Comparative efficiency of detergent-based decellularization procedures in vascular tissue engineering

Abstract

Introduction Atherosclerosis is the leading cause of mortality and morbidity across the world. Conventional therapies available today offer good clinical results, however, the „gold standard” treatment is considered surgical bypass grafting. It assumes blood flow improvement by reconnecting blood vessels with specific vascular conduits, biological or synthetic ones. Studies concerning the evaluation of different grafts’ patency rates have been conducted already. They have shown satisfactory results for replacement of large- and medium-diameter arteries. Nevertheless, the optimal vascular substitutes appliable to small-diameter vessels are still in the research and development. Considering these limitations, high attention has been focused on manufacturing vascular grafts by tissue engineering techniques (TEVGs). A number of different approaches have been taken in this research field. They can be broadly categorized into scaffold-based methods using synthetic or natural materials and decellularized natural matrix techniques (Fig. 1). Purpose To evaluate the effect of the detergents widely used in tissue decellularization on histology of blood vessels and to understand their potential impact on functional changes. Material and methods Fresh porcine aortas (PAs: length 70–170 mm, lumen diameter 14–25mm, wall thickness 2–3mm) were obtained from a local meat. The samples were cleaned off excess connective tissue and fats and rinsed in ddH2O for 24h (Fig. 2). The research protocol included 5 experimental and one control group (Tab. 1) Material and methods For statistical calculations, SPSS software was used. Data were expressed as DNA mean in ng per milligram (ng/mg) ± SD. Statistical significance was obtained by one-way ANOVA in conjunction with Turkey post-hoc procedure. The significance level was set up at p=0,05. Results Hematoxylin/eosin staining of native and decellularized PA (magnification, X10 / X40). Native tissue: normal aorta with normal tunica media and tunica intima in adult pig Group A: complete decellularization with few endothelial cells remaining; mostly of collagen fibers preserved Group B: decellularization quality >60%; elastic fibers structure significantly altered Group C: remnant cells visible over all the layers Group D: decellularization quality >60% Group E: almost no nuclear material visible (>80%). DNA quantification All DC groups had significantly lower DNA content (p<0,05) compared to native tissue. Conclusions Detergent-based decellularization technique were found to be more effectively in cellular components elimination. However, biocompatibility and mechanical properties assessments should be carried out in future studies.