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PubMed2002

The sintered microsphere matrix for bone tissue engineering: in vitro osteoconductivity studies.

A tissue engineering approach has been used to design three-dimensional synthetic matrices for bone repair. The osteoconductivity and degradation profile of a novel polymeric bone-graft substitute was evaluated in an in vitro setting. Using the copolymer poly(lactide-co-glycolide) [PLAGA], a sintering technique based on microsphere technology was used to fabricate three-dimensional porous scaffolds for bone regeneration. Osteoblasts and fibroblasts were seeded onto a 50:50 PLAGA scaffold. Morphologic evaluation through scanning electron microscopy demonstrated that both cell types attached and spread over the scaffold. Cells migrated through the matrix using cytoplasmic extensions to bridge the structure. Cross-sectional images indicated that cellular proliferation had penetrated into the matrix approximately 700 microm from the surface. Examination of the surfaces of cell/matrix constructs demonstrated that cellular proliferation had encompassed the pores of the matrix by 14 days of cell culture. With the aim of optimizing polymer composition and polymer molecular weight, a degradation study was conducted utilizing the matrix. The results demonstrate that degradation of the sintered matrix is dependent on molecular weight, copolymer ratio, and pore volume. From this data, it was determined that 75:25 PLAGA with an initial molecular weight of 100,000 has an optimal degradation profile. These studies show that the sintered microsphere matrix has an osteoconductive structure capable of functioning as a cellular scaffold with a degradation profile suitable for bone regeneration.

Animals↗

PubMed2004

Optimization of bone-tissue engineering in goats.

Successful bone-tissue engineering (TE) has been reported for various strategies to combine cells with a porous scaffold. In particular, the period after seeding until implantation of the constructs may vary between hours and several weeks. Differences between these strategies can be reduced to (a) the presence of extracellular matrix, (b) the differentiation status of the cells, and (c) the presence of residual potentially immunogenic serum proteins. These parameters are investigated in two types of calcium phosphate scaffolds in a goat model of ectopic bone formation. Culture-expanded bone-marrow stromal cells from eight goats were seeded onto two types of hydroxyapatite granules: HA60/400 (60% porosity, 400-microm average pore size) and HA70/800. Scaffolds seeded with cells and control scaffolds were cultured for 6 days in medium containing autologous or semisynthetic serum, in the presence or absence of dexamethasone. Other scaffolds were seeded with cells just before implantation in medium with or without serum. All conditions were implanted autologously in the paraspinal muscles. After 12 weeks, bone had formed in 87% of all TE constructs, as demonstrated by histology. Histomorphometry indicated significantly more bone in the HA70/800 scaffolds. Furthermore, a significant advantage in bone formation was found when the constructs had been cultured for 6 days. In conclusion, both scaffold characteristics (porosity) and TE strategy (culturing of the constructs) were demonstrated to be important for bone TE.

Animals↗

PubMed2003

Preparation of degradable porous structures based on 1,3-trimethylene carbonate and D,L-lactide (co)polymers for heart tissue engineering.

Biodegradable porous scaffolds for heart tissue engineering were prepared from amorphous elastomeric (co)polymers of 1,3-trimethylene carbonate (TMC) and D,L-lactide (DLLA). Leaching of salt from compression-molded polymer-salt composites allowed the preparation of highly porous structures in a reproducible fashion. By adjusting the salt particle size and the polymer-to-particle weight ratio in the polymer-salt composite preparation the pore size and porosity of the scaffolds could be precisely controlled. The thermal properties of the polymers used for scaffold preparation had a strong effect on the morphology, mechanical properties and dimensional stability of the scaffolds under physiological conditions. Interconnected highly porous structures (porosity, 94%; average pore size, 100 microm) based on a TMC-DLLA copolymer (19:81, mol%) had suitable mechanical properties and displayed adequate cell-material interactions to serve as scaffolds for cardiac cells. This copolymer is noncytotoxic and allows the adhesion and proliferation of cardiomyocytes. During incubation in phosphate-buffered saline at 37 degrees C, these scaffolds were dimensionally stable and the number average molecular weight (Mn) of the polymer decreased gradually from 2.0 x 10(5) to 0.3 x 10(5) in a period up to 4 months. The first signs of mass loss (5%) were detected after 4 months of incubation. The degradation behavior of the porous structures was similar to that of nonporous films with similar composition and can be described by autocatalyzed bulk hydrolysis.

Biocompatible Materials↗

PubMed2005

A comparative qualitative histological analysis of tissue-engineered bone using bone marrow mesenchymal stem cells, alveolar bone cells, and periosteal cells.

For tissue-engineered bone formation, autogenous osteogenic cells are of paramount importance for successful bone formation. In order to investigate the donor cell-related differences in tissue-engineered bone, cultured bone marrow mesenchymal stem cells, cultured alveolar bone cells, and cultured periosteal cells were examined for their in vivo potential to form bone. These cells were isolated from dogs, expanded in vitro, mixed with autologous fibrin glue and BMP-2, and then injected into the subcutaneous space on the dorsum of nude mice. Bone formation was evaluated at 12 weeks. Histomorphometric analysis demonstrated that the subcutaneous nodules formed in nude mice contained 26.9% newly formed bone when using the bone marrow mesenchymal stem cells, 41.1% newly formed bone when using the alveolar bone cells, and 58.2% newly formed bone when using the periosteal cells. The results suggest that periosteal cells are the best choice for enhancing bone formation in tissue engineering of bone regeneration.

Alveolar Process↗

PubMed2002

[Repair of porcine full-thickness skin defects with autologous tissue engineered skin].

OBJECTIVE: To explore a feasible method to repair full-thickness skin defects with tissue engineered techniques. METHODS: The skin specimens were cut from the Changfeng hybrid swines' abdomen, then keratinocytes and fibroblasts were isolated and harvested by trypsin, EDTA and type II collagenase. The cells were seeded in petri dishes for primary culture. When the cells were in logarithmic growth phase, they were treated with dispase II (keratinocytes) or trypsin (fibroblasts) to separate them from the floor of the tissue culture dishes. A biodegradable material-pluronic F-127 was prefabricated and mixed with these cells, and then the cells-pluronic compounds were seeded evenly into polyglycolic acid (PGA). Tinally the constructs were replanted to autologous animals to repair full-thickness skin defects. Histological changes were observed in 1, 2, 4 and 8 weeks postsurgery. RESULTS: The cells-pluronic F-127-PGA compounds could repair autologous full-thickness skin defects. Histologically, the tissue engineered skin was similar to normal skin with stratified epidermis overlying a moderately thick collageneous dermis. CONCLUSION: Tissue engineered skin can repair autologous full-thickness skin defects with primary-cultured keratinocytes and fibroblasts as seed cells and PGA as a cell carrier.

Animals↗

PubMed2006

MR assessment of osteogenic differentiation in tissue-engineered constructs.

Bone marrow stromal cells (MSC) are a promising source of osteoprogenitor cells for bone tissue engineering. However, the population of the osteoprogenitor cells and their differentiation potentials change with the gender, age, and health of the donor. Development of a noninvasive method to assess osteogenic progression is critical for successful bone tissue regeneration. High-resolution magnetic resonance imaging (MRI) (at 11.7 T, with spatial resolution of 62.5 x 62.5 microm in 500 microm slices) is used in the present study to monitor osteogenic differentiation of tissue-engineered constructs prepared by seeding human bone MSCs on gelatin sponge scaffolds. Quantitative measurements of the MR relaxation times (T1, T2) and the apparent diffusion coefficient (ADC) were performed for four successive weeks on control tissue constructs and constructs exposed to osteogenic differentiation medium. The T1 and T2 relaxation times and ADC were found to decrease as osteogenic progression proceeded in samples exposed to osteogenic differentiation medium. At week 4, the T1, T2, and ADC of TE constructs were 1.81 +/- 0.11 s, 19.5 +/- 11.02 ms, and 1.01 +/- 0.47 x 10(3) mm(2)/s, respectively, for osteogenic differentiated constructs, significantly different from control constructs 2.22 +/- 0.08 s, 50.39 +/- 5.57 ms, and 1.86 +/- 0.18 x 107(3) mm(2)/s (p < 0.05). The MR parameters were also highly correlated with the cell seeding densities and alkaline phosphatase (ALP) activities of the osteogenic constructs. In conclusion, periodic measurements of MR parameters (T1, T2, and ADC) provide a promising method for noninvasive monitoring of the status of tissue-engineered bone growth and differentiation.

Alkaline Phosphatase↗

PubMed1998

Videofetoscopically assisted fetal tissue engineering: bladder augmentation.

BACKGROUND/PURPOSE: Treatment of several congenital anomalies is frequently hindered by lack of enough tissue for surgical reconstruction in the neonatal period. Minimally invasive harvest of fetal tissue, which is then processed through tissue engineering techniques in vitro while pregnancy is allowed to continue so that at delivery a newborn with a prenatally diagnosed congenital anomaly can benefit from having autologous, expanded tissue promptly available for surgical reconstruction at birth. This concept was applied to a bladder defect. METHODS: Bladder exstrophy was surgically created in ten 90- to 95-day gestation fetal lambs, which were divided in two groups. In group I, a small fetal bladder specimen was harvested through a minimally invasive technique (videofetoscopy). Urothelial and smooth muscle cells were then separately cultivated and expanded in vitro for 55 to 60 days, resulting in a total of approximately 200 million cells. Seven to 10 days before delivery, the cells were seeded in two layers in a 16- to 20-cm2, 3-mm thick biodegradable polyglycolic acid polymer matrix. One to 4 days after delivery, autologous engineered tissue was used for surgical augmentation of the exstrophic bladder. In group II, no harvest was performed, and the bladder exstrophy was primarily closed after delivery. In both groups, a catheter was left inside the bladder for 3 weeks, at which time a cystogram was performed and the catheter then removed. In all animals, at 60 days, another cystogram was performed and urodynamic studies of the bladder were performed. The bladder was then removed for histological analysis. RESULTS: Fetal survival rate was 100%. One newborn died immediately after the implantation of the engineered bladder from an anesthetic accident. The other nine (four in group I and five in group II) survived. One of the animals from group I lost its bladder catheter prematurely and had a urinary leak detected only at the time of death. There were no other complications. The engineered bladders were more compliant (P < .05) and had greater capacity pressures greater than 20 mm Hg (P < .05) than those closed primarily. Histological analysis of the engineered tissue showed a multilayered urothelial lining on the luminal side and overlying layers of smooth muscle cells surrounded by connective tissue. CONCLUSIONS: Videofetoscopically assisted fetal bladder engineering may be a viable alternative for prompt bladder reconstruction at birth. The architecture of autologous engineered fetal bladder tissue resembles that of native bladder. This concept may prove useful for the treatment of certain human neonatal conditions such as bladder and cloacal exstrophies.

Animals↗

PubMed2003

[Maxillary growth following tissue engineered oral mucosal implantation on mucoperiosteal denudated palate process in young rat].

OBJECTIVE: To observe the effectiveness of prevention and cure for maxillary growth deformity following tissue engineered oral mucosa implantation on mucoperiosteal denuded palate process in young rat. METHODS: Hard palate mucoperiosteum of a SD baby rat were excised and oral keratinocytes were isolated and cultured. Tissue engineered oral mucosa was fabricated with the cultured oral keratinocytes and the membrane made of sodium alginate (SA). 80 female three-week-old SD rats were used as subjects in this study. The animals were divided randomly into a normal control group and 3 experimental groups, each group included 20 rats. Normal control group (NG) were not operated. Hard palate mucoperiosteum on left side in all experimental groups were excised, exposed bone were not treated in denuded group (DG), but repaired with membrane in material group (MG) and repaired with the tissue engineered oral mucosa in mucosal group (MUG). All the animals were sacrificed at 9th week postoperatively (12 weeks old), and the clean widths of right and left hard palatal were measured under a dissection microscope. The difference between palatal widths of two sides and the asymmetry ratio between the different groups were compared and analyzed. RESULTS: No significant difference in asymmetry was discovered between the DG and the MG, but the asymmetry in MUG was less than DG or MG. CONCLUSION: Tissue engineered oral mucosal implantation in palatoplasty is an effective method in preventing and curing secondary maxilla deformity by repairing denuded bone wound.

Animals↗

PubMed2002

Tissue-engineered repair of osteochondral defects: effects of the age of donor cells and host tissue.

Transplantation of a tissue-engineered construct containing cells of a chondrocytic phenotype into an osteochondral defect provides a biological solution to this type of injury. Among the factors that affect cell proliferation and phenotypic expression, age is one that has not been well characterized. In this study adult and aged male donor cells, derived from perichondrium, were cultured and adsorbed into a polylactic acid (PLA) scaffold and transplanted into osteochondral defects created in adult (8- to 10-month-old) and aged (4- to 5-year-old) female rabbits. Three groups were investigated: (1) adult cells transplanted into aged defects, (2) aged cells transplanted into aged defects, and (3) aged cells transplanted into adult defects. In vitro characterization of adult and aged cells and in vivo assessment of osteochondral repair tissue at 12 weeks posttransplantation were carried out. The in vitro studies demonstrated that the proliferation rate of aged cells was less than that of mature cells during the earliest stage of culture. Also, the chondrocytic phenotype was reduced in aged cells compared with mature cells. The in vivo results showed that donor (SRY-positive) cell survival differed among the three groups: survival of adult cells into aged defect > survival of aged cells into aged defect > survival of aged cells into adult defect. The biological acceptability of the repair, defined as smooth firm cartilaginous tissue filling the defect, was < 25% of the operated specimens in each of the three groups. This repair tissue contained only 20-25% of the amounts of type II collagen and glycosaminoglycans found in normal articular cartilage. These data suggest that the outcome of tissue-engineered repair of osteochondral defects is affected by both the age of donor cells and the age of the host.

Age Factors↗

PubMed2006

[The extracellular matrix in bone tissue engineering].

OBJECTIVE: To review the research progress of osteoblast extracellular matrix (ECM) and its application in bone tissue engineering. METHODS: The recent related literatures were extensively reviewed. RESULTS: The ECM was complex in its components. The configuration of cell and cell's adhesion, migration, proliferation, and differentiation were subject to the ECM. The bioactivity of the tissue engineering products was revealed by ECM, which predicted the product's efficiency in clinic application. CONCLUSION: ECM has the potential to become the effective index in evaluating tissue engineered products.

Extracellular Matrix↗

PubMed2006

[Tooth movement in tissue-engineered bone].

OBJECTIVE: To explore the effect and practical value of repairing the alveolar defects with tissue engineering technique and investigate the influence of the generated new bone on the orthodontic tooth movement. METHODS: The marrow stromal cells of rats were separated, cultured in vitro and induced to osteoblast-like cells. The osteoblast-like cells were implanted onto the ceramic bovine bone. Then the complex was implanted into the alveolar defect in one side of the rat's mandible. The other side of the mandible served as control. Eight weeks later, the orthodontic appliances were placed between the first molar and incisors of SD rats to move the first molar forward. The tooth movement and root resorption of the molar were observed. RESULTS: We found that the tooth movement in the experimental area was faster than that in the normal alveolar bone (P < 0.05). The root resorption and the alveolar bone height loss were less than that in the control area (P < 0.05). CONCLUSIONS: The tissue-engineered bone did not have negative influences on tooth movement. The repair of alveolar bone defect by tissue engineering approach may be used in craniofacial surgery and orthodontics.

Animals↗

PubMed2000

Morphogenesis and tissue engineering of bone and cartilage: inductive signals, stem cells, and biomimetic biomaterials.

Morphogenesis is the developmental cascade of pattern formation, body plan establishment, and the architecture of mirror-image bilateral symmetry of many structures and asymmetry of some, culminating in the adult form. Tissue engineering is the emerging discipline of design and construction of spare parts for the human body to restore function based on principles of molecular developmental biology and morphogenesis governed by bioengineering. The three key ingredients for both morphogenesis and tissue engineering are inductive signals, responding stem cells, and the extracellular matrix. Among the many tissues in the human body, bone has considerable powers for regeneration and is a prototype model for tissue engineering based on morphogenesis. Implantation of demineralized bone matrix into subcutaneous sites results in local bone induction. This model mimics sequential limb morphogenesis and permitted the isolation of bone morphogens. Although it is traditional to study morphogenetic signals in embryos, bone morphogenetic proteins (BMPs), the inductive signals for bone, were isolated from demineralized bone matrix from adults. BMPs and related cartilage-derived morphogenetic proteins (CDMPs) initiate, promote, and maintain chondrogenesis and osteogenesis and have actions beyond bone. The symbiosis of bone inductive and conductive strategies are critical for tissue engineering, and is in turn governed by the context and biomechanics. The context is the microenvironment, consisting of extracellular matrix, which can be duplicated by biomimetic biomaterials such as collagens, hydroxyapatite, proteoglycans, and cell adhesion proteins including fibronectins. Thus, the rules of architecture for tissue engineering are an imitation of the laws of developmental biology and morphogenesis, and thus may be universal for all tissues, including bones and joints.

Adult↗

PubMed2003

[Experimental study on constructing small-caliber artery by tissue engineering approach].

OBJECTIVE: To investigate the possibility of constructing small-caliber artery by means of tissue engineering. METHODS: Cell-PGA mixtures were made by separately seeding 1 x 10(7) smooth muscle cells and 5 x 10(6) endothelial cells isolated from neonate umbilicus onto PGA scaffold, the cell-PGA constructs were wrapped around a silicone tube before its implantation subcutaneously to nude mice and the mice were sacrificed in 2 and 6 weeks. The tissue engineered artery (TEA) were examined both grossly and immunohistochemically. RESULTS: The gross appearance of TEA was similar to that of the natural counterparts; histologic and immunohistochemical analyses of the neoformed tissues revealed a typical artery structure, including the presence of EC at the luminal surface and the presence of SMC and collagen in the wall. CONCLUSION: TEA with histology similar to natural vessel can be constructed by tissue engineering.

Animals↗

PubMed1999

Tissue engineering: orthopedic applications.

Because of an aging population and increased occurrence of sports-related injuries, musculoskeletal disorders have become one of the major health concerns in the United States. Current treatments, although fairly successful, do not provide the optimum therapy. These treatments typically rely on donor tissues obtained either from the patient or from another source. The former raises the issue of supply, whereas the latter poses the risk of rejection and disease transfer. This has prompted orthopedic surgeons and scientists to look for viable alternatives. In recent years, tissue engineering has gained increasing support as a method to treat orthopedic disorders. Because it uses principles of engineering, biology, and chemistry, tissue engineering may provide a more effective approach to the treatment of musculoskeletal disorders than traditional methods. This chapter presents a review of current methods and new tissue-engineering techniques for the treatment of disorders affecting bone, ligament, and cartilage.

Animals↗

PubMed2004

Cartilage and bone tissue engineering for reconstructive head and neck surgery.

The loss of cartilage and bone because of congential defects, trauma and after tumor resection is a major clinical problem in head and neck surgery. The most prevalent methods of tissue repair are through autologous grafting or using implants. Tissue engineering applies the principles of engineering and life sciences in order to create bioartificial cartilage and bone. Most strategies for cartilage tissue engineering are based on resorbable biomaterials as temporary scaffolds for chondrocytes or precursor cells. Clinical application of tissue-engineered cartilage for reconstructive head and neck surgery as opposed to orthopedic applications has not been well established. While in orthopedic and trauma surgery engineered constructs or autologous chondrocytes are placed in the immunoprivileged region of joints, the subcutaneous transplant site in the head and neck can lead to strong inflammatory reactions and resorption of the bioartificial cartilage. Encapsulation of the engineered cartilage and modulation of the local immune response are potential strategies to overcome these limitations. In bone tissue engineering the combination of osteoconductive matrices, osteoinductive proteins such as bone morphogenetic proteins and osteogenic progenitor cells from the bone marrow or osteoblasts from bone biopsies offer a variety of tools for bone reconstruction in the craniofacial area. The utility of each technique is site dependent. Osteoconductive approaches are limited in that they merely create a favorable environment for bone formation, but do not play an active role in the recruitment of cells to the defect. Delivery of inductive signals from a scaffold can incite cells to migrate into a defect and control the progression of bone formation. Rapid osteoid matrix production in the defect site is best accomplished by using osteoblasts or progenitor cells.

Bone and Bones↗

PubMed1999

Porous chitosan scaffolds for tissue engineering.

The wide array of tissue engineering applications exacerbates the need for biodegradable materials with broad potential. Chitosan, the partially deacetylated derivative of chitin, may be one such material. In this study, we examined the use of chitosan for formation of porous scaffolds of controlled microstructure in several tissue-relevant geometries. Porous chitosan materials were prepared by controlled freezing and lyophilization of chitosan solutions and gels. The materials were characterized via light and scanning electron microscopy as well as tensile testing. The scaffolds formed included porous membranes, blocks, tubes and beads. Mean pore diameters could be controlled within the range 1-250 microm, by varying the freezing conditions. Freshly lyophilized chitosan scaffolds could be treated with glycosaminoglycans to form ionic complex materials which retained the original pore structure. Chitosan scaffolds could be rehydrated via an ethanol series to avoid the stiffening caused by rehydration in basic solutions. Hydrated porous chitosan membranes were at least twice as extensible as non-porous chitosan membranes, but their elastic moduli and tensile strengths were about tenfold lower than non-porous controls. The methods and structures described here provide a starting point for the design and fabrication of a family of polysaccharide based scaffold materials with potentially broad applicability.

Biocompatible Materials↗

PubMed2004

Scaffold-based tissue engineering: rationale for computer-aided design and solid free-form fabrication systems.

One of the milestones in tissue engineering has been the development of 3D scaffolds that guide cells to form functional tissue. Recently, mouldless manufacturing techniques, known as solid free-form fabrication (SFF), or rapid prototyping, have been successfully used to fabricate complex scaffolds. Similarly, to achieve simultaneous addition of cells during the scaffold fabrication, novel robotic assembly and automated 3D cell encapsulation techniques are being developed. As a result of these technologies, tissue-engineered constructs can be prepared that contain a controlled spatial distribution of cells and growth factors, as well as engineered gradients of scaffold materials with a predicted microstructure. Here, we review the application, advancement and future directions of SFF techniques in the design and creation of scaffolds for use in clinically driven tissue engineering.

Biocompatible Materials↗

PubMed2003

[Research advances in gene-enhanced tissue engineering].

Gene-enhanced engineering deals with the scientific and technologic endeavour to produce cultured cells or polymer matrices transduced with multiple gene vectors encoding cytokine cDNA by means of genetic engineering technique, to make transduced cells or gene activated matrices highly express according cytokine, and then to enhance certain abilities of the artificial tissue. Up to now, various genes encoding modulatory species of ribonucleic of proteins such as growth factors, receptors, and transcription factors have been used in the context of gene-enhanced tissue engineering and expressed within numerous tissues, including artificial blood vessels, bone, cartilage, skin and urinary system, etc. Many experiments in vitro or in vivo have begun to show good prospects and great potential application of the new approach. We believe great changes will take place in the research field of tissue engineering due to the induction the of genetic engineering, and the new approach will become a very promising and valuable tool for therapy.

Blood Vessel Prosthesis↗