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PubMed2003

Effect of pulse rate on collagen deposition in the tissue-engineered blood vessel.

The effect of mechanical stimulation on the development of tissue-engineered blood vessels was examined. In particular, three different rates of radial distension were chosen to produce a nonpulsed environment (0 beats per minute [bpm]), an adult heart rate (90 bpm), and a fetal heart rate (165 bpm). Engineered vessels were cultured for an average of 7 weeks. Vessel walls were then analyzed for collagen content and distribution. In addition, extracellular matrix remodeling was assessed through measurement of active matrix metalloproteinase type 1 (MMP-1) and tissue inhibitor of metalloproteinases type 1 (TIMP-1) levels. Vessels grown at a distension rate of 165 bpm had significantly higher collagen levels than those grown under static conditions. MMP-1 and TIMP-1 levels were also higher under pulsed conditions as compared with nonpulsed conditions. For the 90- and 165-bpm conditions, collagen and MMP-1 levels were not significantly different. TIMP-1 levels were significantly elevated at 165 bpm, indicating an increased cellular response to mechanical stimulation. Mechanical forces and their transduction represent a means to enhance the physical properties of artificial blood vessels, possibly by affecting the rate of extracellular matrix deposition and remodeling.

Animals↗

PubMed2006

Porogen-based solid freeform fabrication of polycaprolactone-calcium phosphate scaffolds for tissue engineering.

Drop on demand printing (DDP) is a solid freeform fabrication (SFF) technique capable of generating microscale physical features required for tissue engineering scaffolds. Here, we report results toward the development of a reproducible manufacturing process for tissue engineering scaffolds based on injectable porogens fabricated by DDP. Thermoplastic porogens were designed using Pro/Engineer and fabricated with a commercially available DDP machine. Scaffolds composed of either pure polycaprolactone (PCL) or homogeneous composites of PCL and calcium phosphate (CaP, 10% or 20% w/w) were subsequently fabricated by injection molding of molten polymer-ceramic composites, followed by porogen dissolution with ethanol. Scaffold pore sizes, as small as 200 microm, were attainable using the indirect (porogen-based) method. Scaffold structure and porosity were analyzed by scanning electron microscopy (SEM) and microcomputed tomography, respectively. We characterized the compressive strength of 90:10 and 80:20 PCL-CaP composite materials (19.5+/-1.4 and 24.8+/-1.3 Mpa, respectively) according to ASTM standards, as well as pure PCL scaffolds (2.77+/-0.26 MPa) fabricated using our process. Human embryonic palatal mesenchymal (HEPM) cells attached and proliferated on all scaffolds, as evidenced by fluorescent nuclear staining with Hoechst 33258 and the Alamar Blue assay, with increased proliferation observed on 80:20 PCL-CaP scaffolds. SEM revealed multilayer assembly of HEPM cells on 80:20 PCL-CaP composite, but not pure PCL, scaffolds. In summary, we have developed an SFF-based injection molding process for the fabrication of PCL and PCL-CaP scaffolds that display in vitro cytocompatibility and suitable mechanical properties for hard tissue repair.

Calcium Phosphates↗

PubMed2002

Expression of transforming growth factor beta 1 in mesenchymal stem cells: potential utility in molecular tissue engineering for osteochondral repair.

The feasibility of using gene therapy to treat full-thickness articular cartilage defects was investigated with respect to the transfection and expression of exogenous transforming growth factor (TGF)-beta 1 genes in bone marrow-derived mesenchymal stem cells (MSCs) in vitro. The full-length rat TGF-beta 1 cDNA was transfected to MSCs mediated by lipofectamine and then selected with G418, a synthetic neomycin analog. The transient and stable expression of TGF-beta 1 by MSCs was detected by using immunohistochemical staining. The lipofectamine-mediated gene therapy efficiently transfected MSCs in vitro with the TGF-beta 1 gene causing a marked up-regulation in TGF-beta 1 expression as compared with the vector-transfected control groups, and the increased expression persisted for at least 4 weeks after selected with G418. It was suggested that bone marrow-derived MSCs were susceptible to in vitro lipofectamine mediated TGF-beta 1 gene transfer and that transgene expression persisted for at least 4 weeks. Having successfully combined the existing techniques of tissue engineering with the novel possibilities offered by modern gene transfer technology, an innovative concept, i.e. molecular tissue engineering, are put forward for the first time. As a new branch of tissue engineering, it represents both a new area and an important trend in research. Using this technique, we have a new powerful tool with which: (1) to modify the functional biology of articular tissue repair along defined pathways of growth and differentiation and (2) to affect a better repair of full-thickness articular cartilage defects that occur as a result of injury and osteoarthritis.

Animals↗

PubMedDate not supplied

Tissue engineering of cartilage using a hybrid scaffold of synthetic polymer and collagen.

A biodegradable hybrid scaffold of synthetic polymer, poly (DL-lactic-co-glycolic acid) (PLGA), and naturally derived polymer, collagen, was prepared by forming collagen microsponges in the pores of PLGA sponge. This was then used as the three-dimensional scaffold for tissue engineering of bovine articular cartilage, both in vitro and in vivo. In vitro studies show that hybridization with collagen facilitated cell seeding in the sponge and raised seeding efficiency. Chondrocytes adhered to the collagen microsponges, where they proliferated and secreted extracellular matrices with time, filling the space within the sponge. Hematoxylin and eosin staining revealed that most of the chondrocytes after 4 weeks of culture, and almost all cell types after 6 weeks of culture, maintained their phenotypically rounded morphology. While new tissue formed, the scaffold degraded and lost almost 36.9% of its original weight after 10 weeks. Subcutaneous implantation studies in nude mice demonstrated more homogeneous tissue formation in hybrid sponge than in PLGA sponge. The new tissue formed maintained the original shape of the hybrid sponge. The synthetic PLGA sponge, serving as a skeleton, facilitated easy formation into desired shapes and provided appropriate mechanical strength to define the ultimate shape of engineered tissue. Incorporation of collagen microsponges facilitated cell seeding and homogeneous cell distribution and created a favorable environment for cellular differentiation. The hybrid sponge could therefore represent a promising candidate as a three-dimensional scaffold for articular cartilage tissue engineering.

Animals↗

PubMed2005

Tissue engineered ceramic artificial joint--ex vivo osteogenic differentiation of patient mesenchymal cells on total ankle joints for treatment of osteoarthritis.

Total joint arthroplasty is the common treatment of severe cases of osteoarthritis. However, complications involving failure of the bone-prosthesis interface are significant, especially in ankle arthroplasty. To prevent this complication, we attempted a tissue engineering approach using the mesenchymal cells of the patient. We collected a small amount of fresh bone marrow cells from the patient's iliac crest and expanded the number of mesenchymal cells. We then applied the mesenchymal cells to a ceramic ankle prosthesis and cultured them to form an osteoblasts/bone matrix on the prosthesis. We used tissue engineered prostheses on three patients suffering from ankle arthritis and followed their progress for at least 2 years. Follow-up X-ray examinations revealed early radiodense appearance (bone formation) around the cell-seeded areas of the prostheses about 2 months after the operation after which a stable host bone-prosthesis interface was established. All patients showed high clinical scores after the operation and did not exhibit inflammatory reactions. These preliminary results indicate that the tissue engineering approach using autologous cultured marrow mesenchymal cells might prevent aseptic loosening of the total ankle arthroplasty.

Ankle Joint↗

PubMed2005

[Experimental study on gelatin-chondroitin sulfate-sodium hyaluronate tri-copolymer as novel scaffolds for cartilage tissue engineering].

OBJECTIVE: To fabricate a novel gelatin-chondroitin sulfate-sodium hyaluronate tri-copolymer scaffold and to confirm the feasibility of serving as a scaffold for cartilage tissue engineering. METHODS: Different scaffolds was prepared with gelatin-chondroitin sulfate-sodium hyaluronate tri-copolymer by varying the freezing temperatures (-20 degrees C, - 80 degrees C and liquid nitrogen). Pore size, porosity, inter pores and density were observed with light microscopy and scanning electron microscopy (SEM). The load-stiffness curves were compared between different scaffolds and normal cartilage. The number of MSCs attaching to different scaffolds and the function of cells were also detected with MTT colorimetric microassay. RESULTS: The pore size was 300 +/- 45, 230 +/- 30 and 45 +/- 10 microm; the porosity was 81%, 79% and 56%; the density was 9.41+/-0.25, 11.50 +/- 0. 36 and 29.50 +/- 0. 61 microg/mm3 respectively in different scaffolds fabricated at - 20 degrees C, - 80 degrees C and liquid nitrogen; the latter two scaffolds had nearly the same mechanical property with normal cartilage; the cell adhesion rates were 85.0%, 87.5% and 56.3% respectively in different scaffolds and the scaffolds can mildly promote the proliferation of MSCs. CONCLUSION: Gelatin-chondroitin sulfate-sodium hyaluronate tri-copolymer scaffold fabricated at -80 degrees C had proper pore size, porosity and mechanical property. It is a novel potential scaffold for cartilage tissue engineering.

Animals↗

PubMed2006

Reference models for mitral valve tissue engineering based on valve cell phenotype and extracellular matrix analysis.

The advance of mitral valve repair techniques through tissue engineering is impeded by the lack of information regarding the cellular and extracellular components of the mitral valve. The present study aims to expand our understanding of the mitral valve structure by analysing the synthesis of extracellular matrix (ECM) proteins and the expression of nitric oxide synthase (NOS). Valvular endothelial cells (VECs) and valvular interstitial cells (VICs) were isolated from porcine mitral valves. Immunochemical staining of ECM components, including type I, II, III, IV and V collagen, laminin, fibronectin, elastin and chondroitin sulphate (CS), was performed on both mitral valve tissue and cell cultures. Reverse transcription polymerase chain reaction and immunochemistry were used to analyse NOS expression in native valve and in culture. Both VECs and VICs synthesised the basement membrane components, laminin and type IV collagen both in vivo and in vitro, amongst other fibrous ECM proteins. Synthesis of type I collagen and CS was absent in VEC cultures. Each cell type had a characteristic profile of NOS expression. VECs synthesised endothelial NOS both in vivo and in vitro, with a minority of VICs expressing neuronal NOS in vitro. The present study reports newly recognised aspects of the mitral valve structure and the in vitro behaviour of mitral valve cell populations based on ECM synthesis and NOS expression. The presented profiles can be used as base tools for the generation of data necessary for the selection of ideal cell sources and for the design of appropriate scaffolds for the development of effective tissue-engineered mitral valves.

Actins↗

PubMed2004

Glutamate signalling and its potential application to tissue engineering of bone.

Mechanical loading of the skeleton is important for maintenance of adequate bone mass and defined mechanical stimuli are highly osteogenic. The identification of mechanoresponsive signalling molecules in bone may allow osteogenic signals to be mimicked. This approach would be useful in the treatment of bone pathologies where the skeleton is too weak to withstand osteogenic forces and to tissue engineering of bone where the mechanical environment of bone cells is disrupted. Glutamate has been implicated as a mediator of mechanical signalling in bone. Evidence for glutamate signalling in bone, its role in mechanotransduction and potential applications of this pathway to tissue engineering of bone is considered in this review. Glutamate receptors, transporters and proteins that regulate glutamate release, are all expressed in bone cells. Glutamate receptor activation affects both osteoblast and osteoclast phenotypes revealing a potential for therapeutic manipulation of glutamate signalling to enhance bone formation. Glutamate transporters contribute to this system by regulating extracellular glutamate concentrations and acting as glutamate-gated ion channels. Artificial regulation of glutamate receptors or transporters may be used to increase the bone forming capacity of osteoblasts. This novel approach may potentially enhance bone tissue engineering strategies.

Animals↗

PubMed1999

In vivo tissue engineering: Part I. Concept genesis and guidelines for its realization.

A loss of function of an organ often represents a life-threatening situation. Transplantations are successful, but "replacement" availability, its compatibility with the host, and subsequent healing often pose serious questions. Tissue engineering, where a carefully prepared scaffold is populated, in vitro, by cells to form an artificial organ, addresses some of the problems mentioned above. Trauma associated with the implant introduction to the host often complicates the process. The novel concept of in vivo tissue engineering which is designed to mediate the healing and tissue regeneration process by providing an in vitro formed porous, microcellular scaffold is proposed. The scaffold (part or entire organ) is then populated by cells either spontaneously (the surrounding cells will spread and populate to inhabit the scaffold) or by cellular augmentation (encapsulated cells are delivered to this in statu nascendi scaffold). Minimally traumatic arthroscopic surgery combined with a unique polymer delivery system is envisioned for the introduction of this implant to a site to be repaired. Such an approach will require the formation of polymer in-situ, in a reasonable time. The scaffold-forming polymers will be, in principle, biodegradable. We propose to utilize biodegradable polyurethane systems for in vivo tissue engineering. Diversity of their structure/property relationships, relative "ease" of their preparation, and excellent biocompatibility predetermine polyurethanes to be the materials of choice. This paper describes the genesis of this concept and potentials for its realization. It is intended to initiate and stimulate discussion among the related scientific disciplines to form a basis for this field. The synthesis, application, and biodegradation of selected polyurethanes and variety of its medical utilization will be discussed in upcoming papers.

Artificial Organs↗

PubMed2002

Interaction of chondrocytes, extracellular matrix and growth factors: relevance for articular cartilage tissue engineering.

UNLABELLED: The abundant extracellular matrix of articular cartilage has to be maintained by a limited number of chondrocytes. Vice versa, the extracellular matrix has an important role in the regulation of chondrocyte function. OBJECTIVE: In this review we discuss the role of the extracellular matrix in the regulation of chondrocyte function and the relevance for cartilage tissue engineering. To reach this goal the international literature on this subject has been searched with a major focus on the last 5 years. RESULTS: Structural matrix macromolecules (e.g. collagen, hyaluronate), but also growth factors (e.g. IGF-I, TGF beta) entrapped in the matrix and released under specific conditions affect chondrocyte behavior. These factors communicate with the chondrocyte via specific membrane receptors. In this way there is a close interaction between the extracellular and intracellular milieu. Articular cartilage has a limited capacity of intrinsic repair, which has resulted in the development of tissue engineering approaches to repair damaged cartilage. Successful application of scaffolds has to take into account the important role of both soluble and insoluble matrix-derived factors in cartilage homeostasis. CONCLUSION: Functional tissue engineering will only be realized when the scaffolds used will provide cartilage cells with the correct extracellular signals.

Annexins↗

PubMed2001

[Advances of construction of tissue engineered blood vessels].

OBJECTIVE: To discuss the approaches of tissue engineered blood vessels (TEBV) reconstruction. METHODS: The recent literatures about TEBV were widely reviewed. We summarized various types of biomaterials served as scaffold for TEBV and evaluated the construction model of TEBV. And the biological properties of some TEBV were compared. RESULTS: Although the final model of construction of TEBV was not clear, reports in the last two years had shown several important advances in this exciting field. CONCLUSION: Mimicry of some or all of the properties of three layers of natural healthy blood vessels has been the strategy of all TEBV approaches.

Biocompatible Materials↗

PubMed2002

Scaffolds for tissue engineering of cartilage.

Articular cartilage lesions resulting from trauma or degenerative diseases are commonly encountered clinical problems. It is well-established that adult articular cartilage has limited regenerative capacity, and, although numerous treatment protocols are currently employed clinically, few approaches exist that are capable of consistently restoring long-term function to damaged articular cartilage. Tissue engineering strategies that focus on the use of three-dimensional scaffolds for repairing articular cartilage lesions offer many advantages over current treatment strategies. Appropriate design of biodegradable scaffold conduits (either preformed or injectable) allow for the delivery of reparative cells bioactive factors, or gene factors to the defect site in an organized manner. This review seeks to highlight pertinent design considerations and limitations related to the development, material selection, and processing of scaffolds for articular cartilage tissue engineering, evidenced over the last decade. In particular, considerations for novel repair strategies that use scaffolds in combination with controlled release of bioactive factors or gene therapy are discussed, as are scaffold criteria related to mechanical stimulation of cell-seeded constructs. Furthermore, the subsequent impact of current and future aspects of these multidisciplinary scaffold-based approaches related to in vitro and in vivo cartilage tissue engineering are reported herein.

Animals↗

PubMed2006

Application of stem cells for cardiovascular grafts tissue engineering.

Congenital and acquired heart diseases are leading causes of morbidity and mortality world-wide. Currently, the synthetic materials or bioprosthetic replacement devices for cardiovascular surgery are imperfect and subject patients to one or more ongoing risks including thrombosis, limited durability and need for reoperations due to lack of growth in children and young adults. Suitable replacement grafts should have appropriate characteristics, including resistance to infection, low immunogenicity, good biocompatability and thromboresistance, with appropriate mechanical and physiological properties. Tissue engineering is a new scientific field aiming at fabrication of living, autologous grafts having structure or function properties that can be used to restore, maintain or improve tissue function. The use of autologous stem cells in cardiovascular tissue engineering is quite promising due to their capacity of self-renewal, high proliferation, and differentiation into specialized progeny. Progress has been made in engineering the various components of the cardiovascular system, including myocardial constructs, heart valves, and vascular patches or conduits with autologous stem cells. This paper will review the current achievements in stem cell-based cardiovascular grafts tissue engineering, with an emphasis on its clinical or possible clinical use in cardiovascular surgery.

Animals↗

PubMed2005

In vitro tissue engineering of a cardiac graft using a degradable scaffold with an extracellular matrix-like topography.

OBJECTIVE: Cardiac tissue engineering has been proposed as a treatment to repair impaired hearts. Bioengineered cardiac grafts are created by combining autologous cell transplantation with a degradable scaffold as a temporary extracellular matrix. Here we present a system for engineered myocardium combining cultured cardiomyocytes and a novel biodegradable scaffold with a unique extracellular matrix-like topography. METHODS: Cardiomyocytes were harvested from neonatal rats and cultured in vitro on biodegradable electrospun nanofibrous poly(epsilon-caprolactone) meshes. Between days 5 and 7, the meshes were overlaid to construct 3-dimensional cardiac grafts. On day 14 of in vitro culture, the engineered cardiac grafts were analyzed by means of histology, immunohistochemistry, and scanning electron microscopy. RESULTS: The cultured cardiomyocytes attached well to the meshes, and strong beating was observed throughout the experimental period. The average fiber diameter of the scaffold is about 250 nm, well below the size of an individual cardiomyocyte. Hence the number of cell-cell contacts is maximized. Constructs with up to 5 layers could be formed without any incidence of core ischemia. The individual layers adhered intimately. Morphologic and electrical communication between the layers was established, as verified by means of histology and immunohistochemistry. Synchronized beating was also observed. CONCLUSIONS: This report demonstrates the formation of thick cardiac grafts in vitro and the versatility of biodegradable electrospun meshes for cardiac tissue engineering. It is envisioned that cardiac grafts with clinically relevant dimensions can be created by using this approach and combining it with new technologies to induce vascularization.

Animals↗

PubMed2006

Biomaterials-based tissue engineering and regenerative medicine solutions to musculoskeletal problems.

Tissue engineering and regenerative medicine offer solutions to a number of compelling clinical problems that have not been adequately addressed through the use of permanent replacement devices. The challenge will be to select the optimal combination of a biomaterial scaffold, cells, and soluble regulators for a particular clinical problem. For many connective tissues of the musculoskeletal system, with microstructures that reflect the mechanical environment, it may be more advantageous to regenerate the tissue (in such a way that they retain their phenotypic characteristics) and in the genetic engineering that has led to the production of growth factors and cloning of their genes.

Biocompatible Materials↗

PubMed2006

Endocrine cell and nerve regeneration in autologous in situ tissue-engineered small intestine.

BACKGROUND: The purpose of this study was to regenerate a larger size of small intestinal tissue than that of our previous study and to evaluate the regeneration of the endocrine cells (ECC) and nerve system of autologous tissue-engineered small intestine. The effect of implantation of large numbers of smooth muscle cells (SMC) for the regeneration of small intestine was also investigated. METHODS: Two types of scaffolds with different cell densities were fabricated: low density (LD) of SMC in the scaffold and high density (HD) of SMC in the scaffold. Both scaffolds were implanted into defects of isolated ileum in a canine model. Animals were sacrificed at 8, 12, 18, and 24 weeks. RESULTS: The area of engineered small intestine in the HD group was four times larger than that in the LD group, although that was smaller in size than the original size of the defect. There were no significant changes in the thickness of regenerated smooth muscle layer (SML) in the LD and HD groups. The numbers of endocrine cells gradually increased after implantation. At 18 weeks of regeneration, the number of ECC reached levels comparable to that of normal mucosa. The nerve fibers extended to the center of the graft area and were observed in regenerated SML and regenerated villi at 24 weeks. CONCLUSIONS: The ECC and nerve fibers were regenerated in autologous in situ tissue-engineered small intestine. Seeding a large number of SMC was not sufficient for the regeneration of the small intestine in a tubular configuration.

Animals↗

PubMed2003

Tissue-engineered heart valve on acellular aortic valve scaffold: in-vivo study.

The feasibility of constructing a tissue-engineered heart valve on an acellular porcine aortic valve leaflet was evaluated. A detergent and enzymatic extraction process was developed to remove the cellular components from porcine aortic valves. The acellular valve leaflets were seeded for 7 days in vitro with cells from canine arterial wall and endothelial cells. The constructs were implanted into the lumens of 6 canine abdominal aortas to assess the reconstruction of the valve leaflets. It was found that all cellular components had been removed from the porcine aortic valves. The valve leaflets were completely reconstructed at the end of the 10th week in vivo. Scanning electron microscopy showed that the valve leaflets were partially covered with endothelial cells. It was concluded that porcine aortic valves can be decellularized by the detergent and enzymatic extraction process and it is feasible to construct a tissue-engineered heart valve in vivo on an acellular valve scaffold.

Animals↗

PubMed1999

[Interaction of cell adhesion to materials in tissue engineering].

OBJECTIVE: To investigate the adhesive interactions of cells with materials and the effects of material properties on cell adhesion in tissue engineering. METHODS: By looking up the recent literatures dealt with adhesive interactions of cells with materials and reviewing previous work on the adhesion of tissue-derived cells to materials. RESULTS: The adhesion characteristics of cells to materials not only depend on the nature of materials, including bulk and surface properties, surface modification, surface morphology, net charge, porosity and degradation rate, but also on the expression of cell surface molecules and their interaction with the material. CONCLUSION: The quantitative measure and biophysical mechanisms of cell adhesion to materials might be very important in tissue engineering.

Biocompatible Materials↗