Experimental Study On The Clinical Feasibility Of All Bio-based Small-caliber Vascular

Background:Coronary artery bypass grafting (CABG) surgery is currently the most effective means of treating coronary heart disease. There are more than 800,000 cases of patients with coronary artery bypass grafting each year in the world, so the grafts of the demand is very large. With coronary artery bypass grafting surgery is growing maturity and popularity, the bridge vascular graft material studies have become an important topic of people's attention. Clinical autologous blood vessels are the most commonly used as the preferred material, but the source is limited. The varicose veins, malformations, atherosclerosis and other factors limit their use. There are about 30% of patients had a lack of autologous blood vessels for transplantation. Blood vessel tissue engineering is the best way to solve the shortage of clinical vascular graft, artificial blood vessels and tissue-engineered blood vessels of non-biological materials for building a small-caliber blood vessel's results are unsatisfactory. All bio-based xenograft blood vessels are similar to the normal body vascular structure and function, with good mechanical properties. It also can be self-renewal, and avoid rejection, reduce infection rates, help to improve the long-term graft patency. In this study, we use the method of decellularize and heparinization to prepare the small-caliber blood vessels full of biological material in a short time, thus providing a new source of graft.Objective:To build a small-caliber vascular scaffold by enzymatic-detergent extraction and heparin linkage procedure treatment canine carotid artery off, thus providing a new source of graft.Method:1. Take warm ischemia time of less than 30min canine bilateral carotid arteries in Laboratory of Forensic Medicine, Shanxi Medical University.Use the saline which contain antibiotics (penicillin potassium 1000 U/ml) to flush the carotid arteries and lumen. Gentle divest its outer loose connective tissue. Then the blood vessels cut into 5cm as the experimental material, and put them into 4℃Hanks solution to save. All vasculars were divided into three groups:A Group:Fresh canine carotid artery Group, B Group:Single-off cells Group and C group:heparin-binding group.2. Observed indicatrix:Three groups vascular specimens were did HE, VG staining and electron microscopy separately, then to evaluat the overall morphology of the specimen and collagen fibers and elastic fibers composition on histology. Measuring the mechanical properties of specimens of blood vessels on the Material Properties Testing Machine. B, C two sets of specimens were stained toluidine blue (0.05mg/ml) to observe heparin-binding and binding effect of the degree, and determin the clotting time of the two groups in vitro, and of combined effects of heparin anticoagulation.Results:HE and VG staining:A group:the cells in the vessel wall can be seen uniformly distributed, and the fibers are dense and clear. The cellular components of blood vessels have been completely removed in B,C group. The integrity of extracellular matrix components retain completely, the fiber did not breakage did not too. Compared with the A group, B, C groups scaffold fibers arranged more loosely. VG staining showed that stent inside the material can clearly show the complete structure of the internal elastic membrane. Scanning electron microscope:A group of vascular integrity of the inner surface covered with a monolayer endothelial cells. After acellular treatment, endothelial cell layer was completely removed without any residual cells and cellular debris components. The inner surface has a complete mesh arranged elastin, and there is a regular pore structure and single fiber without significant expansion of or fracture. Mechanical performance test showed between the three groups tensile strength, maximum load, maximum deformation, strain rate, sutures no significant difference in endurance (P>0.05), only the B, C the elastic modulus of the two groups is less than A group (P<0.05). Toluidine blue staining:C group of full-thickness wall stained, B group were negative. Determination of clotting time:the B group all had thrombosis within 60min, while Group C were not the formation of thrombus within 60min.Conclusion:The canine common carotid arteries decellularized by enzyme-detergent have not cellular components, and have acellular matrix and three-dimensional structure completely. The mechanical properties of the grafts before and after decellularization were no significant change, and heparin cover on the wall full thickness. After heparin treatment, the blood vessel specimens can effectively carry out anticoagulation. Vascular graft patency can be maintained, and meet the demands of vascular grafting.