Investigation of the Suitability of Decellularised Porcine Pericardium for Mitral Valve Reconstruction

Abstract

The aim of this study was to investigate the suitability of decellularised porcine pericardium for heterotopic repair of the mitral valve (MV) leaflets, and its potential to regenerate through endogenous cell repopulation in vivo, or in vitro seeding and bioreactor conditioning. Anterior and posterior MV leaflets and pericardia were excised from porcine hearts within. The pericardia were decellularised according to the in-house protocol. Anterior and posterior leaflet, and decellularised and fresh pericardial samples were subjected to histology (H and E, Masson trichrome, Sirius Red, Miller’s elastin, Alcian blue-PAS), immunohistochemistry (collagen type I, III, IV, fibronectin, laminin, and chondroitin sulfate labelling), SEM, and uniaxial tensile testing. Samples were isolated along the radial and circumferential direction (leaflets), and perpendicular and parallel to the collagen fibres (pericardium). Biochemical assays for quantification of the sulphated GAG and collagen content of the tissues were also performed. Contact and extract cytotoxicity testing, and DNA quantification was performed to assess the decellularised pericardia. Histology revealed the trilaminar structure of the pericardium and quadrilaminar structure of the leaflets. Collagen type I and III was found in the fibrosa layers of both pericardium and leaflets, whereas fibronectin and laminin were found throughout the tissues. Decellularisation produced a completely acellular pericardial scaffold, which retained the histoarchitecture of the natural tissue. The biomechanics showed the anterior leaflets being stiffer along the circumferential direction. No significant anisotropy was observed in the biomechanics of the posterior leaflets, or fresh and decellularised pericardium. The anisotropy of the anterior leaflet was attributed to the orientation of the collagen (aligned along the circumferential direction). Biochemistry showed a significant increase in sulphated GAGs between the fresh leaflets and pericardium. No difference was found between the collagen content of the fresh leaflets and the fresh or decellularised pericardium. The decellularised pericardium showed a 99% reduction in DNA and a high loss in the GAG content compared to the fresh pericardium. The study showed that the MV leaflets and pericardium share similar histoarchitectures and comparable biomechanics. The similarity was more pronounced in the case of the posterior leaflet which was more isotropic both in terms of histoarchitecture and biomechanics. Apart from a decreased GAG content, the similarity was also apparent between the leaflets and the pericardial scaffolds. The decellularised pericardium has the potential to deliver the necessary biological and biomechanical cues to seeded or migrating cells, representing a plausible scaffold option for the regeneration of the MV leaflets in vitro or in vivo.

Biomaterial characterization of off-the-shelf decellularized porcine pericardial tissue for use in prosthetic valvular applications

“…These findings match similar testing done in bovine pericardium that showed that the UTS of native and fixed pericardium was over 50% stronger than native cusps (Arbeiter, Grabow, Wessarges, Sternberg, & Schmitz, 2012). Additionally, our decellularized tissues showed similar UTS to those reported (Morticelli, Korossis, Thomas, Roberts, & Ingham, 2013); yet we observed anisotropy in our material as is seen in other published literature (Gauvin et al, 2012;Labrosse et al, 2016). Our results show that the decellularization process lowers the GAGs compared with the native scaffolds (Figure 3), likely due to the caustic nature of SDS.…”

Feasibility of pig and human‐derived aortic valve interstitial cells seeding on fixative‐free decellularized animal pericardium

“…Before testing, all the samples were maintained in PBS at 4 C. Dog-bone shaped (4-mm width and 20-mm height) specimens were dissected from pericardial tissue sheets and no preferential orientation was considered when cutting the specimens, assuming porcine pericardium isotropy. 27 The ends of the specimens were glued with cyanoacrylate adhesive in between of frames made of finegrained emery article to facilitate uniform gripping and to avoid sample slippage. Then the specimens were mounted onto the clamps of the testing machine (MTS hydraulic testing machine equipped with 100-N loading cell, MTS System Corporation) and the sides of the sandpaper frame were cut.…”

In Vivo Assessment of Novel Stentless Valve in the Mitral Position

“…13- 15 Glutaraldehyde fixation has been used to treat both homogeneic and xenogeneic pericardium to reduce immunogenicity, as well as to treat autologous pericardium in order to increases stiffness, making it easier to handle. 16 The disadvantage of chemically treated pericardium is a lack of in vivo regeneration related to the presence of tissue cross-links. 15 In addition, the toxicity of residual glutaraldehyde prevents cellular infiltration, repair, and growth.…”

“…Chemically treated autologous and xenogeneic (bovine and porcine) pericardial tissue has been extensively used for aortic and MV reconstructions. 11- 16 Autologous pericardium is preferred because it has several advantages including low cost, lack of donor-derived pathogens, and nonimmunogenic characteristics, although it has disadvantages such as limited availability and inflammatory changes that occur after pericardiotomy. 12 A current biological substitute for autologous human pericardium is glutaraldehyde-fixed…”