In vitro study in the endothelial cell compatibility and endothelialization of genipin-crosslinked biological tissues for tissue-engineered vascular scaffolds

Abstract

To overcome the cytotoxicity of the chemical reagents used to fix bioprostheses, genipin, a naturally occurring crosslinking agent, was used to fix biological tissues in present study. We prepared the biological vascular scaffolds through cell extraction and fixing the porcine thoracic arteries with 1% (by w/v) genipin solution for 3 days, and then examined their mechanical properties and microstructures; glutaraldehyde- and epoxy-fixed counterparts were used as controls. HUVECs were seeded on the type I collagen-coated surface of different modified acellular vascular tissues (fixed with different crosslinking agents), and the growths of HUVECs on the specimens were demonstrated by means of MTT test, the secretion of PGI2 and vWF by HUVECs on the various specimens was also measured. Finally, HUVECs were seeded on the luminal surface of acellular biological vascular scaffolds (<6 mm internal diameter) which were, respectively, treated in the same manner described above, and then cultured for 9 days. On the ninth day, the HUVECs on the luminal surface of these vascular scaffolds were examined morphologically and by immunohistochemistry. Genipin-fixation can markedly diminish antigenicity of the vascular tissues through partially getting rid of cell or reducing the level of free amino groups in the vascular tissues. Genipin-fixed acellular vascular tissues mimicked the natural vessels due to the maintenance of the integrity of total structure and the large preservation of the microstructures of collagen fibers and elastic fibers; therefore, it appeared suitable to fabricate vascular scaffolds in mechanical properties. Compared to controls, the genipin-fixed acellular vascular tissues were characterized by low cytotoxicity and good cytocompatibility. The HUVECs can not only proliferate well on the genipin-fixed acellular vascular tissues, but also preserve the activities and function of endothelial cells, and easily make it endothelialized in vitro. The results showed that the genipin-fixed acellular porcine vascular scaffolds should be promising materials for fabricating vascular grafts or the scaffolds of tissue-engineered blood vessels.