Shrinkage temperature versus protein extraction as a measure of stabilization of photooxidized tissue

A rise in thermal denaturation temperature has been utilized as an indication of stabilization of collagen-containing materials such as pericardial tissue and porcine heart-valve leaflets following treatment with glutaraldehyde, Denacol, or other chemical agents. In contrast, stabilization of bovine pericardial tissue by dye-mediated photooxidation does not result in a significant rise in shrinkage temperature comparable with these treated materials. It was therefore hypothesized that a rise in shrinkage temperature is not a necessary indication for tissue stabilization. A sensitive protein extraction assay has been developed which can be used to monitor the stabilization of pericardial tissue by a variety of treatment methods, including photooxidation. A reduction in extractable protein, as analyzed by polyacrylamide gel electrophoresis, is noted for pericardial tissue treated with photooxidation, glutaraldehyde, or Denacol. Loss of extractable protein, as a function of treatment time, correlates well with a significant rise in shrinkage temperature for pericardium treated with glutaraldehyde or Denacol but not with photooxidation. This difference is attributed to the stabilization processes of glutaraldehyde and Denacol, which involve extensive crosslinking and polymer formation within and in addition to the native pericardial matrix, leading to a rise in matrix complexity and thermal stability. In contrast, photooxidation is a catalytic process involving modification and crosslink formation within existing matrix components, resulting in a material with little added matrix complexity.