Trilayer anisotropic structure versus randomly oriented structure in heart valve leaflet tissue engineering

It has been hypothesized that leaflet substrates with a trilayer structure and anisotropicmechanical properties could be useful for the production of functional and long-lasting tissue-engineered leaflets.To investigate the influence of the anisotropic structural and mechanical characteristics of a substrate on cells,in this study,we electrospun trilayer anisotropic fibrous substrates and randomly oriented isotropic fibrous substrates(used as controls)from polycaprolactone polymers.Consequently,the random substrates had higher radial and lower circumferential tensile properties than the trilayer substrates;however,they had similar flexural properties.Porcine valvular interstitial cells cultured on both substrates produced random and trilayer cell-cultured constructs,respectively.The trilayer cell-cultured constructs had more anisotropic mechanical properties,17%higher cellular proliferation,14%more extracellular matrix(i.e.,collagen and glycosaminoglycan)production,and superior gene and protein expression,suggesting that more cells were in a growth state in the trilayer constructs than in the random constructs.Furthermore,the random and radial layers of the trilayer constructs had more vimentin,collagen,transforming growth factor-beta 1(TGF-ß1),transforming growth factor-beta 3(TGF-ß3)gene expression than in the circumferential layer of the constructs.This study verifies that the differences in structural,tensile,and anisotropic properties of the trilayer and random substrates influence the characteristics of the cells and ECM in the constructs.

Fabrication of a Novel Hybrid Scaffold for Tissue Engineered Heart Valve

The aim of this study was to fabricate biomatrix/polymer hybrid scaffolds using an electrospinning technique. Then tissue engineered heart valves were engineered by seeding mesenchymal stromal cells （MSCs） onto the scaffolds. The effects of the hybrid scaffolds on the proliferation of seed cells, formation of extracellular matrix and mechanical properties of tissue engineered heart valves were investigated. MSCs were obtained from rats. Porcine aortic heart valves were decellularized, coated with poly（3-hydroxybutyrate-co-4-hydroxybutyrate） using an electrospinning technique, and reseeded and cultured over a time period of 14 days. In control group, the decellularized valve scaffolds were reseeded and cultured over an equivalent time period. Specimens of each group were examined histologically （hematoxylin-eosin [HE] staining, immunohistostaining, and scanning electron microscopy）, biochemically （DNA and 4-hydroxyproline） and mechanically. The results showed that recellularization was comparable to the specimens of hybrid scaffolds and controls. The specimens of hybrid scaffolds and controls revealed comparable amounts of cell mass and 4-hydroxyproline （P〉0.05）. However, the specimens of hybrid scaffolds showed a significant increase in mechanical strength, compared to the controls （P〈0.05）. This study demonstrated the superiority of the hybrid scaffolds to increase the mechanical strength of tissue engineered heart valves. And compared to the decellularized valve scaffolds, the hybrid scaffolds showed similar effects on the proliferation of MSCs and formation of extracellular matrix. It was believed that the hybrid scaffolds could be used for the construction of tissue engineered heart valves.

Application of modified polyethylene glycol hydrogels in the construction of tissue engineered heart valve

To enhance the adhesion of seeding-cells to the biomaterial scaffolds, the PEG-hydrogels were modified. Porcine aortic valves were decellularized with Triton X-100 and trypsin. The cells were encapsulated into the PEG-hydrogels to complete the process of the cells attaching to the acellular porcine aortic valves. Herein, the autologous mesenchymal stem cells （MSCs） of goats were selected as the seeding-cells and the tendency of MSCs toward differentiation was observed when the single semilunar TEHV had been implanted into their abdominal aortas. Furthermore, VEGF, TGF-β1, and the cell adhesive peptide motif RGD were incorporated. Light and electron microscopy observations were performed. Analysis of modified PEG-hydrogels TEHV＇s （PEG-TEHV） tensile strength, and the ratio of reendothelial and mural thrombosis revealed much better improvement than the naked acellular porcine aortic valve （NAPAV）. The data illustrated the critical importance of MSC differentiation into endothelial and myofibroblast for remodeling into native tissue. Our results indicate that it is feasible to reconstruct TEHV efficiently by combining modified PEG-hydrogels with acellular biomaterial scaffold andautologous MSCs cells.

Expression of matrix metalloproteinase-9 was effected by epoxy chloropropan on creating tissue engineered heart valves

Objectives To investigate the effects of epoxy chloropropan on the expression of matrix metalloproteinases-9 （MMP-9）in creating tissue engineered heart valves（TEHV）,on the tissue structures of TEHV,and to study the effects of epoxy chloropropan on the calcification of TEHV.Methods The porcine aortic valve leaflets were digested and decellularized by using detergent and trypsin.Those treated with 0.3% glutaraldehyde for 48 hours were the control group;those treated with 3% epoxy choloropropan for 24 hours were the experimental group.The cultured human bone marrow mesenchymal stem cells（hBMSCs）were seeded onto the decellularized scaffolds of TEHV.The histological studies were done with pathological sections and scanning electron microscopy and reverse transcriptase-polymerase chain reaction（RT-PCR）were used to detect the expression of MMP-9.Results In the experimental group.the histology showed that the BMSCs grew well into the pores and formed a confluent layer in decellularized scaffolds;RT-PCR indicated significantly attenuated expressions of MMP-9,compared with the control（P〈0.05）.Conclusion The decellularized porcine aortic valves treated with 3% epoxy chloropropan may inhibit the expression of MMP-9;therefore epoxy chloropropan may prevent the calcification of tissue engineered heart valves.

Application of Decellularized Scaffold Combined with Loaded Nanoparticles for Heart Valve Tissue Engineering in vitro

The purpose of this study was to fabricate decelluarized valve scaffold modified with polyethylene glycol nanoparticles loaded with transforming growth factor-β1（TGF-β1）,by which to improve the extracellular matrix microenvironment for heart valve tissue engineering in vitro.Polyethylene glycol nanoparticles were obtained by an emulsion-crosslinking method,and their morphology was observed under a scanning electron microscope.Decelluarized valve scaffolds,prepared by using trypsinase and TritonX-100,were modified with nanoparticles by carbodiimide,and then TGF-β1 was loaded into them by adsorption.The TGF-β1 delivery of the fabricated scaffold was measured by asing enzyme-linked immunosorbent assay.Whether unseeded or reseeded with myofibroblast from rats,the morphologic,biochemical and biomechanical characteristics of hybrid scaffolds were tested and compared with decelluarized scaffolds under the same conditions.The enzyme-linked immunosorbent assay revealed a typical delivery of nanoparticles.The morphologic observations and biological data analysis indicated that fabricated scaffolds possessed advantageous biocompatibility and biomechanical property beyond decelluarized scaffolds.Altogether this study proved that it was feasible to fabricate the hybrid scaffold and effective to improve extracellular matrix microenvironment,which is beneficial for an application in heart valve tissue engineering.

Development of Biofunctionalized Cellulose Acetate Nanoscaffolds for Heart Valve Tissue Engineering

Currently-used mechanical and biological heart valve prostheses have a satisfactory short-term performance, but may exhibit several major drawbacks on the long-term. Mechanical prostheses, based on carbon, metallic and polymeric components, require permanent anticoagulation treatment, and their usage often leads to adverse reactions, e.g. thromboembolic complications and endocarditis. In recent years, there is a need for a heart valve prosthesis that can grow, repair and remodel. The concept of tissue engineering offers good prospects into the development of such a device. An ideal scaffold should mimic the structural and purposeful profile of materials found in the natural extracellular matrix (ECM) architecture. The goal of this study was to develop cellulose acetate scaffolds (CA) for valve tissue regeneration. After their thorough physicochemical and biological characterization, a biofunctionalization process was made to increase the cell proliferation. Especially, the surface of scaffolds was amplified with functional molecules, such as RGD peptides (Arg-Gly-Asp) and YIGSRG laminins (Tyrosine-Isoleucine-Glycine-Serine-Arginine-Glycine) which immobilized through biotin-streptavidin bond, the strongest non-covalent bond in nature. Last step was to successfully coat an aortic metallic valve with CA biofunctionallized nanoscaffolds and cultivate cells in order to create an anatomical structure comparable to the native valve. Promising results have been obtained with CA-based nanoscaffolds. We found that cells grown successfully on the biofunctionalized valve surface thereby scaffolds that resemble the native tissues, elaborated with bioactive factors such as RGD peptides and laminins not only make the valve’s surface biocompatible but also they could promote endothyliazation of cardiac valves causing an anti-coagulant effect

Amniotic membrane as a potent source of stem cells and a matrix for engineering heart tissue

Existing therapies for the treatment of chronic heart failure still have some limitations and there is a pressing need for the development of new therapeutic modalities. The amniotic membrane has been used for the treatment of various diseases, such as conjunctive defects;however, the mechanisms behind its repair functions are still unclear. Regenerative medicine is seeking newer alternatives and among them, biomaterials have emerged in recent years for developing and manipulating molecules, cells, tissues or organs grown in laboratories in order to replace human body parts. Many such materials have been used for this purpose, either synthetically or biologically, in order to provide new medical devices. This review provides a wider view of the regeneration potential of the use of amniotic membrane as a potential biomaterial to facilitate the implementation of new research in surgical procedures. Amniotic membrane appears to be an alternative source of stem cells as well as an excellent biomaterial for cell-based therapeutic applications in engineering heart tissue.

The recent advance in heart tissue engineering research

Nowadays coronary heart disease has become one of the principal： diseases which harms papular health. In the ease of existing procedures,none of them can thoroughly cure the myocardial damage and cardiac function deterioration caused by coronary heart disease. After 1990s,tbe concept of Heart Tissue Engineering has been proposed. By means of construction of engineering heart tissue in vitro, it can replace the damaged tissue in vivo and improve the cardiac function. The purpose of our review is to describe the principles,advance,and challenges of Heart Tissue Engineering.

Large-Scale Surface Modification of Decellularized Matrix with Erythrocyte Membrane for Promoting In Situ Regeneration of Heart Valve

In situ regeneration is a promising strategy for constructing tissue engineering heart valves(TEHVs).Currently,the decellularized heart valve(DHV)is extensively employed as a TEHV scaffold.Nevertheless,DHV exhibits limited blood compatibility and notable difficulties in endothelialization,resulting in thrombosis and graft failure.The red blood cell membrane(RBCM)exhibits excellent biocompatibility and prolonged circulation stability and is extensively applied in the camouflage of nanoparticles for drug delivery;however,there is no report on its application for large-scale modification of decellularized extracellular matrix(ECM).For the first time,we utilized a layer-by-layer assembling strategy to immobilize RBCM on the surface of DHV and construct an innovative TEHV scaffold.Our findings demonstrated that the scaffold significantly improved the hemocompatibility of DHV by effectively preventing plasma protein adsorption,activated platelet adhesion,and erythrocyte aggregation,and induced macrophage polarization toward the M2 phenotype in vitro.Moreover,RBCM modification significantly enhanced the mechanical properties and enzymatic stability of DHV.The rat models of subcutaneous embedding and abdominal aorta implantation showed that the scaffold regulated the polarization of macrophages into the anti-inflammatory and pro-modeling M2 phenotype and promoted endothelialization and ECM remodeling in the early stage without thrombosis and calcification.The novel TEHV exhibits excellent performance and can overcome the limitations of commonly used clinical prostheses.

Distribution and characteristics of telocytes as nurse cells in the architectural organization of engineered heart tissues

Interstitial Cajal-like cells are a distinct type of interstitial cell with a wide distribution in mammalian organs and tissues,and have been given the name"telocytes".Recent studies have demonstrated the potential roles of telocytes in heart development,renewal,and repair.However,further research on the functions of telocytes is limited by the complicated in vivo environment.This study was designed to construct engineered heart tissue(EHT)as a three-dimensional model in vitro to better understand the role of telocytes in the architectural organization of the myocardium.EHTs were constructed by seeding neonatal cardiomyocytes in collagen/Matrigel scaffolds followed by culture under persistent static stretch.Telocytes in EHTs were identified by histology,toluidine blue staining,immunofluorescence,and transmission electron microscopy.The results from histology and toluidine blue staining demonstrated widespread putative telocytes with compact toluidine blue-stained nuclei,which were located around cardiomyocytes.Prolongations from the cell bodies showed a characteristic dichotomous branching pattern and formed networks in EHTs.Immunofluorescence revealed positive staining of telocytes for CD34 and vimentin with typical moniliform prolongations.A series of electron microscopy images further showed that typical telocytes embraced the cardiomyocytes with their long prolongations and exhibited a marked appearance of nursing cardiomyocytes during the construction of EHTs.This finding highlights the great importance of telocytes in the architectural organization of EHTs.It also suggests that EHT is an appropriate physical and pathological model system in vitro to study the roles of telocytes during heart development and regeneration.

Construction of tissue-engineered heart valves by using decellularized scaffolds and endothelial progenitor cells

Background Tissue-engineered heart valves have the potential to overcome the limitations of present heart valve replacements. This study was designed to develop a tissue engineering heart valve by using human umbilical cord blood-derived endothelial progenitor cells （EPCs） and decellularized valve scaffolds. Methods Decellularized valve scaffolds were prepared from fresh porcine heart valves. EPCs were isolated from fresh human umbilical cord blood by density gradient centrifugation, cultured for 3 weeks in EGM-2-MV medium, by which time the resultant cell population became endothelial in nature, as assessed by immunofluorescent staining. EPC-derived endothelial cells were seeded onto the decellularized scaffold at 3 × 10^6 cells/cm^2 and cultured under static conditions for 7 days. Proliferation of the seeded cells on the scaffolds was detected using the MTT assay. Tissue-engineered heart valves were analyzed by HE staining, immunofluorescent staining and scanning electron microscopy. The anti-thrombogenic function of the endothelium on the engineered heart valves was evaluated by platelet adhesion experiments and reverse transcription-polymerase chain reaction （RT-PCR） analysis for the expression of endothelial nitric oxide synthase （eNOS） and tissue-type plasminogen activator （t-PA）.Results EPC-derived endothelial cells showed a histolytic cobblestone morphology, expressed specific markers of the endothelial cell lineage including von Willebrand factor （vWF） and CD31, bound a human endothelial cell-specific lectin, Ulex Europaeus agglutinin-1 （UEA-1）, and took up Dil-labeled low density lipoprotein （Dil-Ac-LDL）. After seeding on the decellularized scaffold, the cells showed excellent metabolic activity and proliferation. The cells formed confluent endothelial monolayers atop the decellularized matrix, as assessed by HE staining and immunostaining for vWF and CD31. Scanning electron microscopy demonstrated the occurrence of tight junctions between cells forming the confluent monolayer. Platelets adhesion experiments suggested that the neo-endothelium was non-thrombogenic. The expression levels of eNOS and t-PA genes in the neo-endothelium were quite similar to those in human umbilical vein endothelial cells. Conclusions EPCs isolated from the human umbilical cord blood can differentiate into endothelial cells in vitro and form a functional endothelium atop decellularized heart valve scaffolds. Thus, EPCs may be a promising cell source for constructing tissue-engineered heart valves.

Early in vivo experience with tissue-engineered heart valve leaflets from autologous bone marrow-derived cells

Background Currently used heart valve prostheses are associated with anticoagulation complications or limited durability. The advancement of stem cell study and tissue-engineered heart valve research may offer a relatively ideal solution to these problems. Methods Bone marrow was aspirated from sternum of lamb goats to isolate BMCs. Cells were identified by flow cytometry and its capacity of differentiation. Cellular viability was assessed with Rhdomine 123 staining. 1 × 10^7cells were seeded on a patch of PGA sheet. After two-day in vitro culture, autologous cell/ scaffold sheets were used to replace the right posterior pulmonary valve leaflets under cardiopulmonary bypass. The leaflets were explanted at 2 days, 2, 6, 8 and 10 weeks after implantation. The samples were examined macroscopically, histologically, immunohistochemically, and by Scanning Electron Microscope （SEM）. Two goats were implanted with acellular sheets and established as a control group. Results BMCs exhibited fibroblastoid morphology with good viability. Flow cytometry showed negative CD14 and CD45 expression. In vitro cultured BMCs demonstrated the potential to differentiate into adipocytes. The explanted leaflets resembled the characteristics of native extracellular matrix was leaflets macroscopicaIly in the cellular group. Histology showed synthesized and cells were distributed in the single-layered leaflets.Immunohistochemistry revealed positive staining for yon Willebrand factor, α-SMA, vimentin. A confluent cell surface was formed on the explanted TEHLs. No calcium deposited on the leaflets. In control group, the acellular scaffolds were completely degraded, without leaflet remained at 8 weeks. Conclusions It is possible to create tissue-engineered heart valves in vivo using autologous bone marrow-derived cells.

Current progress on scaffolds of tissue engineering heart valves

Tissue engineering heart valves(TEHV)may be the most promising valve substitute,but the study has been relatively stagnant in the recent five years due to the special position,function and mechanical property of heart valves.It is one of the key factors to select an ideal scaffold material in the construction of TEHV.And this article will briefly review the current research and progress on the scaffolds of TEHV,especially based on Chinese works.

In vitro calcification studies on bioprosthetic and decellularized heart valves under quasi-physiological flow conditions

The lifespan of biological heart valve prostheses available in the market is limited due to structural alterations caused by calcium phosphate deposits formed from blood plasma in contact with the tissues.The objective of this work is to present a comparative methodology for the investigation of the formation of calcium phosphate deposits on bioprosthetic and tissue-engineered scaffolds in vitro and the influence of mechanical forces on tissue mineralization.Based on earlier investigations on biological mineralization at constant supersaturation,a circulatory loop simulating dynamic blood flow and physiological pressure conditions was developed.The system was appropriately adapted to evaluate the calcification potential of decellularized(DCV)and glutaraldehyde-fixed(GAV)porcine aortic valves.Results indicated that DCV calcified at higher,statistically nonsignificant,rates in comparison with GAV.This difference was attributed to the tissue surface modifications and cell debris leftovers from the decellularization process.Morphological analysis of the solids deposited after 20 h by scanning electron microscopy in combination with chemical microanalysis electron-dispersive spectroscopy identified the solid formed as octacalcium phosphate(Ca8(PO4)6H2·5H2O,OCP).OCP crystallites were preferentially deposited in high mechanical stress areas of the test tissues.Moreover,GAV tissues developed a significant transvalvular pressure gradient increase past 36 h with a calcium deposition distribution similar to the one found in explanted prostheses.In conclusion,the presented in vitro circulatory model serves as a valuable prescreening methodology for the investigation of the calcification process of bioprosthetic and tissue-engineered valves under physiological mechanical load.

组织工程技术治疗缺血性心脏病的新方向

背景:干细胞疗法是目前研究的热点,已经在各种临床学科中得到应用,但在治疗缺血性心脏病方面收效甚微,主要是因为移植到缺血心肌部位的干细胞存活率较低,新型组织工程材料的发现和应用使人间充质干细胞在心肌缺血区域的存活率得到了提高,这让人们看到了人间充质干细胞在治疗缺血心肌中的广阔应用前景。目的:归纳并总结目前常见的3种不同来源组织工程材料在干细胞治疗缺血性心脏病的研究进展,提出未来的应用展望。方法:应用英文检索词“mesenchymal stem cells,biological hydrogel nanometer material,3D printing,nanostructured material”检索PubMed、Web of Science数据库,应用中文检索词“干细胞,缺血性心脏病,组织工程,水凝胶,3D打印,纳米材料”检索CNKI、万方、维普数据库;检索2010-2021年期间有关组织工程技术治疗缺血性心肌病的相关文献,排除重复研究、个案报告或Meta分析类文章,纳入符合标准的74篇文献进行综述。结果与结论:①可供选择的明胶水凝胶基底材料和制备技术多种多样,同时也是目前研究最成熟的支架材料,是未来最有望率先成为搭载干细胞治疗缺血性心脏病的支架材料。②纳米材料有着独特的理化性质,借助纳米材料构建的支架,不但能促进人间充质干细胞在缺血心肌处的存活和增殖,并且能使其更好地发挥旁分泌作用,从而起到治疗心肌缺血的效果。③新兴的3D打印技术,可以以水凝胶、纳米材料、干细胞为原材料,打印出所需要的细胞、血管、心肌、心室和心房甚至是“完整”心脏,但是距离临床的应用还需进一步发展。④在未来展望方面,大量的体外组织细胞实验有助于建立更加完善的支架体系,寻找最适合人间充质干细胞生存的细胞外基质环境;其次,构建标准化的心肌缺血动物模型有助于在基础研究方面深入了解干细胞在不同支架材料中的生理学表现,而临床应用需要未来多中心大样本的动物实验数据支撑。

可注射水凝胶在心肌梗死中的应用与进展

心肌梗死后随着负性心室重塑的进行,心力衰竭是大量患者难以避免的终末状态,其5年死亡率50%以上。作为一种新型生物材料,越来越多的证据表明可注射水凝胶能抑制梗死后心室重塑,改善心功能,为心肌梗死患者提供了一种新的治疗方式,具有广阔的应用前景。现对可注射水凝胶在心肌梗死治疗中的应用及进展做一综述。

心脏类器官技术要点及特征分析

心脏类器官技术通过在体外3D培养获得具备心脏部分生理及结构特征的细胞结构,是在科研领域对传统的体外2D培养及动物实验技术的重要补充。其在药物筛选、疾病模型构建等多领域存在巨大潜力。但因心脏发育的复杂性,心脏类器官技术的发展相较于其他器官更缓慢。目前已有的心脏类器官技术主要依赖于组织工程技术和干细胞自组织性。本文将就不同心脏类器官技术的特点进行论述,总结已有的技术进展及待完善的方向。

Current state of myocardial tissue engineering

Ischemic heart disease and dilated cardiomyopathy followed by heart failure are a worldwide problem,which seriously challenge clinical outcomes and quality of life of patients. Heart failure is one of the major causes of morbility and mortality. The human heart cannot regenerate significantly because adult cardiomyocytes are terminally differentiated and cannot replicate after injury. The loss of cardiomyocytes accounts for a decrease in myocardial function, which leads to heart failure. Conservative treatment for cardiac conditions has focused on the reduction of workload, and protection from risk factors and has little therapeutic effect on patients in end-stage heart failure. Heart transplantation represents a life-saving and life-extending treatment modality for end-stage heart failure. In spite of advances in surgical techniques, the shortage of availability of donor organs has limited this treatment modality and has prompted researchers to develop alternative approaches. Cardiomyocyte regeneration is a prospective treatment modality, of which, in vitro engineering of myocardial tissue has had promising outcomes that should help cope with failing hearts in the future. Over the past years, much progress has been made to replace infarcted, non-functioning myocardium with newly formed tissue by means of cell-grafting techniques. Our country has made substantial progress in this field and promises a bright future for clinical management of heart failure.

人工瓣环置入和迷宫Ⅲ型手术治疗退行性二尖瓣环扩张伴发心房颤动

背景:对退行性二尖瓣关闭不全合并心房颤动患者,需要在行二尖瓣人工瓣环置入治疗时同期处理心房颤动问题,迷宫Ⅲ型手术虽是治疗心房颤动的金标准,但其安全性遭到质疑。目的:评价人工瓣环置入和迷宫Ⅲ型手术治疗退行性二尖瓣环扩张伴发心房颤动的安全性。方法:纳入退行性二尖瓣关闭不全合并心房颤动患者43例,按随机数字表法分两组治疗,试验组(n=21)施行二尖瓣成形和迷宫Ⅲ型手术,对照组(n=22)仅施行二尖瓣成形。结果与结论:(1)术中安全性指标:试验组体外循环时间、主动脉阻断时间、血红蛋白尿发生率高于对照组(P<0.001),两组尿量无差异。(2)术后安全性指标:试验组房性心律失常发生率、临时起搏器应用比例和住院时间高于对照组(P<0.05),两组间二次开胸止血率、脑卒中发生率无差异。(3)术后疗效:试验组术后当天与出院时的窦性心律转复率均明显高于对照组(P<0.001)。出院时,两组心功能均恢复为NYHAⅠ级或Ⅱ级,超声心动图显示二尖瓣反流均在微量或轻度以下,三尖瓣均关闭良好,两组间左心房内径、左心室内径、左心室容积和射血分数无差异,但试验组跨二尖瓣A峰和跨三尖瓣A峰比例明显高于对照组(P<0.001)。结果说明人工瓣环置入和迷宫Ⅲ型手术治疗退行性二尖瓣瓣环扩张伴发心房颤动在围手术期安全有效。

组织工程心脏瓣膜体外脉动反应器的初步研制

目的:探讨自行研制的体外脉动反应器应用于组织工程心脏瓣膜应力预适应的可行性。方法:根据流体力学原理,采用特殊材料自行设计制作了体外脉动反应器和预适应应力培养系统;将生物瓣置于反应器中运转,观察生物瓣的工作状态和系统的稳定性。以脱细胞猪主动脉瓣作支架,新生牛主动脉内皮细胞作种子细胞构建组织工程心脏瓣叶,静态培养4 d后,置入反应器内进行预适应处理24 h(流量由30 ml/min调至400 ml/min,频率30次/min),观察内皮细胞残留情况,并与静态培养的瓣膜相比较。结果:预适应应力培养系统能产生搏动性液体流,生物瓣正常工作,运转7 d,系统状态稳定。静态培养成功构建组织工程心脏瓣膜,内皮细胞与瓣叶支架黏附紧密,沿瓣叶表面形成单层细胞层,生长状态良好;动态培养的瓣叶内皮细胞部分被冲刷脱落,但仍有少部分细胞(6.26％)残留黏附。结论:自行设计和构建的体外脉动反应器基本能模拟体内心脏瓣膜的血流动力学,用于组织工程心脏瓣膜预适应研究具有可行性。