Acellular Endocardium as a Novel Biomaterial for the Intima of Tissue-Engineered Small-Caliber Vascular Grafts.

We aimed to investigate whether acellular endocardium can be used as a useful biomaterial for the intima of engineered small-caliber vascular grafts. Fresh endocardium was harvested from the swine left atrium and was decellularized by digestion with the decellularization solution of Triton X-100 and SDS containing DNase I and RNase A. Surface morphological characteristics and Young's modulus were evaluated. To analyze the effect of mechanical characteristics on cell adhesion, the decellularized endocardium was stiffened with 2.5% glutaraldehyde. Small-caliber vascular grafts were constructed using decellularized endocardium treated with or without glutaraldehyde as the intima. CD34+ cells were seeded onto the luminal surface of the vascular grafts and linked to bioreactors that simulate a pulsatile blood stream. Acellular endocardium had distinct surface morphological characteristics, which were quite different from those of other materials. The compliance of acellular endocardium was higher than that of other materials tested by Young's modulus. CD34+ cells formed a monolayer structure and adhered to the inner face of the acellular endocardium. The glutaraldehyde treatment stiffened the acellular endocardium but had little impact on the surface morphological characteristics or static adhesiveness of the cells. Data from the bioreactor study showed that the detachment of the cells from the surface of glutaraldehyde-treated acellular endocardium increased dramatically when the pressure was equal or higher than 40 mm Hg, while the cells on the untreated acellular endocardium remained well and formed confluent monolayers and tight junctions under the same pressure. Acellular endocardium has distinct structures and mechanical characteristics that are beneficial for CD34+ cell adhesion and retention under dynamic fluid perfusion. Thus, it can be used as a useful biomaterial for the construction of the intima of engineered small-caliber vascular grafts.

Production of novel elastic bacterial nanocellulose/polyvinyl alcohol conduits via mercerization and phase separation for small-caliber vascular grafts application.

Size and properties of tubular bacterial nanocellulose (BNC) can be regulated by controllable mercerization with thinner tube walls, better mechanical properties, and improved biocompatibility. Although mercerized BNC (MBNC) conduits have considerable potential as small-caliber vascular grafts (<6 mm), poor suture retention and lack of compliance that cannot match natural blood vessels increase the difficulty of surgery and limit potential clinical application. Polyvinyl alcohol (PVA) is a kind of hydrophilic polymer with good biocompatibility and elasticity, which can precipitate in alkaline solutions. In this study, novel elastic mercerized BNC/PVA conduits (MBP) are manufactured combining mercerization of BNC tubes with precipitation and phase separation of PVA with thinner tube wall, improved suture retention, better elasticity, good hemocompatibility and great cytocompatibility. The MBP obtained with 12.5 % PVA is selected for transplantation in a rat abdominal aorta model. For 32 weeks, normal blood flow is observed using Doppler sonographic inspection, which demonstrates long-term patency. Immunofluorescence staining results also indicate the formation of endothelium and smooth muscle layers. The results indicate the introduction of PVA, and its phase separation into mercerization of tubular BNC can endow MBP conduits with better compliance and suture retention, making it a promising candidate for blood vessel replacement.

Vascular cells responses to controlled surface structure and properties of bacterial nanocellulose artificial blood vessel after mercerization.

Therapeutic benefits of small caliber artificial blood vessels to cure cardio and cerebrovascular diseases are mainly limited by their low patency during long-term transplantation. Bacterial nanocellulose (BNC), as a natural polysaccharide mainly synthesized by a bacterium Komagataeibatacter xylinus, has shown great potential in small-caliber vascular graft applications due to its shape controllability, and furthermore its physical surface structure can be adjusted with different treatments. However, influences of physical surface structure and properties of BNC conduits on behaviors of vascular cells have not been investigated. In this work, mercerized BNC conduits (MBNC) with different surface roughness and stiffness were constructed by controlled alkali (NaOH) treatment. The changes of surface structures and properties significantly affected the behaviors of vascular cells and gene expression; meanwhile, the cell seeding density also affected the cell responses. After mercerization with NaOH concentration > 10 %, it was observed that the increased stiffness of MBNC decreased several functional gene expressions of human vascular endothelial cells, and the pathological transformation of smooth muscle cells was inhibited. This study demonstrates physical surface structure of MBNC conduits will critically regulate functions and behaviors of vascular cells and it also provides important designing parameters to improve the long-term patency of BNC-based conduits.

Development of a dual-component infection-resistant arterial replacement for small-caliber reconstructions: A proof-of-concept study.

Introduction: Synthetic vascular grafts perform poorly in small-caliber (<6mm) anastomoses, due to intimal hyperplasia and thrombosis, whereas homografts are associated with limited availability and immunogenicity, and bioprostheses are prone to aneurysmal degeneration and calcification. Infection is another important limitation with vascular grafting. This study developed a dual-component graft for small-caliber reconstructions, comprising a decellularized tibial artery scaffold and an antibiotic-releasing, electrospun polycaprolactone (PCL)/polyethylene glycol (PEG) blend sleeve. Methods: The study investigated the effect of nucleases, as part of the decellularization technique, and two sterilization methods (peracetic acid and γ-irradiation), on the scaffold's biological and biomechanical integrity. It also investigated the effect of different PCL/PEG ratios on the antimicrobial, biological and biomechanical properties of the sleeves. Tibial arteries were decellularized using Triton X-100 and sodium-dodecyl-sulfate. Results: The scaffolds retained the general native histoarchitecture and biomechanics but were depleted of glycosaminoglycans. Sterilization with peracetic acid depleted collagen IV and produced ultrastructural changes in the collagen and elastic fibers. The two PCL/PEG ratios used (150:50 and 100:50) demonstrated differences in the structural, biomechanical and antimicrobial properties of the sleeves. Differences in the antimicrobial activity were also found between sleeves fabricated with antibiotics supplemented in the electrospinning solution, and sleeves soaked in antibiotics. Discussion: The study demonstrated the feasibility of fabricating a dual-component small-caliber graft, comprising a scaffold with sufficient biological and biomechanical functionality, and an electrospun PCL/PEG sleeve with tailored biomechanics and antibiotic release.

Construction of spider silk protein small-caliber tissue engineering vascular grafts based on dynamic culture and its performance evaluation.

Tissue engineering is an alternative method for preparing small-caliber (<6 mm) vascular grafts. Dynamic mechanical conditioning is being researched as a method to improve mechanical properties of tissue engineered blood vessels. This method attempts to induce unique reaction in implanted cells that regenerate the matrix around them, thereby improving the overall mechanical stability of the grafts. In this study, we used a bioreactor to seed endothelial cells and smooth muscle cells into the inner and outer layers of the electrospun spider silk protein scaffold respectively to construct vascular grafts. The cell proliferation, mechanical properties, blood compatibility and other indicators of the vascular grafts were characterized in vitro. Furthermore, the vascular grafts were implanted in Sprague Dawley rats, and the vascular grafts' patency, extracellular matrix formation, and inflammatory response were evaluated in vivo. We aimed to construct spider silk protein vascular grafts with the potential for in vivo implantation by using a pulsating flow bioreactor. The results showed that, when compared with the static culture condition, the dynamic culture condition improved cell proliferation on vascular scaffolds and enhanced mechanical function of vascular scaffolds. In vivo experiments also showed that the dynamic culture of vascular grafts was more beneficial for the extracellular matrix deposition and anti-thrombogenesis, as well as reducing the inflammatory response of vascular grafts. In conclusion, dynamic mechanical conditioning aid in the resolution of challenges impeding the application of electrospun scaffolds and have the potential to construct small-caliber blood vessels with regenerative function for cardiovascular tissue repair.

Silk Vascular Grafts with Optimized Mechanical Properties for the Repair and Regeneration of Small Caliber Blood Vessels.

As the incidence of cardiovascular diseases has been growing in recent years, the need for small-diameter vascular grafts is increasing. Considering the limited success of synthetic grafts, vascular tissue engineering/repair/regeneration aim to find novel solutions. Silk fibroin (SF) has been widely investigated for the development of vascular grafts, due to its good biocompatibility, tailorable biodegradability, excellent mechanical properties, and minimal inflammatory reactions. In this study, a new generation of three-layered SF vascular scaffolds has been produced and optimized. Four designs of the SILKGraft vascular prosthesis have been developed with the aim of improving kink resistance and mechanical strength, without compromising the compliance with native vessels and the proven biocompatibility. A more compact arrangement of the textile layer allowed for the increase in the mechanical properties along the longitudinal and circumferential directions and the improvement of the compliance value, which approached that reported for the saphenous and umbilical veins. The higher braid density slightly affected the grafts' morphology, increasing surface roughness, but the novel design mimicked the corrugation approach used for synthetic grafts, causing significant improvements in kink resistance.

Three-Layered Silk Fibroin Tubular Scaffold for the Repair and Regeneration of Small Caliber Blood Vessels: From Design to in vivo Pilot Tests.

Silk fibroin (SF) is an eligible biomaterial for the development of small caliber vascular grafts for substitution, repair, and regeneration of blood vessels. This study presents the properties of a newly designed multi-layered SF tubular scaffold for vascular grafting (SilkGraf). The wall architecture consists of two electrospun layers (inner and outer) and an intermediate textile layer. The latter was designed to confer high mechanical performance and resistance on the device, while electrospun layers allow enhancing its biomimicry properties and host's tissues integration. In vitro cell interaction studies performed with adult Human Coronary Artery Endothelial Cells (HCAECs), Human Aortic Smooth Muscle Cells (HASMCs), and Human Aortic Adventitial Fibroblasts (HAAFs) demonstrated that the electrospun layers favor cell adhesion, survival, and growth. Once cultured in vitro on the SF scaffold the three cell types showed an active metabolism (consumption of glucose and glutamine, release of lactate), and proliferation for up to 20 days. HAAF cells grown on SF showed a significantly lower synthesis of type I procollagen than on polystyrene, meaning a lower fibrotic effect of the SF substrate. The cytokine and chemokine expression patterns were investigated to evaluate the cells' proliferative and pro-inflammatory attitude. Interestingly, no significant amounts of truly pro-inflammatory cytokines were secreted by any of the three cell types which exhibited a clearly proliferative profile. Good hemocompatibility was observed by complement activation, hemolysis, and hematology assays. Finally, the results of an in vivo preliminary pilot trial on minipig and sheep to assess the functional behavior of implanted SF-based vascular graft identified the sheep as the more apt animal model for next medium-to-long term preclinical trials.