Biaxial mechanical/structural effects of equibiaxial strain during crosslinking of bovine pericardial xenograft materials.

We have investigated the effect of biaxial constraint during glutaraldehyde crosslinking on the equibiaxial mechanical properties of bovine pericardium. Crosslinking of cruciate samples was carried out with: (i) no applied load, (ii) an initial 25 g ( approximately 30 kPa) equibiaxial load, or (iii) an initial 200 g (approximately 250 kPa) equibiaxial load. All loading during crosslinking was done under a defined initial equibiaxial load and subsequently fixed biaxial strain. Load changes during crosslinking were monitored. Mechanical testing and constraint during crosslinking were carried out in a custom-built biaxial servo-hydraulic testing system incorporating four actuators with phase-controlled waveform synthesis, high frame-rate video dimension analysis, and computer-interfaced data acquisition. The paired biaxial stress strain responses under equibiaxial loading at 1 Hz (before and after treatment) were evaluated for changes in anisotropic extensibility by calculation of an anisotropy index. Scanning electron microscopy (SEM) was performed on freeze-fractured samples to relate collagen crimp morphology to constraint during crosslinking. Fresh tissue was markedly anisotropic with the base-to-apex direction of the pericardium being less extensible and stiffer than the circumferential direction. After unconstrained crosslinking, the extensibility in the circumferential direction, the stiffness in the base-to-apex direction, and the tissue's anisotropy were all reduced. Anisotropy was preserved in the tissue treated with an applied 25 g load; however, tissue treated with an applied 200 g load became extremely stiff and nearly isotropic. SEM micrographs correlated well with observed extensibility in that the collagen fibre morphology changed from very crimped (unconstrained crosslinking), to straight (200 g applied load). Biaxial stress-fixation may allow engineering of bioprosthetic materials for specific medical applications.

Thickness measurement of soft tissue biomaterials: a comparison of five methods.

Thickness measurement in soft connective tissues is a continuing problem due to the apparent compression of the tissue by micrometer-type gauges. We have compared five methods for the measurement of thickness: (1) a Mitutoyo non-rotating thickness gauge; (2) a custom-built, instrumented thickness gauge which was strain-gauged to measure contact force; (3) a commercial Hall effect probe (Panametrics Magna-Mike); (4) a custom-built electrical resistance probe; and (5) measurement of fresh frozen histological sections under polarized light. Using bovine pericardium as a test material, all the methods examined were adequate to assess sample-to-sample and location-to-location differences in thickness. The resistance gauge gave significantly greater thicknesses than did the other methods, with little or no compression; indeed, extrapolation to zero load of thickness readings from the instrumented gauge yielded identical thickness. Thicknesses measured by frozen sections were indistinguishable from those measured with the non-rotating gauge, the instrumented gauge under 0.5-1.2 g compressive load, or the Hall effect probe. With the correct technique, the simple and inexpensive non-rotating gauge remains a pragmatic choice for thickness measurement in planar soft tissue.

Biomechanical and ultrastructural comparison of cryopreservation and a novel cellular extraction of porcine aortic valve leaflets.

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.