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To address the need of alternatives to autologous vessels for small-calibre vascular applications (e.g. cardiac surgery), a bio-hybrid semi-degradable material composed of silk fibroin (SF) and polyurethane (Silkothane®) was herein used to fabricate very small-calibre grafts (Ø

To address the need of alternatives to autologous vessels for small-calibre vascular applications (e.g. cardiac surgery), a bio-hybrid semi-degradable material composed of silk fibroin (SF) and polyurethane (Silkothane®) was herein used to fabricate very small-calibre grafts (Øin = 1.5 mm) via electrospinning. Bio-hybrid grafts were in vitro characterized in terms of morphology and mechanical behaviour, and compared to similar grafts of pure SF. Similarly, two native vessels from a rodent model (abdominal aorta and vena cava) were harvested and characterized. Preliminary implants were performed on Lewis rats to confirm the suitability of Silkothane® grafts for small-calibre applications, specifically as aortic insertion and femoral shunt. The manufacturing process generated pliable grafts consisting of a randomized fibrous mesh and exhibiting similar geometrical features to rat aortas. Both Silkothane® and pure SF grafts showed radial compliances in the range from 1.37 ± 0.86 to 1.88 ± 1.01% 10−2 mmHg−1, lower than that of native vessels. The Silkothane® small-calibre devices were also implanted in rats demonstrating to be adequate for vascular applications; all the treated rats survived the surgery for three months after implantation, and 16 rats out of 17 (94%) still showed blood flow inside the graft at sacrifice. The obtained results lay the basis for a deeper investigation of the interaction between the Silkothane® graft and the implant site, which may deal with further analysis on the potentialities in terms of degradability and tissue formation, on longer time-points.

To solve the problem of vascular access failure, a novel semi-degradable hybrid vascular graft, manufactured by electrospinning using silk fibroin and polyurethane (Silkothane), has been previously developed and characterized in vitro. This proof-of-concept animal study aims at evaluating the performances of Silkothane grafts in a sheep model of arteriovenous shunt, in terms of patency and short-term remodeling. Nine Silkothane grafts are implanted between the common carotid artery and the external jugular vein of nine sheep, examined by palpation three times per week, by echo-color Doppler every two weeks, and euthanized at 30, 60, and 90 days (N = 3 per group). At sacrifice, grafts are harvested and submitted for histopathology and/or scanning electron microcopy (SEM). No cases of graft-related complications are recorded. Eight of nine sheep (89%) show 100% primary unassisted patency at the respective time of sacrifice (flow rate 1.76 ± 0.61 L min-1 , one case of surgery-related thrombosis excluded). Histopathology and SEM analysis evidence signs of inflammation and pseudointima inside the graft lumen, especially at the venous anastomosis; however, endoluminal stenosis never impairs the functionality of the shunt and coverage by endothelial cells is observed. In this model, Silkothane grafts grant safety and 100% patency up to 90 days.

Several attempts made so far to combine silk fibroin and polyurethane, in order to prepare scaffolds encompassing the bioactivity of the former with the elasticity of the latter, suffer from critical drawbacks concerning industrial and clinical applicability (e.g., separation of phases upon processing, use of solvents unaddressed by the European Pharmacopoeia, and use of degradable polyurethanes). Overcoming these limitations, in this study, we report the successful blending of regenerated silk fibroin with a medical-grade, non-degradable polyurethane using formic acid and dichloromethane, and the manufacturing of hybrid, semi-degradable electrospun tubular meshes with different ratios of the two materials. Physicochemical analyses demonstrated the maintenance of the characteristic features of fibroin and polyurethane upon solubilization, blending, electrospinning, and postprocessing with ethanol or methanol. Envisioning their possible application as semidegradable substrates for haemodialysis arteriovenous grafts, tubular meshes were further characterized, showing submicrometric fibrous morphologies, tunable mechanical properties, permeability before and after puncture in the same order of magnitude as commercial grafts currently used in the clinics. Results demonstrate the potential of this material for the development of hybrid, new-generation vascular grafts with disruptive potential in the field of in situ tissue engineering. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 807-817, 2019.

Clinically available alternatives of vascular access for long-term haemodialysis - currently limited to native arteriovenous fistulae and synthetic grafts - suffer from several drawbacks and are associated to high failure rates. Bioprosthetic grafts and tissue-engineered blood vessels are costly alternatives without clearly demonstrated increased performance. In situ tissue engineering could be the ideal approach to provide a vascular access that profits from the advantages of vascular grafts in the short-term (e.g. early cannulation) and of fistulae in the long-term (e.g. high success rates driven by biointegration). Hence, in this study a three-layered silk fibroin/polyurethane vascular graft was developed by electrospinning to be applied as long-term haemodialysis vascular access pursuing a 'hybrid' in situ engineering approach (i.e. based on a semi-degradable scaffold). This Silkothane ® graft was characterized concerning morphology, mechanics, physical properties, blood contact and vascular cell adhesion/viability. The full three-layered graft structure, influenced by the polyurethane presence, ensured mechanical properties that are a determinant factor for the success of a vascular access (e.g. vein-graft compliance matching). The Silkothane ® graft demonstrated early cannulation potential in line with self-sealing commercial synthetic arteriovenous grafts, and a degradability driven by enzymatic activity. Moreover, the fibroin-only layers and extracellular matrix-like morphology, presented by the graft, revealed to be crucial in providing a non-haemolytic character, long clotting time, and favourable adhesion of human umbilical vein endothelial cells with increasing viability after 3 and 7 d. Accordingly, the proposed approach may represent a step forward towards an in situ engineered hybrid vascular access with potentialities for vein-graft anastomosis stability, early cannulation, and biointegration.

BACKGROUND In an attempt at answering the need for a valuable and durable hemodialysis vascular access, a novel semi‐degradable hybrid vascular graft, manufactured by electrospinning using silk fibroin and polyurethane (Silkothane®), has been developed and characterized in vitro according to standards. OBJECTIVES In this proof‐of‐principle animal study, we aimed at evaluating the performances of Silkothane® grafts in a sheep model of arteriovenous shunt, with respect to patency and short‐term graft remodeling. MATERIALS AND METHODS Nine Silkothane® grafts (6mm inner diameter, ≈7cm long, ≈400μm thick) were surgically placed into the left cervical fossa of nine female sheep, between the common carotid artery and the external jugular vein. During the housing period the animals were treated with an inhibitor of platelets aggregation. Grafts were examined by palpation three times per week, and by duplex ultrasound every two weeks to exclude significant stenosis, dilation, presence of thromboses, neointimal proliferation or other complicating features. The sheep were euthanized at 30, 60 and 90 days (N=3 for each group). At sacrifice, grafts were harvested, fixed, and submitted for histologic, immunohistochemical and Scanning Electron Microscopy examination. RESULTS No cases of graft‐related complications (e.g. seroma formation, transgraft bleeding, aneurismal dilation, calcification, structural yielding) were recorded in this study. One graft failed due to a thrombotic event likely related to a technical error (11%). One sheep had post‐operative wound infection, surgically resolved (11%). Four out of nine sheep (44%) showed post‐operative edema, usually disappearing in 7 days. Eight of nine sheep (89%) showed 100% primary unassisted patency at the respective time of sacrifice, as confirmed by palpation and ultrasound evaluation (average flow rate 1.76 ± 0.61 l/min). Histological, immunoistochemical and SEM analysis evidenced signs of inflammation and formation of neointimal hyperplasia at the venous anastomosis, as it typically happens with arteriovenous fistulae and synthetic grafts. CONCLUSIONS In our ovine model of arteriovenous shunt, Silkothane® grafts provided evidences of safety and efficacy in granting 100% patency up to 90 days. Further studied need to be conducted in order to directly compare the novel grafts with commercially available synthetic grafts.

“Bioreactors”, a term generally associated with classical industrial bioprocesses such as fermentation, was initially used in tissue engineering applications to describe little more than simple mixing of a Petri dish. Over the last two decades, bioreactors used in tissue engineering research evolved, not only for the function of engineering in vitro various types of biological tissues (e.g., skin, tendons, blood vessels, cartilage, and bone), but also to serve as defined model systems supporting investigations on cell function and tissue development. In recent years, as bioreactors continued to progress in sophistication, the term has gradually become synonymous with sophisticated devices enabling semiautomated, closely monitored, and tightly controlled cell and tissue culture. In particular, by controlling specific physicochemical culture parameters at defined levels, bioreactors provide the technological means with which to perform controlled studies aimed at understanding the effects of specific biological, chemical, or physical cues on basic cell functions in a three-dimensional spatial arrangement. Moreover, bioreactors successfully make up for limitations of conventional manual methods when driving the development of structurally uniform and functionally effective three-dimensionally engineered constructs.

Over the last decade, we have witnessed an increased recognition of the importance of 3D culture models to study various aspects of cell physiology and pathology, as well as to engineer implantable tissues. As compared to well-established 2D cell-culture systems, cell/tissue culture within 3D porous biomaterials has introduced new scientific and technical challenges associated with complex transport phenomena, physical forces, and cell-microenvironment interactions. While bioreactor-based 3D model systems have begun to play a crucial role in addressing fundamental scientific questions, numerous hurdles currently impede the most efficient utilization of these systems. We describe how computational modeling and innovative sensor technologies, in conjunction with well-defined and controlled bioreactor-based 3D culture systems, will be key to gain further insight into cell behavior and the complexity of tissue development. These model systems will lay a solid foundation to further develop, optimize, and effectively streamline the essential bioprocesses to safely and reproducibly produce appropriately scaled tissue grafts for clinical studies.