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J. R. Soc.

Interface (2009) 6, S311–S324
doi:10.1098/rsif.2008.0448.focus
Published online 4 March 2009

REVIEW

Biomaterial technology for tissue
engineering applications
Yasuhiko Tabata*
Department of Biomaterials, Field of Tissue Engineering,
Institute for Frontier Medical Sciences, Kyoto University, 53 Kawara-cho Shogoin,
Sakyo-ku, Kyoto 606-8507, Japan

Tissue engineering is a newly emerging biomedical technology and methodology to assist and
accelerate the regeneration and repairing of defective and damaged tissues based on the
natural healing potentials of patients themselves. For the new therapeutic strategy, it is
indispensable to provide cells with a local environment that enhances and regulates their
proliferation and differentiation for cell-based tissue regeneration. Biomaterial technology
plays an important role in the creation of this cell environment. For example, the biomaterial
scaffolds and the drug delivery system (DDS) of biosignalling molecules have been
investigated to enhance the proliferation and differentiation of cell potential for tissue
regeneration. In addition, the scaffold and DDS technologies contribute to develop the basic
research of stem cell biology and medicine as well as obtain a large number of cells with a high
quality for cell transplantation therapy. A technology to genetically engineer cells for their
functional manipulation is also useful for cell research and therapy. Several examples of tissue
engineering applications with the cell scaffold and DDS of growth factors and genes are
introduced to emphasize the significance of biomaterial technology in new therapeutic and
research fields.

Keywords: biomaterials; drug delivery system; biosignalling molecules; tissue engineering;
tissue regeneration

1. SIGNIFICANCE OF BIOMATERIAL In this clinical situation, a new therapeutic trial, in
TECHNOLOGIES IN TISSUE which disease healing can be achieved based on the
ENGINEERING APPLICATIONS natural healing potential of patients, has been explored.
This trial is termed tissue regeneration therapy where
Advanced surgical therapies currently available consist
of reconstruction surgery and organ transplantation. the regeneration of tissues and organs is naturally
Although there is no doubt that these therapies have induced to therapeutically treat diseases by artificially
saved and improved countless lives, they have several promoting the potential of cell proliferation and
therapeutic and methodological limitations. In the case differentiation. To realize this cell-induced regeneration
of reconstruction surgery, biomedical devices cannot therapy, there are two approaches. One is cell
completely substitute the biological functions even for a transplantation where cells with a high potential of
single tissue or organ, and consequently cannot prevent proliferation and differentiation are transplanted to
the progressive deterioration of injured or damaged induce tissue regeneration based on their potentials.
tissues and organs. One of the biggest issues for organ The other is the therapeutic approach with biomater-
transplantation is the shortage of donor tissues or ials and technologies. In the latter approach, an in vivo
organs. Additionally, the continuous and permanent local environment that enables cells to promote their
use of immunosuppressive agents to prevent immuno- proliferation and differentiation is created by making
logical rejection responses often causes side effects, such use of biomaterials and technologies. If the environ-
as the high possibility of bacterial infection, carcino- ment efficiently manipulates the cells inherently
genesis and virus infection. To resolve these issues in present in the body to enhance the biological potentials
the two advanced therapies, a new therapeutic solution of tissue regeneration, cell-induced natural healing of
that is clinically mild to patients is required. tissues and organs will be achieved without cell
transplantation. This approach is called tissue engin-
*yasuhiko@frontier.kyoto-u.ac.jp eering. This basic concept of biomaterial-based tissue
One contribution of 10 to a Theme Supplement ‘Japanese engineering was originally introduced by Langer &
biomaterials’. Vacanti (1993). Cell scaffold and biosignalling molecule

Received 14 October 2008
Accepted 26 January 2009 S311 This journal is q 2009 The Royal Society

practically possible that combination with the tech. Kuo et al. It is possible that transplanted site and their grafted rate is very low this tissue occupation causes the physical impairment because of their excretion and death. 2003. As one trial to overcome the problem. generally. Biomaterials-based tissue engineering Y. patients to shorten the healing period and suppress the and additionally the natural healing potential of the deterioration process of disease even under inflam- surrounding healthy tissue can be augmented. which consequently rats with liver cirrhosis (Oe et al. in expected as the third therapeutic choice. are surgically applied to the tissue defect Biomaterials need to assist the approach of cell for generation therapy. The bFGF release promoted the cell proliferation third objective is to suppress the progressive deterio. cells infused are hardly accumulated and grafted at the natural healing potential for tissue regeneration still the site to be regenerated. cirrhosis. if the fibrosis can be loosened and digested to disappear To overcome these problems. The intraperitoneal release of hepatocyte growth create new therapeutic strategies as well as increase the factor (HGF) histologically cured the liver fibrosis of therapeutic choice of clinicians. R. it is mation and infection conditions. If the fibrosis can be loosened and digested. 2006. Even in adults. At present. For the former approach. A disadvantage of this J. and has the medulla of chronic renal sclerosis induced the advantages and disadvantages. However. For conventional catheter suffer from other diseases. Tabata dilated cardiomyopathy and chronic nephritis. 2003. the scaffold and the growth factors. Bach natural manner. possible therapy for chronic fibrosis diseases and pared from biomaterials. such as diabetes and treatment. it is problematic. Chen & Mooney 2003. an aneurysm occlusion with blood clots has hyperlipaemia. Bannasch et al. et al. it will enable us to 2003). or their nutrients and oxygen to the transplanted cells. take advantage of the natural healing potentials of ally available.S312 Review. combination. For 2008). A biodegradable brings about large therapeutic benefits for patients who hydrogel could achieve the controlled release of small have not been able to receive clinically effective interfering RNA (siRNA) for transforming growth therapies. medicine (figure 1). even for patients who have been clinically treated. This will enable disease site. but cannot operate naturally for certain transplanted and their consequent poor functions often reasons in disease conditions. We This new regeneration therapy cannot always have demonstrated that the controlled release of a therapeutically substitute the reconstructive surgery matrix metalloproteinase (MMP)-1 plasmid DNA at and organ transplantation clinically available. in chronic fibrosis diseases. regeneration therapy is realized. is generally well known. cause low therapeutic efficacy of cell transplantation. Biomaterials play a key role in based on the natural regeneration potential of the creating the environment for cells. the of healing processes at the disease site. there is no effective medical therapy for Hubbell 2004. There are three objectives of regeneration factor (TGF)-b1 type II receptor and regenerate and therapy. the fibrous accelerate the natural healing of body injury through tissue of excessive collagen fibres and fibroblasts causes promoted angiogenesis or the infiltration and recruit- the impairment of natural healing processes at the ment of key cells at the injured site. In addition. therefore. few cells are retained at the fibroblasts excessively proliferate. For example. 2003). based on the natural healing potential of patients. which et al. been achieved by using coils incorporating basic healing potentials improves the therapeutic efficacy fibroblast growth factor (bFGF. Langer & Tirrell 2004. this is applicable for another always effective in treating patients who are aged or therapy of internal medicine. repair the fibrosis of chronic renal sclerosis (Kushibiki peutic strategy of surgery and internal medicine. The scaffold to surrounding healthy tissue. or cannot be applied because the been clinically performed. 2006). because both approaches are transplantation and enhance the therapeutic efficacy. aneurysm occlusion with tissue organization has nology and methodology to promote cell-based self. it is necessary to give the by any method of drug treatment. promoting cell potentials to induce tissue regeneration. Silva & Mooney chronic fibrosis diseases. while the biomaterial is used defined as ‘tissue regeneration of internal medicine’ as the delivery carrier of biosignalling molecules as the because of the drug treatment application of internal cell nutrients to biologically activate cells. The tissue regeneration strategy will be disease conditions are suppressed by artificially applied to the therapy of internal medicine. However. Tabata delivery technologies with biomaterials have been highly expected that the disease deterioration and demonstrated to create cell environments suitable for progression can be suppressed in a physiologically tissue regeneration (Saltzman & Olbricht 2002. This therapeutic approach is bination with biomaterials enables an angiogenic factor similar to the surgical regeneration therapy where the to efficiently induce in vivo angiogenesis. The first objective is to create a new thera. The 2005). it is highly expected cells an environment suitable to their survival and that the disease site will be regenerated and repaired functional achievement. One of the therapeutic advantages is the ability to For example. In this case. Com. Interface (2009) . This trial is a new and promote the proliferation and differentiation is pre. However. This low grafting rate of cells remains. Soc. sometimes the number of key cells is small and their potential for recurrence of aneurysm due to clot lysis is clinically proliferation and differentiation is low. such as lung fibrosis. The second objective is to The basic idea of tissue regeneration therapy is to enlarge the clinical application of therapies convention. the injured site is normally occupied transplanted into the body by the bolus injection or with fibrous tissue of excessive collagen fibres and the infusion method. it is clinically histological regeneration of kidney structure. cells are these diseases. 2004. inside the aneurysm to allow occlusion by the tissue ration of diseases. The deterioration and progress of organized. Thus. If this tissue contrast to the plasmid DNA solution (Aoyama et al. Kawakami et al. Leo & Grande 2006. which gives cell. Conventional surgical therapy is not patients themselves.

various synthetic and natural proliferation and differentiation (natural scaffold) as materials. it is necessary for better medical poly(L-lactic acid) (PLLA) collagen treatment to combine the conventional therapies with poly(glycolic acid) (PGA) gelatin the tissue regeneration strategy. There are some cases where tissue different manners. However. metals and ceramics except for calcium carbonate since successful tissue regeneration cannot always be and tricalcium phosphate are not biodegradable. Depending on the clinical situation. biomaterials are also useful in the progress material is chemically cross-linked to form a hydrogel of research and development of stem cell biology insoluble in water. Tabata S313 regeneration and repairing of HGF: hepatocyte growth factor chronic fibrosis based on the natural anti-fibrotic therapy of HGF release healing potential promoted by the on dilated cardiomyopathy drug treatment of internal medicine HGF solution HGF released fibrosis tissue fibrotic tissue is loosened or anti-fibrotic therapy of HGF release digested by the drug treatment. generally. R. poly(e-caprolactone) (PCA) fibrin copoly(LL-GA) copoly(LL-CA) hyaluronic acida copoly(LLA-ethylene glycol (EG)) alginatea 2. body tissue is composed of two components: cells and the surrounding environment. at least a few days are Table 1. solubilized by any process in the body to disappear from Biomaterials play a key role in designing and creating the site implanted. the applications. The latter includes the extracellular matrix (ECM) for cell As the biomaterials. such as polymers. the J. chitin TECHNOLOGY AND METHODOLOGY FOR a There are no enzymes in the body to directly degrade these TISSUE ENGINEERING-BASED polymers. Biomaterials-based tissue engineering Y. degraded to generate water-soluble fragments. regeneration. A new therapeutic strategy for chronic fibrotic diseases based on the natural healing potentials of patients themselves. When the cross-linking bond is and medicine. Consequently. point. proper and positive assistance of phenomenon where a material is degraded or water biomaterial technology will be practically promising. and finally disappears. ceramics and metals or the living place of cells and biosignalling molecules as their composites. Review. Soc. (table 1). it cannot be expected that tissue regeneration therapy alone will achieve the synthetic polymers natural polymers rapid healing of wounds or diseases. In addition to therapeutic the molecular weight. FUNDAMENTAL BIOMATERIAL copoly(fumarate-EG) chitosan. biodegradable bio- regeneration is achieved by the single or combinational materials are explained here. On the expected only by their simple combination. Basically. have been investigated and used in the nutrients of cells. The potential is assisted and promoted for tissue regeneration by DDS technology. Biodegradable polymers used for tissue engineering required to induce and activate cell-based tissue of cell scaffold and biosignalling molecule release. From the practical view- use of the components in an appropriate way. therapy is that. it is other hand. There are two ways of material substitutes for ECM and the drug delivery system disappearance. The word ‘biodegradation’ is defined to be the To this end. on liver cirrhosis Tissue regeneration at the fibrotic tissue is achieved HGF solution based on the natural healing DDS potential of surrounding technology healthy tissue HGF released regeneration and repairing of fibrotic tissue Figure 1. the main chain of the material (DDS) of biosignalling molecules to enhance their is hydrolysed or enzymatically digested to decrease biological activities. They are washed out by body fluids to disappear REGENERATIVE THERAPY from the implanted site. some polymers are biodegradable materials necessary to biomedically contrive the way to combine. First. Among them. Interface (2009) . Second.

(iii) with growth factors. (ii) prolongation of signalling molecule lifetime. (iii) absorption acceleration of signalling molecule. Natural polymers of proteins. (iv) signalling molecule targeting. it is difficult derivatives. the mechanical around the implanted scaffold infiltrate into the scaffold strength of materials weakens as their degradation and consequently proliferate and differentiate therein becomes faster. fragments are washed out from the site implanted. Therefore. (iv) with cells and growth factors. but morphogenesis. to induce with natural ones. R. (a) Biomaterials for cell scaffold to induce in vivo tissue regeneration. The synthetic polymers can be modified therapy of cell transplantation. Synthetic that are necessary for biomaterial-based tissue polymers are generally degraded by simple hydrolysis regeneration therapy and the basic stem cell research while natural polymers are mainly degraded enzy. (e) Biomaterials for engineering biological functions of cells. (b) Biomaterials to protect a space and induce angiogenesis for in vivo tissue regeneration. Their support for cells. Biomaterials play an important role in the preparation J. Biomaterials-based tissue engineering Y. Generally. the synthetic polymers are because both the cells and the ECM as well as the hydrophobic and mechanically strong when compared surrounding environment are lost. the which is a three-dimensional scaffold of artificial ECM retention of biomaterials implanted in the body often to initially assist their attachment and subsequent causes the physical impairment of tissue regeneration. Tabata presence of membrane to the cells cannot survive (a) (b) provide a space for cells transplanted without any supplies of bioabsorbable scaffold tissue regeneration nutrients and oxygen defect (i) (ii) fibrous tissue the scaffold is combined to use with cells and/or growth factors depending on the a space providing membrane in-advance angiogenesis in site to be regenerated (iii) (iv) the site to be transplanted fibrous tissue regenerated tissue regenerated tissue space providing space for cell-based tissue regeneration and supply of nutrients and oxygen to cells by angiogenesis (c) (d) modification of (i) target cells (ii) signalling molecule with a water-soluble cells and growth (e) scaffold factors conventional gene transfection from polymer gene transfection DNA-immobilized DNA– carrier substrates free release carrier complex (reverse transfection) signalling of signalling molecule molecule (iii) (iv) normal cells activation of biological cell sheet preparation by function by gene isolation and tissue construct temperature-responding transfection proliferation of stem cells in bioreactor substrate transplantation diseased cells cell-specific cell cell construct biological barrier by cells and recognition transplantation transplantation tissue Figure 2. (ii) with cells. (c) Biomaterials for DDS of biosignalling molecules (growth factors and genes): (i) controlled release of signalling molecule. cross-linking. in other words. The two opposite properties should be if the artificial ECM is biologically compatible. For the purpose of way is to artificially build a local environment for cells. Generally. The natural polymers are normally used in to naturally regenerate and repair a large-size tissue the formulation of hydrogels prepared by chemical defect only by supplying cells to the defective site. There are five key technologies or methodologies resulting in the disappearance of material. inducing cell-based On the other hand. Soc. one possible rates are comparatively slow. Role of biomaterials in tissue engineering-based regeneration therapy. regeneration (figure 2a). ECM is not only a physical polysaccharides and nucleic acids are available. but also provides a natural environ- degree of freedom for property modification is small ment for cell proliferation and differentiation or when compared with that of synthetic polymers. their degradation tissue regeneration at the defective site. that scientifically supports the future regeneration matically. which contributes to cell-based tissue they can be chemically modified to produce various regeneration and organogenesis. Interface (2009) .S314 Review. material application to tissue regeneration. Bioabsorbable scaffold: (i) without cells and growth factors. It is expected that cells residing of materials is also required. an appropriate mechanical strength tissue regeneration. proliferation and differentiation. The first key tech- with ease to change their chemical composition and nology is for the preparation of cell scaffolds to promote molecular weight. balanced by material design and combination. Generally. which affect the physicochemical cell proliferation and differentiation for in vivo tissue property of the materials. (d ) Biomaterials for in vitro cell manipulation to obtain cells and cell constructs for transplantation.

The et al. cell-based tissue regeneration and supply nutrients and In addition to the controlled release. the regeneration of bulky tissue cannot be and genes has been mainly applied to induce tissue always expected. 2006. The release Takahashi et al. When a of signalling molecule stabilization. 2005. The cell scaffold the washout of cell wastes. To expect cell-based of cells to induce tissue and organ regeneration. has been little investigation of DDS-based tissue To prevent tissue ingrowth. tissue regeneration. it is necessary to prepare cells superior to that of matured cells. An appropriate mechanical design is regeneration. growth factors. If the mechanical strength is Although every DDS technology is available for tissue low. The long-term the potential to accelerate cell-induced tissue regen- retention of cell scaffold sometimes causes physical eration. There of a target tissue at the space can no longer be expected. the incorporation of fibres growth factor at the site of action over an extended and ceramic granules enabled biomaterial sponges to period of time is achieved by incorporating the factor into an appropriate biomaterial carrier. The scaffold is a (e. to prolong the activity retention in vivo. R. the controlled release of much required. it is promising to cell biology and medicine is actively ongoing to select induce in vivo angiogenesis by biomaterial technology candidate cells suitable for cell therapy. several stem cells with a high hindrance against the natural process of tissue regen. as one for transplantation therapy. Biomaterials-based tissue engineering Y. the regeneration and repairing logically applied to tissue regeneration therapy. tiation potentials higher than matured cells. Chan & nology is required. their potential for proliferation and differentiation is As mentioned earlier. For example. biosignalling molecules and cell. if there are no supplies of molecules. the supply of cannot always be expected if only the scaffold or the oxygen and nutrients to cells proliferated therein and space-providing membrane is supplied. a new tech- tissue regeneration (Lutolf & Hubbell 2005. It is also mechanically reinforce and enhance in vivo tissue possible that when incorporated in the release carrier. stem cells. Therefore. Tambara et al. once this on fixed ideas and historical background. Once the cell-induced tissue and membrane of biomaterials should be used in regeneration is naturally initiated. in solution into the site to be regenerated is generally a new type of cell scaffold has been investigated to not effective. Basically. 2008. the release technology of growth factors result. the improvement oxygen to cells by angiogenesis (figure 2b). scaffolds is also important. However. it is not practically expected that activity to make use of the DDS technology and the cells transplanted would survive and function well methodology. For Therefore. cells eventually combination with the cells or/and signalling molecules produce the ECM of natural scaffold. transplanted should be physiologically arranged. fibroblasts.b. so-called Takehara et al. the scaffold readily deforms in the body. they are biologically unstable in vivo. basic research of possible strategy to tackle the issue. Tabata S315 of cell scaffolds. cytokines. the scaffold should be porous When the tissue around a defect does not have the and biodegradable. Fernyhough defect as well as the three-dimensional structure. 2005). Considering that drugs applicable for provide a space for tissue regeneration is required. Tabata 2003a. This is one of the typical developed as the only technology or methodology to wound healing processes to temporarily fill and repair a enhance the in vivo efficacy of therapeutic drugs. Interface (2009) . it is necessary for enhanced in vivo biological nutrients and oxygen. Review. Mountford 2008). 2003. the prolongation of body defect is generated. A similar problem is used for DDS applications. tissue regeneration the infiltration of cells into the scaffold. the defect space is generally the in vivo half-life and the targeting to the site of action occupied rapidly by the fibrous tissue produced by will enhance the factor-induced tissue regeneration. For example. Among the and methodology (Tabata & Ikada 1999. carrier should be degraded in the body. regeneration following the implantation of sponges the growth factor is protected against proteolysis so as combined with stem cells ( Hiraoka et al. candidates. potential for proliferation and differentiation are being eration. The pore structure is necessary for inherent potential to regenerate. Soc. which are ubiquitously present in the body Over time. Soto-Gutierrez et al. the site at which the cells are to be generally. This is because the growth factor rapidly combine biomaterials with biological ligands reactive diffuses from the injected site and is enzymatically for cell receptors. Many types of biomaterials have been to induce tissue regeneration. For example. are promising from the viewpoint of cell-based J. Based body defect in case of emergency. it has been ingrowth of fibrous tissue into the space to be thought that DDS cannot be scientifically and techno- regenerated takes place. The application of DDS to observed for the transplantation of stem cells although tissue regenerative therapy will be described later. Although there are some cell scaffold developed initially is a physical entity that cases where a growth factor is required to promote does not have any functions to positively promote tissue regeneration. As a engineering. To enable the growth factor to adhesive substances for artificially promoted cell-based efficiently exert its biological function. 2006. Currently. The regenerative therapy include proteins and genes effec- scaffold (figure 2a) sometimes functions to provide the tive in promoting the proliferation and differentiation space for tissue regeneration. However. This comes in the form of the third Mooney 2008). chemokines) that have temporary platform of cell activities. a barrier membrane to regeneration. 2008). It is a key for successful tissue regeneration to prepared and applied to a tissue defect to induce tissue control the time profile of scaffold biodegradation at the regeneration therein (Salgado et al. upon in vivo administration of biosignalling the transplantation of cells. cells that have proliferation and differen- 2005a.g. The mechanical property of biomaterial key technology of tissue engineering: DDS (figure 2c). digested or deactivated. the direct injection of growth factor the proliferation and differentiation of cells. because it The second key technology is to provide the space for becomes useless after the release function is completed. DDS has been investigated and and can proliferate rapidly.

The three-dimensional substrate can three-dimensional tissue-like construct is prepared be designed and prepared from biomaterials of cyto. Docheva et al. Presently. cytokines. the construct does not always connect culture methods and bioreactors are required (Holtorf with surrounding natural tissue biologically. which are achieved by able. but also to clinical experiments have been begun. The cells are combined function well to induce cell-based tissue regeneration. Kumagai 2002. It is necessary for the genetic engineering of et al. Schaffler & Buchler 2007) have reconstruction by cell culture methods and organ been extensively investigated. non-viral Distinct from the in vitro tissue engineering. isolation. supplied to patients when required. engineering is the substitution of organ functions by the There are some cases where cells transplanted do not use of allo. In addition. Soc. technologies is practically promising to create an poietic stem cells. At present. Kobayashi et al.S316 Review. Kornblum 2007) and the site where tissue regeneration or organ substitution stem cells isolatable from fat tissue (Strem & Hedrick is performed. R. in addition to haemo. chondrogenic. oxygen supplies. 2006. mesenchymal stem cells (MSC) are environment that promotes the proliferation and present in adult bone marrow. induction and in vitro culture difficult to reproduce the in vivo event completely technologies of stem cells are required. 2007. research and development of cell In addition. almost all the approaches of tissue J. vein and bone substitutes for human treatment. 1999) while research of stem cell biology and medicine. space protection and DDS this advantage. Interface (2009) . it can be cell candidates for regenerative medical therapy. Olle et al. which gives important for tissue regeneration. 2007). one of the problems is the shortage of cells cells of blood vessel and bone marrow-derived stem cells clinically available. However. it is practically difficult to allow the construct compatibility. From the viewpoint of nutrients and to survive and function in vivo after grafting. The cell scaffold for in vivo tissue regeneration biological environment to induce cell-based tissue mentioned previously can be used for the purpose of reconstruction is practically impossible. It has been elucidated differentiation of cells for cell-induced tissue regen- that the MSC of adult stem cells have a proliferative eration. 2001) and cubic hydroxyl large number of stem cells of high quality. Shanti et al. If it is possible to realize cell-based tissue regeneration therapy. Takemoto et al. For example. 1991. Another application of in vitro tissue for their functional manipulation and basic biology. In vitro tissue engineering involves tissue 2005. 2008). This com. the However. Yamashita functions. The et al. 2001. adipogenic and myocardial available for cell therapy. The fourth in vitro by using the basic knowledge of biology and technology is for the efficient preparation and prolifer. Many researchers of tissue regeneration 3. Embryonic stem (ES. cells to develop a carrier of gene transfection and cell Biomaterials have been investigated to design and culture system for efficient gene expression. 2003. 2006). 2007. Generally. be classified as either in vitro or in vivo depending on neural stem cells (Hsu et al. However. Tissue engineering for clinical regeneration therapy can 2007. in vitro. clinically use a differentiated type of a patient’s own MSC. medicine or cell culture technologies currently avail- ation of cells (figure 2d ). To prepare pulmonary of drugs. Mountford 2008) and inducible human skin fibroblasts are cultured in a collagen pluripotent stem (iPS) cells (Yamanaka 2007) have sponge to prepare an artificial dermis for skin grafting been established and expected as cell sources for (Kuroyanagi et al. It is technology is also applicable for the basic research of likely that most of the biological components necessary stem cell biology and medicine. have reported ENGINEERING-BASED TISSUE their therapeutic feasibility in tissue regeneration REGENERATION (Leo & Grande 2006. it is difficult to realize in vitro tissue providing a cell culture substrate as the artificial engineering because the artificial arrangement of a ECM. Therefore. Mironov et al. Ehashi et al. Even if a culture substrate. human MSC are isolated and are technology is important not only to develop the basic commercially available (Pittenger et al. it is purpose. CLINICAL ASPECTS OF TISSUE therapy with stem cells. transplantation therapy and the research and development 2005. 2008). their biological functions. If a tissue can be reconstructed in vitro in for cell differentiation and are expected to be promising factories or laboratories on a large scale. immunological rejection will no longer be a question. Based on bination of cell scaffold. For this apatite (Ohgushi & Caplan 1999). create a biological environment that can assist the viral vectors have been scientifically used for gene proliferation and differentiation of cells and maintain transfection because of their high efficiency. especially MSC. Picinich et al. Tabata tissue regeneration. cells are genetically substitute the physiological functions of liver and engineered with biomaterials to activate the biological pancreas (Falqui et al. They can be prepared substitution with functional cells—termed bioartificial from the foetus and adult and have exhibited plasticity hybrid organ. thus. Ichioka et al. Every biomaterial-based cell lineages. Gimble et al. 2007. Biomaterials-based tissue engineering Y. viruses cannot be used to treat patients. are naturally supplied by the body. such as growth factors and knowledge and results for cell therapy. in vivo environment for the construct implanted should The fifth technology is to genetically engineer cells be designed. with an immunoisolation membrane of biomaterials to As one trial to tackle this issue. Fibbe et al. 1996. Both the culturing and genetic engineering of ability and an inherent potential to differentiate into cells are key technologies to prepare cells clinically osteogenic.or xenogeneic cells. 2002. For example. This of the environment to induce tissue regeneration. 2007. it is necessary to develop are cultured in the porous scaffolds of tube-shape a technology or methodology for the preparation of a polylactide (Shin’oka et al. in vivo gene carrier biomaterials are required to be developed tissue engineering is advantageous from the viewpoint from the clinical viewpoint of cell therapy.

because it has been clinically used in medical a low concentration of cells and growth factors. Fujioka a high potential to induce regeneration. Patel et al. extensively investigated to demonstrate the potential to However. fore. Kaken Pharmaceutical Co. There. Physical As a first example. from the viewpoint of examples where in vivo tissue regeneration is achieved therapeutic applications. This result of hydrogel biodegradation. resulting in the formation of new tissue. 2008). or if the regeneration to prepare a biodegradable hydrogel protein release potential of tissue is low as a result of. skin. It has been demonstrated that this activity loss mainly the controlled release of bioactive growth factors over a results from denaturation and deactivation of protein time range of 5 days to three months was possible. 2003b. Zisch et al. Tabata & Ikada 1999). interaction with protein. 2003) or leg is not released by simple diffusion. carrier. One of the largest problems in the release technology Gelatin is easily chemically derivatized to change the of growth factor protein is the loss of biological activity molecular nature. it is necessary to make use of the accompanied with the degradation of carrier hydrogel DDS technology. 1998. cations (Tabata et al. the hydrogel of gelatin of regeneration for cell differentiation and proliferation immobilized the growth factor through the physico- in a controllable fashion. a biodegradable hydrogel. In general. The requirements of a hydrogel polymer angiogenic therapy by using the DDS technology of for this release system are to have biodegradability and growth factor without any cell transplantation. such as requirement described above. Tessmar & Gopferich immature cells infiltrate the matrix of an implanted 2007). it is practically impossible to achieve protein 2008). it is preferable to use bFGF for angiogenic therapy necessary to contrive a method of protein release. and the tissue to be repaired has release of proteins (Andrianov & Payne 1998. 2005b. In such a release angiogenic therapy for leg ischaemia has been clinically system. it will be spray. This is the first report on clinical degradation. to chemical interaction between the gelatin and factor. because the chemical reaction to growth factor and ‘in-advance angiogenesis’ for cell transplantation. Considering protein. As expected. in marked contrast to the injection phase inside the hydrogel and the release is generally of bFGF solution even at higher doses (Ikada & Tabata diffusion controlled through the water pathway of 1998). Using the gelatin hydrogel. there is a limitation for the regulation of induce angiogenesis in different forms of DDS modifi- protein diffusion by the cross-linking density.kaken. 2007. From this viewpoint. Biomaterials-based tissue engineering Y. but only by the ischaemia ( Nakajima et al. 1994. 2004). As described previously. Gombotz & Wee 1998. 2007. Based on the are aged and/or suffer from other diseases. Van Tomme & Hennink 2007). J.co. nerve and fat tissues (Tabata release carrier because of its biosafety and its high 2007. allow a growth factor of in vivo instability to efficiently The growth factor could be released from the hydrogel function in the body. Tokyo. Tabata S317 engineering have been performed in vivo with or the ability to physically interact with protein for without biodegradable cell scaffolds. Therefore. figure 4). The during the preparation process of the formulation controlled release of various growth factors has (Gombotz & Wee 1998. cartilage.jp) was incorporated into a gelatin of protein over a long period of time from hydrogels only hydrogel and subcutaneously implanted into a mouse when the protein is simply combined in the hydrogels. gelatin microspheres solubilization of the factor in water accompanied with incorporating bFGF induced angiogenesis to a signi- the generation of water-soluble hydrogel fragments as a ficantly greater extent than the bFGF solution. succeeded in the regeneration therapy of various tissues a method to prepare the formulation of protein release (figure 3). active and et al. it is preferable that the by use of the scaffold with or without cells (Tabata polymer has been in previous clinical use. In addition.. Vascular endothelial growth factor (VEGF) is hydrogels. Review. When human recombinant bFGF (Fibrast inertness towards protein drugs. Comparing the for at least 7 days or longer is required. Some hydrogels were effective in enhancing the biodegradable scaffold from the surrounding healthy in vivo biological activity of growth factors to induce tissue. Coviello However. 1999. http:// practically impossible to achieve the controlled release www. The and pharmaceutical applications and proven to be simplest method is to supply a growth factor to the site biocompatible. back. which is susceptible to physical of protein released from a protein-carrier formulation. by VEGF are fine when compared with those of bFGF induced activation of cell functions. Thus. protein release and VEGF often causes tissue oedema. 2003. It should be noted that There are two important objectives of angiogenesis chemical immobilization is not suitable for this in tissue engineering: the therapy of ischaemic disease purpose. if patients There have been several reports on the controlled are young and healthy. The immobilized factor of myocardial infarction (Iwakura et al. As described previously. However. Tabata 2003b). from the viewpoint of the therapeutic necessity of wide A possible approach is to immobilize a growth factor in blood vessels. however. bFGF is one of the angiogenic factors and has with biomaterials should be exploited to minimize the ability to enhance wound healing through an protein denaturation. it is points. we have selected gelatin diabetes and hyperlipaemia. Interface (2009) . (Tabata et al. blood vessels regenerated release for more than a few days. Soc. 2000a. often causes a loss of biological activity. The release profile is regulated by modifying another angiogenic biosignalling molecule and has been the cross-linking density of hydrogel polymers. the time profile of growth factor release is shown to demonstrate good therapeutic efficacy (Marui governed only by that of the in vivo hydrogel et al. tissue regeneration (Lutolf & Hubbell 2005. a polymer induction of angiogenesis and induce the regeneration hydrogel may be a preferable candidate as a protein of bone. for example. There are several immobilization. R. additional means are required if patients et al. 2008). significant angiogenic effect was observed around The protein combined is normally localized in the water the implanted site. when injected into the ischaemic site immobilization is preferable.

**p!0. Biomaterials-based tissue engineering Y. We have performed the bFGF-induced angio- blood circulation was detected. (ii) gelatin microspheres incorporating CTGF. the patients could not walk due to their severe pain. Tabata (a) (b) (i) (c) 6 (i) (ii) *.2 2. (vi) systole (LAD. bFGF was locally released over two weeks at the site injected to induce in vivo angiogenesis resulting in promoted wound healing. the walking distance of ischaemic leg. left circumflex coronary artery). Intractable diseases could be repaired only by the intramuscular injection or implantation of hydrogel granules incorporating bFGF. In addition. (iv) systole. LCX. Examples of clinical tissue regeneration for ischaemic ASO and diabetic foot ulcer of intractable disease with biodegradable hydrogels for bFGF release. approximately 360 collaborations with the release technology of various growth factors are being performed by clinicians and researchers. When compared before genic clinical treatment for 25 patients in four and after the bFGF treatment. male/female J.0 20 in a water in a released (d ) -soluble form form (i) VEGF injected (mg/mouse) (e) (iv) (vi) (ii) Figure 3. The first clinical case worldwide: (a) 27 years. But angiogenic therapy allowed them to walk without any problem. the university hospitals in Japan (ages: 27–73. Currently. and (iii) diastole. male. (b) Bone regeneration: (i) BMP-2 solution. (c) Promotion of hair shaft elongation (*p!0. (b) four weeks later. (a) (b) (c) (d ) (e) (f) Figure 4. left arterior descending coronary artery. and (v) diastole.05 versus water-soluble form. (ii) hydrogel incorporating BMP-2. (e) four weeks later. ( e ) Fat tissue regeneration: gelatin microspheres incorporating bFGF. bFGF is released locally around the patients treated increased by a clinically significant injected site over two weeks and no bFGF in the extent. Soc. Before treatment.05 versus other VEGF concentrations in a released form). the pain score. (d ) 73 years. (c) 12 weeks later.** 5 hair length (mm) LCX LCX 4 LAD LAD 3 (ii) 2 1 0 (iii) (v) 0 20 0 0. The granules of gelatin hydrogels incorporating bFGF tissue oxygen and the blood flow were all significantly were injected into the femoral muscle of the patient’s improved. (a) Regeneration of coronary artery: (i) bFGF solution. (d ) Articular cartilage regeneration: (i) CTGF solution. Examples of tissue regeneration with biodegradable hydrogels for growth factor release. ( f ) 16 weeks later.S318 Review. (ii) gelatin microspheres incorporating bFGF. Interface (2009) . female. R.

cardio. the therapeutic efficacy induced by cation of culture medium and cell substrate. Yamada. manipulate their potentials for tissue regeneration. When the molecular mechanism of stem cell niche is no bone tissue was regenerated at the defect. in marked contrast with the use of of sternum. it has been demonstrated marrow (Tabata et al. angiogenesis 1 plastic surgery of soft tissues bFGF angiogenesis 1 facial nerve paralysis bFGF nerve function recovery 1 ratioZ15/10). 2005). It is well known that that the direction of cell differentiation can be modified insulin-like growth factor (IGF)-1 suppresses the apop. applicable for cell therapy and the research develop- myocytes (Sakakibara et al. the combination with the conven- the simultaneous release of two factors ( L. there have been reports (Iwai et al. ment of stem cells to naturally regulate their prolifer- Upon applying a hydrogel incorporating a low dose of ation and differentiation in vivo (Arai & Suda 2007). knee healing of the intractable foot ulcer of diabetes and meniscus (Ishida et al. by using the bFGF release to the in vivo cell-based tissue regeneration. K. as well as the engrafting of a bioartificial ation and differentiation of stem cells in vitro. Ikada 2000. It has been well not only one type of growth factor but also two or more recognized that a biological niche is the local environ- types in different concentrations and release profiles. unpublished data). FURTHER EXPERIMENTAL ASPECTS OF maintenance of biological functions in vivo (figure 1b). Interface (2009) . 2005). Regeneration therapy (Nagae et al. The culture medium to manipulate cell behaviour. Review. J. Y. Tabata S319 Table 2. 2005) of cell substrates and the surface modifi- gelatin hydrogel inhibited the ageing of acoustic nerve. The acceptable. by the softness (Engler et al. biologically clarified and the key components can be a synergistic effect on bone regeneration was observed by elucidated and used. In addition. soft tissue regeneration 1 chest plastic surgery bFGF regeneration of sternum bone. However. TGF-b1 and bone and biologically function without attachment on to the morphogenetic protein-2 (BMP-2) to induce bone regen. clinically (table 2). 2008). cation of biosignalling molecules (Nagaoka et al. two scientific and technological approaches: the modifi- 2007). the hydrogel system can release the material (Yamamoto et al. of disease or operation growth factor effect facilities vascular graft surgery for heart bFGF angiogenesis 1 arteriosclerosis obliterans (ASO) bFGF angiogenesis 4 and buerger diseases diabetic skin ulcer bFGF angiogenesis. culture substrate. it is beneficial to REGENERATION THERAPY induce angiogenesis in advance throughout the site The cell scaffolding technology can not only be applied where cells are transplanted. 2002) and kidney cells (Saito ment of stem cell biology. Biomaterials-based tissue engineering Y. Depending on the patients’ conditions. out of seven patients. 2003). dermatogenesis 3 periodontal disease bFGF regeneration of alveolar bone 3 hardness of hearing IGF-1 inhibition of acoustic nerve ageing 2 meniscus injury PRP chondrogenesis 1 facial plastic surgery bFGF chondrogenesis. 2007). Soc. no. This technology of in-advance angiogenesis also assist and promote the basic sciences in vitro efficiently enhanced the grafting rate and improved the proliferation and differentiation of stem cells. Tabata. S. This IGF-1 treatment has been started on a three-dimensional scaffold that has a steric clinically to demonstrate good therapeutic efficacy gradient of bioactive molecule concentration inside (table 2). 2000b). The synergistic angiogenesis of bFGF in the case of diabetic foot ulcer. such as bFGF. fat and meniscus is being proceeded with cocktail alone. but can system. resulting in suppressed deterioration of hearing hardness Benoit et al. Ongoing clinical experiments of tissue regeneration with the release technology of growth factors. the bFGF-induced angiogenic controlled release of the cocktail by the hydrogel enabled therapy was also effective in accelerating regeneration the regeneration of bone (Hokugo et al. TISSUE ENGINEERING-BASED For successful cell transplantation. 2007) and intervertebral disc the defect of periodontal tissue. There have the cells and constructs implanted was significantly been several trials to add various soluble factors in the augmented compared with that of no angiogenesis. To manipulate the prolifer- et al. 2006) and size (Nakamura tosis of nerve cells. Hong. The latter biological functions of pancreatic islets (Balamurugan is for the preparation of cells with a good quality et al. hepatocytes (Ogawa et al. In addition. In addition. Miyamoto. tional cell scaffold will enable stem cells to artificially Y. 2001). either bFGF or TGF-b1 to a bone defect of rabbit skulls. For example. there are dermis–epidermis skin-tissue construct (Tsuji-Saso et al. The platelet four have been healed to recover a condition clinically contains a cocktail of autologous growth factors. release system enabled the enhanced activity of various Considering that normally most cells cannot survive growth factors. 2006). 2003). 2007). 2006. and HGF was observed (Marui et al. as well as to synergistically greatly affects the profiles of cell proliferation and promote bone regeneration induced by MSC of bone differentiation. there is no doubt that the substrate eration and bone healing. Consequently. The sufficient supply of nutrients and oxygen to cells transplanted is indispensable for their survival and 4. Controlled release of IGF-1 from the et al. R.

but the good results cannot be applied to clinical Biosignalling molecules to regulate the functions and therapies because of the virus use. 2003. contriving the methodology of gene transfection culture. For gene than commercially available cationized liposomes. The microspheres incor. it is practically important to enhance the Gene transfection with the cationized polysaccharide efficiency of gene transfection. it is fate of stem cells have been elucidated. ant role in the basic research of stem cell biology. which is applicable for unless a local environment of cells suitable to promote cell transplantation therapy and stem cell biology. substantial genome has been elucidated and disease therapy on the collaborative research between material. the proliferation and differentiation is created and There are some cases where the transplantation of stem provided properly. the efficacy of was effective in enhancing the gene expression of stem viral vectors is higher than that of non-viral carriers. To create the environment. the DNA sequence of the human by use of tissue engineering technologies. The DDS technology and methodology be undoubtedly increased to develop the basic research are very effective in developing a non-viral system of of tissue regeneration and consequently realize stem gene transfection with the efficiency of gene transfec- cell-based regeneration therapy. It is practically tion as high as that of the viral system (Yamamoto & impossible to allow biosignalling proteins to function Tabata 2006. it is necessary to improve the in vivo efficacy than the original cells (Jo et al. Generally. In addition. the gene engineering technology with biomaterials 2004. such lead to a promising strategy of cell therapy. biological and clinical scientists is needed. genes are also unstable in vivo. for their functional activation. a new non-viral carrier has been prepared from regeneration therapy. There are two future directions cationized polysaccharides. in vitro and in vivo. Tabata Future development of stem cell biology and substan. Therefore. J. With the recent development of transplantation. cells to genetically engineer the biological function However. The level of gene of gene transfection. 2004). 2007. In addition. Jo & Tabata 2008). This trial has been of great success. So far. The technology of gene DNA from a biodegradable hydrogel of cationized transfection will be available for non-viral induction of gelatin derivative enhanced the level of gene expression iPS cells and also allow several bioactive substances to as well as prolonged the time period of expression internalize into cells for an investigation of their (Kushibiki et al. Several gene carrier biomaterials. 2003. Using the microspheres incorporating plasmid DNA. The release of plasmid cell biology and medicine. CLOSING REMARKS intracellular controlled release of plasmid DNA was achieved to enhance the efficiency of gene transfection to Tissue regeneration therapy—a new therapy based on a higher level than that of adenovirus transfection. In addition to the microspheres for the intracellular release of plasmid growth factors. The efficient injection of the cationized gelatin hydrogel incorporating action of siRNA by non-viral carriers is effective in plasmid DNA enhanced the in vivo therapeutic effects to genetically regulating the cell function. Nagane et al. is no doubt that the biomaterials and the technology table therapeutic effect. R. Kushibiki & Tabata 2004). The biomaterial a significantly greater extent than plasmid DNA solution carrier enables siRNA to enhance the silencing effect alone (Kushibiki et al. it is no Even if superior stem cells can be practically obtained. therapy following reconstructive surgery and organ Jo & Tabata 2008). Jo & Tabata 2008). pharma- genetic level will develop in the future. The first direction is the conventional fection enabled plasmid DNA to enhance the level of gene therapy where a plasmid DNA and adenovirus are gene expression for stem cells with less cytotoxicity directly injected to give the therapeutic effect. Biomaterials-based tissue engineering Y. the release is effective in enhancing stem cell-based regeneration technology is also effective in enhancing gene therapy and also developing the basic research of stem transfection and expression. doubt that DDS technology will have an important role it is impossible to therapeutically treat patients only by in gene therapy. there cells alone does not always induce a clinically accep. cell activation has been tried by using virus vectors DDS technology and methodology play an import. therapy. genes have been used for tissue DNA. In addition to the in vivo without DDS assistance. The carrier of gene trans- of gene therapy. 2007). the necessity for DDS will transfection. resulting in the artificial modifi- porating plasmid DNA also enhanced the gene expression cation of cell functions. and consequently increased the efficacy of cell therapy. Therefore. In this necessary to develop a system of non-virus gene connection. 2008). to overcome this problem would be to genetically tial collaboration with biomaterials technology will engineer stem cells by gene transfection for the open a new research field of stem cells and consequently activation of their biological functions. As proteins. 5. One of the possible ways is to use expression by the non-viral carrier was also enhanced by DDS technologies. the stem cells because of their toxicity and immunogenicity. which is applicable for stem cell of cells to genetically activate their biological functions biology and future cell therapy. ceutical. for the future. The the natural induction of tissue regeneration through cell cells genetically engineered functioned well to achieve transplantation and tissue engineering—is a third higher therapeutic efficacy (Yamamoto & Tabata 2006. A promising and practical way are needed. The biological functions in stem cell biology. the viral vectors are not clinically suitable ( Jo & Tabata 2008). To achieve the regeneration therapy genomics researches. Soc. Kushibiki & Tabata Thus. transplanting the cells prepared even combined with The second direction is to genetically engineer cells the scientific knowledge of cells and related substances. engineered showed the efficacy of cell therapy greater Therefore. have such as a reverse transfection method and bioreactor been investigated to enhance the level of gene combination (Okazaki et al. such as cationic liposomes and cationic polymers. in press). Interface (2009) . expression (Kushibiki et al.S320 Review. (Lai et al.

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