Neomycin enhances extracellular matrix stability of glutaraldehyde crosslinked bioprosthetic heart valves

Glutaraldehyde (GLUT) crosslinked porcine aortic heart valves are continued to be extensively used in heart valve replacement surgeries. GLUT does not crosslink glycosaminoglycans in the tissue and we have demonstrated that GAG loss is associated with tissue degeneration. In this study, we examined the ability of neomycin to enhance GLUT crosslinking to stabilize GAGs, as well as provide evidence of improved functional integrity. Neomycin enhanced GLUT crosslinked (NG) leaflets exposed to collagenase and elastase enzymes exhibited an increased resistance to proteolytic degradation. Furthermore, NG leaflets exhibited small but significant increases in collagen denaturation temperatures when compared to that of standard GLUT crosslinked BHVs. NG leaflets subjected to storage, accelerated cyclic fatigue, and in vitro enzyme mediated GAG degradation revealed improved GAG stabilization versus standard GLUT crosslinked valves, which sustained substantial decreases in GAG content. Ultrastructural analysis using transmission electron microscopy qualitatively confirmed NG leaflets preserved GAGs after enzymatic degradation. Biomechanical analyses demonstrated that NG leaflets were functionally similar to GLUT tissues but were slightly stiffer under both planar biaxial tension and under flexure. Interestingly, after GAGase treatment, GLUT tissues showed increased areal compliance and reduced hysteresis, while NG leaflets were unchanged. Collectively, NG cross-linking functionally insulated the tissue from GAG digestion, and imparted modest additional matrix stiffness but maintained tissue hysteresis properties.