Supramolecular structure of human aortic valve and pericardial xenograft material: atomic force microscopy study.

Pericardial tissue (bovine or porcine), chemically stabilized with glutaraldehyde (GA), is widely used in cardiovascular surgery in the form of bioprosthetic valves. GA reacts with tissue proteins and creates inter- and intra-molecular cross-links, resulting in improved durability. However, tissue calcification and mechanical damage are still unresolved problems. The purpose of this study was to examine the surface topography of normal human aortic valve and GA-stabilized porcine pericardium tissue in order to gain comparative insight into supramolecular structure of both tissues. The analysis was focused on morphologic evaluation of collagen constituents of the tissues. Atomic force microscopy working in the contact mode in air was employed in the study. Considerable diversity in the spatial orientation of collagen fibrils for the human aortic valve and pericardial tissue were observed. It was found that different forms of collagen fibril packing, i.e. dense and "in phase" or loose, could have an impact on the collagen D-banding pattern. Stabilization with GA introduced significant changes in the surface topography of collagen fibrils and in their spatial organization on the tissue surface. Strong disturbance in the fibril's D-spacing was observed. It was also suggested, that the observed structural changes at the supramolecular level might make an important contribution to the progressive damage and calcification of the tissue. The presented results demonstrate that the AFM method can be useful for non-destructive structural characterization of heart valves and bioprosthetic heart valve material.

Raman spectroscopic study of glutaraldehyde-stabilized collagen and pericardium tissue.

For the first time, Raman spectroscopy has been employed to investigate formation of cross-links in collagen and porcine pericardium tissue upon glutaraldehyde (GA) treatment. GA treatment causes a very high fluorescence background, which overlaps Raman bands. It has been found that short fixation time, i.e. 2 h, reduces background radiation significantly, providing new possibilities for studying changes in molecular structure of collagen upon GA modification. The observed changes in position and intensity of Raman bands allowed us to recognize different types of GA-collagen interactions. Strong spectral evidence has been found for the peptide contribution to the formation of the GA-collagen cross-links and for the formation of secondary amines via Schiff base intermediates, and pyridinium-type cross-links. The results also revealed that different hydration levels and a more complex structure of intact tissue in comparison to collagen preparation strongly influence the formation of a GA cross-linking network, e.g. ether-type bond is preferred to form in a less hydrated collagen preparation. Our results have shown that GA treatment causes an increase in water content of pericardium tissue and collagen.