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PubMed2004

Gene delivery from polymer scaffolds for tissue engineering.

The combination of gene therapy with tissue engineering offers the potential to direct progenitor cell proliferation and differentiation into functional tissue replacements. Many approaches to engineering tissue replacements feature a polymer scaffold to create and maintain a space, support cell adhesion, and organize tissue formation. Polymer scaffolds, either natural, synthetic, or a combination of the two, have also been adapted to serve as delivery vehicles for viral and nonviral vectors, which can induce the expression of tissue inductive factors. Gene delivery is a versatile approach, capable of targeting any cellular process through localized expression of tissue inductive factors. The design and application of tissue engineering scaffolds for localized gene transfer are reviewed. Scaffolds are designed either to release the vector into the local tissue environment or maintain the vector at the polymer surface, which is regulated by the effective affinity of the vector for the polymer. Polymeric delivery can enhance gene transfer locally, promote and extend transgene expression, avoid vector distribution to distant tissues, and reduce the immune response to the vector. Scaffolds capable of controlled DNA delivery can provide a fundamental tool for directing progenitor cell function, which has applications with the engineering of numerous types of tissue. The utility of this approach will increase with the development of design parameters that correlate release and transgene expression, and with continued research into the biology of tissue formation.

Cell Culture Techniques↗

PubMed2004

[Research advance of corneal tissue engineering].

OBJECTIVE: To review research progress of corneal tissue engineering. METHODS: The recent articles on corneal tissue engineering focus on source and selection of corneal cells, the effects of growth factors on culture of corneal cells in vitro. The preparation and selection of three-dimensional biomaterial scaffolds and their strong and weak points were discussed. RESULTS: The corneal tissue engineering cells come from normal human corneal cells. The embryo corneal cell was excellent. Several kinds of growth factors play important roles in culture, growth and proliferation of corneal cell, and incorporated into matrix. Growth factors including basic fibroblast growth factor, keratinocyte growth factor, transforming growth factor beta 1 and epidermal growth factor was favor to corneal cell. Collagen, chitosan and glycosaninoglycans were chosen as biomaterial scaffolds. CONCLUSION: Human tissue engineering cornea can be reconstructed and transplanted. It has good tissue compatibility and can be used as human corneal equivalents.

Biocompatible Materials↗

PubMed2001

Tissue engineering research in oral implant surgery.

In this article, we introduce some of the more extensively evaluated technologies using concepts of tissue engineering. We report on hard tissue engineering and soft tissue engineering and their utility for dental implant therapy. For hard tissue engineering, we evaluated human recombinant bone morphogenetic protein-2 and marrow mesenchymal stem cells using a model of sinus augmentation procedure in rabbit. We also describe distraction osteogenesis as another category for hard tissue engineering. In addition, we evaluate soft tissue management using cultured epithelial grafting for soft tissue engineering. The results of our tissue regeneration materials and methods in this study are positive. When the tissue engineering materials are used in clinics in the future, implant surgery could be the leading field.

Absorbable Implants↗

PubMed2001

Tissue engineering: the biophysical background.

Tissue engineering is the construction, repair or replacement of damaged or missing tissue in humans and other animals. This engineering may take place within the animal body or as tissue constructs to be made in a bioreactor for later grafting into the animal. The minimal set of materials for this are the appropriate types of cell. Usually, however, non-living substrata are used as well. These substrata may be nothing more than materials that bulk up any voids in the damaged tissue and provide the mechanical strength that has been lost when the tissue is damaged or removed. They may serve a similar pair of functions in the bioreactor. They can do much more in terms of pattern formation. The orientations and morphology of the cells, the arrangement of intercellular material as it is laid down and the relationships between different cell types in the repairing or construct tissue are all of importance, for these should resemble the correct normal tissue as closely as possible. Most of these requirements are ones involving pattern formation. This review discusses the various ways in which tissue pattern can be engineered chiefly from a biophysical standpoint. Unpatterned cells are effectively not tissue. This engineering includes the use of topography on the substrata, chemical patterning of adhesive and other cues for the cells, mechanical force application to cause cell orientation and appropriate synthetic responses and electrical fields. The review also discusses the methods used to impart the appropriate cues to and through the materials which are often biodegradable polymers. The article gives particular attention to regions of research and practice where the involvement of the physicist or biophysicist is of importance.

Animals↗

PubMed2003

Biomimetic materials for tissue engineering.

The development of biomaterials for tissue engineering applications has recently focused on the design of biomimetic materials that are capable of eliciting specific cellular responses and directing new tissue formation mediated by biomolecular recognition, which can be manipulated by altering design parameters of the material. Biomolecular recognition of materials by cells has been achieved by surface and bulk modification of biomaterials via chemical or physical methods with bioactive molecules such as a native long chain of extracellular matrix (ECM) proteins as well as short peptide sequences derived from intact ECM proteins that can incur specific interactions with cell receptors. The biomimetic materials potentially mimic many roles of ECM in tissues. For example, biomimetic scaffolds can provide biological cues for cell-matrix interactions to promote tissue growth, and the incorporation of peptide sequences into materials can also make the material degradable by specific protease enzymes. This review discusses the surface and bulk modification of biomaterials with cell recognition molecules to design biomimetic materials for tissue engineering. The criteria to design biomimetic materials such as the concentration and spatial distribution of modified bioactive molecules are addressed. Recent advances for the development of biomimetic materials in bone, nerve, and cardiovascular tissue engineering are also summarized.

Amino Acid Sequence↗

PubMed2001

The design of scaffolds for use in tissue engineering. Part I. Traditional factors.

In tissue engineering, a highly porous artificial extracellular matrix or scaffold is required to accommodate mammalian cells and guide their growth and tissue regeneration in three dimensions. However, existing three-dimensional scaffolds for tissue engineering proved less than ideal for actual applications, not only because they lack mechanical strength, but they also do not guarantee interconnected channels. In this paper, the authors analyze the factors necessary to enhance the design and manufacture of scaffolds for use in tissue engineering in terms of materials, structure, and mechanical properties and review the traditional scaffold fabrication methods. Advantages and limitations of these traditional methods are also discussed.

Animals↗

PubMed2005

Tissue engineering with adult stem cells in reconstructive surgery (review).

Tissue engineering is a multidisciplinary field combining biology and engineering along with clinical application to design, manufacture, modify, grow and maintain living tissue. This field has enjoyed tremendous growth in the past 10 years fueled by its potential role in regenerating new tissues and naturally healing injured or diseased organs. Many approaches to tissue engineering have been explored, including ex vivo de novo construction of tissues and strategies of in vivo induction of tissue regeneration. Interventions are hindered by factors such as rejection by the immune system, limited blood supply or morbidity of the donor site. Regardless of the approach, most researchers and clinicians agree that any successful tissue engineering construct will derive from a single unit, the cell. Because the engineering of tissue necessitates a sufficient number of tissue-specific cells with minimal donor site morbidity, a great deal of scientific effort has been directed towards stem cell research and the use of stem cells as a source of cells for new tissues. This review aims at outlining the role of stem cells in tissue engineering, focusing on the use of adult-derived stem cells as applied to the research and practice of plastic surgery.

Animals↗

PubMed2005

Tissue engineering and regenerative medicine: manufacturing challenges.

Tissue engineering and regenerative medicine are interdisciplinary fields that apply principles of engineering and life sciences to develop biological substitutes, typically composed of biological and synthetic components, that restore, maintain or improve tissue function. Many tissue engineering technologies are still at a laboratory or pre-commercial scale. The short review paper describes the most significant manufacturing and bio-process challenges inherent in the commercialisation and exploitation of the exciting results emerging from the biological and clinical laboratories exploring tissue engineering and regenerative medicine. A three-generation road map of the industry has been used to structure a view of these challenges and to define where the manufacturing community can contribute to the commercial success of the products from these emerging fields. The first-generation industry is characterised by its demonstrated clinical applications and products in the marketplace, the second is characterised by emerging clinical applications, and the third generation is characterised by aspirational clinical applications. The paper focuses on the cost reduction requirement of the first generation of the industry to allow more market penetration and consequent patient impact. It indicates the technological requirements, for instance the creation of three-dimensional tissue structures, and value chain issues in the second generation of the industry. The third-generation industry challenges lie in fundamental biological and clinical science. The paper sets out a road map of these generations to identify areas for research.

Cell Culture Techniques↗

PubMed2000

[Bibliometric analysis on tissue engineering research literatures].

OBJECTIVE: To comprehend the progress of tissue engineering research and speculate its developmental trends. METHODS: MEDLINE search was conducted to retrieve the papers published between 1987 to 1999 under the main headings of tissue engineering. Years, nationalities, languages, journals, authors and heading frequencies of 314 papers were analyzed by bibliometrics. RESULTS: Since 1990, the number of tissue engineering research literatures had doubled, and papers between 1998 and 1999 made up 57.96% of the total papers. All papers came from 15 nations, in 6 languages and 140 journals; 64.97% came from United States and 25.79% from England, Netherlands and Germany; 93.95% was in English; 42.04% was published on 15 journals. Vacanti JP and 19 other authors presented 5 to 24 papers. Heading frequencies were cytology 22.89%, transplantation 13.30%, scaffolds and extracellular matrixes 11.72%, implanting 10.60%, polymers 8.91%, potential applications 8.91%, artificial substitutes 6.88%, tissue culture 6.70% and biogenetics 4.96%. CONCLUSION: Tissue engineering literatures mainly come from United States, England, Netherlands and Germany. English is the major language. J Biomed Mater Res and 14 other journals are important journals about tissue engineering research. Vacanti JP and 19 other authors are prolific authors. Cytology, transplantation, scaffolds and extracellular matrixes and implanting are hot topics and key points on tissue engineering research.

Animals↗

PubMed2001

Fibrin gel -- advantages of a new scaffold in cardiovascular tissue engineering.

OBJECTIVE: The field of tissue engineering deals with the creation of tissue structures based on patient cells. The scaffold plays a central role in the creation of 3-D structures in cardiovascular tissue engineering like small vessels or heart valve prosthesis. An ideal scaffold should have tissue-like mechanical properties and a complete immunologic integrity. As an alternative scaffold the use of fibrin gel was investigated. METHODS: Preliminary, the degradation of the fibrin gel was controlled by the supplementation of aprotinin to the culture medium. To prevent tissue from shrinking a mechanical fixation of the gel with 3-D microstructure culture plates and a chemical fixation with poly-L-lysine in different fixation techniques were studied. The thickness of the gel layer was changed from 1 to 3 mm. The tissue development was analysed by light, transmission and scanning electron microscopy. Collagen production was detected by the measurement of hydroxyproline. Injection molding techniques were designed for the formation of complex 3-D tissue structures. RESULTS: The best tissue development was observed at an aprotinin concentration of 20 microg per cc culture medium. The chemical border fixation of the gel by poly-L-lysine showed the best tissue development. Up to a thickness of 3 mm no nutrition problems were observed in the light and transmission electron microscopy. The molding of a simplified valve conduit was possible by the newly developed molding technique. CONCLUSION: Fibrin gel combines a number of important properties of an ideal scaffold. It can be produced as a complete autologous scaffold. It is moldable and degradation is controllable by the use of aprotinin.

Aprotinin↗

PubMed2002

Mechanical stimulation improves tissue-engineered human skeletal muscle.

Human bioartificial muscles (HBAMs) are tissue engineered by suspending muscle cells in collagen/MATRIGEL, casting in a silicone mold containing end attachment sites, and allowing the cells to differentiate for 8 to 16 days. The resulting HBAMs are representative of skeletal muscle in that they contain parallel arrays of postmitotic myofibers; however, they differ in many other morphological characteristics. To engineer improved HBAMs, i.e., more in vivo-like, we developed Mechanical Cell Stimulator (MCS) hardware to apply in vivo-like forces directly to the engineered tissue. A sensitive force transducer attached to the HBAM measured real-time, internally generated, as well as externally applied, forces. The muscle cells generated increasing internal forces during formation which were inhibitable with a cytoskeleton depolymerizer. Repetitive stretch/relaxation for 8 days increased the HBAM elasticity two- to threefold, mean myofiber diameter 12%, and myofiber area percent 40%. This system allows engineering of improved skeletal muscle analogs as well as a nondestructive method to determine passive force and viscoelastic properties of the resulting tissue.

Biocompatible Materials↗

PubMed2000

[Musculoskeletal tissue engineering with resorbable polymers].

Musculoskeletal tissues can present congenital or acquired defects as a result of disease, accidental trauma or iatrogenous causes. This loss of bony substance is traditionally treated by the replacement of bony tissue (grafts or flaps), or by synthetic materials. Each of these methods of treatment, however, entails its specific disadvantages, limitations and complications. The recent approach for treatment of musculoskeletal defects has been the development of the growing of neotissues derived from autogenous cells, and artificial biodegradable matrixes. This method assumed the name "tissue engineering" in the late 1980s. Tissue Engineering, or TE, has employed advances made in the area of cellular culture, intercellular matrix biology, and also, in the area of biomaterial science. TE is an multi-disciplinary approach. Musculoskeletal TE, although in its preliminary stages, should allow access to treatments of the future.

Biocompatible Materials↗

PubMed2001

Dynamics of extracellular matrix production and turnover in tissue engineered cardiovascular structures.

Appropriate matrix formation, turnover and remodeling in tissue-engineered small diameter vascular conduits are crucial requirements for their long-term patency and function. This complex process requires the deposition and accumulation of extracellular matrix molecules as well as the remodeling of this extracellular matrix (ECM) by matrix metalloproteinases (MMPs) and their endogenous inhibitors (TIMPs). In this study, we have investigated the dynamics of ECM production and the activity of MMPs and TIMPs in long-term tissue-engineered vascular conduits using quantitative ECM analysis, substrate gel electrophoresis, radiometric enzyme assays and Western blot analyses. Over a time period of 169 days in vivo, levels of elastin and proteoglycans/glycosaminoglycans in tissue-engineered constructs came to approximate those of their native tissue counter parts. The kinetics of collagen deposition and remodeling, however, apparently require a much longer time period. Through the use of substrate gel electrophoresis, proteolytic bands whose molecular weight was consistent with their identification as the active form of MMP-2 (approximately 64--66 kDa) were detected in all native and tissue-engineered samples. Additional proteolytic bands migrating at approximately 72 kDa representing the latent form of MMP-2 were detected in tissue-engineered samples at time points from 5 throughout 55 days. Radiometric assays of MMP-1 activity demonstrated no significant differences between the native and tissue-engineered samples. This study determines the dynamics of ECM production and turnover in a long-term tissue-engineered vascular tissue and highlights the importance of ECM remodeling in the development of successful tissue-engineered vascular structures.

Animals↗

PubMedDate not supplied

Measurement of DNA biomarkers for the safety of tissue-engineered medical products, using artificial skin as a model.

To test the hypothesis that the process of tissue engineering introduces genetic damage to tissue-engineered medical products, we employed the use of five state-of-the-art measurement technologies to measure a series of DNA biomarkers in commercially available tissue-engineered skin as a model. DNA was extracted from the skin and compared with DNA from cultured human neonatal control cells (dermal fibroblasts and epidermal keratinocytes) and adult human fibroblasts from a 55-year-old donor and a 96-year-old donor. To determine whether tissue engineering caused oxidative DNA damage, gas chromatography/isotope-dilution mass spectrometry and liquid chromatography/isotope-dilution mass spectrometry were used to measure six oxidatively modified DNA bases as biomarkers. Normal endogenous levels of the modified DNA biomarkers were not elevated in tissue-engineered skin when compared with control cells. Next, denaturing high-performance liquid chromatography and capillary electrophoresis-single strand conformation polymorphism were used to measure genetic mutations. Specifically, the TP53 tumor suppressor gene was screened for mutations, because it is the most commonly mutated gene in skin cancer. The tissue-engineered skin was found to be free of TP53 mutations at the level of sensitivity of these measurement technologies. Lastly, fluorescence in situ hybridization was employed to measure the loss of Y chromosome, which is associated with excessive cell passage and aging. Loss of Y chromosome was not detected in the tissue-engineered skin and cultured neonatal cells used as controls. In this study, we have demonstrated that tissue engineering (for TestSkin II) does not introduce genetic damage above the limits of detection of the state-of-the-art technologies used. This work explores the standard for measuring genetic damage that could be introduced during production of novel tissue-engineered products. More importantly, this exploratory work addresses technological considerations that need to be addressed in order to expedite accurate and useful international reference standards for the emerging tissue-engineering industry.

Biomarkers↗

PubMed2005

Fabrication using a rapid prototyping system and in vitro characterization of PEG-PCL-PLA scaffolds for tissue engineering.

In the field of tissue engineering new polymers are needed to fabricate scaffolds with specific properties depending on the targeted tissue. This work aimed at designing and developing a 3D scaffold with variable mechanical strength, fully interconnected porous network, controllable hydrophilicity and degradability. For this, a desktop-robot-based melt-extrusion rapid prototyping technique was applied to a novel tri-block co-polymer, namely poly(ethylene glycol)-block-poly(epsilon-caprolactone)-block-poly(DL-lactide), PEG-PCL-P(DL)LA. This co-polymer was melted by electrical heating and directly extruded out using computer-controlled rapid prototyping by means of compressed purified air to build porous scaffolds. Various lay-down patterns (0/30/60/90/120/150 degrees, 0/45/90/135 degrees, 0/60/120 degrees and 0/90 degrees) were produced by using appropriate positioning of the robotic control system. Scanning electron microscopy and micro-computed tomography were used to show that 3D scaffold architectures were honeycomb-like with completely interconnected and controlled channel characteristics. Compression tests were performed and the data obtained agreed well with the typical behavior of a porous material undergoing deformation. Preliminary cell response to the as-fabricated scaffolds has been studied with primary human fibroblasts. The results demonstrated the suitability of the process and the cell biocompatibility of the polymer, two important properties among the many required for effective clinical use and efficient tissue-engineering scaffolding.

Adult↗

PubMed2003

Clinical applications of tissue engineered constructs.

The reconstruction of soft tissue defects poses a challenge for plastic surgeons and tissue engineers. The construction of a biologically, functionally, and cosmetically successful replacement part will involve the combination of a composite that contains endoderm, mesoderm, and ectoderm. It will be active in immune surveillance and function. It must be durable to withstand the stress and strain encountered by the skin. Such a composite will involve the use of bone, cartilage, muscle, blood vessels, nerves, connective tissue, dermis, and epidermis. Fortunately, many of these tissues are among the best studied by tissue engineers. The future of this field will likely involve to some degree the co-mingling of current reconstructive modalities, including the techniques of prefabrication and pre-lamination, with more aggressive and successful tissue engineering technology and the rapidly developing science of stem cell biology. Tissues synthesized in vitro with better structure, color, and texture can be pre-laminated to a site that has already been prefabricated. Prefabrication of a bio-absorbable matrix can create a well perfused scaffold onto which larger subunits can be prelaminated. The future of this field of endeavor is exciting, and, with further research, experience, and interdisciplinary collaboration, bioengineered tissue constructs will become a reality.

Burns↗

PubMed2006

Development of a model system for preliminary evaluation of tissue-engineered vascular conduits.

BACKGROUND/PURPOSE: The ability to construct tissue-engineered neovessels for use as arterial or venous grafts holds great promise for the advancement of pediatric surgical disciplines. Although the feasibility of tissue engineering vascular grafts has been demonstrated, the long-term function, safety, and efficacy of these grafts as well as their capacity to grow and adapt remain largely unknown. In an attempt to further characterize and develop this technology, we used severe combined immunodeficiency beige (SCID/bg) mouse recipients, chosen because such animals accept xenogenic human cells, to create a small animal model that would allow a rapid and cost-effective preliminary evaluation of the function of tissue-engineered vascular grafts. METHODS: Eight CB-17 SCID/bg female mice underwent vascular graft placement. Four of these mice received aortic interposition grafts, 1 mouse received an inferior vena cava interposition graft, and 3 mice underwent aortocaval graft insertion. All grafts were fashioned from decellularized ovine arteriole tissue engineering scaffolds. Grafts were evaluated for patency using clinical examination, ultrasound interrogation, and micro-computed tomography. Animals were killed at various time points after implantation, and grafts were harvested and analyzed histologically using standard hematoxylin and eosin staining. RESULTS: All grafts were patent based on clinical examination for up to 35 days. Patency was confirmed in 5 grafts using ultrasound interrogation. Patency was confirmed in 4 grafts using micro-computed tomography. One animal that underwent arteriovenous grafting had to be euthanized secondary to high-output cardiac failure on postoperative day 2. The remaining animals were killed between postoperative days 12 and 35. Histologic evaluation of the specimens demonstrated patent grafts with cellular ingrowth into the tissue engineering scaffold. CONCLUSIONS: From these results, we conclude that the use of the SCID/bg mouse model for preliminary evaluation of new tissue engineering methodologies for construction of vascular conduits is feasible. Use of this model has the added advantage of evaluating nonautologous and even xenograft tissues, including human cells.

Animals↗

PubMed2005

Computer-aided tissue engineering of a human vertebral body.

Tissue engineering is developing into a less speculative science involving the careful interplay of numerous design parameters and multidisciplinary professionals. Problem solving abilities and state of the art research tools are required to develop solutions for a wide variety of clinical issues. One area of particular interest is orthopedic biomechanics, a field that is responsible for the treatment of over 700,000 vertebral fractures in the United States alone last year. Engineers are currently lacking the technology and knowledge required to govern the subsistence of cells in vivo, let alone the knowledge to create a functional tissue replacement for a whole organ. Despite this, advances in computer-aided tissue engineering are continually growing. Using a combinatory approach to scaffold design, patient-specific implants may be constructed. Computer-aided design, optimization of geometry using voxel finite element models or other optimization routines, creation of a library of architectures with specific material properties, rapid prototyping, and determination of a defect site using imaging modalities highlight the current availability of design resources. This study proposes a novel methodology from start to finish which could, in the future, be used to design a tissue-engineered construct for the replacement of an entire vertebral body.

Biocompatible Materials↗