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1.
基于RP的组织工程细胞外基质—载体制造问题研究   总被引:2,自引:0,他引:2  
针对组织工程中急需具有适当尺寸的孔隙及孔隙度结构的细胞外基质一载体和组织载体的骨架材料的问题,本引入了目前正在飞速发展的快速成形(RP)技术,应用这种技术可以在短时间内制出既具有精神解剖学形态,又具有适当尺寸的孔隙及孔隙度的三维立体结构的支架,它可以控制载体骨架材料内部微孔的数量,大小,分布及形状,从而提供给细胞的生存以最适宜的空间和营养条件。  相似文献   

2.
采用多孔的 PCL/ PL A共聚膜 ,对材料进行亲水性和细胞亲和性表面加工后 ,用作组织工程载体材料。通过将 5× 10 6 / ml成骨诱导 10 d的 MSCs与 PCL/ PL A共同培养 7d,扫描电镜观察细胞在材料中的生长状态。发现材料作为细胞生长的支持结构与细胞的相容性良好 ,细胞可以黏附在材料的表面和孔隙中。实验证明该材料具有良好的生物相容性 ,可以在体外与骨髓间质干细胞共同构建组织工程化的细胞 -材料复合物用于引导性骨再生。  相似文献   

3.
组织工程用超结构骨架   总被引:1,自引:0,他引:1  
本文从工程角度分析作为细胞载体和组织载体的骨架材料在支撑设计上的结构。超结构工程通过结构元素(如微孔或纤维等)以改变细胞间接触的顺序,而提供给细胞的生存以最适宜的空间和营养条件。这在以下几种骨架设计中体现出来:三维织状纤维超结构作为骨合成植入物的最适宜骨架:新型可注射性开孔植入系统;血管极座(angiopolar)非降解性陶瓷载体;可注射性或可微手术植入型缠结载体系统。以下讨论到这些骨架在组织工程上的应用。  相似文献   

4.
组织工程用超结构骨架   总被引:5,自引:1,他引:4  
本文从工程角度分析作为细胞载体和组织载体的骨架材料在支撑设计上的结构。超结构工程通过结构元素(如微孔或纤维等)以改变细胞间接触的顺序,而提供给细胞的生存以最适宜的空间和营养条件。这在以下几种骨架设计中体现出来:三维织状纤维超结构作为骨合成植入物的最适宜骨架:新型可注射性开孔植入系统;血管极座非降解性陶瓷载体;可注射性或可微手术植入型缠结载体系统。以下讨论到这些骨架在组织工程上的应用。  相似文献   

5.
基底材料的拓扑形貌是影响细胞行为的重要因素之一,材料表面微纳米图案化不但可以提供规则的结构模版,用以研究细胞对生长环境的响应特性,而且可以为组织再生用支架和植入性器件的设计提供基础数据.以表面具有微纳米沟槽结构的聚氨酯薄膜为基底材料,选择在促进组织修复和再生中起重要作用的成纤维细胞为模型细胞,通过细胞活性检测和免疫荧光分析,探讨材料表面的微纳米图案结构对成纤维细胞黏附、增殖、形态以及细胞骨架发育的作用.实验结果表明,微纳米沟槽结构能够明显促进成纤维细胞在材料表面的黏附和增殖,并诱导细胞响应所生长的微纳米沟槽结构进行骨架重排.  相似文献   

6.
应用先进快速成形技术(RP)制备32枚粒度均匀(尺寸均为4mm×4mm×4mm)的聚乳酸-聚羟乙酸(PLGA)人工载体,该载体经I型胶原表面修饰后均分为A、B两组。A组载体复合人骨形态发生蛋白-2基因转染(rAAV-hBM P-2)的兔骨髓基质细胞(M SC s,2×104个细胞/枚);B组每枚载体复合等量、同代次、未基因转染M SC s。体外培养第5 d,从两组各取12枚细胞-载体复合物植入裸鼠皮下,术后30 d取材观察。结果发现rAAV-hBM P-2转染的M SC s成功表达目的基因。RP制备的PLGA载体具有良好的空间结构,大孔及材料表面微孔孔径分别为300μm和3~5μm。体外培养3~5 d,两组载体均复合生长着大量种子细胞。皮下埋植30 d,A组植入物形成较为典型的软骨细胞及基质,II型胶原蛋白表达阳性;同期B组植入物无软骨组织形成。A组聚酯材料面积百分率显著低于B组(P<0.01)。结果表明RP结合载体材料表面修饰,能制备出兼具理想孔隙结构和良好生物相容性的组织工程支架载体,该载体高效复合rAAV-hBM P-2转染的M SC s为组织工程软骨构建创造有利条件。  相似文献   

7.
田伟  贾长青  柏树令 《解剖学报》2005,36(3):314-316
目的新型重组骨脱细胞细胞外基质(REAECM)的制备及其细胞相容性的初步检测,为骨组织工程寻找一种新型的细胞外支架提供实验依据。方法应用体外细胞培养技术,对鼠成骨细胞和REAECM体外进行联合培养1-4周,通过相差显微镜、光镜、电镜观察细胞在材料中的生长情况。结果成骨细胞可以在REAECM上发生良好的黏附、增殖,并且可以长入REAECM的孔隙内。结论REAECM可作为构建组织工程骨的一种较好的支架材料,具有网状孔隙结构;在体外和成骨细胞复合培养时表现出良好的细胞相容性,可以作为一种天然的骨组织替代材料。  相似文献   

8.
心肌组织工程支架是心肌组织工程研究的重要内容之一.本文通过模仿天然心肌细胞外基质的材料组成和结构特征,采用"胶原-壳聚糖-黏着"蛋白的材料体系,设计并应用定向凝固后热致相分离的成形工艺,构建具有定向大孔和贯通小孔的管状心肌组织工程支架.扫面电镜观察结果表明,该心肌组织工程支架具有100μm左右定向大孔和10μm左右贯通小孔的孔隙特征,定向结构取向明显.生物学实验显示细胞与支架相容性良好,并可贴附支架沿孔隙生长.本文构建的具有定向结构的管状心肌组织工程支架,具有进一步用于组织工程心肌体外构建的潜力.  相似文献   

9.
生物三维打印技术与水凝胶的完美结合,可为制造复杂结构功能的组织器官提供一种极具吸引力的解决方案。定制打印细胞负载水凝胶类组织的内部结构,可以更好地仿生真实组织器官的三维微环境,对打印后细胞生长、组织形成和功能再生至关重要。但水凝胶理化特性多变,精准打印与设计结构匹配的多孔结构仍然极具挑战。提出基于光学相干层析成像技术(OCT)的生物三维打印细胞负载水凝胶类组织的精准优化方法,通过自制的三维扫频OCT系统无损在线成像打印组织块和定量评价结构参数,迭代降低设计与打印间的结构差异,提高细胞负载水凝胶打印的精准性和稳定性。实验结果表明,基于OCT无损定量表征结果反馈优化打印参数设置,指导打印过程,使得细胞负载水凝胶类组织的结构形态参数与设计值的偏差从40%左右控制到7%以内,包括内部孔隙尺寸、支撑尺寸、孔隙率、表面积、体积五项关键参数;细胞培养两周后的存活率从80%左右显著提高到90%以上。研究表明OCT技术为批量定制细胞负载水凝胶类组织、生物三维打印组织和器官等提供了具有潜力的精准化工具。  相似文献   

10.
目的 观察新型三维支架材料胶原复合梯度磷酸三钙在体外与软骨细胞的相容性和黏附性,评价其作为软骨组织工程支架的可行性.方法 取8周龄新两兰大白兔膝关节软骨,以酶消化法获得高纯度软骨细胞,培养3代后与三维支架材料胶原复合梯度磷酸三钙在体外复合培养.用倒置相差显微镜、HE染色、免疫组织化学及扫描电镜观察软骨细胞形态、Ⅱ型胶原表达及成软骨能力,同时观察支架材料与软骨细胞的相容性.结果 扫描电镜观察显示支架材料具有疏松多孔结构,孔隙结构规则,孔径100~150 μm,材料内部孔与孔之间贯通良好.支架亲水性好.软骨细胞吸附于支架表面,增殖并逐渐顺孔隙迁徙至支架内部,在孔壁贴附良好,表型维持稳定,可分泌细胞外基质.结论 胶原复合梯度磷酸三钙三维支架具有良好的细胞相容性.  相似文献   

11.
The zonal organization of cells and extracellular matrix (ECM) constituents within articular cartilage is important for its biomechanical function in diarthroidal joints. Tissue-engineering strategies adopting porous three-dimensional (3D) scaffolds offer significant promise for the repair of articular cartilage defects, yet few approaches have accounted for the zonal structural organization as in native articular cartilage. In this study, the ability of anisotropic pore architectures to influence the zonal organization of chondrocytes and ECM components was investigated. Using a novel 3D fiber deposition (3DF) technique, we designed and produced 100% interconnecting scaffolds containing either homogeneously spaced pores (fiber spacing, 1 mm; pore size, about 680 microm in diameter) or pore-size gradients (fiber spacing, 0.5-2.0 mm; pore size range, about 200-1650 microm in diameter), but with similar overall porosity (about 80%) and volume fraction available for cell attachment and ECM formation. In vitro cell seeding showed that pore-size gradients promoted anisotropic cell distribution like that in the superficial, middle, and lower zones of immature bovine articular cartilage, irrespective of dynamic or static seeding methods. There was a direct correlation between zonal scaffold volume fraction and both DNA and glycosaminoglycan (GAG) content. Prolonged tissue culture in vitro showed similar inhomogeneous distributions of zonal GAG and collagen type II accumulation but not of GAG:DNA content, and levels were an order of magnitude less than in native cartilage. In this model system, we illustrated how scaffold design and novel processing techniques can be used to develop anisotropic pore architectures for instructing zonal cell and tissue distribution in tissue-engineered cartilage constructs.  相似文献   

12.
In this study, a highly porous collagen-based biodegradable scaffold was developed as an alternative to synthetic, non-degradable corneal implants. The developed method involved lyophilization and subsequent stabilization through N-ethyl-N′-[3-dimethylaminopropyl] carbodiimide/N-hydroxy succinimide (EDC/NHS) cross-linking to yield longer lasting, porous scaffolds with a thickness similar to that of native cornea (500 μm). For collagen-based scaffolds, cross-linking is essential; however, it has direct effects on physical characteristics crucial for optimum cell behavior. Hence, the effect of cross-linking was studied by examining the influence of cross-linking on pore size distribution, bulk porosity and average pore size. After seeding the foam with human corneal keratocytes, cell proliferation, cell penetration into the scaffold and ECM production within the scaffold were studied. After a month of culture microscopical and immunohistochemical examinations showed that the foam structure did not undergo any significant loss of integrity, and the human corneal keratocytes populated the scaffold with cells migrating both longitudinally and laterally, and secreted some of the main constituents of the corneal ECM, namely collagen types I, V and VI. The foams had a layer of lower porosity (skin layer) both at the top and the bottom. Foams had an optimal porosity (93.6%), average pore size (67.7 μm), and chemistry for cell attachment and proliferation. They also had a sufficiently rapid degradation rate (73.6 ± 1.1% in 4 weeks) and could be produced at a thickness close to that of the natural corneal stroma. Cells were seeded at the top surface of the foams and their numbers there was higher than the rest, basically due to the presence of the skin layer. This is considered to be an advantage when epithelial cells need to be seeded for the construction of hemi or full thickness cornea.  相似文献   

13.
In this study, a highly porous collagen-based biodegradable scaffold was developed as an alternative to synthetic, non-degradable corneal implants. The developed method involved lyophilization and subsequent stabilization through N-ethyl-N'-[3-dimethylaminopropyl] carbodiimide/N-hydroxy succinimide (EDC/NHS) cross-linking to yield longer lasting, porous scaffolds with a thickness similar to that of native cornea (500 microm). For collagen-based scaffolds, cross-linking is essential; however, it has direct effects on physical characteristics crucial for optimum cell behavior. Hence, the effect of cross-linking was studied by examining the influence of cross-linking on pore size distribution, bulk porosity and average pore size. After seeding the foam with human corneal keratocytes, cell proliferation, cell penetration into the scaffold and ECM production within the scaffold were studied. After a month of culture microscopical and immunohistochemical examinations showed that the foam structure did not undergo any significant loss of integrity, and the human corneal keratocytes populated the scaffold with cells migrating both longitudinally and laterally, and secreted some of the main constituents of the corneal ECM, namely collagen types I, V and VI. The foams had a layer of lower porosity (skin layer) both at the top and the bottom. Foams had an optimal porosity (93.6%), average pore size (67.7 microm), and chemistry for cell attachment and proliferation. They also had a sufficiently rapid degradation rate (73.6+/-1.1% in 4 weeks) and could be produced at a thickness close to that of the natural corneal stroma. Cells were seeded at the top surface of the foams and their numbers there was higher than the rest, basically due to the presence of the skin layer. This is considered to be an advantage when epithelial cells need to be seeded for the construction of hemi or full thickness cornea.  相似文献   

14.
Porosity has been shown to be a key determinant of the success of tissue engineered scaffolds. A high degree of porosity and an appropriate pore size are necessary to provide adequate space for cell spreading and migration as well as to allow for proper exchange of nutrients and waste between the scaffold and the surrounding environment. Electrospun scaffolds offer an attractive approach for mimicking the natural extracellular matrix (ECM) for tissue engineering applications. The efficacy of electrospinning is likely to depend on the interaction between cells and the geometric features and physicochemical composition of the scaffold. A major problem in electrospinning is the tendency of fibers to accumulate densely, resulting in poor porosity and small pore size. The porosity and pore sizes in the electrospun scaffolds are mainly dependent on the fiber diameter and their packing density. Here we report a method of modulating porosity in three dimensional (3D) scaffolds by simultaneously tuning the fiber diameter and the fiber packing density. Nonwoven poly(ε-caprolactone) mats were formed by electrospinning under various conditions to generate sparse or highly dense micro- and nanofibrous scaffolds and characterized for their physicochemical and biological properties. We found that microfibers with low packing density resulted in improved cell viability, proliferation and infiltration compared to tightly packed scaffolds.  相似文献   

15.
Chang CH  Liu HC  Lin CC  Chou CH  Lin FH 《Biomaterials》2003,24(26):4853-4858
The mechanism by which the cell synthesizes and secretes extracellular matrix (ECM) and is, in turn, regulated by the ECM is termed dynamic reciprocity. The aim of the present work was to produce a gelatin/chondoitin-6-sulfate/hyaluronan tri-copolymer to mimic natural cartilage matrix for use as a scaffold for cartilage tissue engineering. The scaffold produced had a uniform pore size of about 180 microm and adequate porosity of 75%. Porcine chondrocytes were seeded onto the tri-copolymer scaffold and cultured in Petri dishes or spinner flasks for 2, 3, 4, or 5 weeks. Chondrocytes were uniformly distributed in the scaffold in the spinner flask cultures, but less so in the Petri dish cultures. Secretion of ECM was found under histology examination. In spinner flask cultures, chondrocytes retained their phenotype for at least 5 weeks, as shown immunohistochemically, and synthesized type II collagen. These results show that gelatin/chondroitin sulfate/hyaluronan tri-copolymer has potential for use as a cartilage tissue engineering scaffold.  相似文献   

16.
In this study, poly(e-caprolactone)/polyglycolic acid (PCL/PGA) scaffolds for repairing articular cartilage were fabricated via solid-state cryomilling along with compression molding and porogen leaching. Four distinct scaffolds were fabricated using this approach by four independent cryomilling times. These scaffolds were assessed for their suitability to promote articular cartilage regeneration with in vitro chondrocyte cell culture studies. The scaffolds were characterized for pore size, porosity, swelling ratio, compressive, and thermal properties. Cryomilling time proved to significantly affect the physical, mechanical, and morphological properties of the scaffolds. In vitro bovine chondrocyte culture was performed dynamically for 1, 7, 14, 28, and 35 days. Chondrocyte viability and adhesion were tested using MTT assay and scanning electron microscopy micrographs. Glycosaminoglycan (GAG) and DNA assays were performed to investigate the extracellular matrix (ECM) formation and cell proliferation, respectively. PCL/PGA scaffolds demonstrated high porosity for all scaffold types. Morphological analysis and poly(ethylene oxide) continuity demonstrated the existence of a co-continuous network of interconnected pores with pore sizes appropriate for tissue engineering and chondrocyte ingrowth. While mean pore size decreased, water uptake and compressive properties increased with increasing cryomilling times. Compressive modulus of 12, 30, and 60 min scaffolds matched the compressive modulus of human articular cartilage. Viable cells increased besides increase in cell proliferation and ECM formation with progress in culture period. Chondrocytes exhibited spherical morphology on all scaffold types. The pore size of the scaffold affected chondrocyte adhesion, proliferation, and GAG secretion. The results indicated that the 12 min scaffolds delivered promising results for applications in articular cartilage repair.  相似文献   

17.
Previously, a strong and bioactive ceramic scaffold consisting of a porous zirconia body coated with apatite double layers (fluorapatite (FA) as an inner layer and hydroxyapatite (HA) as an outer layer) was successfully fabricated. In this contribution, the authors investigate the in vivo performance of the engineered bioceramic scaffolds using a rabbit calvarial defect model. In particular, the porosity and pore size of the scaffolds are varied in order to observe the geometrical effects of the scaffolds on their bone formation behaviors. The scaffolds supported on a zirconia framework can be produced with an extremely high porosity (approximately 84-87%), while retaining excellent compressive strength (approximately 7-8 MPa), which has been unachievable in the case of pure apatite scaffolds (approximately 74% porosity with approximately 2 MPa strength).The experimental groups used in this study include three types of zirconia scaffolds coated with apatite; high porosity (approximately 87%) with large pore size (approximately 500- 700 microm): AZ-HL, high porosity (approximately 84%) with small pore size (approximately 150-200 microm): AZ-HS, and low porosity (approximately 75%) with large pore size (approximately 500-700 microm): AZ-LL, as well as one type of HA porous scaffold: low porosity (approximately 74%) with a large pore size (approximately 500-700 microm) for the purpose of comparison. The scaffolds prepared with dimensions of approximately 10 mm (diameter) x 1.2 mm (thickness) are grafted in rabbit calvaria defects. The histological sections are made at 4 and 12 weeks after surgery and immunohistochemical analyses are performed on the samples.All of the specimens show a good healing response without adverse tissue reactions. Good healing is shown at 4 weeks post-surgery with the ingrowth of new bone into the macropore-channels of the scaffolds. The newly formed bone amounts to approximately 19.9-24.2% of the initial defect area, depending on the scaffold type, but there is no statistical significance between the scaffold groups. However, the defects without the scaffolds (control group) show a significantly lower bone formation ratio (approximately 4.3%). At twelve weeks after surgery, the extent of new bone formation is more pronounced in all of the scaffold groups. All of the scaffold groups show significantly higher bone formation ratios (26.7-46.9%) with respect to the control without the graft. In the comparison between the scaffold groups, those with high porosities (AZ-HL and AZ-HS) exhibit significantly higher bone formation as compared to the scaffold with low porosity (AZ-LL).Based on the present in vivo test performed within a rabbit calvaria defect model, it is concluded that the apatite-coated zirconia scaffolds show good bone forming ability and are considered to be a promising scaffolding material for bone regeneration since they possess a high level of both mechanical and biological properties.  相似文献   

18.
Porous poly(glycerol sebacate) (PGS) scaffolds were prepared using a salt leaching technique and subsequently surface modified by a low oxygen plasma treatment prior to the use in the in vitro culture of human chondrocytes. Condensation polymerization of glycerol and sebacic acid used at various mole ratios, i.e. 1:1, 1:1.25, and 1:1.5, was initially conducted to prepare PGS prepolymers. Porous elastomeric PGS scaffolds were directly fabricated from the mixtures of each prepolymer and 90% (w/w) NaCl particles and then subjected to the plasma treatment to enhance the surface hydrophilicity of the materials. The properties of both untreated and plasma-treated PGS scaffolds were comparatively evaluated, in terms of surface morphology, surface chemical composition, porosity, and storage modulus using scanning electron microscopy (SEM), X-ray photoelectron spectroscopy, micro-computed tomography, and dynamic mechanical analysis, respectively. The responses of chondrocytes cultured on individual PGS scaffolds were assessed, in terms of cell proliferation and ECM production. The results revealed that average pore sizes and porosity of the scaffolds were increased with an increasing sebacic acid concentration used. The storage moduli of the scaffolds were raised after the plasma treatment, possibly due to the further crosslinking of PGS upon treatment. Moreover, the scaffold prepared with a higher sebacic acid content demonstrated a greater capability of promoting cell infiltration, proliferation, and ECM production, especially when it was plasma-treated; the greatest HA, sGAG, uronic acid, and collagen contents were detected in matrix of this scaffold. The H & E and safranin O staining results also strongly supported this finding. The storage modulus of the scaffold was intensified after incubation with the chondrocytes for 21 days, indicating the accretion and retention of matrix ECM on the cell-cultured scaffold.  相似文献   

19.
Hollister SJ  Maddox RD  Taboas JM 《Biomaterials》2002,23(20):4095-4103
Bone tissue engineering scaffolds must shape regenerating tissue, provide temporary mechanical support and enhance tissue regeneration. These requirements result in conflicting design goals. For example, increased temporary mechanical function requires a dense scaffold while enhanced cell/gene delivery requires a porous scaffold. This paper demonstrates an image-based homogenization optimization approach that can design scaffold microstructure, scaffold material and regenerate tissue microstructure to meet conflicting design requirements. In addition, constraints to ensure adequate cell/gene delivery can be introduced using a minimum porosity threshold. Homogenization theory was used to compute relationships between scaffold microstructure and effective stiffness. The functional relationships were used in the MATLAB optimization toolbox to compute optimal pore dimensions and scaffold material such that the scaffold and regenerate tissue effective stiffness matched that of native bone stiffness. The scaffold design was converted into STL format for solid free-form fabrication. Scaffolds were designed that matched mandibular condyle trabecular bone properties. Results showed excellent agreement between native bone properties and designed scaffold properties (all R2 > 0.89). Finally, example scaffolds were built from hydroxyapatite using a SFF casting technique.  相似文献   

20.
The use of bone grafts for orthopedic applications have increased steadily over the past decade. With improvements in surgical technique, combined with an increasing aged population requiring orthopedic treatment, the need for bone grafts substitutes have also increased. To be useful clinically, the bone graft substitute must be biocompatible, bioabsorbable, and have convenient handling properties. In addition, it must possess a microarchitecture that allows cellular ingrowth and remodeling while simultaneously providing mechanical strength. Poly(propylene fumarate) (PPF) has been investigated as an injectable, biodegradable scaffold for orthopedic applications. Various methods to create a porous, interconnected polymer scaffold are available. The foaming technique is a convenient method to accomplish this task. Reactions between bicarbonate salts and weak acids generate CO(2) gas, causing a bubbling reaction during the polymerization process. This technique allows the porosity of the scaffold to be modulated. Image analysis and mechanical testing of porous PPF fabricated using the foaming technique shows that a highly porous, interconnected scaffold can be produced. At approximately 50% porosity, the scaffold has excellent handling properties, contains pore sizes ranging from 50 to 500 mum with an elastic modulus ranging from 20 to 40 MPa. The foaming technique provides an additional method by which clinically useful polymers can be fabricated for use in various bone tissue engineering applications.  相似文献   

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