Preparation Of A Novel Small-Caliber Vascular Graft

To date, no off-the-shelf graft has performed better than the autologous vessel in applications requiring small-bore(<6mm diameter) vascular grafts, synthetic materials such as polytetrafluorethylen(ePTFE) have been used successfully in peripheral vascular reconstruction. But synthetic small-diameter grafts have not produced acceptable results because of rapid thrombotic buildup in the vessel lumen. The properties of compliance and elasticity of synthetic materials are not as good as those of nature biomaterial; and donor sources of allografts and autografts are very limited. In vascular tissue engineering, the search for an ideal small-diameter graft has been compared to the search for the holy grail in respect to its immense clinical importance. In this study, we used the deendothelialized Wistar rat artery treated by super-low temperature as substrate. And reendothelialization of the treated artery with rabbit artery endothelial cells (RAECs) by rotating culture for 2 hours and then static culture for 3-5 days. Afterward, the reendothelializatial artery (xenograft) was implanted in rabbits femoral and the endothelialization, pathology and preliminary patency of the grafts in vivo were explored.Major content and results in this paper are:1. RAECs were isolated by the improved method of collagenase type II digestion and cultured, proliferated with DMEM (including 10μg/ml VEGF, 50μg /ml Heparin). The endothelial cells were identified by light microscope and transmission electron microscopy (TEM). Light microscope revealed that the cells isolated had typical "cobblestone" morphology. Weibel-Palade bodies were found in the cytoplasm of cultured cells in TEM test.2. Wistar rat artery was treated by super low temperature. Silver nitrate stain, Hematixylin and eosin (H&E) stain and scan electron microscope (SEM) showed that its native endothelium were totally removed. The rest construction of the artery was similar with the nature rat artery.3. Subcultured RAECs of 2-3 passage were seeded on the deendothelializatial Wistar rat artery grafts (DRAGs) with the density of 2 × 106 cell/cm2 and the grafts were cultured in DMEM by rotating for 2 hours, then with the static condition of 37℃ and 5% CO2 for 3-5 days. Silver nitrate staining and SEM indicated that the grafts showed an intact endothelium on the inner surface.4. The reendothelializatial Wistar rat arteries grafts (RRAGs) were implanted into the femoral of rabbits, and the rabbit artery grafts (RAGs) and the DRAGs were chosen as contrast groups. Two animals with artery grafts were sacrificed randomly at 1, 2 and 4 weeks after implantation. Harvested prostheses were analyzed histologically, pathologically and morphologically. H&E staining and SEM of the 1-week RRAGs samples revealed there were dense but non-confused endothelial cells on the intima and few inflammatory cells in the adventitia, similarly with RAGs. But for DRAGs, the inner layers had few endothelial cells and many inflammatory cells in the adventitia. The results of 4-weeks RRAGs and RAGs indicated that an intact endothelium on the intima, but, much adherent thrombus occurred in DRAGs. The preliminary patency of 4-weeks of RRAGs, RAGs and DRAGs were 66.7%, 83.3% and 50% respectively.