Bovine pericardium has been extensively applied as the biomaterial for artificial heart valves and may potentially be used as a scaffold for tissue-engineered heart valves after decellularization. Although various methods of decellularization are currently available, it is unknown which method is the most ideal one for the decellularization for bovine pericardium. We compared three decellularization methods, namely, the detergent and enzyme extraction (DEE), the trypsin (TS), and the Triton X-100 and sodium-deoxycholate (TSD) method, to examine their efficacy on cell removal and their preservation of the mechanical function and the tissue matrix structure. Results indicated that decellularization was achieved by all the three methods as confirmed by hematoxylin-eosin staining, scanning electron microscopy, as well as quantitative DNA measurement. However, TS and TSD methods resulted in severe structural destruction of the bovine pericardium as shown by von Gieson staining and Gomori staining. Furthermore, both TS and TSD methods changed the mechanical property of the bovine pericardium, as evidenced by a lower elastic modulus, maximal-stress, maximal-disfiguration, maximal-load, and maximal-strain. In conclusion, the DEE method achieved both a complete decellularization and preservation of the mechanical function and tissue structure of the bovine pericardium. Thus, this method is superior to either the TS or the TSD method for preparing decellularized bovine pericardium scaffold for constructing tissue-engineered heart valves.
Heparin treatment of decellularized xenografts has been reported to reduce graft thrombogenicity. However, little is known about the in vivo comparison of heparin-treated with non-heparin-treated xenografts, especially for small-caliber vascular implants. We implanted either a heparin-treated or a non-heparin-treated canine carotid artery as bilateral carotid xenograft in rabbits (n = 24). Small-caliber xenografts (3 approximately 4 mm) were decellularized by enzymatic and detergent extraction and were further covalently linked with heparin. During implantation, thrombosis rate was 4% in the heparin-treated xenografts and 25% in the non-heparin-treated xenografts after 3 weeks (P < 0.05). After 6 months, it was 8 versus 58%, respectively (P < 0.01). Both heparin-treated and non-heparin-treated xenografts harvested at the end of 3 and 6 months showed a satisfactory cellular reconstruction of either smooth muscle cells or endothelial cells. These results indicate that heparin treatment of the small-caliber decellularized xenograft reduces the in vivo thrombogenicity. Both heparin-treated and non-heparin-treated xenografts seem to undergo a similar cellular remodeling process up to 6 months.
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