Immune responses of implanted bovine pericardium and porcine heart valves in anα1,3-galactosyltransferase knockout and wild type mouse model

Abstract

Introduction: Degenerative calcification is an important mechanism of bioprosthetic failure. α-Gal epitope-induced immune responses have been reported as a potential cause of degenerative calcification in bioprostheses. The objective of this study was to evaluate the α-Gal immune reaction and anti-calcification effect of decellularization with or without α-galactosidase in glutaraldehyde-fixed bovine pericardium and porcine heart valves, using an α-Gal knockout (KO) & wild type mouse subcutaneous implantation model. Methods: To elucidate the anti-calcification effect of decellularization with or without α-galactosidase, bovine pericardium and porcine heart valve tissues were assigned to the following 3 groups according to the tissue preparation methods: glutaraldehyde fixation only (GA), decellularization + GA fixation (Decell), and decellularization + α-galactosidase + GA fixation (Decell+α-galactosidase). The mechanical properties of the bovine pericardial tissue were measured before implantation. Each prepared tissue was subcutaneously implanted into both α-Gal KO and WT mice. Macrophage and T cell infiltration were examined via immunohistochemistry staining in the explanted tissues. Anti-α-Gal immunoglobulin (Ig) G and IgM antibodies were measured prior to implantation and at 4 weeks, 8 weeks, and 12 weeks after implantation using an enzyme-linked immunosorbent assay. The calcium contents in the explanted tissues were measured at 12 weeks after implantation. Results: The Decell and Decell+α-galactosidase groups showed lower tensile strengths than did the GA group (P < 0.001, P = 0.002). Increased macrophage and T cell infiltration were observed in the α-Gal KO mice in all groups relative to WT mice. The bovine pericardium showed more macrophage infiltration than did the porcine heart valve. The α-Gal-KO mice had higher anti-α-Gal IgG antibody titers than did the WT mice over time in response to both bioprosthetic materials, regardless of the tissue preparation methods (all P < 0.001). There were no significant differences in the anti-α-Gal IgG antibody titers according to the type of bioprosthetic material or tissue preparation method in the α-Gal-KO mice (P > 0.05). The calcium content was significantly lower in the porcine heart valves than in the bovine pericardium when implanted in the α-Gal-KO mice (P < 0.001). The calcium contents in the bovine pericardium and porcine heart valves implanted in α-Gal-KO mice were significantly lower in the Decell and Decell+α-galactosidase groups than in the GA group (all P < 0.05). However, there was no significant difference in calcium content between the Decell and Decell+α-galactosidase groups (all P > 0.05). Conclusions: The porcine heart valve induced lower levels of macrophage/T cell infiltration and calcium deposition than did the bovine pericardium, although the anti-α-Gal IgG antibody titers did not significantly differ between the bioprosthetic tissues. Decellularization with or without α-galactosidase had significant anti-calcification effects in both the bovine pericardium and porcine heart valves implanted in the α-Gal KO mice, although there was no significant difference in the anti-α-Gal IgG antibody titers among tissue preparation methods. The discrepancies in the calcium deposition and anti-α-Gal IgG antibody titers observed in our study suggested that bioprosthetic calcium deposition might occur through a multifactorial mechanism.