Suture-reinforced electrospun polydioxanone-elastin small-diameter tubes for use in vascular tissue engineering: a feasibility study

This study characterizes the cross-linking of electrospun elastin and the mechanical properties of suture-reinforced 1.5mm internal diameter electrospun tubes composed of blended polydioxanone (PDO) and soluble elastin. Several tube configurations were tested to assess the effects of reinforcement on tube mechanical properties. Between the electrospun layers of each double-layered prosthetic, zero, one or two 6-0 sutures were wound, maintaining 1mm spacing with a pitch of 9 degrees . Single-layered tubes without suture were also examined. Samples were cross-linked and tested for compliance and burst strength. Compliance decreased significantly (p <0.05) and burst strength significantly increased (p <0.01) with reinforcement. Uncross-linked tubes were also tested to determine the effects of cross-linking. Results demonstrated that cross-linking significantly decreases burst strength (p <0.01), while decreases in compliance for cross-linked tubes were not significant. Cross-linked suture-reinforced PDO-elastin tubes had burst pressures more than 10 times greater than normal systolic pressures and exhibited a range of compliance values, including those matching native artery. These tubes display many characteristics of the "ideal" small-diameter graft, having mechanical properties that can be tailored to match those desired in vascular replacement applications.     

Mechanical property characterization of electrospun recombinant human tropoelastin for vascular graft biomaterials

The development of vascular grafts has focused on finding a biomaterial that is non-thrombogenic, minimizes intimal hyperplasia, matches the mechanical properties of native vessels and allows for regeneration of arterial tissue. In this study, the structural and mechanical properties and the vascular cell compatibility of electrospun recombinant human tropoelastin (rTE) were evaluated as a potential vascular graft support matrix. Disuccinimidyl suberate (DSS) was used to cross-link electrospun rTE fibers to produce a polymeric recombinant tropoelastin (prTE) matrix that is stable in aqueous environments. Tubular 1 cm diameter prTE samples were constructed for uniaxial tensile testing and 4 mm small-diameter prTE tubular scaffolds were produced for burst pressure and cell compatibility evaluations from 15 wt.% rTE solutions. Uniaxial tensile tests demonstrated an average ultimate tensile strength (UTS) of 0.36 ± 0.05 MPa and elastic moduli of 0.15 ± 0.04 and 0.91 ± 0.16 MPa, which were comparable to extracted native elastin. Burst pressures of 485 ± 25 mm Hg were obtained from 4 mm internal diameter scaffolds with 453 ± 74 μm average wall thickness. prTE supported endothelial cell growth with typical endothelial cell cobblestone morphology after 48 h in culture. Cross-linked electrospun rTE has promising properties for utilization as a vascular graft biomaterial with customizable dimensions, a compliant matrix and vascular cell compatibility.     

The use of air-flow impedance to control fiber deposition patterns during electrospinning

Electrospun non-woven structures have the potential to form bioresorbable vascular grafts that promote tissue regeneration in situ as they degrade and are replaced by autologous tissue. Current bioresorbable grafts lack appropriate regeneration potential since they do not have optimal architecture, and their fabrication must be altered by the manipulation of process parameters, especially enhancing porosity. We describe here an air-impedance process where the solid mandrel is replaced with a porous mandrel that has pressurized air exiting the pores to impede fiber deposition. The mandrel design, in terms of air-flow rate, pore size, and pore distribution, allows for control over fiber deposition and scaffold porosity, giving greater cell penetration without a detrimental loss of mechanical properties or structural integrity.     

组织工程血管支架材料的研究进展

组织工程血管支架是血管组织工程学中的一个重要组成部分.过去几十年时间里,组织工程血管支架材料由简单的天然材料发展到高分子可降解材料和生物材料的复合物,设计和加工的方式由单纯的手工发展到电镀旋压成型技术,取得了很大进步.目前组织工程血管支架材料的设计加工方法还不够成熟,性能还有待完善,应主要着眼于支架材料的机械性能和生物活性的完善方面的研究.     

Constructs of electrospun PLGA,compressed collagen and minced urothelium for minimally manipulated autologous bladder tissue expansion

Bladder regeneration based on minced bladder mucosa in vivo expansion is an alternative to in vitro culturing of urothelial cells. Here, we present the design of a hybrid, electrospun poly(lactic-co-glycolide) (PLGA) – plastically compressed (PC) collagen scaffold that could allow in vivo bladder mucosa expansion. Optimisation of electrospinning was performed in order to obtain increased pore sizes and porosity to consolidate the construct and to support neovascularisation and tissue ingrowth. Tensile tests showed an increase in average tensile strength from 0.6 MPa for PC collagen to 3.57 MPa for the hybrid construct. The optimised PLGA support scaffold was placed between two collagen gels, and the minced tissue was distributed either on top or both on top and inside the construct prior to PC; this was then cultured for up to four weeks. Morphology, histology and SEM demonstrated that the construct maintained its integrity throughout cell culture. Cells from minced tissue migrated, expanded and re-organised to a confluent cell layer on the top of the construct after two weeks and formed a multilayered urothelium after four weeks. Cell morphology and phenotype was typical for urothelial mucosa during tissue culture.     

Electrospun scaffolds for tissue engineering of vascular grafts

There is a growing demand for off-the-shelf tissue engineered vascular grafts (TEVGs) for the replacement or bypass of damaged arteries in various cardiovascular diseases. Scaffolds from the decellularized tissue skeletons to biopolymers and biodegradable synthetic polymers have been used for fabricating TEVGs. However, several issues have not yet been resolved, which include the inability to mimic the mechanical properties of native tissues, and the ability for long-term patency and growth required for in vivo function. Electrospinning is a popular technique for the production of scaffolds that has the potential to address these issues. However, its application to human TEVGs has not yet been achieved. This review provides an overview of tubular scaffolds that have been prepared by electrospinning with potential for TEVG applications.     

A comparison of the performance of mono- and bi-component electrospun conduits in a rat sciatic model

Synthetic nerve conduits represent a promising strategy to enhance functional recovery in peripheral nerve injury repair. However, the efficiency of synthetic nerve conduits is often compromised by the lack of molecular factors to create an enriched microenvironment for nerve regeneration. Here, we investigate the in vivo response of mono (MC) and bi-component (BC) fibrous conduits obtained by processing via electrospinning poly(ε-caprolactone) (PCL) and gelatin solutions. In vitro studies demonstrate that the inclusion of gelatin leads to uniform electrospun fiber size and positively influences the response of Dorsal Root Ganglia (DRGs) neurons as confirmed by the preferential extensions of neurites from DRG bodies. This behavior can be attributed to gelatin as a bioactive cue for the cultured DRG and to the reduced fibers size. However, in vivo studies in rat sciatic nerve defect model show an opposite response: MC conduits stimulate superior nerve regeneration than gelatin containing PCL conduits as confirmed by electrophysiology, muscle weight and histology. The G-ratio, 0.71 ± 0.07 for MC and 0.66 ± 0.05 for autograft, is close to 0.6, the value measured in healthy nerves. In contrast, BC implants elicited a strong host response and infiltrating tissue occluded the conduits preventing the formation of myelinated axons.     

Age-related changes in arterial wall mechanics and composition of NIA Fischer rats

Segments of carotid and tail arteries and descending thoracic aorta were obtained from the NIA colony of Fischer rats at ages 3, 12, 24 and 30 months. Measurements of pressure and diameter were made on intact cylindrical segments under conditions of active (147 mM K⁺ and pssive (Ca²⁺ -free and 2 mM EGTA) smooth muscle. These data were used to compute active and passive mechanics. Contiguous segments were used for the analysis of connective tissue, water and electrolyte contents. Passive stiffness of the carotid and tail arteries increased monotonically with increasing age. Collagen content in the aorta and tail artery generally increased with age, while elastin content decreased in the aorta and carotid artery. The ratio of collagen to elastin increased at all sites with age. Maximum values of active stress response (force development) increased from 3 to 12 months for the carotid artery, but decreased with age (at 24 and 30 months compared to 3 and 12 months) for the tail artery. Changes in relative cell content were such that active cellular force development was the same at all ages for the carotid artery but was smaller at 24 and 30 months compared to the younger animals for the tail artery. Decreased cellular force development by arterial smooth muscle is not an anatomically uniform finding in this animal model.     

Modulation of myelin formation by combined high affinity with extracellular matrix structure of electrospun silk fibroin nanoscaffolds

Remyelination is a major therapeutic goal in peripheral nerve regeneration, serving to restore function of demyelinated axons and provide neuroprotection. In order to apply myelin biogenesis strategies to peripheral nerve defects, the tissue engineered substitutes might be amenable to the promotion of this repair process. Electrospun nanofibers are considered as promising scaffolds for tissue engineering due to extracellular matrix mimicking factor and enhanced electrostatic interaction resulting in a controllable 3?D nanofibrous membrane. In order to explore the role of electrospun silk fibroin (SF) membrane in myelination, co-culture of dorsal root ganglion (DRG) neurons and Schwann cells (SCs) in vitro was established and observed. Scanning electron microscopy was used to observe DRG adhesion to the membranes, the electrospinning SF membrane is more favorable to the adhesion of DRG. The immunofluorescence staining of MAG and NF showed considerable amount of myelin were formed, and the myelin was tightly wrapped around the axons of the neurons, which was confirmed under the scanning electron microscope observation. Real-time quantitative PCR technique was used to determine the gene expression level of DRG neurons cultured at different time points. The results showed that the mRNA levels of N-cadherin, laminin, fibronectin were higher than those in the control group. Our results showed that the electrospun SF nanofibers can provide topographical and chemical cues that mimic (to a certain extent) the extracellular matrix.     

Characterisation of structural changes in the arterial elastic matrix by a new fractal feature: directional fractal curve

A new fractal feature, the Directional Fractal Curve (DFC), defined over an arc of 180° and composed of 90 fractal dimensions determined at intervals of arc of 2°, is developed to account for the anisotropic property of a fractal texture. The DFC algorithm is first applied to two images with different textural patterns one without directional preference and one with a well-organised texture. The DFC of these images shows different patterns. The technique is then applied to quantify the structure of the elastic texture in the arterial wall where the elastic network was imaged by scanning electron microscopy following selective tissue digestion. The results suggest: (i) that images of the elastin matrix of the arterial wall exhibit fractal properties with directional preference, (ii) the DFC gives quantitative parameters which allow characterisation of structural changes in the elastin matrix of the arterial wall in terms of disorganisation and fragmentation of elastin fibres—conditions which are associated with medial degeneration due to normal ageing or presence of arterial disease.     

Fabrication of collagen-coated biodegradable polymer nanofiber mesh and its potential for endothelial cells growth

Endothelialization of biomaterials is a promising way to prevent intimal hyperplasia of small-diameter vascular grafts. The aim of this study was to design a nanofiber mesh (NFM) that facilitates viability, attachment and phenotypic maintenance of human coronary artery endothelial cells (HCAECs). Collagen-coated poly(l-lactic acid)-co-poly(ε-caprolactone) P(LLA-CL 70:30) NFM with a porosity of 64–67% and a fiber diameter of 470±130 nm was fabricated using electrospinning followed by plasma treatment and collagen coating. The structure of the NFM was observed by SEM and TEM, and mechanical property was studied by tensile test. The presence of collagen on the P(LLA-CL) NFM surface was verified by X-ray photoelectron spectroscopy (XPS) and quantified by colorimetric method. Spatial distribution of the collagen in the NFM was visualized by labelling with fluorescent probe. The collagen-coated P(LLA-CL) NFM enhanced the spreading, viability and attachment of HCAECs, and moreover, preserve HCAEC's phenotype. The P(LLA-CL) NFM is a potential material for tissue engineered vascular graft.     

Regeneration of elastic fibers by three-dimensional culture on a collagen scaffold and the addition of latent TGF-β binding protein 4 to improve elastic matrix deposition

The objective of this study was to investigate the effects of latent TGF-β binding protein 4 (LTBP-4) on elastic fiber regeneration in three-dimensional cultures of human dermal fibroblasts (HDFs). Appropriate collagen scaffold for elastic fiber regeneration was also examined. Collagen sponges cross-linked at 120 °C and composed of small pores (25 μm on average) was favorable for elastic fiber regeneration by HDFs. Addition of LTBP-4, followed by culture for 21 days, accelerated elastic fiber accumulation within the scaffolds. Conditioned scaffolds containing either HDFs or LTBP-4-built mature elastic fibers were implanted between the dermis and the cutaneous muscle of mice. The combined use of HDFs and LTBP-4 resulted in thicker tissues containing elastic fibers. These results indicate that weakly cross-linked collagen sponges can be used as scaffolds for regenerating elastic fibers both in vitro and in vivo, and that the addition of LTBP-4 accelerates the deposition of both elastin and fibrillin-1, and increases cell proliferation. These techniques may be useful for generating cutaneous or cardiovascular tissue equivalents; furthermore, they may serve as a useful method for the three-dimensional analyses of drugs used to treat skin diseases or to examine the microstructure of elastin networks.     

Characterization of a cultured dermal substitute composed of a spongy matrix of hyaluronic acid and collagen combined with fibroblasts

 The authors have developed an allogeneic cultured dermal substitute (CDS) through cultivation of fibroblasts on a two-layered spongy matrix of hyaluronic acid (HA) and atelocollagen (Col). The Col spongy layer is essential for attachment and proliferation of fibroblasts on the two-layered spongy matrix. The HA spongy layer is necessary for maintaining the moisture environment on the wound surface. The optimal weight ratio of HA/Col is determined by considering the following characteristics: mechanical properties for handling, cell viability after thawing, potency of vascular endothelial growth factor (VEGF) release after thawing, efficacy of wound healing, and manufacturing cost. This study is designed to investigate the physical properties for handling, the growth behavior of fibroblasts on the spongy matrix, and the quantitative analysis of VEGF released from fibroblasts in the fresh or cryopreserved CDS. The results of this study suggest that a CDS composed of Col spongy matrix alone has the highest potency in regard to the release of VEGF. However, taking into account the manufacturing cost, coupled with the potency of VEGF release, a two-layered sponge of HA and Col with a weight ratio of 5/2 is very promising for commercial application. Received: October 4, 2002 / Accepted: March 20, 2003