Effect Of Decellularized Rat Aorta On Reducing Tissue Response After Transplantation

Background:With the increase in the number of cardiovascular diseases and peripheral artery diseases,autologous vascular grafts have been unable to meet the clinical needs,and it has become an urgent clinical need to find commercial small-diameter vascular grafts.The construction technology of in vitro tissue engineered blood vessels(TEBVs)has become more and more mature,but it is difficult for commercial engineered blood vessels to be derived from somatic cells,and more blood vessel tissue grafts are allogeneic or even xenograft.Theoretically,decellularization can reduce the immunogenicity of tissues,but the decellularized allogeneic blood vessel tissue will still have some residual tissue reaction,the extent of which will affect the patency rate of TEBVs transplantation in the medium and long term.At present,there are few studies on the tissue response about decellularized allogeneic vascular tissue after transplantation.Objective:We used rat aortic vascular tissue as material to observe the tissue response of decellularized rat aorta allograft and to evaluate the degree of tissue response after allotransplantation.Methods:The rat aorta was decellularized with CHAPS combined with SDS double-detergents,Hematoxylin-Eosin staining was performed.Detecting and quantitating DNA residue by Picogreen reagent to evaluate the effect of decellularization.The cytotoxicity of the decellularized blood vessels was analyzed by the direct-contact test in vitro.Thirty-two Wistar rats were divided into 4 groups with 8 rats in each group.Surgical treatment on both sides of each animal dorsal tissue to form pouch.Subcutaneous transplanting decellularized Sprague-Dawley rat aorta(experimental group)and non-acellular Sprague-Dawley rat aorta(control group)as allograft,each group has the Wistar rat aorta as autologous transplantation.The whole group of animals were executed in the four time points(1,7,14,28 days).The specimens were examined with histological and morphological analysis.Semi-quantitative methods were used to evaluate the number of inflammatory cells,the presence of multinucleated giant cells,angiogenesis,fat infiltration and fibrosis.CD3 and CD68 immunohistochemical staining were used to determine the type of infiltrated cells,and the tissue responses in different days were analyzed.Results:Hematoxylin-Eosin staining showed that there was no cellular component left in the decellularized vascular scaffold,and the extracellular matrix component was relatively intact.Picogreen analysis showed that the DNA content of decellularized vascular scaffolds was significantly decreased.The direct-contact cytotoxicity test showed that the acellular scaffold had good compatibility with human aortic outer membrane fibroblasts and the cytotoxicity was grade 0.Semi-quantitative score of tissue response after subcutaneous implantation showed that decellularized allogenic rat aorta remained unstimulated all the time,and non-acellular allogenic rat aorta evolved from severe stimulation to unstimulated.The results of tissue response and tissue remodeling in the allogenic decellularized rat aorta were similar to those in the autograft group.The non-acellular rat aorta showed significant acute inflammatory responses(days 1 and 7),multinucleated giant cell infiltration(days 7 and 14),and adipocyte infiltration(days 28).Conclusion:1.The rat aorta decellularized by double-detergents can become an excellent vascular scaffold,with good vascular structure and without cytotoxicity.2.There is still some tissue reaction in the allogeneic decellularized vascular tissue,but the result of this tissue reaction is close to that of autogenous vascular tissue in tissue in the subcutaneous transplantation model.