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We have used detergent and enzymatic extraction of natural arteries to produce an acellular matrix vascular prosthesis (AMVP). Implanted as an allograft in a canine model, this AMVP shows excellent handling characteristics, low thromboreactivity, no evidence of aneurysm, and exceptional graft patency in the peripheral vasculature. As a first step in the development of xenograft AMVPs, we processed caprine carotid arteries to AMVP and implanted them as femoral interposition grafts in dogs. Explanted xenografts at 4 weeks showed multifocal mixed inflammatory infiltrates and focal destruction of the medial elastin in the inflammatory foci. To further study the immune response to xenogenic AMVP, we implanted canine-derived AMVPs and fresh canine arteries for 4 weeks in a Lewis rat model. Extraction to AMVP markedly reduced the circulating antibody response to the xenogenic implants; however, histological analysis revealed that both xenograft arteries and AMVPs produced a marked immune response with penetration of mononuclear cells into the media and adventitia. To modify the immune response, we applied three crosslinking techniques to the canine AMVPs: glutaraldehyde, polyglycidyl ether, and carbodiimide. All crosslinkers significantly reduced degradation and cellular infiltration of the prostheses. However, crosslinking neither eliminated the chronic inflammatory response surrounding the implants nor reduced the humoral response to the xenogenic materials., (Copyright 2001 John Wiley & Sons, Inc. J Biomed Mater Res 55: 576-586, 2001)

Heart valve substitutes of biological origin often fail by degenerative mechanisms. Many authors have hypothesized that mechanical fatigue and structural degradation are instrumental to in vivo failure. Since the properties of the structural matrix at implantation may predetermine failure, we have examined the ultrastructure, fracture, mechanics, and uniaxial high-strain-rate viscoelastic properties of: (1) fresh, (2) cryopreserved, and (3) cellular extracted porcine aortic valve leaflets. The cellular extraction process is being developed in order to reduce immunological attack and calcification. Cryopreservation causes cellular disruption and necrotic changes throughout the tissue, whereas extraction removes all cells and lipid membranes. Both processes leave an intact collagen and elastin structural matrix and preserve the high-strain-rate viscoelastic characteristics of the fresh leaflets. Extraction does cause a 20% reduction in the fracture tension and increases tissue extensibility, with the percent strain at fracture rising to 45.3 +/- 4 (mean +/- SEM) from 31.5 +/- 3 for fresh leaflets. However, extraction does preserve matrix structure and mechanics over the physiological loading range. Glutaraldehyde fixation produces increased extensibility, increased elastic behavior, and, when applied to extracted leaflets, it causes a marked drop in fracture tension, to 50% of that for fresh leaflets. The combination of extraction and fixation may lead to early degenerative failure. The cellular extraction technique alone may be a useful alternative to glutaraldehyde fixation in preparing bioprosthetic heart valves.

There is evidence to suggest that the cellular components of homografts and bioprosthetic xenografts may contribute to calcification or immunogenic reactions. A four-step detergent and enzymatic extraction process has been developed to remove cellular components from bovine pericardial tissue. The process results in an acellular matrix material consisting primarily of elastin, insoluble collagen, and tightly bound glycosaminoglycans. Light and electron microscopy confirmed that nearly all cellular constituents are removed without ultrastructural evidence of damage to fibrous components. Collagen denaturation temperatures remained unaltered. Biochemical analysis confirmed the retention of collagen and elastin and some differential extraction of glycosaminoglycans. Low strain rate fracture testing and high strain rate viscoelastic characterization showed that, with the exception of slightly increased stress relaxation, the mechanical properties of the fresh tissue were preserved in the pericardial acellular matrix. Crosslinking of the material in glutaraldehyde or poly(glycidyl ether) produced mechanical changes consistent with the same treatments of fresh tissue. The pericardial acellular matrix is a promising approach to the production of biomaterials for heart valve or cardiovascular patching applications.

We have examined the tensile viscoelastic properties of circumferential and radial strips of porcine aortic valve leaflets following fixation in glutaraldehyde and formaldehyde, with or without pressure. After aldehyde treatment, the radial strips remained weaker and less stiff than circumferential strips and responded slightly differently to the treatment. After fixation, with or without pressure, the radial strips showed large changes in stress-strain and hysteresis responses due to initial loading, and there was a twofold reduction in tensile strength and final stiffness. For strips in both directions, fixation without pressure produced doubled extensibility and a ramping stress-strain curve. Permanent (plastic) deformation of 5-20% occurred as a result of cyclic loading, stress relaxation, and creep experiments. Pressure fixation, however, produced little change in stress-strain results other than a simple shift to lower strain and produced no plasticity. Both methods of fixation reduced stress relaxation and creep. Mechanical test results are consistent with a loss of ground substance matrix during fixation. Reductions in tensile strength after fixation may be due to "riveting" of collagen geometry, producing local stress concentrations.

The tensile viscoelastic properties of radial and circumferential strips of fresh porcine aortic valve leaflets have been examined and compared with the fiber architecture of the leaflets as seen in the scanning electron microscope. The leaflet material showed nonlinear viscoelasticity, largely independent of strain rate, and dependent on the degree of preconditioning by cyclic loading. Preconditioning to a stable stress-strain response could only be accomplished on 1-cm-long strips above a minimum width of 4 mm (circumferential) and 6 mm (radial). Preconditioning yielded a more elastic and extensible material with reduced hysteresis. The leaflets were markedly anisotropic. Circumferential strips were up to 8 times stronger and stiffer than radial strips, and displayed greater stress relaxation and less creep. The circumferential mechanical properties were due to well-aligned circumferential collagen bundles reinforcing the composite structure, while radial properties were due to a more random collagenous support throughout the leaflet. Despite the presence of a septal shelf on the right coronary leaflet, no mechanical differences could be discerned between leaflets.