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Tri-layered vascular grafts composed of polycaprolactone, elastin, collagen, and silk: Optimization of graft properties
Abstract
The purpose of this study was to create seamless, acellular, small diameter bioresorbable arterial grafts that attempt to mimic the extracellular matrix and mechanical properties of native artery using synthetic and natural polymers. Silk fibroin, collagen, elastin, and polycaprolactone (PCL) were electrospun to create a tri-layered structure for evaluation. Dynamic compliance testing of the electrospun grafts ranged from 0.4-2.5%/100 mmHg, where saphenous vein (1.5%/100 mmHg) falls within this range. Increasing PCL content caused a gradual decrease in medial layer compliance, while changes in PCL, elastin, and silk content in the adventitial layer had varying affects. Mathematical modeling was used to further characterize these results. Burst strength results ranged from 1614-3500 mmHg, where some exceeded the capacity of the pressure regulator. Four week degradation studies demonstrated no significant changes in compliance or burst strength, indicating that these grafts could withstand the initial physiological conditions without risk of degradation. Overall, we were able to manufacture a multi-layered graft that architecturally mimics the native vascular wall and mechanically matches the gold standard of vessel replacement, saphenous vein.
... To biomimetic the structure and function of native blood vessels, designing multi-layered vascular scaffolds is an effective way. Electrospinning has been widely used, because it is easier to blend or mix various materials and build non-delamination layers to develop multi-layered scaffolds [23][24][25][26][27][28][29][30]. McClure et al. fabricated a threelayered electrospun matrix to mimic native arterial architecture using polycaprolactone, elastin, and collagen [27]. ...
... The results indicated that the graft had sufficient tensile strength, dynamic compliance, suture retention, and burst strength by altering layer properties. Then, they further developed a tri-layered vascular graft composed of polycaprolactone, elastin, collagen, and silk [28]. The results revealed the optimization of graft properties and concluded that the multi-layered graft architecturally mimicked the native vascular wall and mechanically matched the gold standard of vessel replacement, saphenous vein [28]. ...
... Then, they further developed a tri-layered vascular graft composed of polycaprolactone, elastin, collagen, and silk [28]. The results revealed the optimization of graft properties and concluded that the multi-layered graft architecturally mimicked the native vascular wall and mechanically matched the gold standard of vessel replacement, saphenous vein [28]. Zhang and Han et al. designed multi-layered small-diameter vascular scaffolds which had dual-loading of vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF) [23,24]. ...
Designing a biomimetic and functional tissue-engineered vascular graft has been urgently needed for repairing and regenerating defected vascular tissues. Utilizing a multi-layered vascular scaffold is commonly considered an effective way, because multi-layered scaffolds can easily simulate the structure and function of natural blood vessels. Herein, we developed a novel tri-layer tubular graft consisted of Poly(L-lactide-co-caprolactone)/collagen (PLCL/COL) fibers and Poly(lactide-co-glycolide)/silk fibroin (PLGA/SF) yarns via a three-step electrospinning method. The tri-layer vascular graft consisted of PLCL/COL aligned fibers in inner layer, PLGA/SF yarns in middle layer, and PLCL/COL random fibers in outer layer. Each layer possessed tensile mechanical strength and elongation, and the entire tubular structure provided tensile and compressive supports. Furthermore, the human umbilical vein endothelial cells (HUVECs) and smooth muscle cells (SMCs) proliferated well on the materials. Fluorescence staining images demonstrated that the axially aligned PLCL/COL fibers prearranged endothelium morphology in lumen and the circumferential oriented PLGA/SF yarns regulated SMCs organization along the single yarns. The outside PLCL/COL random fibers performed as the fixed layer to hold the entire tubular structure. The in vivo results showed that the tri-layer vascular graft supported cell infiltration, scaffold biodegradation and abundant collagen production after subcutaneous implantation for 10 weeks, revealing the optimal biocompatibility and tissue regenerative capability of the tri-layer graft. Therefore, the specially designed tri-layer vascular graft will be beneficial to vascular reconstruction.
... Thus, McKenna et al. developed a tubular vascular scaffold from tropoelastin with mechanical properties similar to those of native vessels, although the mechanical strength of these scaffolds made them unsuitable for implantation (McKenna et al., 2012). To overcome this problem, tropoelastin-based scaffolds can be co-electrospun with other natural or synthetic polymers (Heydarkhan-Hagvall et al., 2008;McClure et al., 2009McClure et al., , 2012Zhang et al., 2010;Rnjak-Kovacina et al., 2012) to create new hybrid materials with improved properties (Lee et al., 2007;Zhang et al., 2007;Han et al., 2011). Such hybrid materials have mainly been used to develop vascular substitutes by electrospinning composite solutions onto a rotating mandrel that acts a cylindrical collector in order to obtain tubular scaffolds. ...
... A layer of smooth muscle cells was added between these two layers to create the third layer of the scaffold. More tri-layered tubular electrospun scaffolds with different compositions and structures have been explored, for instance a synthetic vascular graft comprising an intima layer made exclusively of electrospun tropoelastin and PCL and an adventitia layer made of collagen and a PCL, with the media comprising a mixture of tropoelastin collagen and PCL, was created by McClure et al. (2010McClure et al. ( , 2012. Similarly, Wise et al. developed a bilayer tubular scaffold based on the sequential deposition of electrospun tropoelastin fibers followed by a mixture of tropoelastin and PCL (80:20) onto a rotating mandrel then crosslinked with glutaraldehyde vapor . ...
Current cutting-edge strategies in biomaterials science are focused on mimicking the design of natural systems which, over millions of years, have evolved to exhibit extraordinary properties. Based on this premise, one of the most challenging tasks is to imitate the natural extracellular matrix (ECM), due to its ubiquitous character and its crucial role in tissue integrity. The anisotropic fibrillar architecture of the ECM has been reported to have a significant influence on cell behaviour and function. A new paradigm that pivots around the idea of incorporating biomechanical and biomolecular cues into the design of biomaterials and systems for biomedical applications has emerged in recent years. Indeed, current trends in materials science address the development of innovative biomaterials that include the dynamics, biochemistry and structural features of the native ECM. In this context, one of the most actively studied biomaterials for tissue engineering and regenerative medicine applications are nanofiber-based scaffolds. Herein we provide a broad overview of the current status, challenges, manufacturing methods and applications of nanofibers based on elastin-based materials. Starting from an introduction to elastin as an inspiring fibrous protein, as well as to the natural and synthetic elastin-based biomaterials employed to meet the challenge of developing ECM-mimicking nanofibrous-based scaffolds, this review will follow with a description of the leading strategies currently employed in nanofibrous systems production, which in the case of elastin-based materials are mainly focused on supramolecular self-assembly mechanisms and the use of advanced manufacturing technologies. Thus, we will explore the tendency of elastin-based materials to form intrinsic fibers, and the self-assembly mechanisms involved. We will describe the function and self-assembly mechanisms of silk-like motifs, antimicrobial peptides and leucine zippers when incorporated into the backbone of the elastin-based biomaterial. Advanced polymer-processing technologies, such as electrospinning and additive manufacturing, as well as their specific features, will be presented and reviewed for the specific case of elastin-based nanofiber manufacture. Finally, we will present our perspectives and outlook on the current challenges facing the development of nanofibrous ECM-mimicking scaffolds based on elastin and elastin-like biomaterials, as well as future trends in nanofabrication and applications.
... The studies showed that bilayer scaffolds could simulate the vessel structure with higher cell infiltration during in vivo implantation. In these cases, the scaffold's outer layer should provide a suitable environment for the growth of smooth muscle cells and fibroblasts with appropriate mechanical strength to resist aneurysm formation and blood leakage (McClure et al. 2012). The internal layer should also provide a proper condition for proliferation and ingrowth of endothelial cells with the lowest thrombogenicity . ...
... Native tissue was considered as control group. The proangiogenic actions of miR-126 correlated with its repression of SPRED-1, a negative regulator of MAP kinase signaling (McClure et al. 2012). In the absence of miR-126, the increased expression of SPRED-1 diminishes the transmission of intracellular angiogenic signals generated by VEGFs and FGFs the qualitative comparison between the bilayer scaffolds, PELCL scaffolds and native tissue showed that the fabricated bilayer scaffold loaded with miR-126 could enhance endothelialization in vivo ). ...
Purpose:
This systematic review focus on the application of bilayer scaffolds as an engaging structure for the engineering of multilayered tissues, including vascular and osteochondral tissues, skin, nerve, and urinary bladder. This article provides a concise literature review of different types of bilayer scaffolds to understand their efficacy in targeted tissue engineering.
Methods:
To this aim, electronic search in the English language was performed in PMC, NBCI, and PubMed from April 2008 to December 2019 based on the PRISMA guidelines. Animal studies, including the "bilayer scaffold" and at least one of the following items were examined: osteochondral tissue, bone, skin, neural tissue, urinary bladder, vascular system. The articles which didn't include "tissue engineering" and just in vitro studies were excluded.
Results:
Totally, 600 articles were evaluated; related articles were 145, and 35 full-text English articles met all the criteria. Fifteen articles in soft tissue engineering and twenty items in hard tissue engineering were the results of this exploration. Based on selected papers, it was revealed that the bilayer scaffolds were used in the regeneration of the multilayered tissues. The highest multilayered tissue regeneration has been achieved when bilayer scaffolds were used with mesenchymal stem cells and differentiation medium before implanting. Among the studies being reported in this review, bone marrow mesenchymal stem cells are the most studied mesenchymal stem cells. Among different kinds of multilayer tissue, the bilayer scaffold has been most used in osteochondral tissue engineering in which collagen and PLGA have been the most frequently used biomaterials. After osteochondral tissue engineering, bilayer scaffolds were widely used in skin tissue engineering.
Conclusion:
The current review aimed to manifest the researcher and surgeons to use a more sophisticated bilayer scaffold in combinations of appropriate stem cells, and different can improve multilayer tissue regeneration. This systematic review can pave a way to design a suitable bilayer scaffold for a specific target tissue and conjunction with proper stem cells.
... The predominant compositions are elastin with collagen, or sometimes with PCL. While crosslinking after fiber formation via EDC/NHS is a common strategy to confer adequate mechanical properties (71). The elastin-collagen mesh from electrospinning can be deposited in its formation on an implant tube to favor its grafting. ...
... The elastin-collagen mesh from electrospinning can be deposited in its formation on an implant tube to favor its grafting. Other scaffolding structures consist of double or triple layers of interlocked mesh (71). Even though electrospinning is a technique that is not recent, its peak is evident in literature today. ...
Elastin is one of the main components of the extracellular matrix; it provides resistance and elasticity to a variety of tissues and organs of the human body, besides participating in cellular signaling. On the other hand, elastin-derived peptides are synthetic biopolymers with a similar conformation and structure to elastin, but these possess the advantage of solubility in aqueous mediums. Due to their biological activities and physicochemical properties, elastin and related peptides may be applied as biomaterials to develop diverse biomedical devices, including scaffolds, hydrogels, and drug delivery systems for tissue engineering. Likewise, the combination of elastin with natural or synthetic polymers has demonstrated to improve the mechanical properties of biomedical products and drug delivery systems. Here we comprehensively describe the physicochemical properties and physiological functions of elastin. Moreover, we offer an overview of the use of elastin and its derivative polymers as biomaterials to develop scaffolds and hydrogels for tissue engineering. Finally, we discuss some perspectives on the employment of these biopolymers to fabricate new biomedical products.
... The composite tropoelastin solutions often display more favourable viscosity and electrospinning efficiency [67,68] . The electrospun elastin-based hybrid scaffolds also better mimic the architecture and components of native tissue, and display improved strength, structural integrity, elasticity, and wettability [26,63,66,68,74,[76][77][78] . Moreover, they show increased bioactivity including cell attachment [30,63,66,73,74] , proliferation [30,68,73,74] , maturation [63,68] , migration [67] , and infiltration [64,66,67,74,76] compared to biologically inert synthetic scaffolds. ...
... In other studies, tri-layer grafts have been constructed, in which the intima comprises only elastin and PCL, the media comprised elastin, collagen and PCL, and the adventitia comprised only collagen and PCL. These scaffolds are cross-linked with either 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) and 70% ethanol or genipin [64,77] . Other multi-layered constructs have likewise been formed by sequential electrospinning of elastin, gelatin and polyglyconate blends [60] , or elastin, gelatin, poliglecaprone (PGC) and PCL [73] . ...
Significance
Extracellular matrix proteins have primarily been designated as supporting scaffolds for cells. This work presents the soluble extracellular matrix component tropoelastin as a powerful proproliferative and cell-attractive molecule that surpasses the potency of conventional growth factors and matrix proteins used in a mesenchymal stem cell (MSC) culture. Tropoelastin is also demonstrated to modulate MSCs both as a substrate coating and as a soluble additive in media, which significantly deviates from the classical dogma of cell anchorage-dependent structural roles of the matrix. We show that these activities of tropoelastin can be harnessed and establish a path to boosting the efficacy of and simplifying processes for clinical MSC expansion and therapeutic MSC recruitment.
... The composite tropoelastin solutions often display more favourable viscosity and electrospinning efficiency [67,68] . The electrospun elastin-based hybrid scaffolds also better mimic the architecture and components of native tissue, and display improved strength, structural integrity, elasticity, and wettability [26,63,66,68,74,[76][77][78] . Moreover, they show increased bioactivity including cell attachment [30,63,66,73,74] , proliferation [30,68,73,74] , maturation [63,68] , migration [67] , and infiltration [64,66,67,74,76] compared to biologically inert synthetic scaffolds. ...
... In other studies, tri-layer grafts have been constructed, in which the intima comprises only elastin and PCL, the media comprised elastin, collagen and PCL, and the adventitia comprised only collagen and PCL. These scaffolds are cross-linked with either 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) and 70% ethanol or genipin [64,77] . Other multi-layered constructs have likewise been formed by sequential electrospinning of elastin, gelatin and polyglyconate blends [60] , or elastin, gelatin, poliglecaprone (PGC) and PCL [73] . ...
In this article, tables - contained incorrect citation numbers. The correct citation numbers in the tables are as follows. Mechanical properties and applications of elastin-only electrospun materials (Table presented.) GA: glutaraldehyde; HMDI: Hexamethylene diisocyanate; DSS: disuccinimidyl suberate. Mechanical properties and applications of elastin-only hydrogels (Table presented.) GA: glutaraldehyde; BS3: bis(sulfosuccinimidyl)suberate; MA: Methacrylic anhydride; EGDE: ethylene glycol diglycidyl ether; HMDI: Hexamethylene diisocyanate. Morphological and mechanical properties of composite elastin-containing fibrous scaffolds and their applications in tissue engineering (Table presented.) GA: glutaraldehyde; EDC: 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide; NHS: N-hydroxysuccinimide. Physical and mechanical properties and applications of composite elastin-containing hydrogels fabricated by freeze-drying, casting, or gas goaming (Table presented.) EDC/NHS: 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide/N-hydroxysuccinimide; GA: glutaraldehyde. Mechanical properties of ELP-based polymer networks compared to natural polymers and native tissues (Table presented.) ELP, elastin-like polypeptide; tTG, tissue transglutaminase; DSS; BS3, bis(sulfosuccinimidyl) suberate; TSAT, tris-succinimidyl aminotriacetate; THPP, β-[tris(hydroxymethyl)-phosphine]-propionic acid; HDMI, hexamethylene diisocynate; PQQ, pyrroloquinoline quinone. Composition, form, and potential applications of composite ELP-based materials (Table presented.) This does not affect the original reference list in the article.
... When the obtained compliance values were compared with the values of the native vessels, it was clear that compliance values of the samples of randomly distributed fibers were higher than those of the saphenous vein and coronary artery, with a compliance of 1.5%/100 mmHg and 0.0725%/mmHg, respectively [21,22]. Additionally, oriented samples also had higher compliance values than the coronary artery. ...
Vascular diseases are a major source of fatalities globally. However, the lack of accessibility of autologous vessels and the poor efficacy of commercial small-diameter vascular grafts limit surgical alternatives. Researchers therefore aimed to develop vascular prostheses that meet all requirements. Apart from the benefits of tissue-engineered grafts, significant obstacles that still hinder successful grafting include compliance mismatch, dilatation, thrombus development, and the absence of elastin. Among these issues, compliance mismatch between native vessel and artificial vascular scaffold has been mentioned in the literature as a possible cause of intimal hyperplasia, suture site rupture and endothelial and platelet cell damage. As a result, the usage of suitable materials and optimized fabrication techniques are required to achieve better control over the characteristics and functionality of the grafts. In particular, in the case of electrospun vascular grafts, the compliance can be adjusted throughout a broad range of values by adjusting the electrospinning parameters such as material selection, fiber orientation, porosity, and wall thickness. In this study, the electrospun vascular grafts consisting of pure PCL, PLA, and their blends were produced by using two different rotation speeds to achieve the oriented and non-oriented scaffolds. The impact of polymer type and fiber orientation on the compliance properties was evaluated. The results revealed that both material selection and fiber alignment have a significant effect on the compliance levels. PCL100_R grafts had the highest compliance value whereas the PCLPLA50_O scaffold had the lowest.
... Besides, BV did not influence the mechanical properties of electrospun PCL grafts (Fig. 2F-H). It is noteworthy to mention that all the burst pressures of the electrospun PCL grafts were higher than the clinical threshold (1600 mmHg) (Fig. 2F), which meet the requirements of vascular graft implantation [50]. ...
Synthetic artificial vascular grafts are hampered by their low patency rate in substituting small-caliber arteries (inner diameter <6 mm) because of their poor hemocompatibility. As compared to the heparin, bivalirudin (BV), a direct thrombin inhibitor, is a more reliable anticoagulant. In this study, BV-loaded polycaprolactone (PCL) grafts (PCL-BV) were fabricated by electrospinning. The loading rate of BV was approximately 87%. The PCL-BV grafts achieved the sustained release of BV for up to 30 days in vitro, but mainly released in the first 15 days. Scanning electron microscopy (SEM) and mechanical testing revealed that the BV loading did not affect the structure and mechanical properties of PCL grafts. In vitro coagulation analysis showed that the loading of the BV reduced thrombin activity, prolonged hemocompatibility parameters (activated partial thromboplastin time (APTT), prothrombin time (PT) and thromboplastin time (TT)), reduced blood coagulation, and inhibited the adhesion and activation of fibrinogen (FIB) and platelets. The assessment of vascular grafts in a rabbit arteriovenous (AV)-shunt ex vitro and carotid artery implantation in vivo indicated that BV significantly improved the antithrombogenicity and patency of PCL grafts. Overall, these results suggested that the incorporation of BV into electrospun grafts would be an effective approach to improve the hemocompatibility and patency rate of synthetic vascular grafts.
... Since both natural and synthetic biopolymers have numerous advantages but also noticeable barriers, the blend of natural and synthetic biopolymers for vascular applications is among the research topics of interest to researchers. McClure et al. (2012) designed electrospun trilayered vascular grafts using PCL, elastin (ELAS), collagen (COL), and silk (SF), also accomplished of their mechanical investigations. All the materials were dissolved in HFIP; moreover, ternary polymer systems were used at different rates. ...
... In particular, co-blended materials can be utilized in this multilayered electrospinning approach to provide even more complex combinations of material properties. The advantages of multilayered electrospun grafts were highlighted through demonstrating that co-blended combinations of PCL, elastin, collagen, and silk fibroin grafts possessed more idealistic mechanical properties for vascular applications compared to single material grafts (McClure et al. 2012). This approach also presents interesting opportunities for modularizing the tissueengineered vascular graft. ...
The clinical demand for a superior vascular graft is rising due to the increase in cardiovascular disease with an aging population. Despite decades of research, clinically translatable solutions remain limited. Recent progress in vascular graft engineering has highlighted the significance of biological integration for the success of implanted grafts. Thus there has been an increase in the usage of biological materials in vascular graft manufacture. Elastin, a natural protein that makes up a significant portion of the natural vascular extracellular matrix, has been demonstrated to be particularly important with both mechanical and biological modulatory roles. Progress in understanding elastogenesis, the process by which elastin is naturally synthesized, and increased access to synthetic elastin-based materials, has increased the usage of elastin in vascular graft engineering. In this chapter, we explore recent advances in the utilization of elastin as a material for vascular graft engineering. In particular, we focus on the myriad of methods which incorporate elastin into vascular grafts which demonstrate superior biological functionality and closer resemblance to native blood vessels.
... However, many of these materials have been abandoned because local reactions with tissues increase the risk of meningitis symptoms, excessive scar formation or hemorrhage [7][8][9]. Electrospinning is one of the most commonly used techniques for the fabrication of scaffolds because the final parameters (fiber dimensions, morphology and porosity) can be easily controlled, and the large specific surface areas required for tissue reconstruction can be obtained [10][11][12][13][14]. Scaffolds fabricated using electrospinning based on PLA fibers have excellent mechanical properties that are similar to natural tissues [15]. ...
An optimal medical scaffold should be biocompatible and biodegradable and should have adequate mechanical properties and scaffold architecture porosity, a precise three-dimensional shape, and a reasonable manufacturing method. Polylactic acid (PLA) is a natural biodegradable thermoplastic aliphatic polyester that can be fabricated into nanofiber structures through many techniques, and electrospinning is one of the most widely used methods. Medical fiber mat scaffolds have been associated with inflammation and infection and, in some cases, have resulted in tissue degradation. Therefore, surface modification with antimicrobial agents represents a suitable solution if the mechanical properties of the fiber mats are not affected. In this study, the surfaces of electrospun PLA fiber mats were modified with naturally occurring l-ascorbic acid (ASA) or fumaric acid (FA) via a plasma treatment method. It was found that 30 s of radio-frequency (RF) plasma treatment was effective enough for the wettability enhancement and hydroperoxide formation needed for subsequent grafting reactions with antimicrobial agents upon their decomposition. This modification led to changes in the surface properties of the PLA fiber mats, which were analyzed by various spectroscopic and microscopic techniques. FTIR-ATR confirmed the chemical composition changes after the modification process and the surface morphology/topography changes were proven by SEM and AFM. Moreover, nanomechanical changes of prepared PLA fiber mats were investigated by AFM using amplitude modulation-frequency modulation (AM-FM) technique. A significant enhancement in antimicrobial activity of such modified PLA fiber mats against gram-positive Staphylococcus aureus and gram-negative Escherichia coli are demonstrated herein.
... This is in accordance with previous studies showing that vascular grafts with lower compliance have higher burst pressures. 63,64 EtO grafts were generally stiffer than the untreated control as shown in multiple mechanical property tests such as compliance, Young's modulus, burst pressure, suture retention strength, and MBR. The EtO grafts were also visually more transparent than the other two groups. ...
This study investigated the effects of terminal sterilization of polyvinyl alcohol (PVA) biomaterials using clinically translatable techniques, specifically ethylene oxide and gamma (γ) irradiation. While a few studies have reported the possibility of sterilizing PVA with γ-radiation, the use of ethylene oxide (EtO) sterilization of PVA requires additional study. PVA solutions were chemically crosslinked with trisodium trimetaphosphate and sodium hydroxide. The three experimental groups included untreated control, EtO, and γ-irradiation, which were tested for the degree of swelling and water content, and mechanical properties such as radial compliance, longitudinal tensile, minimum bend radius, burst pressure, and suture retention strength. Additionally, samples were characterized with scanning electron microscopy, differential scanning calorimeter, X-ray photoelectron spectroscopy, and water contact angle measurements. Cell attachment was assessed using the endothelial cell line EA.hy926 and the sterilized PVA cytotoxicity was studied with a live/dead stain. Platelet and fibrin accumulation were measured using an ex vivo shunt baboon model. Lastly, the immune responses of PVA implants were analyzed after a 21-day subcutaneous implantation in rats and a 30-day implantation in baboon. EtO sterilization reduced the PVA graft wall thickness, its degree of swelling and water content compared to both γ-irradiated and untreated PVA. Moreover, EtO sterilization significantly reduced the radial compliance and increased Young's modulus. EtO did not change PVA hydrophilicity, while γ-irradiation increased the water contact angle of the PVA. Consequently, endothelial cell attachment on the EtO sterilized PVA showed similar results to the untreated PVA, while cell attachment significantly improved on the γ-irradiated PVA. When exposing the PVA grafts to circulating whole blood, fibrin accumulation of EtO sterilized PVA was found to be significantly lower than γ-irradiated PVA. The immune response of γ-irradiated PVA, EtO treated PVA, and untreated PVA were compared. Implanted EtO treated PVA showed the least MAC387 reaction. The terminal sterilization methods in this study changed PVA hydrogel properties; nevertheless, based on the characterizations performed, both sterilization methods were suitable for sterilizing PVA. We concluded that EtO can be used as an alternative method to sterilize PVA hydrogel material.
... These findings highlighted the importance of SF as a promising material to develop vascular grafts for smaller caliber blood vessels. Wu et al. (2018) and McClure et al. (2012) engineered three-layered tubular scaffolds using only electrospinning as the manufacturing technology. Combinations of natural (SF, collagen, elastin) and biodegradable synthetic polymers [polycaprolactone, poly(L-lactide-co-caprolactone), and poly(lactide-co-glycolide)], alone or as blends, were used to build layers with finely tuned morphological and mechanical properties. ...
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.
... Concurrently, VG modification has been attempted by loading endothelial growth factors [49,50] that would enhance cell adhesion [51][52][53], various drugs [54,55], and so on. Multilayered grafts have been proposed to improve their biomechanics [56,57]. For example, the earlier designed tissueengineered grafts of GL-loaded polycaprolactone and a poorly permeable inner layer have shown both good mechanical properties and biocompatibility [12,58]. ...
General physicochemical properties of the vascular grafts (VGs) produced of the solutions of Tecoflex (Tec) with gelatin (GL) and bivalirudin (BV) by electrospinning (ES) are studied. The electrospun VGs of Tec-GL-BV and expanded polytetrafluoroethylene (e-PTFE) implanted in the abdominal aorta of 36 Wistar rats have been observed over different time intervals up to 24 weeks. A comparison shows that 94.5% of the Tec-GL-BV VGs and only 66.6% of e-PTFE VGs (р = 0.0438) are free of occlusions after 6-month implantation. At the intermediate observation points, Tec-GL-BV VGs demonstrate severe neovascularization of the VG neoadventitial layer as compared with e-PTFE grafts. A histological examination demonstrates a small thickness of the neointima layer and a low level of calcification in Tec-GL-BV VGs as compared with the control grafts. Thus, the polyurethane-based protein-enriched VGs have certain advantages over the e-PTFE VGs, suggesting their utility in clinical studies.
... When such prostheses are used as a small-diameter replacement, an unfavorable healing process has been described in terms of thrombogenicity as a reason for the lack of endothelium together with anastomotic intimal hyperplasia caused by local changes in blood flow ( Sarkar et al., 2007;Chlupac et al., 2009). Therefore, novel biodegradable materials based on, for example, polyesters (de Valence et al., 2012a,b;Tara et al., 2014), collagen (Menasche et al., 1984), elastin (Wise et al., 2011;Koens et al., 2015) and silk fibroin ( Wang et al., 2010;McClure et al., 2012) are under development of ideal vascular grafts. These materials generally support the endothelialization of the inner surface of the prosthesis, ensuring a long-term nonthrombogenic surface, facilitating healing and enabling the regeneration of functional vessels. ...
Background:
Using animal models in experimental medicine requires mapping of their anatomical variability. Porcine common carotid arteries (CCA) are often preferred for the preclinical testing of vascular grafts due to their anatomical and physiological similarity to human small-diameter arteries. Comparing the microscopic structure of animal model organs to their human counterparts reveals the benefits and limitations of translational medicine.
Methods:
Using quantitative histology and stereology, we performed an extensive mapping of the regional proximodistal differences in the fractions of elastin, collagen, and smooth muscle actin as well as the intima-media and wall thicknesses among 404 segments (every 1 cm) of porcine CCAs collected from male and female pigs (n = 21). We also compared the microscopic structure of porcine CCAs with segments of human coronary arteries and one of the preferred arterial conduits used for the coronary artery bypass grafting (CABG), namely, the internal thoracic artery (ITA) (n = 21 human cadavers).
Results:
The results showed that the histological structure of left and right porcine CCA can be considered equivalent, provided that gross anatomical variations of the regular branching patterns are excluded. The proximal elastic carotid (51.2% elastin, 4.2% collagen, and 37.2% actin) transitioned to more muscular middle segments (23.5% elastin, 4.9% collagen, 54.3% actin) at the range of 2-3 centimeters and then to even more muscular distal segments (17.2% elastin, 4.9% collagen, 64.0% actin). The resulting morphometric data set shows the biological variability of the artery and is made available for biomechanical modeling and for performing a power analysis and calculating the minimum number of samples per group when planning further experiments with this widely used large animal model.
Conclusions:
Comparison of porcine carotids with human coronary arteries and ITA revealed the benefits and the limitations of using porcine CCAs as a valid model for testing bioengineered small-diameter CABG vascular conduits. Morphometry of human coronary arteries and ITA provided more realistic data for tailoring multilayered artificial vascular prostheses and the ranges of values within which the conduits should be tested in the future. Despite their limitations, porcine CCAs remain a widely used and well-characterized large animal model that is available for a variety of experiments in vascular surgery.
... In particular, co-blended materials can be utilized in this multilayered electrospinning approach to provide even more complex combinations of material properties. The advantages of multilayered electrospun grafts were highlighted through demonstrating that co-blended combinations of PCL, elastin, collagen, and silk fibroin grafts possessed more idealistic mechanical properties for vascular applications compared to single material grafts (McClure et al. 2012). This approach also presents interesting opportunities for modularizing the tissueengineered vascular graft. ...
... Solutions for electrospinning were prepared by separately dissolving collagen and elastin in acetic acid (20 mg/mL) [6,13], and PCL in a 90/10 (v/v) chloroform/methanol mixture (150 mg/mL). ...
Polymeric scaffolds prepared from polycaprolactone (PCL), PCL-collagen and PCL-elastin were prepared by electrospinning. The scaffolds were coated by plasma polymerization of pyrrole doped with iodine, to improve cellular adhesion and fibroblast proliferation. The morphology, composition, and crystalline structure of the scaffolds were characterized by scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy, small- and wide-angle X-ray scattering, thermogravimetric analysis and differential scanning calorimetry. The scaffold fibers had average diameters between 90 and 330 nm before coating. After coating by plasma polymerization, the average diameters increased to between 290 and 530 nm. The presence of elastin and collagen in the fibers was corroborated by infrared spectroscopy. X-ray scattering measurements show that neither elastin nor collagen form crystals in the fiber, while PCL maintains its crystallinity and is not affected by the plasma treatment. Fibroblast cell cultures in the presence of the scaffolds were characterized by viability assays and SEM. The results show that the scaffolds modified with plasma polymerized pyrrole provide a suitable environment for fibroblast adhesion, growth and viability.
... Hence, the development of multilayered vascular scaffolds was in progress. (Mcclure et al., 2012). They concluded that multilayered grafts that architecturally mimicked the native vascular wall and mechanically matched the gold standard of vessel replacement could be manufactured. ...
... Together with its exceptional elasticity and minimal immunogenic response [24], silk fibroin (SF) has been universally accepted biopolymer in tissue engineering domain [25,26]. These very intrinsic disparities of both PCL and silk fibroin have been utilized in numerous research field of bone [27][28][29], skin, wound healing [30][31][32], tendon tissue engineering [33], vascular [34][35][36], and a myriad of other tissue engineering applications [37][38][39]. ...
The vast domain of regenerative medicine comprises complex interactions between specific cells’ extracellular matrix (ECM) towards intracellular matrix formation, its secretion, and modulation of tissue as a whole. In this domain, engineering scaffold utilizing biomaterials along with cells towards formation of living tissues is of immense importance especially for bridging the existing gap of late; nanostructures are offering promising capability of mechano-biological response needed for tissue regeneration. Materials are selected for scaffold fabrication by considering both the mechanical integrity and bioactivity cues they offer. Herein, polycaprolactone (PCL) (biodegradable polyester) and ‘nature’s wonder’ biopolymer silk fibroin (SF) are explored in judicious combinations of emulsion electrospinning rather than conventional electrospinning of polymer blends. The water in oil (W/O) emulsions’ stability is found to be dependent upon the concentration of SF (aqueous phase) dispersed in the PCL solution (organic continuous phase). The spinnability of the emulsions is more dependent upon the viscosity of the solution, dominated by the molecular weight of PCL and its concentration than the conductivity. The nanofibers exhibited distinct core-shell structure with better cytocompatibility and cellular growth with the incorporation of the silk fibroin biopolymer.
... Thus, multi-layer designs are more advantageous than single-layer scaffolds in terms of vascular graft production in that they create a composite structure with differing functions and capabilities in separate layers, i.e. in the same way as in native arteries [1]. In addition to single-layer polycaprolactone (PCL) scaffolds [6][7][8], the literature contains references to bi-layer [9,10] and multi-layer scaffolds [11,12] with respect to the construction of an ideal vascular graft that resembles the native vessel. ...
Vascular grafts provide promising scaffolds for patients recuperating from cardiovascular diseases. Since it is necessary to mimic the native vessel in order to overcome the limitations of currently employed synthetic prostheses, researchers are tending to focus on the design of electrospun biodegradable multi-layer scaffolds which involves varying either the polymer type or constructional properties in each layer which, in turn, reveals the importance of layer interactions and their composite effect on the final multi-layer graft. This study describes the fabrication of biodegradable single-layer tubular scaffolds from polycaprolactone and poly(L-lactide)caprolactone polymers composed of either randomly distributed or, preferably, radially oriented fibers. Subsequently, bi-layer scaffolds were fabricated with a randomly distributed inner layer and a radially oriented outer layer from various polymer couple variations. The study focuses on vascular graft production technology including its morphology and mechanical properties. The post-morphologies of single-layer and bi-layer tubular scaffolds designed for vascular grafts were investigated as a continuation of a previously performed analysis of their mechanical properties. The results indicate that the mechanical properties of the final bi-layer grafts were principally influenced by the radially oriented outer layers acting as the
... However, the combination of biocompatibility and mechanical properties is difficult to achieve with a homogeneous construct. To overcome this issue, multilayered grafts composed of different synthetic polymers and natural biomaterials have been widely utilized 6,[16][17][18][19] . Although some success has been reported in resolving mechanical and biological concerns, appropriate microstructure is another critical point which needs to be addressed to improve the outcome of tissue regeneration 20 . ...
Autologous grafts, as the gold standard for vascular bypass procedures, associated with several problems that limit their usability, so tissue engineered vessels have been the subject of an increasing number of works. Nevertheless, gathering all of the desired characteristics of vascular scaffolds in the same construct has been a big challenge for scientists. Herein, a composite silk-based vascular scaffold (CSVS) was proposed to consider all the mechanical, structural and biological requirements of a small-diameter vascular scaffold. The scaffold’s lumen composed of braided silk fiber-reinforced silk fibroin (SF) sponge covalently heparinized (H-CSVS) using Hydroxy-Iron Complexes (HICs) as linkers. The highly porous SF external layer with pores above 60 μm was obtained by lyophilization. Silk fibers were fully embedded in scaffold’s wall with no delamination. The H-CSVS exhibited much higher burst pressure and suture retention strength than native vessels while comparable elastic modulus and compliance. H-CSVSs presented milder hemolysis in vitro and significant calcification resistance in subcutaneous implantation compared to non-heparinized ones. The in vitro antithrombogenic activity was sustained for over 12 weeks. The cytocompatibility was approved using endothelial cells (ECs) and vascular smooth muscle cells (SMCs) in vitro. Therefore, H-CSVS demonstrates a promising candidate for engineering of small-diameter vessels.
... Other material made of collagen/ chitosan/P(LLA-CL) served as vascular graft exhibited similar tensile strength at the ratios of 20:5:75 as the saphenous vein [32]. Some reports indicate that elastin and collagen-based materials had higher resilience than elastin/silk [33]. In addition to these instances, it was found that PCL/elastin fibers enhanced the attachment and growth of neurite cells. ...
Collagen and elastin are the two most abundant proteins in the human body, and as biomaterials offer fascinating properties to composite materials. More detailed investigations including these biomaterials within reinforced composites are still needed. This report describes physicochemical properties of fibers composed of collagen type I, collagen III, elastin and polycaprolactone (PCL). Prior to the electrospinning process, PCL was functionalized through covalent attachment of –NH2 groups by aminolysis reaction with hexamentilendiamine (HMDA). The fibers were fabricated by electrospinning technique set up with a non-conventional collector. A morphological comparative study was developed at different rations of collagen type I, observing in some cases two populations of fibers. The diameters and morphology were analyzed by scanning electros microscopy (SEM), observing a wide array of nanostructures with diameters of ~ 310 to 693 nm. Chemical characterization was assessed by FT-IR spectroscopy and the functionalized PCL was characterized through ninhydrin assay resulting in 0.36 mM NH2/mg fiber. Swelling tests were performed for 24 h, obtaining 320% for the majority of the fibers indicating morphological stability and good water uptake. In addition, contact angle analysis demonstrated adequate permeability and differences for each system depending mainly upon the type of biopolymer incorporated and the functionalization of PCL, ranging the values from 108° to 17°. Moreover, differential scanning calorimetry (DSC) results showed a melting temperature (Tm) of ~ 60 °C. The onset degradation temperatures (Td,onset) ranged between 115 and 148 °C, and were obtained by thermogravimetric analysis (TGA). The local mechanical properties of individual fibers were quantified by atomic force acoustic microscopy (AFAM). These results proposes that the physicochemical and mechanical properties of these scaffolds offer the possibility for enhanced biological activity Thus, they have a great potential as candidate scaffolds in tissue engineering applications.
... Based on muscular arteries collagen/elastin ratio [44] and the studied amount of each of these natural polymers on their layers [45,46], PLLA/PHD 50/50 blend ratio was selected to mimic native "media" layer in a previous work [17]. This corresponds to the inner layer and so the blood contacting surface of the obtained bilayered SDVG [17]. ...
The replacement of small-diameter vessels is one of the main challenges in tissue engineering. Moreover, the surface modification of small-diameter vascular grafts (SDVG) is a key factor in the success of the therapy due to their increased thrombogenicity and infection susceptibility caused by the lack of a functional endothelium. In this work, electrospun scaffolds were prepared from blends of poly(L-lactic acid) (PLLA) and segmented polyurethane (PHD) with a composition designed to perform as SDVG inner layer. The scaffolds were then successfully surface-modified with heparin following two different strategies that rely on grafting of heparin to either PLLA or PHD functional groups. Both strategies afforded high heparin density, being higher for urethane methodology. The functionalized scaffolds did not cause hemolysis and inhibited platelet adhesion to a large extent. However, lysozyme/heparin-functionalized scaffolds obtained through urethane methodology achieved the highest platelet attachment inhibition. The increase in hydrophilicity and water absorption of the surface-functionalized nanostructures favored adhesion and proliferation of human adipose-derived stem cells. Heparinized surfaces conjugated with lysozyme presented microbial hydrolysis activity dependent on heparin content. Overall, a better performance obtained for urethane-modified scaffold, added to the fact that no chain scission is involved in urethane methodology, makes the latter the best choice for surface modification of PLLA/PHD 50/50 electrospun scaffolds. Scaffolds functionalized by this route may perform as advanced components of SDVG suitable for vascular tissue engineering, exhibiting biomimetic behavior, avoiding thrombi formation and providing antimicrobial features.
... A three-layered polycaprolactone-soluble elastin-collagen electrospun matrix was made to further mimic the structure of the native artery, with an intimal layer (98-2-0 blend), medial layer (45-45-10; 55-35-10; 65-25-10 blends) and adventitial layer (70-0-30 blend) [252,253]. ...
Research into elastin-containing biomaterials has for many years been hampered by elastin׳s extreme insolubility and the short in vivo lifetime of its precursor tropoelastin, making the formulation of constructs from intact elastic fibers exceedingly difficult. The increased availability of soluble animal-derived elastin, obtained through alkaline (κ-elastin), acid (α-elastin), and enzymatic hydrolysis, and of elastin-based peptides, and in particular the development of the recombinant full-length elastin precursor tropoelastin now allow for enhanced and improved use of this class of protein in the development of advanced biomaterials.
... However, the combination of biocompatibility and mechanical properties is difficult to achieve with a homogeneous construct. To overcome this issue, multilayered grafts composed of different synthetic polymers and natural biomaterials have been widely utilized 6,[16][17][18][19] . Although some success has been reported in resolving mechanical and biological concerns, appropriate microstructure is another critical point which needs to be addressed to improve the outcome of tissue regeneration 20 . ...
One of main challenges in the development of small diameter vascular scaffolds is achieving a tubular conduit with appropriate mechanical properties that resist thrombosis. Silk-based vascular scaffolds have been prepared by various methods. It is showed that silk-based tubular scaffold can be constructed with appropriate mechanical properties and structure that support endothelial cells (ECs) expansion and proliferation. But in clinical use the luminal surface of the scaffold must have anticoagulant activity until confluent ECs coverage achieved. In this study we fabricated a heparin nanomodified silk-based scaffold via alternating linkage of heparin and Hydroxy Iron complexes via self-assembly. Heparin was firmly linked to and formed nanoscale coatings on the surface of the scaffold. Heparin nanomodification retained cytocompatibility of the scaffolds and extended coagulation time of healthy blood to several times higher regardless of washing duration. Results of this study demonstrate that heparin nanomodification of silk-based scaffold is a promising method to development of small diameter vascular scaffolds.
Silk is a globally renowned abundant biopolymer obtained from various sources of the Lepidoptera family, among which the most commonly used and researched are spider silk and silk worm silk. All varieties of silk have beneficial characteristics such as high tensile strength, biocompatibility, producing a reduced immune response in a biological system, biodegradability, and the ability to withstand environmental stresses as well. These features make silk suitable for a number of applications as a biomaterial. The vast potential of silk and its proteins in cosmetics, oncology, tissue engineering, TOC screenings, for preserving food, cosmetic product as a silk gel and bioremediation makes it a well-sought biopolymer among researchers. Experiments over the years have revealed that biomaterials constituting silk are very potent but are yet to be scaled up for commercial uses, but the various advantageous properties of silk biomaterial far overshadows the impeding problems of production.
The main objective of the present review article is to investigate the in-vitro and in-vivo biocompatibility behavior of hybrid PCL-based scaffolds with collagen, gelatin, and chitosan to improve endothelialization for VTE applications. It is reported that the high-rate failure of small diameters vascular grafts (SDVGs, <6 mm) due to adhesion of platelets and plasma protein and aggregation, over-proliferation of smooth muscle cells (SMCs), and neointimal hyperplasia at the implantation site is main challenge in vascular tissue engineering (VTE). The fast re-establishment of a functional endothelial cell (EC) layer representing a crucial importance strategy has been proposed to reduce these adverse outcomes. Polycaprolactone (PCL), with optimum mechanical properties, it showed great potential in biomedical applications, but its biocompatibility is still highly concerned in VTE. Modifications of the PCL vascular grafts by developing the hybrid structures using natural polymers with optimum hydrophilicity and biocompatibility properties in order to speed up the re-endothelialization process have been proposed over the last years. Analyzing the mentioned results in the present study can offer a better understanding of the benefits and challenges of applying the natural polymer as a thrombotic response upon implantation in clinical trials. Also, it can suggest the PCL-Collagen as the best framework for the fabrication of rationally designed SDVGs for VTE.
Native arteries contain a distinctive intima‐media comprised of organized elastin and an adventitia containing mature collagen fibrils. In contrast, implanted biodegradable small‐diameter vascular grafts do not present spatially regenerated, organized elastin. The elastin‐containing structures within the intima‐media region encompass the elastic lamellae (EL) and internal elastic lamina (IEL) and are crucial for normal arterial function. Here, we describe the development of a novel electrospun small‐diameter vascular graft that facilitates de novo formation of a structurally appropriate elastin‐containing intima‐media region following implantation. The graft comprise a non‐porous microstructure characterized by tropoelastin fibers that are embedded in a PGS matrix. After implantation in mouse abdominal aorta, the graft develops distinct cell and extracellular matrix profiles that approximate the native adventitia and intima‐media by 8 weeks. Within the newly formed intima‐media region there are circumferentially aligned smooth muscle cell layers that alternate with multiple EL similar to that found in the arterial wall. By 8 months, the developed adventitia region contains mature collagen fibrils and the neoartery presents a distinct IEL with thickness comparable to that in mouse abdominal aorta. We propose that this new class of material can generate the critically required, organized elastin needed for arterial regeneration. This article is protected by copyright. All rights reserved
Vascular tissue engineering is a rapidly growing field of regenerative medicine, which strives to find innovative solutions for vascular reconstruction. Considering the limited success of synthetic grafts, research impetus in the field is now shifted towards finding biologically active vascular substitutes bestowing in situ growth potential. In this regard, silk biomaterials have shown remarkable potential owing to their favorable inherent biological and mechanical properties. This review provides a comprehensive overview of the progressive development of silk-based small diameter (<6mm) tissue-engineered vascular grafts (TEVGs), emphasizing their pre-clinical implications. Herein, we first discuss the molecular structure of various mulberry and non-mulberry silkworm silk and identify their favorable properties at the onset of vascular regeneration. The emergence of various state-of-the-art fabrication methodologies for the advancement of silk TEVGs is rationally appraised in terms of their in vivo performance considering the following parameters: ease of handling, long-term patency, resistance to acute thrombosis, stenosis and aneurysm formation, immune reaction, neo-tissue formation, and overall remodeling. Finally, we provide an update on the pre-clinical status of silk-based TEVGs, followed by current challenges and future prospects.
Statement of Significance
: Limited availability of healthy autologous blood vessels to replace their diseased counterpart is concerning and demands other artificial substitutes. Currently available synthetic grafts are not suitable for small diameter blood vessels owing to frequent blockage. Tissue-engineered biological grafts tend to integrate well with the native tissue via remodeling and have lately witnessed remarkable success. Silk fibroin is a natural biomaterial, which has long been used as medical sutures. This review aims to identify several favorable properties of silk enabling vascular regeneration. Furthermore, various methodologies to fabricate tubular grafts are discussed and highlight their performance in animal models. An overview of our understanding to rationally improve the biological activity fostering the clinical success of silk-based grafts is finally discussed.
Electrospinning is an interesting technique widely used in industry to produce complex fibrous structures with tunable porosity and morphology. Yet, little is known about possible delamination between electrospun layers. In the present work, we studied the adhesion between PCL electrospun mats of similar porosity and investigated the effect of different morphologies, solvents and post-treatment on adhesive strength using a T-peel test. The objective is to establish guiding rules for improving the adhesive strength when fabricating multilayered electrospun systems. The effect of fiber diameter was found to be clearly dominant, large diameter fibers (3.6 µm) showing almost 7-fold increase (p<0.0001) of the adhesive strength compared to smaller ones (0.5 µm), while the use of different solvents for the two layers didn’t induce any major change. Heat treatment was also found to improve the adhesive strength. Further study is needed to better understand adhesion mechanisms between electrospun materials. Finally, this T-peel test was found to be an adequate simple tool to test electrospun bilayer materials and evaluate their adhesive strength.
The development of techniques for fabricating vascular wall models will foster the development of preventive and therapeutic therapies for treating cardiovascular diseases. However, the physical and biological complexity of vascular tissue represents a major challenge, especially for the design and the production of off-the-shelf biomimetic vascular replicas. Herein, we report the development of a biocasting technique that can be used to replicate the tunica adventitia and the external elastic lamina of the vascular wall. Type I collagen embedded with neonatal human dermal fibroblast (HDFn) and an elastic click cross-linkable, cell-adhesive and protease-sensitive elastin-like recombinamer (ELR) hydrogel were investigated as readily accessible and tunable layers to the envisaged model. Mechanical characterization confirmed that the viscous and elastic attributes predominated in the collagen and ELR layers, respectively. In vitro maturation confirmed that the collagen and ELR provided a favorable environment for the HDFn viability, while histology revealed the wavy and homogenous morphology of the ELR and collagen layer respectively, the cell polarization towards the cell-attachment sites encoded on the ELR, and the enhanced expression of glycosaminoglycan-rich extracellular matrix and differentiation of the embedded HDFn into myofibroblasts. As a complementary assay, 30% by weight of the collagen layer was substituted with the ELR. This model proved the possibility to tune the composition and confirm the versatile character of the technology developed, while revealing no significant differences with respect to the original construct. On-demand modification of the model dimensions, number and composition of the layers, as well as the type and density of the seeded cells, can be further envisioned, thus suggesting that this bi-layered model may be a promising platform for the fabrication of biomimetic vascular wall models.
The replacement of small-diameter arteries remains an unmet clinical need. Here we investigated the cellular remodeling of fibrotic conduits as vascular grafts. The formation of fibrotic conduit around subcutaneously implanted mandrels involved not only fibroblasts but also the trans-differentiation of inflammatory cells such as macrophages into fibroblastic cells, as shown by genetic lineage tracing. When fibrotic conduits were implanted as vascular grafts, the patency was low, and many fibrotic cells were found in neointima. Decellularization and anti-thrombogenic coating of fibrotic conduits produced highly patent autografts that remodeled into neoarteries, offering an effective approach to obtain autografts for clinical therapy. While autografts recruited mostly anti-inflammatory macrophages for constructive remodeling, allogenic DFCs had more T cells and pro-inflammatory macrophages and lower patency. Endothelial progenitors and endothelial migration were observed during endothelialization. Cell infiltration into DFCs was more efficient than decellularized arteries, and infiltrated cells remodeled the matrix and differentiated into smooth muscle cells (SMCs). This work provides insight into the remodeling of fibrotic conduits, autologous DFCs and allogenic DFCs, and will have broad impact on using fibrotic matrix for regenerative engineering.
Tubular polymer scaffolds based on tissue engineering techniques have been studied as potential alternatives for vascular regeneration implants. The blood vessels of the cardiovascular system are mainly fibrous, composed of collagen (Col) and elastin (El), and its inner layer consists of endothelial cells. In this work, Col and El were combined with polyurethane (PU), a biocompatible synthetic polymer, and Rotary Jet Spinning, a new and highly productive technique, to produce fibrous scaffolds. The scaffolds produced at 18,000 rpm presented homogeneous, bead‐free, and solvent‐free fibers. The blend formation between PU‐Col‐El was identified by chemical composition analysis and enhanced thermal stability up to 324 °C. The hydrophilic nature of the scaffold was revealed by its low contact angle. Cell viability of human umbilical vein endothelial cells (HUVEC) with the scaffold was proven for 72 h. The combined strategy of Rotary Jet Spinning with a polymer blend containing Col and El was verified as an effective and promising alternative to obtain tubular scaffolds for tissue engineering on a large scale production.
En réponse à la faible performance des prothèses synthétiques pour les revascularisations de petit diamètre (≤ 6 mm), de nombreux travaux de recherche ont porté sur le développement de substituts vasculaires biologiques. Bien que des résultats prometteurs aient été obtenus, la production de ces greffons repose principalement sur des méthodes d’ingénierie tissulaire coûteuses. L’objectif de cette thèse était donc de produire, à moindre coût, un substitut vasculaire biologique humain de petit diamètre. Pour cela, la membrane amniotique humaine (MAH) et une méthode d’assemblage sans cellule, basée sur le tissage, ont été associées. Afin de mieux connaître ce tissu, une étude détaillée des propriétés mécaniques a, tout d’abord, permis d’identifier la MAH placentaire comme une zone mécaniquement forte ne devant pas être négligée lors de la production de nos fils. La découpe de la MAH a ensuite permis d’obtenir des fils qui ont été caractérisés, puis assemblés par tissage manuel pour produire des substituts vasculaires. In vitro, ces greffons présentaient des propriétés mécaniques (pression à l’éclatement, rétention à la suture, perméabilité transmurale) compatibles avec une utilisation clinique justifiant le passage à uneimplantation en site artériel chez l’animal.
A great deal of research is focusing nowadays in the development of vascular substitutes to cover the unmet clinical need of small-caliber grafts. A fundamental understanding of the design principles that dictate the functionality of the native tissue and the transfer of such principles to engineer bioinspired vascular grafts represents an appealing approach. The recognition of the native vessels as textile-reinforced entities elevates textile technologies (such as knitting, weaving, electrospinning, melt electrospinning) as an attractive platform for vascular engineering. The combination of biomimetic textiles with cell-interactive matrices constitutes the main pillar of the biohybrid concept. In this chapter, we will review the current state of the art on textile-reinforced vascular grafts in the context of tissue engineering. We will also illustrate how the progress made in biomimicry, biofabrication, and material’s science disciplines constitutes a solid scenario for the development of advanced biohybrid vascular conduits.
Aim:
For the proper function of small diameter vascular grafts their mechanical properties are essential. A variety of testing methods and protocols exists to measure tensile strength, compliance and viscoelastic material behavior. In this study the impact of the measurement protocol in hoop tensile tests on the measured compliance and tensile strength was investigated.
Methods:
Vascular grafts made out of two different materials, a thermoplastic polyurethane (PUR) and polylactid acid (PLLA), with three different wall thicknesses were produced by electrospinning. Samples were tested with a measurement protocol that allowed the comparison of dynamic sample loading to a common quasistatic tensile test. Influence of measurement temperature, preconditioning cycles and the influence of a high number of loading cycles was also investigated. Compliance and tensile strength were evaluated and compared between the different samples and the different load cases.
Results:
In all samples a significant difference in the measured compliance was seen between an unloaded sample and a sample that was already in a preloaded state. For example in the PUR group with 100 μm wall thickness at 37 °C, the first compliance was 32.6 ± 9.6%/100 mmHg, which reduced to 15.4 ± 2.9%/100 mmHg at preloaded state. The PLLA group showed 7.5 ± 4.3%/100 mmHg vs. 0.94 ± 0.11%/100 mmHg respectively. The measurements showed the importance of dynamic testing, as the samples viscoelastic behavior had a considerable influence on the measured compliance. The quasistatic ultimate tensile test alone was not able to predict the sample's in vivo compliance. The measurement temperature had a significant influence on tensile strength and compliance. Both, the number of preconditioning cycles and the high number of loading cycles had a minor influence on the sample's compliance.
Conclusion:
With a quasistatic tensile tests alone, overestimated compliance values are measured in viscoelastic electrospun vascular samples, therefore dynamic loading cycles are required. Measurements at 37 °C are mandatory, as temperature has a significant influence on the mechanical properties.
We compared graft outcome between two types of a novel composite three-layer carp-collagen-coated vascular graft in low-flow conditions in a sheep model. Collagen in group A underwent more cycles of purification than in group B in order to increase the ratio between collagen and residual fat. The grafts were implanted end-to-side in both carotid arteries in sheep (14 grafts in 7 sheep in group A, 18 grafts in 9 sheep in group B) and artificially stenosed on the right side. The flow in the grafts in group A decreased from 297±118 ml/min to 158±159 ml/min (p=0.041) after placement of the artificial stenosis in group A, and from 330±164ml/min to 97±29 ml/min (p=0.0052) in group B (p=0.27 between the groups). From the five surviving animals in group A, both grafts occluded in one animal 3 and 14 days after implantation. In group B, from the six surviving animals, only one graft on the left side remained patent (p=0.0017). Histology showed degradation of the intimal layer in the center with endothelization from the periphery in group A and formation of thick fibrous intimal layer in group B. We conclude that the ratio between collagen and lipid content in the novel three-layer graft plays a critical role in its patency and structural changes in vivo.
Substantial progress in biomaterial research over the years has culminated in revolutionary technological advancements in the healthcare domain. This has triggered the quest for affordable healthcare solutions with focus on sustainable biomaterials with versatile applications endowed with green fabrication strategies. Silk as a biopolymer has garnered special attention which can largely be attributed to the excellent material properties of silk in addition to its affordability and resource ability. Silk fibroin from various silkworm and spider species and sericin from various silkworm species have been researched for their potential applications in the healthcare industry such as tissue-engineered grafts, cancer therapeutics, high-throughput tissue-on-chip models, food preservatives, biomedical imaging, biosensing, biomedical textiles, implants, cosmetics and bioremediation products. The present review mainly focusses on the various sources of silk fibroin and its relevant properties that have been conferred to it by nature. Moreover, recent developments, progress and prevalent modalities of healthcare industry that involve the application of silk fibroin and sericin have been outlined in the present review.
Impaired or damaged blood vessels can occur at all levels in the hierarchy of vascular systems from large vasculatures such as arteries and veins to meso‐ and microvasculatures such as arterioles, venules, and capillary networks. Vascular tissue engineering has become a promising approach for fabricating small‐diameter vascular grafts for occlusive arteries. Vascularized tissue engineering aims to fabricate meso‐ and microvasculatures for the prevascularization of engineered tissues and organs. The ideal small‐diameter vascular graft is biocompatible, bridgeable, and mechanically robust to maintain patency while promoting tissue remodeling. The desirable fabricated meso‐ and microvasculatures should rapidly integrate with the host blood vessels and allow nutrient and waste exchange throughout the construct after implantation. A number of techniques used, including engineering‐based and cell‐based approaches, to fabricate these synthetic vasculatures are herein explored, as well as the techniques developed to fabricate hierarchical structures that comprise multiple levels of vasculature. Impaired or damaged blood vessels can be repaired by tissue engineering approaches. Significant advances have been made in fabricating tissue‐engineered vascular grafts to replace autologous and commercially available grafts. However, persistent hurdles include the provision of sophisticated hierarchical vasculature. Herein, techniques used to fabricate tissue‐engineered vascular graft, mesovasculature, microvasculature, and hierarchical vasculatures are reviewed.
Cardiovascular diseases are a severe threat to human health. Implantation of small‐diameter vascular substitutes is a promising therapy in clinical operations. Polyurethane (PU) is considered one of the most suitable materials for this substitution due to its good mechanical properties, controlled biostability, and proper biocompatibility. According to biodegradability and biostability, in this review, PU small‐diameter vascular substitutes are divided into two groups: biodegradable scaffolds and biostable prostheses, which are applied to the body for short‐ and long‐term, respectively. Following this category, the degradation principles and mechanisms of different kinds of PUs are first discussed; then the chemical and physical methods for adjusting the properties and the research advances are summarized. On the basis of these discussions, the problems remaining at present are addressed, and the contour of future research and development of PU‐based small‐diameter vascular substitutes toward clinical applications is outlined. Polyurethane‐based substitutes for small‐diameter vessels are reviewed from a new perspective, that is, by dividing into biodegradable scaffolds and biostable prostheses according to biostability and biodegradability. The chemical structures, degradation mechanisms, methods for adjusting properties, and suitable fabrication techniques are summarized. Based on these discussions, the problems currently remaining and the contour of future research and development are outlined.
The design and analysis of a device to measure the burst strength (strength under a state of pure radial internal pressure) and compliance of vascular grafts and flexible pressurized tubes is presented. The device comprises three main sections, viz., a clean air-dry pressure controller, a test specimen holder, and automated software for control and data collection. Air pressure is controlled by means of a valve and a dedicated mechanism allowing reaching up to 120 psi in increments of 1 psi, and recording pressure changes with 0.04 psi resolution. The circumferential strain is determined by measuring the radial displacement of the vascular graft using an optical arrangement capable of determining a maximum radial displacement of 10 mm with 0.02 mm resolution. The instrument provides a low uncertainty in compliance (±0.32%/100 mm Hg⁻¹) and burst strength measurements. Due to its simplicity, the device can easily be reproduced in other laboratories contributing to a dedicated instrument with high resolution at low cost. The reliability of the apparatus is further confirmed by conducting finite element analysis, elasticity solutions for pressurized cylinders, and testing of small diameter vascular grafts made of a commercial aliphatic polyurethane tested under radial internal pressure.
Cardiovascular disease is one of the leading causes of death in the world. A characteristic symptom of cardiovascular disease is occlusion of vessels. Once vascular occlusion occurs there is a critical need to re-establish flow to prevent ischemia in the downstream tissues. In the most advanced cases, flow is re-established by creating a secondary flow path around the blockage, bypass grafting. For large diameter applications, synthetic conduits are successfully implanted, however in small diameter applications re-occlusion occurs and there is a critical need for new vascular grafts. There are many strategies and approaches that are being employed to design an effective and successful vascular graft. However, to date, there are no clinically available small diameter vascular grafts that are consistently successful in vivo long term (>7 years). As an effort to develop a successful graft there are several tissue engineering approaches: cell sheets, synthetic and natural biomaterial platforms, and decellularized extracellular matrices that are being investigated. While each area has its advantages, scaffold-based approaches are among the most widely studied. Scaffold based approaches are extensively studied due to tailorability and the availability of synthetic and natural polymers. Within the area of scaffold-based approaches, biomimicry has become an increasingly studied area, and structural biomimicry is one of the many approaches. The focus of this review paper is to analyze scaffold-based approaches. Particularly the advantages and disadvantages of using multi-layer scaffold-based approaches to engineer conduits for small diameter applications.
Despite major advances made in modeling vascular tissue biomechanics, the predictive power of constitutive models is still limited by uncertainty of the input data. Specifically, key measurements, like the geometry of the stress-free (SF) state, involve a definite, sometimes non-negligible, degree of uncertainty. Here, we introduce a new approach for sensitivity analysis of vascular hyperelastic constitutive models to uncertainty in SF measurements. We have considered two vascular hyperelastic models: the phenomenological Fung model, and the structure-motivated Holzapfel-Gasser-Ogden model. Our results indicate up to 160% errors in the identified constitutive parameters for a 5% measurement uncertainty in the SF data. Relative mar- gins of errors of up to 30% in the luminal pressure, 36% in the axial force, and over 200% in the stress predictions, were recorded for 10% uncertainties. These findings are relevant to the large body of studies involving experimentally based modeling and analysis of vascular tissues. The impact of uncertainties on calibrated constitutive parameters is significant in context of studies that use constitutive parameters to draw conclusions about the underlying microstructure of vascular tissues, their growth and remodeling processes, aging and disease states. The propagation of uncertainties into the predictions of biophysical parameters e.g. force, luminal pressure, and wall stresses, is of practical importance in the design and execution of clinical devices and interventions. Furthermore, insights provided by the present findings may lead to more robust parameters identification techniques, and serve as selection criteria in the trade-off between model complexity and sensitivity.
Studies have shown that traditional Chinese medicine Salvianolic acid B (SAB) can promote the proliferation and migration of endothelial cells. Heparin has a good anticoagulant effect. The inner layer of artificial vascular graft is fabricated by coaxial electrospinning method, and loaded with heparin and SAB. The release of heparin and SAB was sustained for almost 30 days, and SAB without initial burst release. Furthermore, the combined effect of SAB and heparin contributed to promoted Human Umbilical Vein Endothelial Cells (HUVECs) growth and improved blood compatibility of graft. In addition, up-regulation of GRP78 by SAB have protecting human endothelial cells from oxidative stress-induced cellular damage. In vivo evaluation through Masson's trichrome and H&E staining were performed after the graft embedded in SD rats' subcutaneously for two weeks, demonstrating that graft has good biocompatibility and without significant immune response. Hence, the functional inner layer is promising to prevent acute thrombosis and promote rapid endothelialization of artificial vascular graft.
Tissue engineering (TE) aims to restore function or replace damaged tissue through biological principles and engineering. Nanofibers are attractive substrates for tissue regeneration applications because they structurally mimic the native extracellular matrix. Composite nanofibers, which are hybrid nanofibers blended from natural and synthetic polymers, represent a major advancement in TE and regenerative medicine, since they take advantage of the physical properties of the synthetic polymer and the bioactivity of the natural polymer while minimizing the disadvantages of both. Although various nanofibrous matrices have been applied to almost all the areas of TE, in this chapter we will focus on nanofiber composites scaffolds for vascular TE.
Collagen and elastin represent the two most predominant proteins in the body and are responsible for modulating important biological and mechanical properties. Thus, the focus of this review is the use of collagen and elastin as biomaterials for the fabrication of living tissues. Considering the importance of both biomaterials, we first propose the notion that many tissues in the human body represent a reinforced composite of collagen and elastin. In the rest of the review, collagen and elastin biosynthesis and biophysics, as well as molecular sources and biomaterial fabrication methodologies, including casting, fiber spinning, and bioprinting, are discussed. Finally, we summarize the current attempts to fabricate a subset of living tissues and, based on biochemical and biomechanical considerations, suggest that future tissue-engineering efforts consider direct incorporation of collagen and elastin biomaterials.
Small diameter (< 6 mm) vascular grafts suffer from serious deleterious effects not encountered with their larger diameter relatives, leading to premature graft failure through restenosis. Platelet activation, inflammation, and smooth muscle cell proliferation are leading contributors to thrombosis and neointimal hyperplasia, both contributors to the progression of restenosis. It may be possible to suppress negative biological responses to vascular implants through the modification of surface properties and incorporation of drug release into blood contacting materials. In this work, bioengineering approaches are presented to improve the biocompatibility of small diameter vascular grafts.
We demonstrate a novel engineering approach for incorporating natural, decollagenized elastin matrices into PEU 1074A reinforced vascular grafts through spray-coating and electrospinning processes in a manner that retains elastin’s excellent blood contacting properties. A vascular construct with excellent mechanical and surgical handling properties demonstrating the suppression of neointimal hyperplasia is presented after 21 days in vivo.
Nitric oxide (NO) has been investigated over the past several decades due to its platelet, inflammation, and smooth muscle cell suppressing effects; and if appropriately delivered, could positively mediate the contributors to restenosis. Here, we characterize a novel macrocyclic NO donor developed by linking S-nitroso-N-acetyl-D-penicillamine (SNAP) directly to 1,4,8,11-tetraazacyclotetradecane (cyclam). Here, we present characterization data for SNAP-Cyclam and demonstrate stable, long term NO release at physiologically relevant levels for more than 90 days when incorporated into poly(˪-lactic acid) films.
Transition metals, such as copper and iron, are known to initiate NO production from S-nitrosothiols. It is reported in this work that additional transition metal ions; Co2+, Ni2+, and Zn2+, which have not been reported to generate NO from RSNOs have the capacity to generate NO from the S-nitrosothiol, S-nitroso-N-acetyl-D-penicillamine. In vivo data alludes to the possibility that Zn2+ may be able to generate NO from endogenous donors and provide beneficial effects.
These three novel developments form the basis for the potential construction of clinically relevant small diameter vascular grafts capable of suppressing the deleterious effects, namely thrombosis and neointimal hyperplasia, commonly encountered in current small diameter vascular grafts.
The development of vascular grafts requires the matching of material and viscoelastic properties to those of native artery. The hypothesis of this study was to subject electrospun tissue engineering scaffolds composed of polydioxanone, elastin, and collagen to cyclic loading in order to quantify the hysteretic properties, uniaxial tensile mechanical properties of conditioned scaffolds, and stress relaxation properties over a period of 400 cycles when compared to ePTFE, one of the most popular vascular prosthetic materials, and decellularized pig artery. In the electrospun graft, polydioxanone would provide a mechanical backbone, providing tensile support and preventing vessel rupture; while the elastin would provide elasticity and collagen would provide bioactivity (promote regeneration in vitro/in situ).
The purpose of this study was to establish whether material properties of elastin co-electrospun with polydioxanone (PDO) would change over time in both the uncross-linked state and the cross-linked state. First, uncross-linked scaffolds were placed in phosphate buffered saline (PBS) for three separate time periods: 15 minutes, 1 hour, and 24 hours, and subsequently tested using uniaxial materials testing. Several cross-linking reagents were then investigated to verify their ability to crosslink elastin: 1-ethyl-3-(dimethylaminopropyl)-carbodiimide (EDC), ethylene glycol diglycidyl ether (EGDE), and genipin. Uniaxial tensile testing was performed on scaffolds cross-linked with EDC and genipin, yielding results that warranted further investigation for PDO-elastin blends. Material properties of the cross-linked scaffolds were then found within range of both pig femoral artery and human femoral artery. These results demonstrate PDO-elastin blends could potentially be favorable as vascular grafts, thus warranting future in vitro and in vivo studies.
Vascular grafts are widely used for a number of medical treatments. Strength, compliance, endothelialization and availability are issues of most concern for vascular graft materials. With current approaches, these requirements are difficult to satisfy simultaneously. To explore an alternative approach, the present study has engineered the collagen gel construct by incorporating mimetic components and crosslinking the construct with different crosslinkers. The effects of component additives, such as chitosan and elastin, have been evaluated in terms of their mechanical and biological properties. Results demonstrate that the incorporation of chitosan and/or elastin alter stress-strain curves in the low stress loading region, and significantly improve the stretching ratio and ultimate stress of gel constructs compared to collagen constructs. Electron microscopy results suggest that the mechanical improvements might be due to microstructural modifications by chitosan sheets and elastin fibers. The effects of crosslinkers, such as formaldehyde, genipin and ethyl-(dimethyl aminopropyl) and carbodiimide hydrochloride (EDAC) have also been evaluated. Results demonstrate that formaldehyde, EDAC and genipin employ different mechanisms to crosslink collagen-based constructs, and use of genipin as a construct crosslinker exhibits improved elongation and endothelial coverage as compared to formaldehyde and EDAC. In addition, extending gelation time increased the elastic modulus but not the ultimate strength. Therefore, this study suggests that the mimicry of natural vessel tissues with properly crosslinked biopolymer composites could be a potential material design strategy for vascular graft materials.
The increasing age and longevity of the American population, coupled with the prevalence of athero- sclerotic cardiovascular disease (ASCVD),1 ensures that the treatment of ASCVD will remain a major public health priority. Although continued advances in endovascular technology hold promise for less in- vasive approaches to arterial occlusive diseases, the mainstay of therapy for patients with coronary and peripheral occlusions remains surgical bypass graft- ing. This is true because surgical bypass is less restric- tive in terms of the anatomic nature of lesions that are amenable to treatment and, in general, offers su- perior long-term patency, albeit at the cost of greater initial risk. Bypass grafts to small-caliber vessels in the coronary or lower extremity circulations are com- monly used procedures that offer important benefits to patients and incur significant economic cost on a national level. If one further considers the problem of long-term vascular access in the hemodialysis pop- ulation, the importance of a durable, versatile, small- caliber vascular graft assumes enormous proportions. The 'ideal' vascular graft would be characterized both by its mechanical attributes and postimplanta- tion healing responses. Mechanical strength is a par- amount issue; grafts placed in the arterial circulation must be capable of withstanding long-term hemody- namic stress without material failure, which might be catastrophic. Availability, suturability, and simplicity of handling are desirable for minimizing operating time, risk, and expense. The graft should be resistant to both thrombosis and infection and, optimally, would be completely incorporated by the body to yield a neovessel resembling a native artery in struc- ture and function. Given the economic considera- tions, low cost and long-term durability are also important issues. For large-caliber arterial reconstructions, currently available synthetic grafts offer a reasonable approxi- mation of these ideals and proven clinical efficacy. Long-term results of synthetic grafts for replacement of the thoracic and abdominal aorta, arch vessels, il- iac, and common femoral arteries for either aneurys- mal or occlusive disease are generally excellent when using any of a number of materials and manufactur- ing processes. While graft infection, occlusion, and dilatation are important clinical problems, the ma- jority of patients can expect durable patency and a low frequency of repeat procedures. Unfortunately, prosthetic grafts have generally proved unfavorable as small-caliber (õ6 mm) arterial substitutes. In these demanding, low-flow environments, the primary fac- tor influencing long-term patency is the type and quality of conduit used, with other patient variables (e.g., clinical indication, outflow resistance, site of distal anastomosis, comorbidities) serving as impor- tant modifiers. In the coronary circulation, the internal mammary artery (IMA) and radial artery have been used as by- pass conduits, and constitute the closest approxima- tion to this ideal graft yet defined. The long-term function of IMA grafts is excellent, making bypass of the left anterior descending artery with an IMA graft the single most durable intervention for coronary re-
Polymers such as Dacron and polytetrafluoroethylene (PTFE) have been used in high flow states with relative success but with limited application at lower flow states. Newer polymers with greater compliance, biomimicry, and ability to evolve into hybrid prostheses, suitable as smaller vessels, are now being introduced. In view of the advances in tissue engineering, this makes possible the creation of an ideal off-the-shelf bypass graft. We present a broad overview of the current state of prosthetic bypass grafts.
To analyze the effects of decellularization on the biomechanical properties of porcine common carotid arteries, decellularization was performed by a detergent-enzymatic procedure that preserves extracellular matrix scaffold. Internal diameter, external diameter, and wall thickness were measured by optical microscopy on neighboring histological sections before and after decellularization. Rupture tests were conducted. Inner diameter and wall thickness were measured by echo tracking during pressure inflation from 10 to 145 mmHg. Distensibility and incremental elastic modulus were computed. At 10 mmHg, mean diameter of decellularized arteries was 5.38 mm, substantially higher than controls (4.1 mm), whereas decellularized and control arteries reached the same internal diameter (6.7 mm) at 145 mmHg. Wall thickness decreased 16% for decellularized and 32% for normal arteries after pressure was increased from 10 to 145 mmHg. Decellularized arteries withstood pressure >2,200 mmHg before rupture. At 145 mmHg, decellularization reduced compliance by 66% and increased incremental elastic modulus by 54%. Removal of cellular elements from media led to changes in arterial dimensions. Collagen fibers engaged more rapidly during inflation, yielding a stiffer vessel. Distensibility was therefore significantly lower (by a factor of 3) in decellularized than in normal vessels: reduced in the physiological range of pressures. In conclusion, decellularization yields vessels that can withstand high inflation pressures with, however, markedly different geometrical and biomechanical properties. This may mean that the potential use of a decellularized artery as a scaffold for the creation of xenografts may be compromised because of geometrical and compliance mismatch.
At autopsy, 13 nonstenotic human left anterior descending coronary arteries [71.5 +/- 7.3 (mean +/- SD) yr old] were harvested, and related anamnesis was documented. Preconditioned prepared strips (n = 78) of segments from the midregion of the left anterior descending coronary artery from the individual layers in axial and circumferential directions were subjected to cyclic quasi-static uniaxial tension tests, and ultimate tensile stresses and stretches were documented. The ratio of outer diameter to total wall thickness was 0.189 +/- 0.014; ratios of adventitia, media, and intima thickness to total wall thickness were 0.4 +/- 0.03, 0.36 +/- 0.03, and 0.27 +/- 0.02, respectively; axial in situ stretch of 1.044 +/- 0.06 decreased with age. Stress-stretch responses for the individual tissues showed pronounced mechanical heterogeneity. The intima is the stiffest layer over the whole deformation domain, whereas the media in the longitudinal direction is the softest. All specimens exhibited small hysteresis and anisotropic and strong nonlinear behavior in both loading directions. The media and intima showed similar ultimate tensile stresses, which are on average three times smaller than ultimate tensile stresses in the adventitia (1,430 +/- 604 kPa circumferential and 1,300 +/- 692 kPa longitudinal). The ultimate tensile stretches are similar for all tissue layers. A recently proposed constitutive model was extended and used to represent the deformation behavior for each tissue type over the entire loading range. The study showed the need to model nonstenotic human coronary arteries with nonatherosclerotic intimal thickening as a composite structure composed of three solid mechanically relevant layers with different mechanical properties. The intima showed significant thickness, load-bearing capacity, and mechanical strength compared with the media and adventitia.
Scaffolds manufactured from biological materials promise better clinical functionality, providing that characteristic features are preserved. Collagen, a prominent biopolymer, is used extensively for tissue engineering applications, because its signature biological and physico-chemical properties are retained in in vitro preparations. We show here for the first time that the very properties that have established collagen as the leading natural biomaterial are lost when it is electro-spun into nano-fibres out of fluoroalcohols such as 1,1,1,3,3,3-hexafluoro-2-propanol or 2,2,2-trifluoroethanol. We further identify the use of fluoroalcohols as the major culprit in the process. The resultant nano-scaffolds lack the unique ultra-structural axial periodicity that confirms quarter-staggered supramolecular assemblies and the capacity to generate second harmonic signals, representing the typical crystalline triple-helical structure. They were also characterised by low denaturation temperatures, similar to those obtained from gelatin preparations (p>0.05). Likewise, circular dichroism spectra revealed extensive denaturation of the electro-spun collagen. Using pepsin digestion in combination with quantitative SDS-PAGE, we corroborate great losses of up to 99% of triple-helical collagen. In conclusion, electro-spinning of collagen out of fluoroalcohols effectively denatures this biopolymer, and thus appears to defeat its purpose, namely to create biomimetic scaffolds emulating the collagen structure and function of the extracellular matrix.
An electrospun cardiovascular graft composed of polydioxanone (PDO) and elastin has been designed and fabricated with mechanical properties to more closely match those of native arterial tissue, while remaining conducive to tissue regeneration. PDO was chosen to provide mechanical integrity to the prosthetic, while elastin provides elasticity and bioactivity (to promote regeneration in vitro/in situ). It is the elastic nature of elastin that dominates the low-strain mechanical response of the vessel to blood flow and prevents pulsatile energy from being dissipated as heat. Uniaxial tensile and suture retention tests were performed on the electrospun grafts to demonstrate the similarities of the mechanical properties between the grafts and native vessel. Dynamic compliance measurements produced values that ranged from 1.2 to 5.6%/100 mmHg for a set of three different mean arterial pressures. Results showed the 50:50 ratio to closely mimic the compliance of native femoral artery, while grafts that contained less elastin exceeded the suture retention strength of native vessel. Preliminary cell culture studies showed the elastin-containing grafts to be bioactive as cells migrated through their full thickness within 7 days, but failed to migrate into pure PDO scaffolds. Electrospinning of the PDO and elastin-blended composite into a conduit for use as a small diameter vascular graft has extreme potential and warrants further investigation as it thus far compares favorably to native vessel.
Significant challenges must be overcome before the true benefit and economic impact of vascular tissue engineering can be fully realized. Toward that end, we have pioneered the electrospinning of micro- and nano-fibrous scaffoldings from the natural polymers collagen and elastin and applied these to development of biomimicking vascular tissue engineered constructs. The vascular wall composition and structure is highly intricate and imparts unique biomechanical properties that challenge the development of a living tissue engineered vascular replacement that can withstand the high pressure and pulsatile environment of the bloodstream. The potential of the novel scaffold presented here for the development of a viable vascular prosthetic meets these stringent requirements in that it can replicate the complex architecture of the blood vessel wall. This replication potential creates an "ideal" environment for subsequent in vitro development of a vascular replacement. The research presented herein provides preliminary data toward the development of electrospun collagen and elastin tissue engineering scaffolds for the development of a three layer vascular construct.
Stress distribution through the wall thickness of the canine carotid artery was analyzed on the basis of the uniform strain hypothesis in which the wall circumferential strain was assumed to be constant over the wall cross-section under physiological loading condition. A newly proposed logarithmic type of strain energy density function was used to describe the wall properties. In contrast with other studies, this hypothesis gave almost uniform distribution of wall stresses under the physiological condition and non-zero residual stresses when all external forces were removed.
The goals of this paper are (i) to re-examine the constitutive law for the description of the (passive) highly nonlinear and anisotropic response of healthy elastic arteries introduced recently by the authors, (ii) to show how the mechanical response of a carotid artery under inflation and extension predicted by the structural model compares with that for a three-dimensional form of Fung-type strain-energy function, (iii) to provide a new set of material parameters that can be used in a finite element program, and (iv) to show that the model has certain mathematical features that are important from the point of view of material and numerical stability.
1. Abstract2. Introduction3. Methods3.1. Electrospinning3.2. Scaffold Characterization3.3. Cell Lines and Cell Culture3.4. Scaffolding Crosslinking, Disinfection, Seeding, and Culture3.5. Preliminary Fabrication of a Three Layered Vascular Construct3.6. Histology4. Results4.1. Electrospinning Collagen4.2. Electrospinning Elastin4.3. Electrospinning Collagen/Elastin Blend4.4. Preliminary Fabrication of a Three Layered Vascular Construct5. Discussion6. Acknowledgements7. References
Elastin-mimetic peptide polymers have been synthesized, and the morphological properties of fabricated small diameter fibers and nonwoven fabrics have been characterized. An 81 kDa recombinant protein based upon the repeating elastomeric peptide sequence of elastin (Val-Pro-Gly-Val-Gly) 4(Val-Pro-Gly-Lys-Gly) was obtained through bacterial expression of an oligomerized gene coding for tandem repeats of the monomer. The protein was processed into fibers by an electrospinning technique and morphology defined by SEM and TEM. The choice of processing parameters influenced both fiber diameter and morphology with diameters varying between 200 and 3000 nm and three morphological patterns noted: beaded fibers, thin filaments, and broad ribbonlike structures. Detailed image analysis of nonwoven textile fabrics produced from elastin-mimetic fibers revealed that the distribution of single fiber orientation was isotropic with an associated unimodal distribution of protein fiber diameter. In a dry state, the ultimate tensile strength of nonwoven fabrics generated from elastin-mimetic peptides was 35 MPa with a material modulus of 1.8 GPa.
The formation of fatty plaques (atherosclerosis) that clog arteries is one of the most common forms of cardiovas-cular disease.
The only permanent solu-tion is surgical replacement of diseased arteries with healthy autologous veins (that is, veins from
the same patient). However, this type of surgery is only possible for patients with healthy veins. Thus, there has been increasing
interest in using biological components to con-struct artificial blood vessels in the laboratory that faithfully mimic the
properties of normal healthy arteries. In a lively TechView article, Laura Niklason discusses several new approaches to growing
arteries in the laboratory and how each type of vessel withstands the rigors of arterial blood pressure when grafted into
different animal models.
Controlling the degradation process of silk is an important and interesting subject in the field of biomaterials. In the present study, silk fibroin films with different secondary conformations and nanostructures were used to study degradation behavior in buffered protease XIV solution. Different from previous studies, silk fibroin films with highest β-sheet content achieved the highest degradation rate in our research. A new degradation mechanism revealed that degradation behavior of silk fibroin was related to not only crystal content but also hydrophilic interaction and then crystal-noncrystal alternate nanostructures. First, hydrophilic blocks of silk fibroin were degraded. Then, hydrophobic crystal blocks that were formerly surrounded and immobilized by hydrophilic blocks became free particles and moved into solution. Therefore, on the basis of the mechanism, which enables the process to be more controllable and flexible, controlling the degradation behavior of silk fibroin without affecting other performances such as its mechanical or hydrophilic properties becomes feasible, and this would greatly expand the applications of silk as a biomedical material.
Throughout native artery, collagen, and elastin play an important role, providing a mechanical backbone, preventing vessel rupture, and promoting recovery under pulsatile deformations. The goal of this study was to mimic the structure of native artery by fabricating a multi-layered electrospun conduit composed of poly(caprolactone) (PCL) with the addition of elastin and collagen with blends of 45-45-10, 55-35-10, and 65-25-10 PCL-ELAS-COL to demonstrate mechanical properties indicative of native arterial tissue, while remaining conducive to tissue regeneration. Whole grafts and individual layers were analyzed using uniaxial tensile testing, dynamic compliance, suture retention, and burst strength. Compliance results revealed that changes to the middle/medial layer changed overall graft behavior with whole graft compliance values ranging from 0.8 to 2.8%/100 mm Hg, while uniaxial results demonstrated an average modulus range of 2.0-11.8 MPa. Both modulus and compliance data displayed values within the range of native artery. Mathematical modeling was implemented to show how changes in layer stiffness affect the overall circumferential wall stress, and as a design aid to achieve the best mechanical combination of materials. Overall, the results indicated that a graft can be designed to mimic a tri-layered structure by altering layer properties.
Long-term patency of conventional synthetic grafts is unsatisfactory below a 6-mm internal diameter. Poly(epsilon-caprolactone) (PCL) is a promising biodegradable polymer with a longer degradation time. We aimed to evaluate in vivo healing and degradation characteristics of small-diameter vascular grafts made of PCL nanofibers compared with expanded polytetrafluoroethylene (ePTFE) grafts.
We prepared 2-mm-internal diameter grafts by electrospinning using PCL (M(n)=80, 000 g/mol). Either PCL (n=15) or ePTFE (n=15) grafts were implanted into 30 rats. Rats were followed up for 24 weeks. At the conclusion of the follow-up period, patency and structural integrity were evaluated by digital subtraction angiography. The abdominal aorta, including the graft, was harvested and investigated under light microscopy. Endothelial coverage, neointima formation, and transmural cellular ingrowth were measured by computed histomorphometry. All animals survived until the end of follow-up, and all grafts were patent in both groups. Digital subtraction angiography revealed no stenosis in the PCL group but stenotic lesions in 1 graft at 18 weeks (40%) and in another graft at 24 weeks (50%) in the ePTFE group. None of the grafts showed aneurysmal dilatation. Endothelial coverage was significantly better in the PCL group. Neointimal formation was comparable between the 2 groups. Macrophage and fibroblast ingrowth with extracellular matrix formation and neoangiogenesis were better in the PCL group. After 12 weeks, foci of chondroid metaplasia located in the neointima of PCL grafts were observed in all samples.
Small-diameter PCL grafts represent a promising alternative for the future because of their better healing characteristics compared with ePTFE grafts. Faster endothelialization and extracellular matrix formation, accompanied by degradation of graft fibers, seem to be the major advantages. Further evaluation of degradation and graft healing characteristics may potentially lead to the clinical use of such grafts for revascularization procedures.
Arteries are not elastic bodies in the usual sense. But when it is subjected to cyclic loading and unloading, the stress-strain relationship is well defined after preconditioning. For either the loading branch or the unloading branch, a pseudo strain energy function can be used to express the stress-strain relationship. An expression which is the most practical approximation for an artery subjected to internal pressure and longitudinal stretching within the physiologic range is shown. The application of this formula is illustrated by the variation of the mechanical properties of rabbit arteries along the arterial tree.
The structure and materials of the blood vessel wall are layered. This article presents the principle of a method to determine the mechanical properties of the different layers in vivo. In vivo measurement begets in vivo data and avoids pitfalls of in vitro tests of dissected specimens. With the proposed method, we can measure vessels of diameters 100 microns and up and obtain data on vascular smooth muscles and adventitia. To derive the full constitutive equations, one must first determine the zero-stress state, obtain the morphometric data on the thicknesses of the layers, and make mechanical measurements in the neighborhood of the zero-stress state. Then eight small perturbation experiments are done on earth blood vessel in vivo to determine eight incremental elastic moduli of the two layers of the blood vessel wall. The calculation requires the morphometric data and the location of the neutral axis. The experiments are simple, the interpretation is definitive, but the analysis is somewhat sophisticated. The method will yield results that are needed to assess the stress and strain in the tissues of the blood vessel. The subject is important because blood vessels remodel themselves significantly and rapidly when their stress and strain deviate from their homeostatic values, and because cell proliferation, differentiation, adhesion, contraction, and locomotion depend on stress and strain in the tissue.
Mechanically challenged tissue-engineered organs, such as blood vessels, traditionally relied on synthetic or modified biological materials for structural support. In this report, we present a novel approach to tissue-engineered blood vessel (TEBV) production that is based exclusively on the use of cultured human cells, i.e., without any synthetic or exogenous biomaterials. Human vascular smooth muscle cells (SMC) cultured with ascorbic acid produced a cohesive cellular sheet. This sheet was placed around a tubular support to produce the media of the vessel. A similar sheet of human fibroblasts was wrapped around the media to provide the adventitia. After maturation, the tubular support was removed and endothelial cells were seeded in the lumen. This TEBV featured a well-defined, three-layered organization and numerous extracellular matrix proteins, including elastin. In this environment, SMC reexpressed desmin, a differentiation marker known to be lost under standard culture conditions. The endothelium expressed von Willebrand factor, incorporated acetylated LDL, produced PGI2, and strongly inhibited platelet adhesion in vitro. The complete vessel had a burst strength over 2000 mmHg. This is the first completely biological TEBV to display a burst strength comparable to that of human vessels. Short-term grafting experiment in a canine model demonstrated good handling and suturability characteristics. Taken together, these results suggest that this novel technique can produce completely biological vessels fulfilling the fundamental requirements for grafting: high burst strength, positive surgical handling, and a functional endothelium.
The study was to investigate the crosslinking characteristics, mechanical properties, and resistance against enzymatic degradation of biological tissues after fixation with genipin (a naturally occurring crosslinking agent) and/or carbodiimide. Fresh tissue was used as a control. It was found that both genipin and carbodiimide are effective crosslinking agents for tissue fixation and genipin crosslinking is comparatively slower than carbodiimide crosslinking. Additionally, tissue fixation in genipin and/or carbodiimide may produce distinct crosslinking structures. Carbodiimide may form intrahelical and interhelical crosslinks within or between tropocollagen molecules, whereas genipin may further introduce intermicrofibrillar crosslinks between adjacent collagen microfibrils. The stability (denaturation temperature and resistance against enzymatic degradation) of the fixed tissue is mainly determined by its intrahelical and interhelical crosslinks. In contrast, intermicrofibrillar crosslinks significantly affect the mechanical properties (tissue shrinkage during fixation, tensile strength, strain at break, and ruptured pattern) of the fixed tissue. Moreover, the degree of enzymatic degradation of the fixed tissue may be influenced by three factors: the availability, to the enzyme, of recognizable cleavage sites, the degree of crosslinking, and the extent of helical integrity of tropocollagen molecules in tissue.
The goals of this paper are (i) to re-examine the constitutive law for the description of the (passive) highly nonlinear and anisotropic response of healthy elastic arteries introduced recently by the authors, (ii) to show how the mechanical response of a carotid artery under inflation and extension predicted by the structural model compares with that for a three-dimensional form of Fung-type strain-energy function, (iii) to provide a new set of material parameters that can be used in a finite element program, and (iv) to show that the model has certain mathematical features that are important from the point of view of material and numerical stability.
The detailed mechanical properties of various layers of the coronary artery are important for understanding the function of the vessel. The present article is focused on the determination of the incremental modulus in different layers and directions in the neighborhood of the in vivo state. The incremental modulus can be defined for any material subjected to a large deformation if small perturbations in strain lead to small perturbations of stresses in a linear fashion. This analysis was applied to the porcine coronary artery, which was treated as a two-layered structure consisting of an inner intima-media layer and an outer adventitia layer. We adopted a theory based on small-perturbation experiments at homeostatic conditions for determination of incremental moduli in circumferential, axial, and cross directions in the two layers. The experiments were based on inflation and axial stretch. We demonstrate that under homeostatic conditions the incremental moduli are layer- and direction dependent. The incremental modulus is highest in the circumferential direction. Furthermore, in the circumferential direction, the media is stiffer than the whole wall, which is stiffer than the adventitia. In the axial direction, the adventitia is stiffer than the intact wall, which is stiffer than the media. Hence, the coronary artery must be treated as a composite, nonisotropic body. The data acquire physiological relevance in relation to coronary artery health and disease.
Although collagen-based tissue-engineered blood vessels (TEBVs) have many interesting properties and have been utilized to study aspects of vascular biology, these constructs are too weak to be implanted as bypass grafts for in vivo investigations. This study presents a method to incorporate organized, intact elastin into collagen-based TEBVs to form hybrid constructs that better mimic arterial physiology and exhibit improved mechanical properties. Porcine carotids were digested with a series of autoclave and chemical treatments to elicit isolated elastin scaffolds. Elastin purity was verified via immunohistochemistry and amino acid analysis. Isolated scaffolds were combined with type I collagen and either human dermal fibroblasts (HDFs) or rat smooth muscle cells (RASMs) to form an elastin hybrid TEBV. Hybrid constructs exhibited increased tensile strengths (11-fold in HDFs; 7.5-fold in RASMs) and linear stiffness moduli (4-fold in HDFs; 1.8-fold in RASMs) compared with collagen control constructs with no exogenous elastin scaffold. Viscoelastic properties of the TEBVs also improved with the addition of an ancillary elastin scaffold as determined through stepwise stress relaxation analysis. Whereas the majority of resistance to deformation in collagen control constructs stemmed from viscous fluidlike effects, elastin hybrid constructs exhibited more ideal elastic solid mechanical behavior. Thus, elastin scaffolds can help recreate the elastic properties of native arteries. Future challenges include stimulating appropriate reorganization or synthesis of the collagen matrix to provide the necessary strength and viscoelastic properties for implantation.
The importance of mechanical stresses and strains has become well recognized in vascular physiology and pathology. To compute the stress and strain on the various components of the vessel wall, we must know the constitutive equations for the different layers of the vessel wall. The objective of the present study is to determine the constitutive equation of the coronary artery treated as a two-layer composite: intima-media and adventitial layers. Twelve hearts were obtained from a local slaughterhouse, and the right coronary artery and left anterior descending artery were dissected free from the myocardium. The vessel wall was initially mechanically tested biaxially (inflation and axial extension) as a whole (intact wall) and subsequently as intima-media or adventitial layer. A Fung-type exponential strain energy function was used to curve fit the experimental data for the intact wall and individual layers for the right coronary artery and left anterior descending artery. Two methods were used for the determination of material constants, including the Marquardt-Levenberg nonlinear least squares method and the genetic algorithm method. Our results show that there were no statistically significant differences in the material constants obtained from the two methods and that either set of elastic constants results in good fit of the data. Furthermore, at an in vivo value of axial stretch ratio, we find that the stiffness is as follows: intima-media > intact > adventitia. These results underscore the composite nature of coronary arteries with different material properties in each layer. The present results are necessary for analysis of coronary artery mechanics and to provide a fundamental understanding of vessel physiology.
Meshes of collagen and/or elastin were successfully prepared by means of electrospinning from aqueous solutions. Flow rate, applied electric field, collecting distance and composition of the starting solutions determined the morphology of the obtained fibres. Addition of PEO (M(w)=8 x 10(6)) and NaCl was always necessary to spin continuous and homogeneous fibres. Spinning a mixture of collagen and elastin resulted in fibres in which the single components could not be distinguished by SEM. Increasing the elastin content determined an increase in fibres diameters from 220 to 600 nm. The voltage necessary for a continuous production of fibres was dependent on the composition of the starting solution, but always between 10 and 25 kV. Under these conditions, non-woven meshes could be formed and a partial orientation of the fibres constituting the mesh was obtained by using a rotating tubular mandrel as collector. Collagen/elastin (1:1) meshes were stabilized by crosslinking with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). This treatment afforded materials with a high thermal stability (T(d)=79 degrees C) without altering their original morphology. Upon crosslinking PEO and NaCl were fully leached out. Smooth muscle cells grew as a confluent layer on top of the crosslinked meshes after 14 d of culture.
The aim of this study was to prepare non-woven materials from a biodegradable polymer, poly(epsilon-caprolactone) (PCL) by electrospinning. PCL was synthesized by ring-opening polymerization of epsilon-caprolactone in bulk using stannous octoate as the catalyst under nitrogen atmosphere. PCL was then processed into non-woven matrices composed of nanofibers by electrospinning of the polymer from its solution using a high voltage power supply. The effects of PCL concentration, composition of the solvent (a mixture of chloroform and DMF with different DMF content), applied voltage and tip-collector distance on fiber diameter and morphology were investigated. The diameter of fibers increased with the increase in the polymer concentration and decrease in the DMF content significantly. Applied voltage and tip-collector distance were found critical to control 'bead' formation. Elongation-at-break, ultimate strength and Young's modulus were obtained from the mechanical tests, which were all increased by increasing fiber diameter. The fiber diameter significantly influenced both in vitro degradation (performed in Ringer solution) and in vivo biodegradation (conducted in rats) rates. In vivo degradation was found to be faster than in vitro. Electrospun membranes were more hydrophobic than PCL solvent-casted ones; therefore, their degradation was a much slower process.
The normal coronary artery consists of two mechanically distinct layers: intima-media and adventitia. The objective of this study is to establish a two-layer three-dimensional (3-D) stress-strain relation of porcine coronary arteries. Experimental measurements were made by a series of biaxial tests (inflation and axial extension) of intact coronary arteries and, subsequently, their corresponding intima-media or adventitia layer. The Fung-type exponential strain energy function was used to describe the 3-D strain-stress relation for each layer and the intact wall. A genetic algorithm was used to determine the material constants in the Fung-type constitutive equation by curve fitting the experimental data. Because one layer must be sacrificed before the other layer can be tested, the material property of the missing layer was computed from the material constants of the intact vessel and the tested layer. A total of 20 porcine hearts were used: one group of 10 hearts for the left anterior descending artery and another group of 10 hearts for the right coronary artery. Each group was further divided into two subgroups of five specimens tested for the intact wall and the intima-media layer and for the intact wall and the adventitia layer. Our results show statistically significant differences in the material properties of the two layers. The mathematical model was validated by experimental stress-strain data for individual layers. The validated 3-D constitutive model will serve as a foundation for formulation of layer-specific boundary value problems in coronary physiology and cardiology.
Dermal regeneration templates arguably represent the first and most clinically successful 'tissue engineering' solution designed for organ reconstruction. Wound healing in the skin normally occurs on a continuum. At one extreme of the continuum lies the promise of tissue regeneration and the complete restoration of normal structure and function. Unfortunately, in the adult, all too often, wound healing occurs at the other extreme of the continuum and the dermis is reconstituted as scar tissue. Dermal regeneration templates are designed to manage the wound-healing process and tip the scales toward regeneration. This review discusses the architecture and molecular composition of the skin and the events that mediate wound healing and scar formation. The development, evolution and commercialization of dermal templates are examined and the clinical and business considerations that drive the product-development cycle are discussed. In the near term, dermal templates cannot be expected to dramatically change in overall composition. Product development will be dominated by continued refinements of existing templates and the field of use will continue to expand as manufacturers seek to increase revenue and capture market share. Continued exploration of novel processing strategies, such as electrospinning, that can be used to fabricate nanoscale biomaterials, may provide a gateway to the next generation of dermal templates.
Fibers with nanoscale diameters provide benefits due to high surface area for biomaterial scaffolds. In this study electrospun silk fibroin-based fibers with average diameter 700+/-50 nm were prepared from aqueous regenerated silkworm silk solutions. Adhesion, spreading and proliferation of human bone marrow stromal cells (BMSCs) on these silk matrices was studied. Scanning electron microscopy (SEM) and MTT analyses demonstrated that the electrospun silk matrices supported BMSC attachment and proliferation over 14 days in culture similar to native silk fibroin (approximately 15 microm fiber diameter) matrices. The ability of electrospun silk matrices to support BMSC attachment, spreading and growth in vitro, combined with a biocompatibility and biodegradable properties of the silk protein matrix, suggest potential use of these biomaterial matrices as scaffolds for tissue engineering.
Numerous crosslinking chemistries and methodologies have been investigated as alternative fixatives to glutaraldehyde (GLUT) for the stabilization of bioprosthetic heart valves (BHVs). Particular attention has been paid to valve leaflet collagen and elastin stability following fixation. However, the stability of glycosaminoglycans (GAGs), the primary component of the spongiosa layer of the BHV, has been largely overlooked despite recent evidence provided by our group illustrating their structural and functional importance. In the present study we investigate the ability of two different crosslinking chemistries: sodium metaperiodate (NaIO(4)) followed by GLUT (PG) and 1-Ethyl-3-(3 dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) followed by GLUT (ENG) to stabilize GAGs within BHV leaflets and compare resulting leaflet characteristics with that of GLUT-treated tissue. Incubation of fixed leaflets in GAG-degrading enzymes illustrated in vitro resistance of GAGs towards degradation in PG and ENG treated tissue while GLUT fixation alone was not effective in preventing GAG loss from BHV leaflets. Following subdermal implantation, significant amounts of GAGs were retained in leaflets in the ENG group in comparison to GLUT-treated tissue, although GAG loss was evident in all groups. Utilizing GAG-targeted fixation did not alter calcification potential of the leaflets while collagen stability was maintained at levels similar to that observed in conventional GLUT-treated tissue.
Elastin is an elastomeric, self-assembling extracellular matrix protein with potential for use in biomaterials applications. Here, we compare the microstructural and tensile properties of the elastin-based recombinant polypeptide (EP) EP20-244 crosslinked with either genipin (GP) or pyrroloquinoline quinone (PQQ). Recombinant EP-based sheets were produced via coacervation and subsequent crosslinking. The micron-scale topography of the GP-crosslinked sheets examined with atomic force microscopy revealed the presence of extensive mottling compared with that of the PQQ-crosslinked sheets, which were comparatively smoother. Confocal microscopy showed that the subsurface porosity in the GP-crosslinked sheets was much more open. GP-crosslinked EP-based sheets exhibited significantly greater tensile strength (P < or = 0.05). Mechanistically, GP appears to yield a higher crosslink density than PQQ, likely due to its capacity to form short-range and long-range crosslinks. In conclusion, GP is able to strongly modulate the microstructural and mechanical properties of elastin-based polypeptide biomaterials forming membranes with mechanical properties similar to native insoluble elastin.
