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Tissue engineering for tendon repair.

Tissue engineering aims to induce tissue self-regeneration in vivo or to produce a functional tissue replacement in vitro to be then implanted in the body. To produce a viable and functional tendon, a uniaxially orientated collagen type I matrix has to be generated. Biochemical and physical factors can potentially alter both the production and the organisation of this matrix, and their combination in a dose- and time-dependent manner is probably the key to in vitro engineered tendons. This review discusses the role of these different factors affecting tenocyte growth in a three-dimensional environment in vivo and in vitro, and underlines the future challenge of tendon tissue engineering.

Biocompatible Materials↗

[Progress in research and application of the tissue engineering blood vessels].

Vascular tissue engineering is a novel approach by which an ideal vascualr graft constructed in vitro that will not be obstructed for a long time without immunological reaction after implantation. This article reviewed the definition of the tissue engineering blood vessel (TEBV), cellular resourses, the selection of biocompatible materials, the devising methods and the research achievements. Furthermore, it also discussed the current problems of TEBV and looked forward to future clinical application.

Biocompatible Materials↗

A pilot study investigating a novel subcutaneously implanted pre-cellularised scaffold for tissue engineering of intestinal mucosa.

Tissue engineering of the small intestine offers an alternative to long-term intravenous nutrition and transplantation in patients with intestinal failure. Initial work, although encouraging, is limited by the volume of neonatal tissue required to produce a small neomucosal cyst. Our novel approach is to implant tubular poly-lactide-co-glycolide (PGLA) foam scaffolds subcutaneously. The aim of this study was to investigate whether these scaffolds would support growth of intestinal neomucosa. PGLA scaffolds were implanted subcutaneously into 8 Lewis rats; after 5 weeks, 'organoid units' were injected into the lumens. Tissue was assessed histologically after harvesting and quantitative immunohistochemistry was performed using antibodies against vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 2 (VEGF-R2), fibroblast growth factor basic (bFGF) and fibroblast growth factor receptor 2 (FGF-R2). At 4 weeks post organoid unit implantation, clearly recognisable mucosa and submucosa was present on the luminal surface of the scaffold. Densities of VEGF and VEGF-R2 positive cells increased with time post organoid unit implantation. This pilot study demonstrates that it is possible to tissue engineer small intestinal neomucosa using subcutaneously implanted PLGA scaffolds. The yield of the process compares favourably to the published literature. Further work is required to optimise the technique.

Animals↗

Harnessing wound healing and regeneration for tissue engineering.

Biomedical science has made major advances in understanding how cells grow into functioning tissue and the signalling mechanisms used to achieve this are slowly being dissected. Tissue engineering is the application of that knowledge to the building or repairing of organs, including skin, the largest organ in the body. Generally, engineered tissue is a combination of living cells and a supporting matrix. Besides serving as burn coverings, engineered skin substitutes can help patients with diabetic foot ulcers. Today, most of these ulcers are treated with an approach that includes antibiotics, glucose control, special shoes and frequent cleaning and bandaging. The results of such treatments are often disappointing and ineffectual, and scarring remains a major problem, mechanically, cosmetically and psychologically. Within our group we are attempting to address this by investigating novel approaches to skin tissue engineering. We are identifying novel therapeutic manipulations to improve the degree of integration between a tissue engineered dermal construct and the host by both molecular manipulation of growth factors but also by understanding and harnessing mechanisms of regenerative biology. For the purpose of this summary, we will concentrate primarily on the latter of these two approaches in that we have identified a novel mouse mutant that completely and perfectly regenerates skin and cartilaginous components following ear injury. This experimental animal will allow us to characterize not only novel genes involved in the regeneration process but also to utilize cells from such animals in artificial skin equivalents to assess their behaviour compared with normal cells. This approach should allow us to create a tissue-engineered substitute, which more closely resembles the normal regional microanatomy and physiology of the skin, allowing better integration to the host with minimal or no scarring.

Animals↗

Tissue engineering of the cornea.

Tissue engineering of the cornea represents a paradigm shift in medical treatment to overcome the present disadvantages of corneal transplantation, primarily immune rejection and the shortage of donor corneas. Transplantation of cultivated corneal epithelial cells expanded ex vivo from corneal epithelial stem cells has been developed and has already entered the clinical realm. However, there remain many hurdles to be overcome. The author and colleagues are developing a method to transplant cultivated cell sheets that uses a temperature-responsive culture dish. This article reviews the present situation regarding tissue-engineered corneal epithelium and introduces the results of this program of research.

Cornea↗

Tissue engineering by modulated gene delivery.

Tissue engineering is a newly emerging biomedical form to create a local environment which enables cells to promote the proliferation and differentiation for regeneration induction. The cell-induced regeneration of tissues and organs is achieved by making use of the tissue engineering technology or methodology. Several genetic approaches with virus and non-viral vectors or genetically engineered cells have been attempted to enhance tissue regeneration. The basic idea is to promote the cell proliferation and differentiation as well as the secretion of biological signal molecules for tissue regeneration from cells by gene transfection. For successful gene transfection and expression, it is important to develop drug delivery system (DDS) which allows a therapeutic gene to be delivered specifically to the target cell at an appropriate timing for a certain time period. This paper overviews the recent development of gene-modified tissue engineering, briefly explaining delivery technologies necessary to modulate the efficiency of gene transfection.

Animals↗

Investigation of optical coherence tomography as an imaging modality in tissue engineering.

Monitoring cell profiles in 3D porous scaffolds presents a major challenge in tissue engineering. In this study, we investigate optical coherence tomography (OCT) as an imaging modality to monitor non-invasively both structures and cells in engineered tissue constructs. We employ time-domain OCT to visualize macro-structural morphology, and whole-field optical coherence microscopy to delineate the morphology of cells and constructs in a developing in vitro engineered bone tissue. The results show great potential for the use of OCT in non-invasive monitoring of cellular activities in 3D developing engineered tissues.

Adolescent↗

Systems for therapeutic angiogenesis in tissue engineering.

The goals in tissue engineering include the replacement of damaged, injured, or missing body tissues with biologically compatible substitutes. To overcome initial tissue-mass loss, improved vascularization of the regenerated tissue is essential. Two pathways of tissue neovascularization are known: vasculogenesis, the in situ assembly of capillaries from undifferentiated endothelial cells (EC), and angiogenesis, the sprouting of capillaries from preexisting blood vessels. Recent advances in our understanding of the process of bloodvessel growth have provided significant tools for the neovascularization of bioengineered tissues. Several growth factors serve as stimuli for EC proliferation and migration as well as the formation of new blood vessels. They convey their effects via specific receptors expressed on the surface of EC. Vascular epithelial growth factor (VEGF) is a major regulator of neovascularization. VEGF plays a major role in the early development of blood-cell progenitors. Basic fibroblast growth factor (bFGF) was identified as the first angiogenic factor. It is a potent inducer of EC proliferation and blood-vessel growth in vitro and in vivo. VEGF and bFGF have been injected into undervascularized ischemic tissues, resulting in new blood-vessel formation and tissue perfusion. Gene-therapy approaches using VEGF cDNA injection into ischemic tissues have augmented the formation of collateral vessels. Angiogenic factors such as VEGF and bFGF have also been incorporated into bioengineered tissues and have facilitated blood-vessel growth. Other approaches such as prevascularization of the matrix prior to cell seeding and incorporation of EC into the bioengineered tissues have produced encouraging results. This article reviews the process of blood-vessel growth and tissue vascularization, placing emphasis on strategies that can be employed for efficient vascularization of engineered tissues in vitro and in vivo.

Angiogenesis Inducing Agents↗

A novel approach to periodontal tissue regeneration with mesenchymal stem cells and platelet-rich plasma using tissue engineering technology: A clinical case report.

Tissue engineering represents one of the most exciting advances in regenerative medicine. However, little has been reported on the application of tissue engineering for regeneration of periodontal tissues. Therefore, the aim of this study was to show how a technique based on tissue engineering principles can be applied to periodontology. Mesenchymal stem cells (MSCs) were isolated from a patient's iliac crest marrow aspirates. Platelet-rich plasma (PRP) was isolated from peripheral blood. Full-thickness periodontal flaps were elevated and the root surfaces were scaled and planed. A MSCs-PRP gel was prepared and applied to the root surface and adjacent defect space. The primary outcome measures were changes in pocket depth, clinical attachment level, bleeding on probing, and defect bone fill. Re-examination demonstrated that the treatment, including the application of MSCs-PRP gel at periodontal sites with angular defects, resulted in a 4-mm reduction in probing depths and a 4-mm clinical attachment gain, while bleeding and tooth mobility disappeared. Radiographic assessments showed that the bone defect had been reduced in depth. Interdental papillae supported by this tissue engineering technology regenerated. The use of MSCs in PRP gel might be helpful for periodontal tissue regeneration, treatment of esthetically sensitive sites, and reduction of patient morbidity.

Alveolar Bone Loss↗

Update on tissue-engineered biological dressings.

Tissue-engineered biological dressings offer promise in the treatment of burns, chronic ulcers, donor site and other surgical wounds, and a variety of blistering and desquamating dermatologic conditions. For example, the prevalence of diabetic foot ulcers ranges from 4.4% to 10.5% of diabetics, resulting in 82,000 lower extremity amputations annually; venous leg ulcers affect 0.18% to 1.35% of the population; and pressure ulcers are found in 5.0% to 8.8% of institutionalized patients and 14.8% of patients in acute care facilities. Despite the large number of potential beneficiaries, cellular tissue-engineered products have suffered setbacks in recent years and have garnered considerably lower market share than commercial promoters anticipated. The mechanism of action of these products is not universally agreed upon, but delivery of growth factors and extracellular matrix components to the wound is thought to be important; graft "take" is not usually considered to occur. These "engineered" products do not specifically match a treatment modality to an underlying pathology. Clinical effect is often modest, and sometimes not justi- fiable from a cost-benefit perspective. Nevertheless, clinical reports in the literature of uses of tissueengineered biological dressings continue to mount, indicating that these products are finding niche applications where clinical utility is high and the cost can be defended. Despite commercial setbacks, the first-approved products, Dermagraft, Apligraf, and Cultured Epidermal Autograft (Epicel) are still being marketed, and new ones, such as OrCel, continue to be developed. The major indications for these products are summarized and a brief review of the available clinical literature is offered.

Biological Dressings↗

Distinction between the extracellular matrix of the nucleus pulposus and hyaline cartilage: a requisite for tissue engineering of intervertebral disc.

Tissue engineering of intervertebral discs (IVD) using mesenchymal stem cells (MSCs) induced to differentiate into a disc-cell phenotype has been considered as an alternative treatment for disc degeneration. However, since there is no unique marker characteristic of discs and since hyaline cartilage and immature nucleus pulposus (NP) possess similar macromolecules in their extracellular matrix, it is currently difficult to recognize MSC conversion to a disc cell. This study was performed to compare the proteoglycan to collagen ratio (measured as GAG to hydroxyproline ratio) in the NP of normal disc to that of the hyaline cartilage of the endplate within the same group of individuals and test the hypothesis that this ratio can be used for in vivo studies to distinguish between a normal NP and hyaline cartilage phenotype. Whole human lumbar spine specimens from fresh cadavers, ranging in age from 12 weeks to 79 years, were used to harvest the IVDs and adjacent endplates. The GAG to hydroxyproline ratio within the NP of young adults is approximately 27:1, whereas the ratio within the hyaline cartilage endplate of the same aged individuals is about 2:1. The production of an extracellular matrix with a high proteoglycan to collagen ratio can be used in vivo to distinguish NP cells from chondrocytes, and could help in identifying a NP-like phenotype in vivo as opposed to a chondrocyte when MSCs are induced to differentiate for tissue engineering of a disc.

Adolescent↗

Tissue engineering in dentistry.

Advances in tissue engineering provide an increased level of understanding of the mechanical and chemical stimuli that regulate tissue responses. Oral tissue engineering can be applied to recreate missing osseous or dental structures or correct orofacial deformities, changing the patient's smile, midfacial height, and the soft tissue drape. Biomechanical principles can also be applied to tissue engineering to enhance the bone/tooth or bone/implant functionality and long-term stability. Advancements are also being achieved in the area of biomimetics that will allow the creation of new biologic replacements for missing oral structures. The opportunity for bioengineering to charter the course of tooth regeneration is an exciting prospect and will improve the quality of life for patients for decades to come.

Biomechanical Phenomena↗

[Tissue engineering in urology. Basic principles and application].

Tissue engineering is a rather new field of science. Despite this fact, some experimental investigations have already been applied in clinical studies. Compared to other medical fields, tissue engineering in urology is well established. Tissue-engineered bulking agents and tissue-engineered bladder augments are being investigated in clinical trials. Even though the knowledge gained in recent years is promising, the results of cellular therapies need to be critically judged before being finally applied in patients. Genetic engineering and stem cell research (adult undifferentiated cells) have had major impact on the field of tissue engineering over the past 2 years. By using the technology of genetic engineering, biochemical and functional qualities of tissues may be modified. Adult stem cells may help to substitute lost tissue in an autologous fashion by isolating undifferentiated cells from the body and by differentiating them into a desired cell type. These cells may be used to form native functional tissue to replace a diseased organ or organ part.

Adult↗

Sustained release of vascular endothelial growth factor from mineralized poly(lactide-co-glycolide) scaffolds for tissue engineering.

Strategies to engineer bone tissue have focused on either: (1) the use of scaffolds for osteogenic cell transplantation or as conductive substrates for guided bone regeneration; or (2) release of inductive bioactive factors from these scaffold materials. This study describes an approach to add an inductive component to an osteoconductive scaffold for bone tissue engineering. We report the release of bioactive vascular endothelial growth factor (VEGF) from a mineralized, porous, degradable polymer scaffold. Three dimensional, porous scaffolds of the copolymer 85 : 15 poly(lactide-co-glycolide) were fabricated by including the growth factor into a gas foaming/particulate leaching process. The scaffold was then mineralized via incubation in a simulated body fluid. Growth of a bone-like mineral film on the inner pore surfaces of the porous scaffold is confirmed by mass increase measurements and quantification of phosphate content within scaffolds. Release of 125I-labeled VEGF was tracked over a 15 day period to determine release kinetics from the mineralized scaffolds. Sustained release from the mineralized scaffolds was achieved, and growth of the mineral film had only a minor effect on the release kinetics from the scaffolds. The VEGF released from the mineralized and non-mineralized scaffolds was over 70% active for up to 12 days following mineralization treatment, and the growth of mineral had little effect on total scaffold porosity.

Biocompatible Materials↗

Tissue engineering of white adipose tissue using hyaluronic acid-based scaffolds. I: in vitro differentiation of human adipocyte precursor cells on scaffolds.

BACKGROUND AND AIM OF THE STUDY: Reconstruction of soft tissue defects is a challenge in plastic surgery and there is clinical need for adequate solutions. Aim of this study was to develop a biohybrid construct consisting of hyaluronic acid-based scaffolds and human adipocyte precursor cells as a soft tissue filler. METHODS: Human adipocyte precursor cells were obtained by collagenase digestion of adipose tissue samples and seeded on hyaluronic acid-based spongy scaffolds of various degrees of esterification and pore size using different techniques. After cell attachment, adipose differentiation was induced by defined adipogenic factors under serum-free culture conditions. RESULTS: Among the five different scaffold types under investigation the highest cell attachment rate was observed for the HYAFF scaffold with 100% esterification and a mean pore size of 400microm (HYAFF 11lp). For inoculation of human adipocyte precursor cells on hyaluronic acid-based scaffolds a "drop-on" technique and low-pressure centrifugation using a Speed Vac airfuge were compared. With respect to efficacy, cell distribution and simpleness the drop-on method proved to be the method of choice. In a serum-free medium supplemented with 66nM insulin, 100nM cortisol and 1microg/ml troglitazone a substantial proportion of cells underwent adipose differentiation as assessed by lipid accumulation and emergence of glycerol-3-phosphate dehydrogenase activity, a lipogenic marker enzyme. CONCLUSION: Hyaluronic acid-based scaffolds appear to be a suitable three-dimensional carrier for the culture and in vitro differentiation of human adipocyte precursor cells.

Adipocytes↗

Tissue engineering of small intestinal tissue using collagen sponge scaffolds seeded with smooth muscle cells.

In a previously reported attempt to regenerate small intestine with autologous tissues, collagen scaffolds were used without cell seeding or with autologous mesenchymal stem cell seeding. However the regenerated intestine lacked a smooth muscle layer. To accomplish regeneration of a smooth muscle layer, this present study used collagen scaffolds seeded with the smooth muscle cells (SMC) in a canine model. Autologous SMC were isolated from stomach wall and cultured. Two types of scaffolds were fabricated: in SMC (+), cultured SMCs were mixed with collagen solution and poured into a collagen sponge; and in SMC (-), SMCs were omitted. Both scaffolds were implanted into defects of isolated ileum as a patch graft. Animals were euthanized at 4, 8, and 12 weeks; for the last time point, the ileal loop had been reanastomosed at 8 weeks. At 12 weeks, the SMC (-) group showed a luminal surface covered by a regenerated epithelial cell layer with very short villi; however only a thin smooth muscle layer was observed, representing the muscularis mucosae. In the SMC (+) group, the luminal surface was covered completely by a relatively well-developed epithelial layer with numerous villi. Implanted SMCs were seen in the lamina propria and formed a smooth muscle layer. Thus, we concluded that collagen sponge scaffolds seeded with autologous SMCs have a potential for small intestine regeneration.

Animals↗