脂肪间充质干细胞分化为内皮细胞并体外构建组织工程心脏瓣膜

背景：组织工程心脏瓣膜是利用组织工程技术将种子细胞种植于瓣膜支架上所构建的一种人工瓣膜,目前国内外研究主要集中于种子细胞来源及支架选择上。 目的：探讨人脂肪间充质干细胞体外向内皮细胞诱导分化后的细胞作为种子细胞,脱细胞猪主动脉瓣膜作为支架体外构建组织工程心脏瓣膜的可行性。 方法：利用吸脂术采集脂肪组织,分离、培养脂肪间充质干细胞,流式细胞仪鉴定细胞表型;免疫细胞化学方法及RT-PCR检测细胞分化标志物;应用Triton X-100联合胰蛋白酶的方法制备脱细胞猪主动脉瓣支架,将体外培养扩增的诱导分化后的内皮细胞种植于支架上构建组织工程心脏瓣膜,光镜及电镜下观察组织工程心脏瓣膜的组织学结构。 结果与结论：脂肪组织分离培养的脂肪间充质干细胞向内皮细胞诱导分化后表达CD31、CD34、CD144、Ⅷ因子和内皮型一氧化氮合成酶等内皮细胞特异性抗原;脱细胞猪主动脉瓣膜支架脱细胞完全,弹力纤维及胶原纤维保持完整;构建的组织工程心脏瓣膜可见支架上排列连续的单细胞层。提示脂肪间充质干细胞在体外向内皮细胞诱导分化后已初步具有内皮细胞功能,在脱细胞猪主动脉瓣膜支架上生长良好,可以在体外初步构建组织工程心脏瓣膜。     

不同方法制备脱细胞猪主动脉瓣膜支架的组织结构

背景：理想的脱细胞方法要求既能完全去除供体细胞,降低免疫原性,又能保留天然瓣膜的胶原纤维、弹力纤维等细胞外基质成分,以保持足够的机械强度。 目的：采用不同洗剂制备脱细胞猪主动脉瓣膜支架,对比其组织结构,探讨最为有效的脱细胞瓣膜支架制备方法。 方法：20个新鲜猪主动脉瓣膜随机分为新鲜对照组、去污剂组、酶消化组和去污剂-酶消化组,后3组分别使用Triton X-100、胰蛋白酶以及二者联合的方法制备脱细胞瓣膜支架,对比支架大体形态、苏木精-伊红染色、Mallory-Heidenhain染色和电镜下超微结构的不同。 结果与结论：经脱细胞处理后,去污剂组瓣叶柔软、光滑,苏木精-伊红染色少量核物质存留、纤维排列规整,Mallory-Heidenhain染色胶原纤维和弹性纤维相交错,电镜下呈波浪状排列、原纤维横纹清楚;酶消化组瓣叶局部塌陷,苏木精伊红染色细胞完全去除、纤维排列较紊乱,Mallory-Heidenhain染色胶原纤维和弹性纤维呈网状排列,电镜下纤维部分断裂、原纤维横纹存在;去污剂-酶消化组瓣叶柔软、光滑,苏木精伊红染色细胞完全去除、纤维完整,Mallory-Heidenhain染色胶原纤维和弹性纤维平行排列,电镜下纤维完好,但排列稀疏,原纤维横纹清晰。说明3种方法均可有效去除供体细胞,保持纤维结构相对完整,在完全清除供体细胞并保持纤维支架完整性方面,Triton X-100联合胰蛋白酶的方法更为有效。     

内皮祖细胞作为组织工程瓣膜内皮种子细胞的可行性研究

目的 探讨利用外周血内皮祖细胞（EPC8）制备组织工程瓣膜的可行性。方法 分离人外周血EPCs，采用酶-去垢剂法去除新鲜猪主动脉瓣细胞制备去细胞瓣膜支架，将培养的人外周血EPCs接种到去细胞瓣膜上。结果 经酶-去垢剂法去除新鲜猪主动脉瓣细胞后，细胞成分全部去除，纤维支架保存完好。去细胞处理后瓣膜无明显细胞毒性。人外周血EPCs与去细胞瓣膜共孵育2周后，细胞紧贴瓣膜表面生长形成一层连续的单细胞层，初步生物力学测定示去细胞前与再内皮化后，瓣膜力学特性无明显改变。结论 外周血分离培养扩增得到的EPCs能够再内皮化去细胞猪主动脉瓣构建组织工程瓣膜，外周血EPCs是组织工程瓣膜内皮种子细胞的新的来源。     

组织工程心脏瓣膜的构建：现状与前景

背景：目前临床上应用的心脏生物瓣和机械瓣都存在一些缺陷和不足,而组织工程心脏瓣膜有可能避免这些问题的出现,成为瓣膜替代物的理想选择。 目的：探讨构建组织工程心脏瓣膜的实验研究进展。 方法：应用数据库检索的方法分析关于组织工程心脏瓣膜的实验研究文献,组织工程心脏瓣膜的三大要素为种子细胞、支架材料和细胞种植。 结果与结论：心脏瓣膜修复和置换是目前治疗心脏瓣膜性疾病的主要外科手段。目前,主要用于构建组织工程心脏瓣膜的种子细胞有血管内皮细胞、内皮祖细胞以及骨髓间充质干细胞等。经脱细胞处理的支架具有良好的生物力学性能和组织相容性,细胞种植后支架表面会形成一层连续的细胞层,其构建的组织工程心脏瓣膜是可行的。组织工程心脏瓣膜有着良好的应用前景,但目前还有很多问题需要解决,还处于研究的初级阶段。 中国组织工程研究杂志出版内容重点：组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程全文链接：     

脱细胞心血管组织的生物力学性能评价

目的比较心血管组织在脱细胞前后的生物力学性质。方法分别测试脱细胞处理前后肺动脉壁和瓣叶的强度、应力-应变、应力松弛等生物力学指标并进行比较。结果肺动脉壁在处理前后各指标无明显差异，瓣叶在脱细胞后其强度和应力-应变明显变化，应力-松弛无明显变化。结论脱细胞的心血管组织与天然组织的生物力学性能一致或接近。     

家犬主动脉窦影像分析和主动脉瓣支架瓣膜的研制

目的:观察家犬主动脉窦造影特点,研制可经导管植入主动脉瓣支架瓣膜的可行性.方法:选用健康杂种犬10只,行左心室造影,对主动脉窦部造影影像分析.将新鲜的猪心包经脱细胞处理后给予0.6%戊二醛浸泡36 h,缝合在瓣膜环上,制成主动脉瓣支架瓣膜.并将瓣膜支架经动脉植入家犬心脏主动脉瓣位置.经胸超声评价瓣膜功能.结果:左室造影可清晰显示主动脉窦部宽度和高度;升主动脉直径(1.73±0.15)mm,主动脉窦部直径(2.15+0.24)mm,冠脉开口至窦部的距离(1.12+0.14)mm.经胸超声检查示人工瓣膜瓣叶功能正常,无返流.结论:主动脉瓣支架瓣膜设计合理,功能正常,可用于经导管主动脉瓣膜置换的实验研究.     

组织工程心脏瓣膜细胞生物学研究进展

由于现有的机械瓣和生物瓣仍存在种种不足，如不具备生长性、需抗凝、易感染、不能生长和自我修复等。组织工程心脏瓣膜是一新兴的研究领域，涉及多门学科。构建组织工程心脏瓣膜应包括支架的制作、细胞的种植、瓣膜的体外培养和最终移植人人体。其中种植的细胞是组织工程心脏瓣膜的基本要素。就组织工程心脏瓣膜的细胞生物学研究进展做一综述。     

心脏瓣膜组织工程支架的制备及细胞种植

为了探讨心脏瓣膜组织工程支架的制备及细胞种植 ,我们将新鲜猪心瓣膜用胰蛋白酶及 DNA酶消化去除细胞 ,光镜和电镜观察其结构 ,并将人脐静脉内皮细胞株、猪颈动脉内皮细胞和狗肌成纤维细胞滴种在脱细胞猪心瓣膜上 ,HE和免疫组化染色观察。结果显示胰蛋白酶联合 DNA酶消化去除了所有细胞 ,而瓣膜的三维结构保持完好。种植的人内皮细胞几乎完全覆盖了瓣膜的表面 ,猪内皮细胞在瓣膜上呈斑块状生长 ,狗肌成纤维细胞不但生长于瓣膜表面 ,且渗透到基质内部生长。这表明 ,胰蛋白酶联合 DNA酶消化是一种较为理想的猪瓣膜脱细胞方法 ;人和猪血管内皮细胞及狗肌成纤维细胞均能在猪瓣膜支架上较好地黏附和生长。     

组织工程心脏瓣膜细胞生物学研究进展

由于现有的机械瓣和生物瓣仍存在种种不足,如不具备生长性、需抗凝、易感染、不能生长和自我修复等。组织工程心脏瓣膜是一新兴的研究领域,涉及多门学科。构建组织工程心脏瓣膜应包括支架的制作、细胞的种植、瓣膜的体外培养和最终移植入人体。其中种植的细胞是组织工程心脏瓣膜的基本要素。就组织工程心脏瓣膜的细胞生物学研究进展做一综述。     

脱细胞猪膀胱支架的构建

文题释义： 脱细胞：生物移植或修补材料制备过程中常用到脱细胞方法,脱细胞能很好的保留组织的成分,降低移植或修补后的免疫原性,同时保持良好的机械性能,在临床上得到较为广泛的应用。 猪膀胱支架：利用不同的脱细胞方法去除猪膀胱组织中的细胞后制备为猪膀胱支架,既保留了细胞外基质的完整性,又可以降低移植后排斥反应,为进一步的对组织工程支架材料的研究奠定了可行的基础。 背景：膀胱修补术是目前临床上治疗膀胱缺损的主要方法之一,同源性组织因各种因素的影响来源较少,组织工程膀胱脱细胞基质受到人们越来越多的关注。猪膀胱脱细胞外基质来源广泛,具备天然的细胞外支架结构,成为组织工程膀胱替代材料研究的热点。 目的：研究脱细胞猪膀胱作为组织工程支架材料的可行性。 方法：取新鲜的猪膀胱,联合液氮冻融、十二烷基硫酸钠、胰蛋白酶脱细胞方法制猪膀胱无细胞基质。按不同脱细胞方法分组：①正常对照组：不做任何处理;②实验组：0.6%胰蛋白酶+5%十二烷基硫酸钠(pH 8.0);③脱细胞对照组：分别用0.75%胰蛋白酶(pH 8.0)、1%胰蛋白酶(pH 8.0)、5%十二烷基硫酸钠(pH 7.6)或10%十二烷基硫酸钠(pH 7.6)处理。通过苏木精-伊红染色和VG染色、DNA定量、α-Gal抗原检测,观察猪膀胱脱细胞效果。 结果与结论：苏木精-伊红染色显示实验组猪膀胱细胞成分已基本去除;VG染色显示实验组完整保留了猪膀胱组织的细胞外基质成分;实验组的DNA残留量仅为(49.84±30.13) μg/g,显著低于几个脱细胞对照组(P < 0.05);同时其α-Gal抗原残留也明显低于脱细胞对照组。提示：应用0.6%胰蛋白酶+5%十二烷基硫酸钠处理猪膀胱,在完整保留猪膀胱组织细胞外基质的同时,可有效去除其细胞成分,为构建脱细胞猪膀胱支架提供一定参考价值。 ORCID: 0000-0003-1004-7390(李芹) 中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程     

利用人骨髓基质干细胞构建组织工程心脏瓣膜的体外实验

BACKGROUND:Nowadays, mechanical or biological valve recipients used in the clinic are still at the risk of infection, hemorrhage, thrombosis and reoperation owing to valve stenosis. Tissue-engineered heart valve with biological activity can overcome the disadvantages above. While, the optimal choice of scaffolds and seeding cells remains disputable. OBJECTIVE:To explore the feasibility to construct tissue-engineered heart valve with acellularized porcine aortic valve scaffold and human bone marrow stromal stem cells in vitro. METHODS:The porcine aortic valves were decellularized with the detergent and enzymatic extraction process to remove the cellular components. Human bone marrow stromal stem cells were aspirated from sternum of the patients with simple congenital heart malformation, and then the cells were seeded on the acellularized porcine aortic valve scaffold and cultured for 5 days. RESULTS AND CONCLUSION:Flow cytometry identified that the characteristics of surface antigen of the inoculated seed cells were in line with those of human bone marrow stromal stem cells. Light microscopy and electron microscopy confirmed that the cellular components in the porcine aortic valves could be removed to obtain the complete acellular fiber mesh stent. There was no significant difference in biomechanical property between before and after acellularization. The human bone marrow stromal stem cells implanted on the acellularized porcine aortic valve scaffold could form a continuous cell layer on the surfaces of the scaffold. The inoculated bone marrow stromal stem cells could be differentiated into fibroblasts. The implantation of human bone marrow stromal stem cells on the acellularized porcine aortic valve scaffold can construct the tissue-engineered heart valve.     

Mechanical properties of a porcine aortic valve fixed with a naturally occurring crosslinking agent.

The study investigates the mechanical properties of porcine aortic valve leaflets fixed with a naturally occurring crosslinking agent, genipin, at distinct pressure heads. Fresh and the glutaraldehyde-fixed counterparts were used as controls. Subsequent to fixation, the changes in leaflet collagen crimps and its surface morphology were investigated by light microscopy and scanning electron microscopy (SEM). Also, the crosslinking characteristics of each studied group were determined by measuring its fixation index and denaturation temperature. In the mechanical testing, tissue strips made from each studied group were examined in both the circumferential and radial directions. Histological and SEM comparisons between fresh porcine aortic valve leaflet and those fixed at medium or high pressure revealed that the following changes may occur: elimination of the natural collagen crimping, and extensive loss of the endothelial layer. The denaturation temperatures of the glutaraldehyde-fixed leaflets were significantly greater than the genipin-fixed leaflets; however, their fixation indices were comparable. Generally, fixation pressure did not affect the crosslinking characteristics of the genipin- and glutaraldehyde-fixed leaflets. It was found that fixation of porcine aortic valves in genipin or glutaraldehyde did not alter the mechanical anisotropy observed in fresh valve leaflets. This indicated that the intramolecular and intermolecular crosslinks introduced into the collagen fibrils during fixation is of secondary importance to the presence of structural and mechanical anisotropy in fresh leaflet. Tissue fixation in genipin or glutaraldehyde may produce distinct crosslinking structures. However, the difference in crosslinking structure between the genipin- and glutaraldehyde-fixed leaflets did not seem to cause any significant discrepancies in their mechanical properties when compared at the same fixation pressure. Nevertheless, regardless of the crosslinking agent used, changes in mechanical properties and ruptured patterns were observed when the valve leaflets were fixed at distinct pressures.     

Tissue-engineered acellularized valve xenografts: a comparative animal study between plain acellularized xenografts and autologous endothelial cell seeded acellularized xenografts

BACKGROUND: Acellularized valve xenografts are considered a promising way of overcoming the inherent limitations of current prosthetic valves. The aim of this study was to compare the biological responses of an autologous endothelial cell seeded acellularized xenograft (AAX) and a plain acellularized xenograft (PAX) implanted in the pulmonary valve leaflet in the same animal. METHODS: Endothelial cells were isolated and cultured from the jugular vein of goats. Porcine valve leaflets were acellularized with Nacl-SDS, and for AAX, leaflets were then seeded with autologous endothelial cells. A PAX and an AAX were implanted as double pulmonary valve leaflet replacement in the same animal in a goat model (n = 6). After sacrifice, the implanted valve leaflet tissues were retrieved and analyzed visually and under a light microscope. RESULTS AND CONCLUSIONS: Six animals were sacrificed as scheduled during the short-term (6 and 24 hours), mid-term (1 week and 1 month) and long-term (3 and 6 months). Gross and ultrasonographic examinations revealed good valve function with no thrombosis but with slight thickening. Microscopic analysis of the leaflets showed abundant cellular ingrowth into the acellularized leaflets over time. The role of endothelial cell seeding remains controversial. This animal experiment demonstrates the practical feasibility of using acellularized valve xenografts.     

Biomechanical characterization of aortic valve tissue in humans and common animal models

Aortic valve disease develops in an escalating fashion in elderly patients. Current treatments including total valve replacement and valve repair techniques are still suboptimal. A thorough understanding of the animal and human valve tissue properties, particularly their differences, is crucial for the establishment of preclinical animal models and strategies for evaluating new valve treatment techniques, such as transcatheter valve intervention and tissue engineered valves. The goal of this study was to characterize and compare the biomechanical properties and histological structure of healthy ovine, porcine, and human aortic valve leaflets. The biaxial mechanical properties of the aortic valve leaflets of 10 ovine (～1 year), 10 porcine (6-9 months), and 10 aged human (80.6 ± 8.34) hearts were quantified. Tissue microstructure was analyzed via histological techniques. Aged human aortic valve leaflets were significantly less compliant than both ovine and porcine leaflets, with the ovine leaflets being the most compliant. Histological analysis revealed structural differences between the species: the human and porcine leaflets contained more collagen and elastin than the ovine leaflets. Significant mechanical and structural differences in the aortic valve tissues of 6- to 9-month-old porcine models and 1-year-old ovine models with respect to those of aged humans, suggest that these animal models may not be representative of the typical patient undergoing aortic valve replacement.     

Normal Physiological Conditions Maintain the Biological Characteristics of Porcine Aortic Heart Valves: An <Emphasis Type="Italic">Ex Vivo</Emphasis> Organ Culture Study

The aortic valve functions in a complex mechanical environment which leads to force-dependent cellular and tissue responses. Characterization of these responses provides a fundamental understanding of valve pathogenesis. The aim of this work was to study the biological characteristics of native porcine aortic valves cultured in an ex vivo pulsatile organ culture system capable of maintaining physiological pressures (120/80 mmHg) and cardiac output (4.2 l/min). Collagen, sGAG and elastin contents of the valve leaflets were measured and cusp morphology, cell phenotype, cell proliferation and apoptosis were examined. Presence of endothelial cells (ECs) on the leaflet surface was also evaluated. The differences in collagen, sGAG and elastin contents were not significant (p > 0.05) between the cultured and fresh valve leaflets. The cultured valves maintained the native ECM composition of the leaflets while preserving the morphology and cell phenotype. Cell phenotype in leaflets incubated statically under atmospheric conditions decreased compared to fresh and cultured valve leaflets, indicating the importance of mechanical forces in maintaining the natural biology of the valve leaflets. ECs were retained on the surfaces of cultured leaflets with no remodeling of the leaflets. The number of apoptotic cells in the cultured leaflets was significantly (p < 0.05) less than in the statically incubated leaflets and comparable to fresh leaflets. The sterile ex vivo organ culture system thus maintained the viability and native biological characteristics of the aortic valves that were cultured under dynamic conditions for a period of 48 h.     

Tissue-engineered heart valve leaflets: an effective method of obtaining acellularized valve xenografts

To determine the most effective method of producing the acellularized xenograft heart valve leaflets, we compared pathological findings of the xenograft heart valve leaflets produced by three methods; freeze-thawing, Triton and NaCl-SDS treatment and further analyzed the pattern of endothelial cells seeded onto them. Materials and methods: Two pigs were sacrificed and three pulmonary valve leaflets were harvested from each animal. They were immediately stored in a tissue preservation solution and assigned in one of the three preparation methods for acellularization. Endothelial cells from the jugular vein of a goat were isolated and seeded onto the acellularized xenograft heart valve leaflets. Light and Electron microscopic analyses were performed. Result and conclusion: H & E stain showed that cells were almost absent in the leaflet treated with NaCl-SDS, while cells were partly present in the leaflets treated, one with Triton and the other Freeze-thawing. Transmission microscopic analyses showed cell-free matrix with well preserved collagen architecture under the seeded endothelial cells in the leaflets treated with NaCl-SDS. In conclusion, the valve leaflets treated with NaCl-SDS among three representative methods of acellularization of tissues (freeze-thawing, Triton X-100, and NaCl-SDS) showed the better results than the others in terms of the efficacy of decellularization and response to endothelial cell seeding.     

The role of collagen cross-links in biomechanical behavior of human aortic heart valve leaflets--relevance for tissue engineering

A major challenge in tissue engineering of functional heart valves is to determine and mimic the dominant tissue structures that regulate heart valve function and in vivo survival. In native heart valves, the anisotropic matrix architecture assures sustained and adequate functioning under high-pressure conditions. Collagen, being the main load-bearing matrix component, contributes significantly to the biomechanical strength of the tissue. This study investigates the relationship between collagen content, collagen cross-links, and biomechanical behavior in human aortic heart valve leaflets and in tissue-engineered constructs. In the main loading direction (circumferential) of native valve leaflets, a significant positive linear correlation between modulus of elasticity and collagen cross-link concentration was found, whereas no correlation between modulus of elasticity and collagen content was found. Similar findings were observed in tissue-engineered constructs, where cross-link concentration was higher for dynamically strained constructs then for statically cultured controls. These findings suggest a dominant role for collagen cross-links over collagen content with respect to biomechanical tissue behavior in human heart valve leaflets. They further suggest that dynamic tissue straining in tissue engineering protocols can enhance cross-link concentration and biomechanical function.     

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.     

A knitted, fibrin-covered polycaprolactone scaffold for tissue engineering of the aortic valve

State-of-the-art tissue engineered heart valves are not strong enough to withstand aortic blood pressure levels. When a strong and slowly degrading scaffold is used, the starting position of valvular tissue engineering is a stronger valve and seeded cells are allowed more time to create a strong extracellular matrix. A polycaprolactone knitted patch with leaflets was developed as a valvular scaffold. It was sutured into a tube and covered with fibrin gel. The opening and closing behavior and leakage of knitted scaffolds without cells were studied and compared to those of stentless porcine valves. An MTT test was performed on polycaprolactone and fibrin. A loading device was developed to study the durability of the knitted scaffold. The scaffold showed proper opening and it showed coaptation upon closing, but a 39 +/- 3% (n = 3) leakage, compared to a 8 +/- 1% (n = 3) leakage of tested porcine valves. MTT tests showed that polycaprolactone and fibrin are biocompatible materials. Durability testing of the knitted scaffold (n = 1) did not show rupture after ten million loading cycles. A tissue engineering process that includes cell culture will have to show whether this scaffold, besides mechanically reliable and biocompatible, is suitable to lead to a functional, nonregurgitant, durable aortic valve.     

Time related histopathologic changes of acellularized xenogenic pulmonary valved conduits

A biologically engineered xenogenic heart valve based on the concept of acellularization offers promise as a means of overcoming the limitations of current prosthetic valves. We evaluated the process of repopulation by recipient cells on an acellularized xenograft treated with a saline (NaCl)-sodium dodecyl sulfate (SDS) solution. Porcine pulmonary valved conduits acellularized with NaCl-SDS solution were implanted in the right ventricular outflow tract of goats under cardiopulmonary bypass. The goats were sacrificed at 1 week and at 1, 3, 6, and 12 months after implantation. Echocardiographic evaluations of the valves were performed before sacrifice. Visual inspections and histopathologic examinations of the explanted xenogenic valved conduits were performed. Immunohistochemical staining was used to identify the composition of the recellularized cells. Fibroblasts and myofibroblasts were stained by vimentin and smooth muscle actin, and endothelial cells were stained by factor VII related antigen and CD31. Echocardiographic examinations of implanted acellularized xenogenic pulmonary valves revealed satisfactory function except for mild regurgitation and stenosis. On gross examination, all leaflets were well preserved, and no calcification was observed. Microscopic analysis of the acellularized leaflets showed progressive cellular ingrowth over time. The recellularization started from the leaflet base and progressed toward the free margin. Microthrombi were detected on unrecellularized portions. Inflammatory response was detected in the early phase, although it gradually subsided. At 6 months after implantation, cellular ingrowth by fibroblasts, myofibroblasts, and endothelial cells progressed up to 50% of the leaflet's height and progressively increased to 70% after 12 months. Extracellular matrices were regenerated by repopulating cells on the recellularized portion. This study suggests that acellularized xenogenic porcine valved conduits are biocompatible heart valve prostheses that are repopulated with autologous cells and extracellular matrices over a reasonable span of time and preserve the functional integrity without degeneration or calcification of leaflets.