The Study Of Heparin-grafted Decellularized Vascular Scaffolds Used For Arteriovenous Fistula In Animal Experiments

Objective:1.To evaluate the effect of decellularization and heparin grafting on the tissue structure of vascular decellularized scaffolds;2.To evaluate the short-term anticoagulant effect of heparin-grafted decellularized vascular scaffolds;3.To observe the long-term performance of heparin-grafted decellularized vascular scaffolds used for arteriovenous fistula.Methods:1.Using the porcine carotid arteries as the source of vascular material,the porcine arteries were decellularized by detergents,and heparin grafting was carried out in the lumen of the decellularized vascular scaffolds by layer-by-layer self-assembly technique to obtain heparin-grafted decellularized vascular scaffolds.2.Hematoxylin-eosin(HE)staining,Masson collagen fiber staining,Verhoeff's Van Gieson(EVG)staining,toluidine blue staining and scanning electron microscopy were performed to analyze the structure of the vascular scaffolds and the effect of heparin grafting.3.Heparin-grafted decellularized porcine vascular scaffolds were evaluated for anticoagulant performance for three hours by the experiment of in vitro porcine arteriovenous fistula.The harvested vascular scaffolds were analyzed by tissue staining(HE,Masson,EVG,toluidine blue),scanning electron microscopy;4.The heparin-grafted decellularized porcine carotid arteries were used as vascular scaffolds in the experiment of arteriovenous fistula in vivo.The vascular scaffolds were removed 4 weeks after the implantation,and Color Doppler ultrasound were performed before and after the implantation.The removed vascular scaffolds were stained with HE,Masson,EVG and scanning electron microscopy.Results:1.(1)Relevant histological staining results showed that the porcine carotid artery had no obvious structure change or damage on collagen fibers and elastic fibers before and after decellularized by detergents as well as heparin grafting by layer-by-layer self-assembly technique.(2)Toluidine blue staining showed that layer-by-layer self-assembly technology could form a blue-stained layer of heparin in the lumen of the vascular scaffolds;(3)Scanning electron microscopy image showed that the lumen of the vessel became rough after decellularization,The continuous fiber arrangement formed a pore structure with different unequalities.After heparin grafting,the lumen of the blood vessel became relatively flat,and the pores of the fibrous network were reduced.2.(1)The tissue staining of harvested vascular scaffold material after the experiment of in vitro porcine arteriovenous fistula revealed that obvious thrombosis was found in the lumen of non heparin-grafted decellularized vascular scaffolds,while the heparin-grafted decellularized vascular scaffolds showed a small amount of nucleated cell adhesion in the lumen;(2)The inner layer of the heparin-grafted decellularized vascular sacffolds were still stained blue by Toluidine blue staining after the experiment,while non heparin-grafted decellularized vascular sacffolds were still negative for staining;(3)Scanning electron microscopy showed that a large number of spheroid cells adhered to the inner surface of the non heparin-grafted decellularized vascular sacffolds,while the inner surface of the heparin-grafted decellularized vascular sacffolds were smooth and no cell adhesion was observed.3.Four weeks after the implantation of the arteriovenous fistula of the pigs,the three groups of heparin-grafted decellularized porcine carotid arteries were completely occluded.Conclusion:1.Using the layer-by-layer self-assembled heparin grafting technique to modified the decellularized vascular scaffolds could form a favorable heparin coating without causing damage to the vascular structure.2.Heparin-grafted decellularized vascular scaffold materials had certain effectiveness in short-term anticoagulant ability.3.The long-term performance of the heparin-grafted decellularized vascular scaffold material was not ideal while using it as a vascular graft in the animal model of pig.