Tissue Engineering of Human Heart Valve Leaflets: A Novel Bioreactor for a Strain-Based Conditioning Approach

Current mechanical conditioning approaches for heart valve tissue engineering concentrate on mimicking the opening and closing behavior of the leaflets, either or not in combination with tissue straining. This study describes a novel approach by mimicking only the diastolic phase of the cardiac cycle, resulting in tissue straining. A novel, yet simplified, bioreactor system was developed for this purpose by applying a dynamic pressure difference over a closed tissue engineered valve, thereby inducing dynamic strains within the leaflets. Besides the use of dynamic strains, the developing leaflet tissues were exposed to prestrain induced by the use of a stented geometry. To demonstrate the feasibility of this strain-based conditioning approach, human heart valve leaflets were engineered and their mechanial behavior evaluated. The actual dynamic strain magnitude in the leaflets over time was estimated using numerical analyses. Preliminary results showed superior tissue formation and non-linear tissue-like mechanical properties in the strained valves when compared to non-loaded tissue strips. In conclusion, the strain-based conditioning approach, using both prestrain and dynamic strains, offers new possibilities for bioreactor design and optimization of tissue properties towards a tissue-engineered aortic human heart valve replacement.     

The in vitro development of autologous fibrin-based tissue-engineered heart valves through optimised dynamic conditioning

Our group has previously demonstrated the synthesis of a completely autologous fibrin-based heart valve structure using the principles of tissue engineering. The present approach aims to guide more mature tissue development in fibrin-based valves based on in vitro conditioning in a custom-designed bioreactor system. Moulded fibrin-based tissue-engineered heart valves seeded with ovine carotid artery-derived cells were subjected to 12 days of mechanical conditioning in a bioreactor system. The bioreactor pulse rate was increased from 5 to 10 b.p.m. after 6 days, while a pressure difference of 20 mmH(2)O was maintained over the valve leaflets. Control valves were cultured under stirred conditions in a beaker. Cell phenotype and extracellular matrix (ECM) composition were analysed in all samples and compared to native ovine aortic valve tissue using routine histological and immunohistochemical techniques. Conditioned valve leaflets showed reduced tissue shrinkage compared to stirred controls. Limited ECM synthesis was evident in stirred controls, while the majority of cells were detached from the fibrin scaffold. Dynamic conditioning increased cell attachment/alignment and expression of alpha-smooth muscle actin, while enhancing the deposition of ECM proteins, including types I and III collagen, fibronectin, laminin and chondroitin sulphate. There was no evidence for elastin synthesis in either stirred controls or conditioned samples. The present study demonstrates that the application of low-pressure conditions and increasing pulsatile flow not only enhances seeded cell attachment and alignment within fibrin-based heart valves, but dramatically changes the manner in which these cells generate ECM proteins and remodel the valve matrix. Optimised dynamic conditioning, therefore, might accelerate the maturation of surgically feasible and implantable autologous fibrin-based tissue-engineered heart valves.     

组织工程心脏瓣膜支架材料的选择与应用

背景：支架材料的选择在组织工程心脏瓣膜中起着至关重要的作用,支架材料的选择也就影响着组织工程心脏瓣膜的构建效果。 目的：评价组织工程心脏瓣膜支架材料的的优缺点,并对其选择进行总结。 方法：以 “组织工程,心脏瓣膜,支架材料,生物相容性”,为中文关键词;以:“tissue engineering,heart valves, scaffold material, biocompatibility” 为英文关键词,采用计算机检索1993-01/2009-10相关文章。纳入与有关生物材料与组织工程心脏瓣膜的相关的文章;排除重复研究及Meta分析类文章。 结果与结论：人工合成高分子材料有更大的可控性,可预先塑性,大量制备,孔径和孔隙率较容易控制,成本低廉;天然生物材料和合成高分子材料都存在一定不足,将人工可降解材料与天然材料相结合构建瓣膜支架,发挥两者各自的优势构建出性能良好的组织工程心脏瓣膜。组织工程心脏瓣膜的研究前景广阔。但距离临床应用还有很长的路要走,相信随着研究的不断深入以及支架材料的不断优化对组织工程心脏瓣膜构建方法的改进,在不远的将来造福于广大心脏瓣膜病患者。     

Tubular Heart Valves from Decellularized Engineered Tissue

A novel tissue-engineered heart valve (TEHV) was fabricated from a decellularized tissue tube mounted on a frame with three struts, which upon back-pressure cause the tube to collapse into three coapting “leaflets.” The tissue was completely biological, fabricated from ovine fibroblasts dispersed within a fibrin gel, compacted into a circumferentially aligned tube on a mandrel, and matured using a bioreactor system that applied cyclic distension. Following decellularization, the resulting tissue possessed tensile mechanical properties, mechanical anisotropy, and collagen content that were comparable to native pulmonary valve leaflets. When mounted on a custom frame and tested within a pulse duplicator system, the tubular TEHV displayed excellent function under both aortic and pulmonary conditions, with minimal regurgitant fractions and transvalvular pressure gradients at peak systole, as well as well as effective orifice areas exceeding those of current commercially available valve replacements. Short-term fatigue testing of one million cycles with pulmonary pressure gradients was conducted without significant change in mechanical properties and no observable macroscopic tissue deterioration. This study presents an attractive potential alternative to current tissue valve replacements due to its avoidance of chemical fixation and utilization of a tissue conducive to recellularization by host cell infiltration.     

Pediatric tubular pulmonary heart valve from decellularized engineered tissue tubes

Pediatric patients account for a small portion of the heart valve replacements performed, but a pediatric pulmonary valve replacement with growth potential remains an unmet clinical need. Herein we report the first tubular heart valve made from two decellularized, engineered tissue tubes attached with absorbable sutures, which can meet this need, in principle. Engineered tissue tubes were fabricated by allowing ovine dermal fibroblasts to replace a sacrificial fibrin gel with an aligned, cell-produced collagenous matrix, which was subsequently decellularized. Previously, these engineered tubes became extensively recellularized following implantation into the sheep femoral artery. Thus, a tubular valve made from these tubes may be amenable to recellularization and, ideally, somatic growth.The suture line pattern generated three equi-spaced leaflets in the inner tube, which collapsed inward when exposed to back pressure, per tubular valve design. Valve testing was performed in a pulse duplicator system equipped with a secondary flow loop to allow for root distention. All tissue-engineered valves exhibited full leaflet opening and closing, minimal regurgitation (<5%), and low systolic pressure gradients (<2.5 mmHg) under pulmonary conditions. Valve performance was maintained under various trans-root pressure gradients and no tissue damage was evident after 2 million cycles of fatigue testing.     

New pulsatile bioreactor for fabrication of tissue-engineered patches.

To date, one approach to tissue engineering has been to develop in vitro conditions to ultimately fabricate functional cardiovascular structures prior to final implantation. In our current experiment, we developed a new pulsatile flow system that provides biochemical and biomechanical signals to regulate autologous patch-tissue development in vitro. The newly developed patch bioreactor is made of Plexiglas and is completely transparent (Mediport Kardiotechnik, Berlin). The bioreactor is connected to an air-driven respirator pump, and the cell culture medium continuously circulates through a closed-loop system. We thus developed a closed-loop, perfused bioreactor for long-term patch-tissue conditioning, which combines continuous, pulsatile perfusion and mechanical stimulation by periodically stretching the tissue-engineered patch constructs. By adjusting the stroke volume, the stroke rate, and the inspiration/expiration time of the ventilator, it allows various pulsatile flows and different levels of pressure. The whole system is a highly isolated cell culture setting, which provides a high level of sterility, gas supply, and fits into a standard humidified incubator. The bioreactor can be sterilized by ethylene oxide and assembled with a standard screwdriver. Our newly developed bioreactor provides optimal biomechanical and biodynamical stimuli for controlled tissue development and in vitro conditioning of an autologous tissue-engineered patch.Copyright 2001 John Wiley & Sons, Inc.     

Bioreactors for Development of Tissue Engineered Heart Valves

Millions of people worldwide are diagnosed each year with valvular heart disease, resulting in hundreds of thousands of valve replacement operations. Prosthetic valve replacements are designed to correct narrowing or backflow through the valvular orifice. Although commonly used, these therapies have serious disadvantages including morbidity associated with long-term anticoagulation and limited durability necessitating repeat operations. The ideal substitute would be widely available and technically implantable for most cardiac surgeons, have normal hemodynamic performance, low risk for structural degeneration, thrombo-embolism and endocarditis, and growth potential for pediatric patients. Tissue engineered heart valves hold promise as a viable substitute to outperform existing valve replacements. An essential component to the development of tissue engineered heart valves is a bioreactor. It is inside the bioreactor that the scaffold and cells are gradually conditioned to the biochemical and mechanical environment of the valve to be replaced.     

Application of tissue-engineering principles toward the development of a semilunar heart valve substitute

Heart valve disease is a significant medical problem worldwide. Current treatment for heart valve disease is heart valve replacement. State of the art replacement heart valves are less than ideal and are associated with significant complications. Using the basic principles of tissue engineering, promising alternatives to current replacement heart valves are being developed. Significant progress has been made in the development of a tissue-engineered semilunar heart valve substitute. Advancements include the development of different potential cell sources and cell-seeding techniques; advancements in matrix and scaffold development and in polymer chemistry fabrication; and the development of a variety of bioreactors, which are biomimetic devices used to modulate the development of tissue-engineered neotissue in vitro through the application of biochemical and biomechanical stimuli. This review addresses the need for a tissue-engineered alternative to the current heart valve replacement options. The basics of heart valve structure and function, heart valve disease, and currently available heart valve replacements are discussed. The last 10 years of investigation into a tissue-engineered heart valve as well as current developments are reviewed. Finally, the early clinical applications of cardiovascular tissue engineering are presented.     

Tissue engineering of a genitourinary tubular tissue graft resistant to suturing and high internal pressures

The aim of this study was to evaluate the possibility of constructing a fully autologous tissue-engineered tubular genitourinary graft (TTGG) and to determine its mechanical and physiological properties. Dermal fibroblasts (DFs) were expanded and cultured in vitro with sodium ascorbate to form fibroblast sheets. The sheets were then wrapped around a tubular support to form a cylinder. After maturation, urothelial cells (UCs) were seeded inside the DF tubes, and the constructs were placed in a bioreactor. The TTGGs were then characterized according to histology, immuno-histochemistry, Western blot, cell viability, resistance to suture, and burst pressure. Results obtained were encouraging on all levels. All layers of the TTGGs had merged, and a pluristratified urothelium coated the luminal surface of the tubes. The burst pressure of non-sutured TTGGs was measured and found to be, on average, three times as resistant as that of porcine urethras. Suturing was accomplished without difficulty. Results have shown that our construct can sustain an entire week of pulsatile stimulation without loss of mechanical or histological integrity. The tissue-engineering technique used to produce this model seems promising for bioengineering a urethra or ureter graft and could open a doorway to new possibilities for their reconstruction.     

6-Month aortic valve implantation of an off-the-shelf tissue-engineered valve in sheep

Diseased aortic valves often require replacement, with over 30% of the current aortic valve surgeries performed in patients who will outlive a bioprosthetic valve. While many promising tissue-engineered valves have been created in the lab using the cell-seeded polymeric scaffold paradigm, none have been successfully tested long-term in the aortic position of a pre-clinical model. The high pressure gradients and dynamic flow across the aortic valve leaflets require engineering a tissue that has the strength and compliance to withstand high mechanical demand without compromising normal hemodynamics. A long-term preclinical evaluation of an off-the-shelf tissue-engineered aortic valve in the sheep model is presented here. The valves were made from a tube of decellularized cell-produced matrix mounted on a frame. The engineered matrix is primarily composed of collagen, with strength and organization comparable to native valve leaflets. In vitro testing showed excellent hemodynamic performance with low regurgitation, low systolic pressure gradient, and large orifice area. The implanted valves showed large-scale leaflet motion and maintained effective orifice area throughout the duration of the 6-month implant, with no calcification. After 24 weeks implantation (over 17 million cycles), the valves showed no change in tensile mechanical properties. In addition, histology and DNA quantitation showed repopulation of the engineered matrix with interstitial-like cells and endothelialization. New extracellular matrix deposition, including elastin, further demonstrates positive tissue remodeling in addition to recellularization and valve function. Long-term implantation in the sheep model resulted in functionality, matrix remodeling, and recellularization, unprecedented results for a tissue-engineered aortic valve.     

Novel pulse duplicating bioreactor system for tissue-engineered vascular construct

Cell culture in a biomimetic environment is known to improve the mechanical endurance of tissue-engineered cardiovascular components. Our goal was to generate a bioreactor that can reproduce a wide range of pulsatile flows with a completely physiological pressure profile. The morphology and biochemical properties of tissue-engineered products were also studied to test the usefulness of this novel bioreactor. The combination of an outflow valve, compliance chamber, and resistant clamps together with a balloon pumping system was able to successfully reproduce both physiological systolic and diastolic pressures. The compliance chamber was especially effective in transforming the original peaky pressure waveform into a physiological pressure profile. The tissues, cultured under a physiological pressure waveform with pulsatile flow, presented widely distributed cells in close contact with each other. They also showed significantly higher cell numbers, total protein content, and proteoglycan-glycosaminoglycan content than cultured tissues under a peaky pressure wave or under static conditions. This new bioreactor system is suitable for evaluating a favorable environment for tissue-engineered cardiovascular components.     

A Bioreactor with Compliance Monitoring for Heart Valve Grafts

The drawbacks of state-of-the-art heart valve prostheses lead researchers to explore the prospect of using tissue-engineered constructs as possible valve substitutes. It is widely accepted that the mechanical properties of the construct are improved with mechanical stimulation during in vitro growth. We designed a new dynamic bioreactor with the perspective of using decellularized valve homografts as scaffolds in order to produce tissue-engineered valve substitutes. The design guidelines were (a) compatibility with the procedures for the treatment of homografts; (b) delivery of finely controlled pulsatile pressure loads, which induce strain stimuli that may drive cells toward repopulation of and integration with the natural scaffold; and (c) monitoring the construct’s biomechanical status through a comprehensive index, i.e., its compliance. The handling needs during the set-up of the homograft and the use of the bioreactor were minimized. The bioreactor and its automated control system underwent tests with a compliant phantom valve. The estimated compliances are in good agreement with the measured ones. Tests were also carried out with porcine aortic samples in order to assess the hydrodynamic and biomechanical reliability. In the future, monitoring the construct’s compliance might represent a key factor in controlling the recellularization of the valve homografts, which provides awareness of the construct’s biomechanical status by real-time, non-destructive, and non-invasive means.     

New pulsatile bioreactor for in vitro formation of tissue engineered heart valves

Two potential obstacles to the creation of implantable tissue engineered heart valves are inadequate mechanical properties (ability to withstand hemodynamic stresses) and adverse host-tissue reactions due to the presence of residual nondegraded polymer scaffold. In an attempt to address these problems, we developed an in vitro cell culture system that provides physiological pressure and flow of nutrient medium to the developing valve constructs. It is anticipated that in vitro physical stress will stimulate the tissue engineered heart valve construct to develop adequate strength prior to a possible implantation. Long-term in vitro development will be realized by an isolated and thereby contamination-resistant system. Longer in vitro development will potentially enable more complete biodegradation of the polymeric scaffold during in vitro cultivation. This new dynamic bioreactor allows for adjustable pulsatile flow and varying levels of pressure. The system is compact and easily fits into a standard cell incubator, representing a highly isolated dynamic cell culture setting with maximum sterility, optimal gas supply and stable temperature conditions especially suited for long-term experiments.     

Fabrication of a trileaflet heart valve scaffold from a polyhydroxyalkanoate biopolyester for use in tissue engineering

Previously, we reported the implantation of a single tissue engineered leaflet in the posterior position of the pulmonary valve in a lamb model. The major problems with this leaflet replacement were the scaffold's inherent stiffness, thickness, and nonpliability. We have now created a scaffold for a trileaflet heart valve using a thermoplastic polyester. In this experiment, we show the suitability of this material in the production of a biodegradable, biocompatible scaffold for tissue engineered heart valves. A heart valve scaffold was constructed from a thermoplastic elastomer. The elastomer belongs to a class of biodegradable, biocompatible polyesters known as polyhydroxyalkanoates (PHAs) and is produced by fermentation (Metabolix Inc., Cambridge, MA). It was modified by a salt leaching technique to create a porous, three-dimensional structure, suitable for tissue engineering. The trileaflet heart valve scaffold consisted of a cylindrical stent (1 mm X 15 mm X 20 mm I.D.) containing three valve leaflets. The leaflets were formed from a single piece of PHA (0.3 mm thick), and were attached to the outside of the stent by thermal processing techniques, which required no suturing. After fabrication, the heart valve construct was allowed to crystallize (4 degrees C for 24 h), and salt particles were leached into doubly distilled water over a period of 5 days to yield pore sizes ranging from 80 to 200 microns. Ten heart valve scaffolds were fabricated and seeded with vascular cells from an ovine carotid artery. After 4 days of incubation, the constructs were examined by scanning electron microscopy. The heart valve scaffold was tested in a pulsatile flow bioreactor and it was noted that the leaflets opened and closed. Cells attached to the polymer and formed a confluent layer after incubation. One advantage of this material is the ability to mold a complete trileaflet heart valve scaffold without the need for suturing leaflets to the conduit. Second advantage is the use of only one polymer material (PHA) as opposed to hybridized polymer scaffolds. Furthermore, the mechanical properties of PHA, such as elasticity and mechanical strength, exceed those of the previously utilized material. This experiment shows that PHAs can be used to fabricate a three-dimensional, biodegradable heart valve scaffold.     

Estimation of the Shear Stress on the Surface of an Aortic Valve Leaflet

The limited durability of xenograft heart valves and the limited supply of allografts have sparked interest in tissue engineered replacement valves. A bioreactor for tissue engineered valves must operate at conditions that optimize the biosynthetic abilities of seeded cells while promoting their adherence to the leaflet matrix. An important parameter is shear stress, which is known to influence cellular behavior and may thus be crucial in bioreactor optimization. Therefore, an accurate estimate of the shear stress on the leaflet surface would not only improve our understanding of the mechanical environment of aortic valve leaflets, but it would also aid in bioreactor design. To estimate the shear stress on the leaflet surface, two-component laser-Doppler velocimetry measurements have been conducted inside a transparent polyurethane valve with a trileaflet structure similar to the native aortic valve. Steady flow rates of 7.5, 15.0, and 22.5 L/min were examined to cover the complete range possible during the cardiac cycle. The laminar shear stresses were calculated by linear regression of four axial velocity measurements near the surface of the leaflet. The maximum shear stress recorded was 79 dyne/cm², in agreement with boundary layer theory and previous experimental and computational studies. This study has provided a range of shear stresses to be explored in bioreactor design and has defined a maximum shear stress at which cells must remain adherent upon a tissue engineered construct. © 1999 Biomedical Engineering Society. PAC99: 8719Rr, 8768+z, 8719Hh, 4262Be, 4727Nz, 0630Gv     

Creation of myocardial tubes using cardiomyocyte sheets and an in vitro cell sheet-wrapping device

Regenerative medicine involving injection of isolated cells and transplantation of tissue-engineered myocardial patches, has received significant attention as an alternative method to repair damaged heart muscle. In the present study, as the next generation of myocardial tissue engineering we demonstrate the in vitro fabrication of pulsatile myocardial tubes using cell sheet engineering technologies. Three neonatal rat cardiomyocyte sheets, which were harvested from temperature-responsive culture dishes, were wrapped around fibrin tubes using a novel cell sheet-wrapping device. The tubular constructs demonstrated spontaneous, synchronized pulsation within 3h after cell sheet wrapping. Contractile force measurements showed that the contractile force increased in accordance with both increasing rest length (Starling mechanism) and increasing extracellular Ca(2+) concentration. Furthermore, the tissue-engineered myocardial tubes presented measurable inner pressure changes evoked by tube contraction (0.11+/-0.01mmHg, max 0.15mmHg, n=5). Histological analyses revealed both well-differentiated sarcomeres and diffuse gap junctions within the myocardial tissues that resembled native cardiac muscle. These data indicate that tissue-engineered myocardial tubes have native heart-like structure and function. These new myocardial tissue constructs should be useful for future applications in physiological studies and pharmacological screening, and present a possible core technology for the creation of engineered tissues capable of independent cardiac assistance.     

Tissue engineering in the cardiovascular system: progress toward a tissue engineered heart

Achieving the lofty goal of developing a tissue engineered heart will likely rely on progress in engineering the various components: blood vessels, heart valves, and cardiac muscle. Advances in tissue engineered vascular grafts have shown the most progress to date. Research in tissue-engineered vascular grafts has focused on improving scaffold design, including mechanical properties and bioactivity; genetically engineering cells to improve graft performance; and optimizing tissue formation through in vitro mechanical conditioning. Some of these same approaches have been used in developing tissue engineering heart valves and cardiac muscle as well. Continued advances in scaffold technology and a greater understanding of vascular cell biology along with collaboration among engineers, scientists, and physicians will lead to further progress in the field of cardiovascular tissue engineering and ultimately the development of a tissue-engineered heart.     

组织工程心脏瓣膜支架材料的研究和展望

背景：当前应用于临床的生物瓣和机械瓣都存在着一定缺陷,而组织工程心脏瓣膜将避免这些问题成为理想的生物瓣膜替代物。 目的：综述近年来组织工程心脏瓣膜支架材料的研究进展及面临的问题。 方法：应用计算机检索1990至2011年万方数据库相关文章,检索词为“组织工程,心脏瓣膜,支架材料”,并限定文章语言种类为中文。同时计算机检索1990至2011年 PubMed数据库相关文章,检索词为“tissue engineering,heart valve,scaffold materials”,并限定文章语言种类为English。共检索到文献147篇,最终纳入符合标准的文献61篇。 结果与结论：人工心脏瓣膜置换是目前治疗心脏瓣膜性病变的主要外科治疗手段,但现有机械瓣和生物瓣都不是理想的心脏瓣膜置换物,在耐久性,增长潜力,相容性,抗感染方面有着显著的缺陷。组织工程心脏瓣膜是一个活体器官,并具有和自体心脏瓣膜同样的生长,修复和重建能力,这一新概念的提出为构建理想的心脏瓣替换物带来了希望。     

Application of stereolithography for scaffold fabrication for tissue engineered heart valves

A crucial factor in tissue engineering of heart valves is the functional and physiologic scaffold design. In our current experiment, we describe a new fabrication technique for heart valve scaffolds, derived from x-ray computed tomography data linked to the rapid prototyping technique of stereolithography. To recreate the complex anatomic structure of a human pulmonary and aortic homograft, we have used stereolithographic models derived from x-ray computed tomography and specific software (CP, Aachen, Germany). These stereolithographic models were used to generate biocompatible and biodegradable heart valve scaffolds by a thermal processing technique. The scaffold forming polymer was a thermoplastic elastomer, a poly-4-hydroxybutyrate (P4HB) and a polyhydroxyoctanoate (PHOH) (Tepha, Inc., Cambridge, MA). We fabricated one human aortic root scaffold and one pulmonary heart valve scaffold. Analysis of the heart valve included functional testing in a pulsatile bioreactor under subphysiological and supraphysiological flow and pressure conditions. Using stereolithography, we were able to fabricate plastic models with accurate anatomy of a human valvular homograft. Moreover, we fabricated heart valve scaffolds with a physiologic valve design, which included the sinus of Valsalva, and that resembled our reconstructed aortic root and pulmonary valve. One advantage of P4HB and PHOH was the ability to mold a complete trileaflet heart valve scaffold from a stereolithographic model without the need for suturing. The heart valves were tested in a pulsatile bioreactor, and it was noted that the leaflets opened and closed synchronously under subphysiological and supraphysiological flow conditions. Our preliminary results suggest that the reproduction of complex anatomic structures by rapid prototyping techniques may be useful to fabricate custom made polymeric scaffolds for the tissue engineering of heart valves.     

Controlled cyclic stretch bioreactor for tissue-engineered heart valves

A tissue-engineered heart valve (TEHV) represents the ultimate valve replacement, especially for juvenile patients given its growth potential. To date, most TEHV bioreactors have been developed based on pulsed flow of culture medium through the valve lumen to induce strain in the leaflets. Using a strategy for controlled cyclic stretching of tubular constructs reported previously, we developed a controlled cyclic stretch bioreactor for TEHVs that leads to improved tensile and compositional properties. The TEHV is mounted inside a latex tube, which is then cyclically pressurized with culture medium. The root and leaflets stretch commensurately with the latex, the stretching being dictated by the stiffer latex and thus controllable. Medium is also perfused through the lumen at a slow rate in a flow loop to provide nutrient delivery. Fibrin-based TEHVs prepared with human dermal fibroblasts were subjected to three weeks of cyclic stretching with incrementally increasing strain amplitude. The TEHV possessed the tensile stiffness and stiffness anisotropy of leaflets from sheep pulmonary valves and could withstand cyclic pulmonary pressures with similar distension as for a sheep pulmonary artery.