Vascular Graft Applications Research Articles

2,5-Dihydro-2,5-dimethoxyfuran crosslinked silk-chitosan blend tubular construct for vascular graft application

The doctrine of vascular tissue engineering guides through the fabrication of biopolymer based vascular scaffolds in this study with favorable hemocompatibility, biological interaction and degradation profile as the decisive factors for polymer selection and processing.2,5-Dihydro-2,5-dimethoxyfuran (DMDF) cross-linked porous tubular scaffolds were fabricated from Silk fibroin and Chitosan blend. The scaffolds were then subjected to structural, mechanical and chemical analysis followed by in vitro seeding of human mesenchymal stem cells (hMSC) and immunolabeling for endothelial differentiation markers. Animal studies were performed to elucidate biocompatibility and in vivo degradation profile.Nano-indentation profiling shows decreased displacement in cross-linked samples resulting from the increased rigidity when compared to uncross-linked ones. In vitro cell seeding shows hMSC differentiation into endothelial cell like lineage with positive immunocytochemical profiling for PECAM-1 and von Willebrand factor. Promising haemocompatibility and bio compatibility on in vivo implantation was noticed. Histological studies revealed minimal inflammatory reaction and favorable tissue remodelling with the gradual replacement of the scaffold.Silk-Chitosan blend, as a potential alternative to existing biopolymers for blood vessel engineering, offers ample mechanical properties along with promising biocompatibility. Primary in vitro and in vivo studies warrant elaborative long term assessment for intended application.

Fabrication of endothelial progenitor cell capture surface via DNA aptamer modifying dopamine/polyethyleneimine copolymer film

Endothelial progenitor cells (EPCs) are mainly located in bone marrow and circulate, and play a crucial role in repairmen of injury endothelium. One of the most promising strategies of stents designs were considered to make in-situ endothelialization in vivo via EPC-capture biomolecules on a vascular graft to capture EPCs directly from circulatory blood. In this work, an EPC specific aptamer with a 34 bases single strand DNA sequence was conjugated onto the stent surface via dopamine/polyethyleneimine copolymer film as a platform and linker. The assembled density of DNA aptamer could be regulated by controlling dopamine percentage in this copolymer film. X-ray photoelectron spectroscopy (XPS), water contact angle (WCA) and fluorescence test confirmed the successful immobilization of DNA aptamer. To confirm its biofunctionality and cytocompatibility, the capturing cells ability of the aptamer modified surface and the effects on the growth behavior of human umbilical vein endothelial cells (HUVECs), smooth muscle cells (SMCs) were investigated. The aptamer functionalized sample revealed a good EPC-capture ability, and had a cellular friendly feature for both EPC and EC growth, while not stimulated the hyperplasia of SMCs. And, the co-culture experiment of three types of cells confirmed the specificity capturing of EPCs to aptamer modified surface, rather than ECs and SMCs. These data suggested that this aptamer functionalized surface may have a large potentiality for the application of vascular grafts with targeted endothelialization.

Impact of elastin incorporation into electrochemically aligned collagen fibers on mechanical properties and smooth muscle cell phenotype

Application of tissue-engineered vascular grafts (TEVGs) for the replacement of small-diameter arteries is limited due to thrombosis and intimal hyperplasia. Previous studies have attempted to address the limitations of TEVGs by developing scaffolds that mimic the composition (collagen and elastin) of native arteries to better match the mechanical properties of the graft with the native tissue. However, most existing scaffolds do not recapitulate the aligned topography of the collagen fibers found in native vessels. In the current study, based on the principles of isoelectric focusing, two different types of elastin (soluble and insoluble) were incorporated into highly oriented electrochemically aligned collagen (ELAC) fibers and the effect of elastin incorporation on the mechanical properties of the ELAC fibers and smooth muscle cell (SMC) phenotype was investigated. The results indicate that elastin incorporation significantly decreased the modulus of ELAC fibers to converge upon that of native vessels. Further, a significant increase in yield strain and decrease in Young’s modulus was observed on all fibers post SMC culture compared with before the culture. Real-time polymerase chain reaction results showed a significant increase in the expression of α-smooth muscle actin and calponin on ELAC fibers with insoluble elastin, suggesting that incorporation of insoluble elastin induces a contractile phenotype in SMCs after two weeks of culture on ELAC fibers. Immunofluorescence results showed that calponin expression increased with time on all fibers. In conclusion, insoluble elastin incorporated ELAC fibers have the potential to be used for the development of functional TEVGs for the repair and replacement of small-diameter arteries.

Approaches to Fabricating Multiple-Layered Vascular Scaffolds Using Hybrid Electrospinning and Thermally Induced Phase Separation Methods

Fabrication of small-diameter vascular scaffolds has been a challenge in recent years, especially scaffolds with multiple layers. In this study, two approaches were proposed to fabricate triple-layered vascular scaffolds based on the electrospinning method and the thermally induced phase separation (TIPS) method. It was found that the electrospun fibers had a compact fibrous structure that provided good mechanical properties. The porous TIPS layer had high porosity and pore interconnectivity to facilitate cell penetration; however, this structure alone could not ensure sufficient mechanical properties for surgical applications. The triple-layered scaffolds, which consisted of electrospun TPU, TIPS TPU, and electrospun PPC layers, showed the highest mechanical properties and best structure and dimensions for vascular graft applications. Preliminary endothelial cell culture results showed that the cells could attach to and proliferate on the inside surface of the scaffolds with >95% viability.

Composite vascular scaffold combining electrospun fibers and physically-crosslinked hydrogel with copper wire-induced grooves structure

While the field of tissue engineered vascular grafts has greatly advanced, many inadequacies still exist. Successfully developed scaffolds require mechanical and structural properties that match native vessels and optimal microenvironments that foster cell integration, adhesion and growth. We have developed a small diameter, three-layered composite vascular scaffold which consists of electrospun fibers and physically-crosslinked hydrogel with copper wire-induced grooves by combining the electrospinning and dip-coating methods. Scaffold morphology and mechanics were assessed, quantified and compared to native vessels. Scaffolds were seeded with Human Umbilical Vein Endothelial Cells (HUVECs), cultured in vitro for 3 days and were evaluated for cell viability and morphology. The results showed that composite scaffolds had adjustable mechanical strength and favorable biocompatibility, which is important in the future clinical application of Tissue-engineered vascular grafts (TEVGs).

Fabrication of triple-layered vascular scaffolds by combining electrospinning, braiding, and thermally induced phase separation

Triple-layered small diameter vascular scaffolds, which consisted of thermoplastic polyurethane (TPU) and silk, were fabricated in this study for the first time by combining electrospinning, braiding, and thermally induced phase separation methods. These novel vascular scaffolds, which possess three layers of different structures (nanofibrous inner layer, woven silk filament middle layer, and porous outer layer), can mimic the structure of native blood vessels. The vascular scaffolds have a desirable toe region in tensile tests and sufficient suture retention and burst pressure for vascular graft applications. Endothelial cell culture tests showed that a cell layer could form on the inner surface of the scaffold with high cell viability and favorable morphology.

Development of a Polyester Coating Combining Antithrombogenic and Cell Adhesive Properties: Influence of Sequence and Surface Density of Adhesion Peptides.

Biofunctionalization strategies have been developed to improve small-diameter vascular grafts. However, a fully successful coating featuring antithrombogenic properties while allowing for endothelialization has not been achieved yet. In this report, we explored the combination of low-fouling polyethylene glycol (PEG) and adhesion peptides, namely, RGD, YIGSR, and REDV, grafted on top of polyvinylamine (PVAm)-coated polyester. The peptides were grafted over a wide range of density (ca. 20-2000 pmol/cm(2)) on top of a dense PEG underlayer. The coating performances were assessed through HUVEC adhesion, platelet attachment, and protein adsorption, which were all drastically diminished on PEG-coated samples. RGD exhibited the expected high adhesive properties, toward both HUVEC and platelets. REDV had no effect neither on platelet attachment, as expected, nor on HUVEC adhesion, in contrast with previous reports. YIGSR was the most promising sequence even though its combination with other agents should be explored to further decrease thrombogenicity for vascular graft applications.

Electrospinning thermoplastic polyurethane/graphene oxide scaffolds for small diameter vascular graft applications.

Fabrication of small diameter vascular grafts plays an important role in vascular tissue engineering. In this study, thermoplastic polyurethane (TPU)/graphene oxide (GO) scaffolds were fabricated via electrospinning at different GO contents as potential candidates for small diameter vascular grafts. In terms of mechanical and surface properties, the tensile strength, Young's modulus, and hydrophilicity of the scaffolds increased with an increase of GO content while plasma treatment dramatically improved the scaffold hydrophilicity. Mouse fibroblast (3T3) and human umbilical vein endothelial cells (HUVECs) were cultured on the scaffolds separately to study their biocompatibility and potential to be used as vascular grafts. It was found that cell viability for both types of cells, fibroblast proliferation, and HUVEC attachment were the highest at a 0.5wt.% GO loading whereas oxygen plasma treatment also enhanced HUVEC viability and attachment significantly. In addition, the suture retention strength and burst pressure of tubular TPU/GO scaffolds containing 0.5wt.% GO were found to meet the requirements of human blood vessels, and endothelial cells were able to attach to the inner surface of the tubular scaffolds. Platelet adhesion tests using mice blood indicated that vascular scaffolds containing 0.5% GO had low platelet adhesion and activation. Therefore, the electrospun TPU/GO tubular scaffolds have the potential to be used in vascular tissue engineering.

Fabrication and characterisation of an electrospun tubular 3D scaffold platform of poly(vinylidene fluoride-co-hexafluoropropylene) for small-diameter blood vessel application

In this research, nanofibrous 3D tubular (~4-mm-diameter tube) scaffolds of poly (vinylidene fluoride-co-hexafluoropropylene) were fabricated by electrospinning. The role of surface charge in the success of these scaffolds for potential small-diameter artificial vascular grafts has been investigated using streaming potential study. Prior to endothelial cell culture, surface properties such as wettability and the surface charge of these tubular scaffolds were evaluated using unmodified and fibrinogen-adsorbed surfaces to understand their interaction with surrounding environment. The tubular scaffolds constructed using electrospinning show similar mechanical properties such as tensile strength and elastic modulus as those of native vessels. Whilst endothelial cell proliferation on unmodified tubes, as analysed by scanning electron microscopy, was found to be moderate, a simple process of dynamic fibrinogen adsorption was seen to enhance the endothelialisation of these tubular grafts. The high negative zeta potential values, high strength, robustness and structural reliability of the scaffolds represent them to be promising biomaterials for vascular graft applications.

Chondroitin sulfate coatings display low platelet but high endothelial cell adhesive properties favorable for vascular implants.

This study highlights the advantages of chondroitin sulfate (CS) as a sublayer combining selective low-fouling properties, low-platelet adhesion and pro-adhesive properties on endothelial cells, making CS promising for vascular graft applications. These properties were evaluated by comparing CS with well-known low-fouling coatings such as poly(ethylene glycol) (PEG) and carboxymethylated dextran (CMD), which were covalently grafted on primary amine-rich plasma polymerized (LP) films. Protein adsorption studies by quartz crystal microbalance with dissipation monitoring (QCM-D) and fluorescence measurements showed that CS is as effective as PEG in reducing fibrinogen adsorption (~90% reduction). CS also largely reduced adsorption of bovine serum albumin (BSA) as well as fetal bovine serum (FBS) but to a lower extent than PEG and CMD surfaces (72% vs 85% for BSA and 66% vs 89% for FBS). Whole blood perfusion assays indicated that, while LP surfaces were highly reactive with platelets, PEG, CMD, and CS grafted surfaces drastically decreased platelet adhesion and activation to levels significantly lower than polyethylene terephthalate (PET) surfaces. Finally, while human umbilical vein endothelial cell (HUVEC) adhesion and growth were found to be very limited on PEG and CMD, they were significantly increased on CS compared to that on bare PET and reached similar values as those for tissue culture polystyrene positive controls. Interestingly, HUVEC retention during perfusion with blood was found to be excellent on CS but poor on PET. Overall, our results suggest that the CS surface has the advantage of promoting HUVEC growth and resistance to flow-induced shear stress while preventing fibrinogen and platelet attachment. Such a nonthrombogenic but endothelial-cell adhesive surface is thus promising to limit vascular graft occlusion.

The application of a thermoresponsive chitosan/β‐GP gel to enhance cell repopulation of decellularized vascular scaffolds

Decellularized arterial scaffolds have achieved success in advancing towards clinical use for small diameter vascular graft applications. Issues remain with effectively cell seeding these scaffolds, which may result in slow remodeling in vivo and reduced patency rates. This study aims to efficiently bulk load decellularized arterial scaffolds with cells and/or growth factors to promote cell migration within the scaffold. A chitosan/β-glycerophosphate (β-GP) hydrogel was used as a delivery vehicle for scaffold repopulation with rat mesenchymal stem cells (rMSCs) and/or murine hepatocyte growth factor (HGF). The thermoresponsivity of the chitosan/β-GP HGF gel was determined by rheological analysis and the release of HGF by ELISA. The chitosan/β-GP gel was fully injectable with gelation at 30°C in ∼30 min and demonstrated sustained controlled HGF release up to 28 days. Encapsulation of rMSCs allowed for consistent efficient cell delivery within the scaffold. Directional migration toward the released HGF was demonstrated within the medial layer of the scaffold up to a distance of 3 mm. The gel/cell/growth factor combination demonstrates a means of accelerating scaffold repopulation, without costly excessive in vitro maturation times.

Engineering Interaction between Bone Marrow Derived Endothelial Cells and Electrospun Surfaces for Artificial Vascular Graft Applications

The aim of this investigation was to understand and engineer the interactions between endothelial cells and the electrospun (ES) polyvinylidene fluoride-co-hexafluoropropylene (PVDF-HFP) nanofiber surfaces and evaluate their potential for endothelialization. Elastomeric PVDF-HFP samples were electrospun to evaluate their potential use as small diameter artificial vascular graft scaffold (SDAVG) and compared with solvent cast (SC) PVDF-HFP films. We examined the consequences of fibrinogen adsorption onto the ES and SC samples for endothelialisation. Bone marrow derived endothelial cells (BMEC) of human origin were incubated with the test and control samples and their attachment, proliferation, and viability were examined. The nature of interaction of fibrinogen with SC and ES samples was investigated in detail using ELISA, XPS, and FTIR techniques. The pristine SC and ES PVDF-HFP samples displayed hydrophobic and ultrahydrophobic behavior and accordingly, exhibited minimal BMEC growth. Fibrinogen adsorbed SC samples did not significantly enhance endothelial cell binding or proliferation. In contrast, the fibrinogen adsorbed electrospun surfaces showed a clear ability to modulate endothelial cell behavior. This system also represents an ideal model system that enables us to understand the natural interaction between cells and their extracellular environment. The research reported shows potential of ES surfaces for artificial vascular graft applications.

Fluorescent imaging of endothelial cells in bioengineered blood vessels: the impact of crosslinking of the scaffold

Fluorescent imaging is a useful tool to monitor and evaluate bioengineered tissues and organs. However, autofluorescence emitted from the scaffold can be comparable or even overwhelm signals generated by fluorescently labelled cells and biomarkers. Using standard fluorescent microscopy techniques, a simple and easy-to-measure signal to noise ratio metric was developed, which can facilitate the selection of fluorescent biomarkers and the respective biomaterials for tissue engineering studies. Endothelial cells (MS1) expressing green-fluorescent protein and red fluorescent protein (mKate) were seeded on poly(epsilon-caprolactone)-collagen hybrid scaffolds that were prepared by crosslinking with glutaraldehyde, genipin and ethyl(dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide. All scaffolds had comparable mechanical properties, which could meet the requirements for vascular graft applications. ethyl(dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide crosslinked scaffolds had a high signal to noise ratio value because of its low autofluorescence in green and red channels. Genipin crosslinked scaffolds had a high signal to noise ratio only in the green channel, while glutaraldehyde crosslinked scaffolds had a low signal to noise ratio in both green and red channels. The signal to noise ratio was independent of the exposure time. The data show that although similar in their mechanical properties and ability to support cell growth, scaffolds crosslinked with different agents have significant differences in causing autofluorescence of the scaffolds. This result indicates that scaffold's preparation method may have a significant impact on direct imaging of fluorescently labelled cells on scaffolds used for tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.

Blood vessel-derived acellular matrix for vascular graft application.

To overcome the issues connected to the use of autologous vascular grafts and artificial materials for reconstruction of small diameter (<6 mm) blood vessels, this study aimed to develop acellular matrix- (AM-) based vascular grafts. Rat iliac arteries were decellularized by a detergent-enzymatic treatment, whereas endothelial cells (ECs) were obtained through enzymatic digestion of rat skin followed by immunomagnetic separation of CD31-positive cells. Sixteen female Lewis rats (8 weeks old) received only AM or previously in vitro reendothelialized AM as abdominal aorta interposition grafts (about 1 cm). The detergent-enzymatic treatment completely removed the cellular part of vessels and both MHC class I and class II antigens. One month after surgery, the luminal surface of implanted AMs was partially covered by ECs and several platelets adhered in the areas lacking cell coverage. Intimal hyperplasia, already detected after 1 month, increased at 3 months. On the contrary, all grafts composed by AM and ECs were completely covered at 1 month and their structure was similar to that of native vessels at 3 months. Taken together, our findings show that prostheses composed of AM preseeded with ECs could be a promising approach for the replacement of blood vessels.

Endothelial cell thrombogenicity is reduced by ATRP-mediated grafting of gelatin onto PCL surfaces.

Reducing the thrombogenicity of a tissue-engineered vascular graft prior to implantation is important for improving graft patency. As functionalization of synthetic materials with cell-adhesive proteins is routinely utilized as a means to promote endothelial cell (EC) growth, we conducted detailed investigation on the proliferation and thrombogenicity of ECs on such functionalized surfaces. We observed that polycaprolactone (PCL) surfaces functionalized with poly(glycidyl methacrylate) [(P(GMA)] brushes via atom transfer radical polymerization (ATRP) alone resulted in the enhancement of an activated EC profile characterized by low production of nitric oxide (NO), platelet activation and elevated expression levels of von Willebrand factor (vWF) and matrix metalloproteinase-2 (MMP-2). When gelatin was conjugated onto the PCL-g-P(GMA) surfaces, not only were EC proliferation and endothelial coverage significantly improved, but an anti-thrombogenic profile was also observed. We demonstrated that PCL can be successfully functionalized by a controllable surface-initiated polymerization method and importantly, the thrombogenic profile of the endothelial cells can be influenced by material surface chemistry (e.g. the presence of polymer graft chains). Our findings emphasize the importance of a careful consideration of materials for vascular graft applications, as well as differential endothelial cell physiology on surfaces with different material chemistry.

Plasma treatment for improving cell biocompatibility of a biodegradable polymer scaffold for vascular graft applications

Biodegradable synthetic scaffolds are being evaluated by many groups for the application of vascular tissue engineering. In addition to the choice of the material and the structure of the scaffold, tailoring the surface properties can have an important effect on promoting adequate tissue regeneration. The objective of this study was to evaluate the effect of an increased hydrophilicity of a polycaprolactone vascular graft by treatment with a cold air plasma. To this end, treated and untreated scaffolds were characterized, evaluated in vitro with smooth muscle cells, and implanted in vivo in the rat model for 3weeks, both in the subcutaneous location and as an aortic replacement. The plasma treatment significantly increased the hydrophilicity of the scaffold, with complete wetting after a treatment of 60sec, but did not change fiber morphology or mechanical properties. Smooth muscle cells cultured on plasma treated patches adopt a spread out morphology compared to a small, rounded morphology on untreated patches. Subcutaneous implantation revealed a low foreign body reaction for both types of scaffolds and a more extended and dense cellular infiltrate in the plasma treated scaffolds. In the vascular position, the plasma treatment induced a better cellularization of the graft wall, while it did not affect endothelialization rate or intimal hyperplasia. Plasma treatment is therefore an accessible tool to easily increase the biocompatibility of a scaffold and accelerate tissue regeneration without compromising mechanical strength, which are valuable advantages for vascular tissue engineering.

Differential support of cell adhesion and growth by copolymers of polyurethane with hyaluronic acid

Mechanical mismatch, along with inadequate hemocompatibility and endothelialization, contribute to the high failure rate of many synthetic vascular grafts. However, due to the dueling nature of these requirements (i.e., inhibiting platelet adhesion frequently means inhibiting endothelial cell (EC) adhesion), the creation of materials that simultaneously satisfy the mechanical and biological design criteria needed for small diameter vascular grafts has been an elusive goal. In this work, we demonstrate the ability of polyurethane (PU) containing hyaluronic acid (HA) in its backbone structure to reduce protein adsorption, platelet and bacterial adhesion, and fibroblast and macrophage proliferation while allowing the retention of both ECs and vascular-appropriate mechanics. Irrespective of HA molecular weight (MW), PU-HA materials selectively supported the growth of ECs relative to fibroblasts, reduced platelet adhesion, and performed comparably to negative controls with respect to bactericidal activity. The extent of EC growth on the PU-HA materials did differ with HA MW, with a lower HA MW yielding improved EC growth in both two-dimensional (2-D) films and 3-D electrospun fibrous scaffolds. These findings illustrate that HA incorporated into the backbone of a synthetic polymer structure can retain bioactivity, with subtle differences in HA MW significantly impacting the physical and biological properties of the biomaterial; in particular, PU modified with low-MW HA appears promising for vascular graft applications.

Potential of Nanocrystalline Cellulose–Fibrin Nanocomposites for Artificial Vascular Graft Applications

The potential of synthesizing new nanocomposites from nanocrystalline cellulose (NCC) and fibrin for small-diameter replacement vascular graft (SDRVG) application was demonstrated. Periodate oxidation of NCC can augment reactive carbonyl groups on NCC and facilitate its cross-linking with fibrin. NCC-fibrin nanocomposites were synthesized, composed of homogeneously dispersed oxidized NCC (ONCC) in a fibrin matrix, with fibrin providing elasticity and ONCC providing strength. The maximum strength and elongation of the nanocomposites were determined by Atomic Force Microscopy (AFM) and compared with a native blood vessel. The manipulation of degree of oxidation of NCC and the NCC-to-fibrin ratio resulted in the variation of strength and elongation of the nanocomposites, indicating that the nanocomposites can be tailored to conform to the diverse mechanical properties of native blood vessels. A mechanistic understanding of the molecular interactions of ONCC and fibrin was illustrated. This study established fundamental information to utilizing NCC for SDRVG applications.

Experience with PTFE ‘Ecoflon’ (Russia) vascular grafts in surgery of aortoiliac lesions

Introduction The development of aortoiliac lesions surgery was impossible without creation and application of the synthetic vascular grafts because the use of autological materials or endarterectomy is limited or impossible. Since 1952, the Vinyon synthetic vascular grafts have been used in aortoiliac surgery.(1) The

A rare anatomical variation of the Berrettini anastomosis and third common palmar digital branch of the median nerve

Variations in the origin and distribution of Berrettini anastomosis between the digital branches of the ulnar and median nerves exist and are well described in the literature. During regular dissections by medical students, we encountered a rare variation in the left hand of an approximately 50-year-old male cadaver. Berrettini anastomosis connecting the third common palmar digital branch of the median nerve with the fourth common palmar and proper palmar digital branches of the ulnar nerve presented a plexiform nature. This communicating branch and the third common palmar digital branch of the median nerve were perforated by the superficial palmar arch. Further, the superficial palmar arch was incomplete, and it was solely formed by the superficial branch of the ulnar artery. The unusual relationship of Berrettini anastomosis with the superficial palmar arch is very rare, and knowledge about such a variation is important when performing carpal tunnel release, flexor tendon surgery, and Dupuytren's fasciectomy and when dealing with arterial repairs and vascular graft applications in the hand.