue

Fig. 5

Electrophoresis analysis of nuclei acids compared to a marker (1). The image demonstrates the presence of DNA in non-decellularized samples (2, 4, 6) and the absence of nucleic acids fragments in decellularized pericardia samples (3, 5, 7). A total of three different batches (three different donors) of pericardia were included in the analysis

Fig. 6

Representative experimental stress–strain curve and identification of linear region for elastic modulus estimation

Fig. 7

Treatment comparison along Direction 1 (a) and Direction 2 (b). The elastic modulus is not statistically different between decellularized samples (DPCs) and non-decellularized ones (NDPCs)

Fig. 8

Donor comparison along Direction 1 (a) and Direction 2 (b). Non statistical differences in the elastic modulus were observed between the three different batches