USPIO-labeled textile materials for non-invasive MR imaging of tissue-engineered vascular grafts

Non-invasive imaging might assist in the clinical translation of tissue-engineered vascular grafts (TEVG). It can e.g. be used to facilitate the implantation of TEVG, to longitudinally monitor their localization and function, and to provide non-invasive and quantitative feedback on their remodeling and resorption. We here incorporated ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles into polyvinylidene fluoride (PVDF)-based textile fibers, and used them to prepare imageable tissue-engineered vascular grafts (iTEVG). The USPIO-labeled scaffold materials were molded with a mixture of fibrin, fibroblasts and smooth muscle cells, and then endothelialized in a bioreactor under physiological flow conditions. The resulting grafts could be sensitively detected using T1-, T2- and T2*-weighted MRI, both during bioreactor cultivation and upon surgical implantation into sheep, in which they were used as an arteriovenous shunt between the carotid artery and the jugular vein. In vivo, the iTEVG were shown to be biocompatible and functional. Post-mortem ex vivo analyses provided evidence for efficient endothelialization and for endogenous neo-vascularization within the biohybrid vessel wall. These findings show that labeling polymer-based textile materials with MR contrast agents is straightforward and safe, and they indicate that such theranostic tissue engineering approaches might be highly useful for improving the production, performance, personalization and translation of biohybrid vascular grafts.     

Tissue-engineered acellular small diameter long-bypass grafts with neointima-inducing activity

Researchers have attempted to develop efficient antithrombogenic surfaces, and yet small-caliber artificial vascular grafts are still unavailable. Here, we demonstrate the excellent patency of tissue-engineered small-caliber long-bypass grafts measuring 20–30 cm in length and having a 2-mm inner diameter. The inner surface of an acellular ostrich carotid artery was modified with a novel heterobifunctional peptide composed of a collagen-binding region and the integrin α4β1 ligand, REDV. Six grafts were transplanted in the femoral–femoral artery crossover bypass method. Animals were observed for 20 days and received no anticoagulant medication. No thrombogenesis was observed on the luminal surface and five cases were patent. In contrast, all unmodified grafts became occluded, and severe thrombosis was observed. The vascular grafts reported here are the first successful demonstrations of short-term patency at clinically applicable sizes.     

Development of sutureless vascular connecting system for easy implantation of small-caliber artificial grafts

A novel sutureless vascular connecting system, an assembly with a delivery rod, an introducing sheath, and a connecting device, was developed for easy implantation of small-caliber vascular grafts less than 2 mm in internal diameter. A microporous stainless tube (length 2 mm, external diameter 1.6 mm, wall thickness 65 µm, pore diameter 400 µm, pore-to-pore distance 500 µm) was designed to serve as a connecting device. The feasibility of the system was tested using two types of preliminary animal experiments. One animal model consisted of graft implantation into the rat abdominal aorta (1.5 mm in diameter). The connecting device was inserted into the proximal and distal ends of the aorta through the introducing sheath by pushing the delivery rod with the connecting device placed over it. Subsequently, the aortic segments were inserted into both ends of model grafts made of segmented polyurethane (1.8 mm in internal diameter) and were fixed with banding silk threads from the exterior. The procedure was completed within 20 min without requiring specialized microsurgery techniques. Blood leakage and obstruction did not occur. The second model consisted of an end-to-end anastomosis between rabbit common carotid arteries (2 mm in diameter), which was performed within several minutes of blood flow interruption. Scanning electron microscopy demonstrated that the luminal surface of the device was fully covered with endothelial cells (ECs) after 1 week as a result of transluminal ingrowth of native ECs through the micropores in the device. This endothelialization may prevent early thrombus-induced occlusion. This simple and “easy-to-learn” technique will promote the development of small-caliber arterial grafts, and furthermore, it may have potential for clinical application.     

Decellularized bovine reinforced vessels for small-diameter tissue-engineered vascular grafts

The aim of the present study was to investigate the influence of a decellularization protocol on the structure and the mechanical behavior of small-diameter (<6 mm) tibial calf arteries and veins. Calf vessels were decellularized by a detergent-enzymatic method (DEM), partially hydrolyzed with trypsin and subsequently cross-linked using poly(ethylene glycol) diglycidyl ether. Our results showed that i) the DEM can be considered a simple and valuable procedure for the preparation of complete acellular arteries and veins able to preserve a high degree of collagen and elastic fibers, and ii) poly(ethylene glycol) diglycidyl ether cross-linking treatment provides appropriate mechanical reinforcement of blood vessels. Histologically, the decellularized vessels were obtained employing the detergent-enzymatic procedure and their native extracellular matrix histoarchitecture and components remained well preserved. Moreover, the decellularization protocol can be considered an effective method to remove HLA class I antigen expression from small-diameter tibial calf arteries and veins. Cytocompatibility of decellularized cross-linked vessels was evaluated by endothelial and smooth muscle cell seeding on luminal and adventitial vessel surfaces, respectively.     

Usefulness of polyurethane for small-caliber vascular prostheses in comparison with autologous vein graft

 For long-term patency of small-caliber vascular prostheses, antithrombogenicity and microporous structure are very important. We have developed a new technique to give a microporous structure to a polyurethane vascular prosthesis that has favorable antithrombogenicity. A solution of tetrahydrofuran/dimethylformamide (1 : 1) containing 13 wt% of segmented polyurethane (PTMG + MDI) and calcium carbonate (mean particle size, 8 μm) was dipcoated on a glass mandrel 3 mm in diameter and placed into distilled water. After the glass mandrel was removed, the polyurethane tube was placed into hydrochloric acid, and a microporous polyurethane vascular prosthesis was produced. Prostheses made in this fashion, and autologous jugular vein grafts were implanted into the femoral artery and the carotid artery of mongrel dogs. Patency was recognized on the arteriogram and duplex scanning (ultrasonography), and the removed grafts were inspected macroscopically and microscopically. This prosthesis was similar in elasticity to a vein graft. Patency was defined 8 weeks after implantation, and this prosthesis showed less intimal hyperplasia than the autologous vein graft. The new polyurethane prosthesis might be useful for small-caliber vascular reconstruction. Received: December 18, 2000 / Accepted: January 28, 2002     

Clinical results of the HDS vascular access graft for hemodialysis

A total of 104 HDS vascular access grafts were implanted in patients with end-stage renal failure. Fortyseven grafts were cannulated within 7 days after implantation (early cannulation), and 57 grafts were cannulated after postoperative day 7 (late cannulation). Comparison of primary and secondary patency rates showed no significant difference between the early and late cannulation groups. The results indicate that early cannulation of the HDS graft is safe and does not adversely affect patency.     

Materials and surface modification for tissue engineered vascular scaffolds

Although vascular implantation has been used as an effective treatment for cardiovascular disease for many years, off-the-shelf and regenerable vascular scaffolds are still not available. Tissue engineers have tested various materials and methods of surface modification in the attempt to develop a scaffold that is more suitable for implantation. Extracellular matrix-based natural materials and biodegradable polymers, which are the focus of this review, are considered to be suitable materials for production of tissue-engineered vascular grafts. Various methods of surface modification that have been developed will also be introduced, their impacts will be summarized and assessed, and challenges for further research will briefly be discussed.     

Application of a vascular graft material (Solcograft-P) in experimental surgery

The implantation and post-implantation behaviour of a Solcograft-P vascular prosthesis in the aortic, aorto-iliac, carotid and vena caval positions in dogs was studied up to 100 d post-surgery in order to assess the suitability of this vascular material for use in man. Solcograft-P is prepared from the carotid arteries of calves by crosslinking the collagen stroma using adipyl dichloride. During the postoperative follow-up period of 3 month, 100% of the aortal grafts, 80% of the aorto-iliac bypasses, 60% of the vena caval grafts and 35% of the carotid implants remained patent. The biochemical properties of the Solcograft-P are better than those of Solcograft, its predecessor. The intimal lining was consistently smooth and homogeneous in grafts of biological origin, and no aneurysm was observed. Infection and early thrombosis occured no more frequently than with other grafts. The new Solcograft-P, crosslinked via ester and amide groups, seems to represent a real improvement over Solcograft. Our results suggest that Solcograft-P should prove valuable in various cases of reconstructive vascular surgery of the lower limb, especially when the autologous vena saphena magna is not available, and its mechanical properties may well prove suitable for both arterial and venous replacement.     

丝素蛋白在小口径人工血管中的应用

背景：国内外众多学者将丝素蛋白应用到小口径人工血管中,得到的人工血管拥有较好的抗血栓性及优秀的机械强度,动物实验也取得了满意的远期通畅率。目的：概述丝素蛋白的生物化学性质,目前在血管中常用的改性手段,以及国内外小口径人工血管中丝素蛋白的具体应用。方法：应用计算机检索中国知网、Medline数据库及ScienceDirect数据库1980年1月至2013年12月有关小口径人造血管血液相容性方面的文献,中文检索词为“人工血管,丝素蛋白”,英文检索词为“Vascular grafts,silk fibroin”。结果与结论：丝素蛋白拥有可控的机械性能,可通过醇化等处理获得不同的顺应性。同时,丝素蛋白拥有良好的生物相容性,分解产物已知、无毒,满足生物医药材料的安全需求。此外,丝素蛋白易于整合抗凝集团,效果明显可靠,且易于内皮细胞定植生长,而短期内能否形成稳定牢固的内皮层是保证血管假体拥有理想远期通畅率的关键,因此丝素蛋白是制备小口径人工血管的理想材料。但是由于制作工艺等问题,中国早期应用丝素蛋白制作的人工血管并未得到长远的发展。近年来,随着纳米材料、仿生技术、组织工程类血管、静电纺丝等技术的研究发展,以及众多学者为应对小口径血管中复杂血流动力学环境对多层血管的研究尝试,为丝素蛋白在人工血管中的应用开辟了新的空间领域。中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程全文链接：     

Histological maturation of vascular smooth muscle cells in in situ tissue-engineered vasculature

The goal of regenerative medicine is to achieve histological and functional recovery to the level of the original tissue. For this purpose, we have developed a biodegradable scaffold to create cell-free in-situ tissue-engineered vasculature (iTEV) with good long-term results. However, the regeneration process of vascular smooth muscle cells (VSMCs) over time has yet to be examined. To evaluate the regeneration ability of VSMCs, the inferior vena cava of experimental animals was replaced with iTEV, and tested at 1, 3, 6, 12, and 24 months (n = 6 each) after implantation. Six animals were enrolled to compare 24-month iTEV and native vasculature in single individual samples. There were no complications throughout the study. Immunohistology, protein expression analysis, and biochemical findings indicate that iTEV can gradually regenerate and develop into a mature vessel within 24 months using our biodegradable scaffold. These results provide a time course for the regeneration of VSMCs within the tissue-engineered vascular autograft constructed using a biodegradable scaffold.     

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.     

Characteristics and prognosis in patients with prosthetic vascular graft infection: a prospective observational cohort study

Prosthetic vascular graft infection (PVGI) is a devastating complication, with a mortality rate of up to 75%, which is especially caused by aortic graft infection. The purpose of this study was to evaluate factors associated with in-hospital mortality of patients with definite graft infection, and with long-term outcome. We reviewed medical records of 85 patients treated for PVGIs defined by positive bacterial culture of intraoperative specimens or blood samples, and/or clinical, biological and radiological signs of infection. In-hospital patient mortality was defined as any death occurring during the initial treatment of the graft infection. Cure was defined as the absence of evidence of relapsing infection during long-term follow-up (≥1 year). Eighty-five patients (54 aortic and 31 limb graft infections) treated by surgical debridement and removal of the infected prosthesis (n=41), surgical debridement without removal of prosthesis (n=34) or antimicrobial treatment without surgery (n=10) were studied. The only microbiological difference observed between patients with early (occurring within 4 months after surgery) vs. late PVGI and between those with aortic vs. limb PVGI was the incidence of PVGI caused by Staphylococcus aureus, which was greater in patients with limb PVGI. Overall cure was observed in 93.2% of 59 patients with a follow-up of a minimum of 1 year. Overall in-hospital mortality was 16.5% (n=14). Two variables were independently associated with mortality: age >70 years (OR 9.1, 95% CI 1.83-45.43, p 0.007) and aortic graft infection (OR 5.6, 95% CI 1.1-28.7, p 0.037).     

A compound arterial prosthesis: the importance of the sterilization procedure on the healing and stability of albuminated polyester grafts

In response to the demand for a vascular prosthesis which achieves reliable haemostasis without preclotting, a new compound albumin/polyester prosthesis has been developed. In order to optimize the sterilization procedures for this device, two series of implantations in the thoracic aorta of dogs were undertaken to compare the effects of ethylene oxide (EtO) and gamma-radiation. Preclotted polyester prostheses were implanted in two additional control series. Pathological analysis of the explanted grafts indicated that gamma-radiation is to be preferred over EtO because it results in faster rates of healing. While the albumin coating delayed the thrombotic response and fibrinolytic activity, the extent of healing of the radiation sterilized graft was equivalent to that achieved by preclotted polyester prostheses in the medium and long term. Measurements of the strength and dimensional changes of the graft demonstrated that, in addition to reducing the risks of acute thrombosis and postoperative haemorrhage, the albumin coating improves the dimensional stability of the knitted structure.     

Patient-derived endothelial progenitor cells improve vascular graft patency in a rodent model

Late outgrowth endothelial progenitor cells (EPCs) derived from the peripheral blood of patients with significant coronary artery disease were sodded into the lumens of small diameter expanded polytetrafluoroethylene (ePTFE) vascular grafts. Grafts (1 mm inner diameter) were denucleated and sodded either with native EPCs or with EPCs transfected with an adenoviral vector containing the gene for human thrombomodulin (EPC + AdTM). EPC + AdTM was shown to increase the in vitro rate of graft activated protein C (APC) production 4-fold over grafts sodded with untransfected EPCs (p < 0.05). Unsodded control and EPC-sodded and EPC + AdTM-sodded grafts were implanted bilaterally into the femoral arteries of athymic rats for 7 or 28 days. Unsodded control grafts, both with and without denucleation treatment, each exhibited 7 day patency rates of 25%. Unsodded grafts showed extensive thrombosis and were not tested for patency over 28 days. In contrast, grafts sodded with untransfected EPCs or EPC + AdTM both had 7 day patency rates of 88-89% and 28 day patency rates of 75-88%. Intimal hyperplasia was observed near both the proximal and distal anastomoses in all sodded graft conditions but did not appear to be the primary occlusive failure event. This in vivo study suggests autologous EPCs derived from the peripheral blood of patients with coronary artery disease may improve the performance of synthetic vascular grafts, although no differences were observed between untransfected EPCs and TM transfected EPCs.     

The decellularized vascular allograft as an experimental platform for developing a biocompatible small-diameter graft conduit in a rat surgical model

Purpose

The present study was aimed to assess the feasibility of using decellularized aortic allograft in a rat small animal surgical model for conducting small diameter vascular tissue engineering research.

Materials and Methods

Decellularized aortic allografts were infra-renally implanted in 12 Sprague-Dawley (SD) adult rats. The conduits were harvested at 2 (n = 6) and 8 weeks (n = 6), and assessed by hematoxylin and eosin (H&E), van Gieson, Masson Trichrome staining, and immunohistochemistry for von Willebrand factor, CD 31⁺, and actin.

Results

Consistent, predictable, and reproducible results were produced by means of a standardized surgical procedure. All animals survived without major complications. Inflammatory immune reaction was minimal, and there was no evidence of aneurysmal degeneration or rupture of the decellularized vascular implants. However, the aortic wall appeared thinner and the elastic fibers in the medial layer showed decreased undulation compared to the normal aorta. There was also minimal cellular repopulation of the vascular media. The remodeling appeared progressive from 2 to 8 weeks with increased intimal thickening and accumulation of both collagen and cells staining for actin. Although the endothelial like cells appeared largely confluent at 8 weeks, they were not as concentrated in appearance as in the normal aorta.

Conclusion

The results showed the present rat animal model using decellularized vascular allograft implants to be a potentially durable and effective experimental platform for conducting further research on small diameter vascular tissue engineering.     

旋转生物反应器培养对组织工程气管软骨力学强度的影响

目的研究旋转生物反应器培养对组织工程气管软骨力学强度的影响，探索适宜的组织工程气管软骨培养方法。 方法分离2周龄Lewis大鼠剑突软骨细胞传代培养，收集第3代软骨细胞种植到DegraPol管状支架上，静态培养7d，然后将软骨细胞-支架复合物分别置于旋转生物反应器内培养（生物反应器组）或继续静态培养培养3周（静态培养组）。取出软骨细胞-支架复合物，以噻唑蓝（MTT）法测定软骨细胞增殖活性，结果以吸光度（A）值表示（每组n=6）；以Zwick1445型材料试验机测定软骨细胞-支架复合物的最大应变值和应力值（每组n=4）；并制备扫描电镜标本，观察软骨细胞在DegraPol支架中培养后的超微结构变化。 结果不同条件下培养3周，生物反应器组和静态培养组A值分别0.17±0.05、0.12±0.01，最大应力值分别为（0.33±0.04）和（0.26±0.01）MPa，最大应变值分别为（3.53±0.91）和（1.71±0.13）mm/mm，2组间3项指标的差异均有统计学意义（均P〈0.05）。扫描电镜观察显示生物反应器组获得更好的软骨样结构和更多的细胞外基质。 结论旋转生物反应器能够提供更好的体外培养条件，有利于组织工程气管软骨的形成。     

组织工程化血管的构建及其功能评价

组织工程化血管的出现为治疗血管缺损提供了新的希望。本文综合国内外最新文献,就组织工程化血管的种子细胞来源、细胞外基质生物材料的选择以及用于培养工程化血管的生物发生器的设计等方面进行了综述。同时,讨论了不同培养条件对组织工程化血管发育和功能的影响,并提出了组织工程化血管的功能评价方法。最后,指出了目前尚需解决的问题和对未来研究的展望。     

电磁场与组织工程组织血管形成

组织工程的血管化问题是制约组织工程产品的关键问题之一.为了解决这一问题,一些研究者使用电磁技术作用于内皮细胞或动物体观察血管形成情况.就电磁场对组织工程组织血管形成的影响作一综述.     

组织工程化口腔黏膜研究进展

口腔黏膜或身体其他部位病变导致较大面积黏膜缺损,都需要有适合的移植材料来修复.组织工程方法可以提供解决这一问题的有效途径,而且通过组织工程方法构建的等效全层黏膜组织还可以成为评价口腔材料、黏膜用药以及口腔黏膜癌变的理想模型.探讨了黏膜组织工程的种子细胞来源以及脱细胞真皮基质、胶原类、纤维蛋白凝胶以及生物高分子电纺丝网状支架材料等黏膜组织工程基本问题,也对用于黏膜修复和黏膜模型的组织工程化口腔黏膜应用研究进行了阐述.     

血管内皮细胞促进组织工程骨血管化的研究进展

组织工程骨的血管化问题严重制约了大块组织工程骨的发展和运用.血管内皮细胞对组织工程骨血管化具有重要促进作用,根据血管化过程中血管内皮细胞的来源不同,将血管化过程分血管发生和血管生成两种形式.成骨细胞和血管内皮细胞共同培养时可以相互促进生长.目前对组织工程骨血管化进行了血管内皮细胞新的来源,细胞混合培养和体内试验等方面的探索.