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PubMed2006

Biomaterials for adipose tissue engineering.

There is high clinical need for an adequate reconstruction of soft tissue defects as found after tumor resections, deep burns or severe trauma. A promising solution for these defects is adipose tissue engineering, with adult stem cells of the adipose tissue, implanted on 3D biomaterials. These adipogenic precursor cells survive ischemia better than mature adipocytes and have the potency to proliferate and differentiate into fat cells after transplantation. They can be yielded from excised adipose tissue or liposuction material. When preadipocytes are seeded on carriers for the generation of adipose tissue, chemical composition, mechanical stability and 3D architecture of the construct are crucial factors. They ensure cellular penetration into the construct, sufficient proliferation on the material and full differentiation inside the construct after transplantation. In hydrogels, it is especially the use and combination of growth factors that determine the overall outcome of the applied biopolymer. Over recent years, in vivo trials in particular have allowed significant insights into the potential, the perspectives, but also the current difficulties and draw-backs in adipose tissue engineering. This review focuses on the main strategies in adipose tissue regeneration, compares the various materials that have been used as carrier matrices so far and considers them in light of the challenges they have yet to meet.

Adipocytes↗

PubMed2004

The effect of PEGT/PBT scaffold architecture on oxygen gradients in tissue engineered cartilaginous constructs.

Repair of articular cartilage defects using tissue engineered constructs composed of a scaffold and cultured autologous cells holds promise for future treatments. However, nutrient limitation (e.g. oxygen) has been suggested as a cause of the onset of chondrogenesis solely within the peripheral boundaries of larger constructs. In the present study, oxygen gradients were evaluated by microelectrode measurements in two porous polyethylene glycol terephthalate/polybutylene terephthalate (PEGT/PBT) scaffold architectures, a compression-molded and particle-leached sponge (CM) and a 3D-deposited fiber (3DF) scaffold. During the first 14 days in vitro, gradients intensified, after which a gradual decrease of the gradients was observed in vitro. In vivo, however, gradients changed instantly and became less pronounced. Although similar gradients were observed regardless of scaffold type, significantly more cells were present in the center of 3DF constructs after 2 weeks of in vivo culture. Our results stress the importance of a rationally designed scaffold for tissue-engineering applications. Organized structures, such as the 3DF PEGT/PBT polymer scaffolds, offer possibilities for regulation of nutrient supply and, therefore, hold promise for clinical approaches for cartilage repair.

Animals↗

PubMedDate not supplied

Different substitute biomaterials as potential scaffolds in tissue engineering.

PURPOSE: To find the optimal scaffold for tissue-engineered bone, one approach is to test existing biomaterials on their suitability as scaffolds. In this study, the suitability of different alloplastic and xenogenic biomaterials as scaffolds for ex vivo osteoblast cultivation was investigated. MATERIALS AND METHODS: Normal human osteoblast cells were cultured on the surface of bovine collagenous materials, bovine hydroxyapatite, porcine gelatin, synthetic polymer, and collagen-containing bovine hydroxyapatite, and the investigation of proliferation was performed after 24, 72, and 120 hours. Measurement of the differentiation marker alkaline phosphatase and osteocalcin was made after 20 days of incubation. RESULTS: The obtained data showed significantly higher proliferation and differentiation rates in cells cultivated on collagen-rich biomaterials in comparison to noncollagenous or collagen-poor biomaterials (P < .05). DISCUSSION: In tissue engineering the scaffold should be biocompatible and serve as a proper matrix for the cells to produce the new structural environment of extracellular matrix ex vivo. Collagen supports initial cell attachment and cell proliferation, allowing immature osteogenic cells to differentiate into mature osteoblasts, but collagen may not be the only dominating factor for cell-matrix interaction during ex vivo bone formation. CONCLUSION: These data suggest that a 3-dimensional collagen matrix can provide a more favorable environment for the attachment, proliferation, and differentiation of in vitro osteoblastlike cells, at least until the initial stage of differentiation, than noncollagenous biomaterials.

Absorbable Implants↗

PubMed1998

Host response to tissue engineered devices.

The two main components of a tissue engineered device are the transplanted cells and the biomaterial, creating a device for the restoration or modification of tissue or organ function. The implantation of polymer/cell constructs combines concepts of biomaterials and cell transplantation. The interconnections between the host responses to the biomaterial and transplanted cells determines the biocompatibility of the device. This review describes the inflammatory response to the biomaterial component and immune response towards transplanted cells. Emphasis is on how the presence of the transplanted cell construct affects the host response. The inflammatory response towards a biomaterial can impact the immune response towards transplanted cells and vice versa. Immune rejection is the most important host response towards the cellular component of tissue engineered devices containing allogeneic, xenogeneic or immunogenic ex vivo manipulated autologous cells. The immune mechanisms towards allografts and xenografts are outlined to provide a basis for the mechanistic hypotheses of the immune response towards encapsulated cells, with antigen shedding and the indirect pathway of antigen presentation predominating. A review of experimental evidence illustrates examples of the inflammatory response towards biodegradable polymer scaffold materials, examples of devices appropriately integrated as assessed morphologically with the host for various applications including bone, nerve, and skin regeneration, and of the immune response towards encapsulated allogeneic and xenogeneic cells.

Journal Article↗

PubMed1999

Effects of anastomosis of tissue-engineered neointestine to native small bowel.

BACKGROUND: Our laboratory is investigating the tissue engineering of small intestine using intestinal epithelial organoid units seeded onto highly porous biodegradable polymer matrices. This study investigated the effects of anastomosis of tissue-engineered intestine to native small bowel alone or combined with small bowel resection on neointestinal regeneration. METHODS: Intestinal epithelial organoid units harvested from neonatal Lewis rats were seeded onto biodegradable polymer tubes and implanted into the omentum of adult Lewis rats as follows: (1) implantation alone (n = 9); (2) implantation followed by anastomosis to native small bowel at 3 weeks (n = 11); and (3) implantation after small bowel resection and anastomosis to native small bowel at 3 weeks (n = 8). All constructs were harvested at 10 weeks and examined by histology. Morphometric analysis of the neomucosa was obtained using a computer image analysis program. RESULTS: Cyst development was noted in all animals. All anastomoses were patent at 10 weeks. Histology revealed the development of a vascularized tissue with a neomucosa lining the lumen of the cyst with invaginations resembling crypt-villus structures. Morphometric analysis demonstrated significantly greater villus number, villus height, crypt number, crypt area, and mucosal surface length in groups 2 and 3 compared with group 1, and significantly greater villus number, villus height, crypt area, and mucosal surface length in group 3 compared with group 2 (P < 0.05, ANOVA, Tukey test). CONCLUSION: Intestinal epithelial organoid units transplanted on biodegradable polymer tubes can regenerate into complex tissue resembling small intestine. Anastomosis to native small bowel combined with small bowel resection and anastomosis alone contribute significant regenerative stimuli for the morphogenesis and differentiation of tissue-engineered neointestine.

Anastomosis, Surgical↗

PubMed2004

[Tissue engineering of the urinary bladder].

In tissue engineering of the urinary bladder with autologous cell transplantation, high differentiation of the cells cultivated in vitro on biocompatible membranes is essential for the functionality of the tissue constructs after implantation. The terminal differentiation of superficial urothelial cells has a key role because of the barrier function of these cells against urine. The aim of this study was to determine optimized conditions for the creation of terminally differentiated urothelium to cover large membrane surfaces. This can bring us closer to the goal of using functioning tissue constructs in clinical trials.

Absorbable Implants↗

PubMed2001

Craniofacial tissue engineering.

There is substantial need for the replacement of tissues in the craniofacial complex due to congenital defects, disease, and injury. The field of tissue engineering, through the application of engineering and biological principles, has the potential to create functional replacements for damaged or pathologic tissues. Three main approaches to tissue engineering have been pursued: conduction, induction by bioactive factors, and cell transplantation. These approaches will be reviewed as they have been applied to key tissues in the craniofacial region. While many obstacles must still be overcome prior to the successful clinical restoration of tissues such as skeletal muscle and the salivary glands, significant progress has been achieved in the development of several tissue equivalents, including skin, bone, and cartilage. The combined technologies of gene therapy and drug delivery with cell transplantation will continue to increase treatment options for craniofacial cosmetic and functional restoration.

Adipose Tissue↗

PubMed2001

Tissue engineering of small caliber vascular grafts.

OBJECTIVE: Previous tissue engineering approaches to create small caliber vascular grafts have been limited by the structural and mechanical immaturity of the constructs. This study uses a novel in vitro pulse duplicator system providing a 'biomimetic' environment during tissue formation to yield more mature, implantable vascular grafts. METHODS: Vascular grafts (I.D. 0.5 cm) were fabricated from novel bioabsorbable polymers (polyglycolic-acid/poly-4-hydroxybutyrate) and sequentially seeded with ovine vascular myofibroblasts and endothelial cells. After 4 days static culture, the grafts (n=24) were grown in vitro in a pulse duplicator system (bioreactor) for 4, 7, 14, 21, and 28 days. Controls (n=24) were grown in static culture conditions. Analysis of the neo-tissue included histology, scanning electron microscopy (SEM), and biochemical assays (DNA for cell content, 5-hydroxyproline for collagen). Mechanical testing was performed measuring the burst pressure and the suture retention strength. RESULTS: Histology showed viable, dense tissue in all samples. SEM demonstrated confluent smooth inner surfaces of the grafts exposed to pulsatile flow after 14 days. Biochemical analysis revealed a continuous increase of cell mass and collagen to 21 days compared to significantly lower values in the static controls. The mechanical properties of the pulsed vascular grafts comprised supra-physiological burst strength and suture retention strength appropriate for surgical implantation. CONCLUSIONS: This study demonstrates the feasibility of tissue engineering of viable, surgically implantable small caliber vascular grafts and the important effect of a 'biomimetic' in vitro environment on tissue maturation and extracellular matrix formation.

Absorbable Implants↗

PubMed2005

[Bone defect repair with a new tissue-engineered bone carrying bone morphogenetic protein in rabbits].

OBJECTIVE: To construct a new tissue-engineered bone with poly (D, L-lactide-co-glycolide) (PLGA), bone morphogenetic protein (BMP) and bone marrow-derived stem cells (BMSCs) and observe its effect in repairing segmental bone defects. METHODS: A 15-mm bone defect in the right radius was induced in New Zealand white rabbits, and the models were randomized into three groups to receive implantation of the tissue-engineered bone grafts constructed with PLGA carrying 5 mg BMP and about 1 x 10(6) BMSCs (experimental group), grafts of PLGA with about 1 x 10(6) BMSCs (control group), or grafts of exclusive PLGA (blank control group), respectively. The osteogenesis in the bone defect after the implantation on was evaluated X-ray films, and the histological changes of the tissues sampled from the bone defect 4, 8, and 12 weeks after operation were observed and new bone formation was measured by image analysis. RESULTS: The bone defect was completely repaired in the experimental group 12 weeks after the implantation, showing the best results among the 3 groups. The bone defects in the blank control group was filled with only fibrous and connective tissues at 12 weeks. CONCLUSION: This tissue-engineered bone constructed with PLGA, BMP and BMSCs possesses good ability in repairing segmental bone defect.

Animals↗

PubMedDate not supplied

Vitrification of tissue engineered pancreatic substitute.

Despite significant advances, some critical issues remain for the long-term storage of an engineered pancreas. In this study we employed a tissue engineered pancreatic substitute model-insulin-secreting betaTC3 cells entrapped in calcium alginate/poly-L-lysine/alginate beads-to demonstrate that a prototype vitrification method can prevent ice formation and maintain cell viability/function. The results showed that the structure of the frozen samples was distorted by ice crystals throughout the matrix. In marked contrast, the vitrified samples appeared to be free of ice. Morphologic studies demonstrated extensive fractures and vacuolation in frozen specimens while there were no fractures in vitrified TEPSs. Both vitrified and frozen constructs showed some vacuolization compared to the control samples. Frozen beads showed a significantly decreased viability compared to fresh controls and the VS55 group (P < .001). There was no significant difference between the vitrified and fresh samples. Vitrification using the VS55 protocol shows similar viability and secretion properties to the control group of fresh beads. Vitrification using the PEG 400 protocol resulted in slightly lower viability and secretion properties relative to the control group; conventional freezing resulted in even significantly lower viability and secretion properties. These results combine to demonstrate feasibility of vitrification as a storage method for a tissue engineered pancreas.

Animals↗

PubMed2004

Preincubation of tissue engineered constructs enhances donor cell retention.

Cartilage tissue engineering has been the focus of considerable research. However, the fate of transplanted donor cells rarely is explored directly. In the current study, the effect of preincubating perichondrial cells into a polylactic acid scaffold before implantation into an osteochondral defect was studied. The extracellular matrix produced during preincubation was characterized; the viability of the donor cells was assessed; and the retention of the donor cells in the repair tissue was determined using a gene marker on the Y chromosome, the gender-determining region Y gene. During in vitro incubation, the cells produced an extracellular matrix consisting of glycosaminoglycans, and Types I and II collagen, and the cell viability remained great. In vivo, preincubated constructs had significantly greater retention of donor cells in the host repair tissue in the short term when compared with nonincubated controls. This study shows the value of preincubating engineered constructs before implantation, and additionally validates the gender-determining region Y gene as an effective tool for assessing the fate of donor cells in cartilage tissue engineering.

Animals↗

PubMed2005

The use of pimonidazole to characterise hypoxia in the internal environment of an in vivo tissue engineering chamber.

UNLABELLED: The distribution of hypoxic cells in an in vivo tissue engineering chamber was investigated up to 28 days post-implantation. METHODS: Arteriovenous loops were constructed and placed into bi-valved polycarbonate chambers containing 2 x 10(6) rat fibroblasts in basement membrane gel (BM gel). Chambers were inserted subcutaneously in the groin of male rats and harvested at 3 (n = 6), 7 (n = 6), 14 (n = 4) or 28 (n = 4) days. Ninety minutes before harvest, pimonidazole (60 mg/kg) was injected intraperitoneally. Chamber tissue was removed, immersion fixed, paraffin embedded, sectioned and stained immunohistochemically using hypoxyprobe-1 Mab that detects reduced pimonidazole adducts forming in cells, where pO2 < 10 mmHg. RESULTS: At 3 days a fibrin clot/BM gel framework filled the chamber. Seeded fibroblasts had largely died. The majority of 3 day chambers did not demonstrate tissue growth from the AV loop nor was pimonidazole binding present in these chambers. In one chamber in which tissue growth had occurred strong pimonidazole binding was evident within the new tissue. In four out of six 7 day chambers a broader proliferative zone existed extending up to 0.4 mm (approximately) from the AV loop endothelium which demonstrated intense pimonidazole binding. The two remaining 7 day chambers displayed even greater tissue growth (leading edge > 0.7 mm from the AV loop endothelium), but very weak or no pimonidazole binding. At 14 and 28 days the fibrin/BM gel matrix was replaced by mature vascularised connective tissue that did not bind pimonidazole. CONCLUSION: Employing a tissue engineering chamber, new tissue growth extending up to 0.4 mm from the AV loop endothelium (chambers < or = 7 days) demonstrated intense pimonidazole binding and, therefore, hypoxia. Tissue growth greater than 0.5 mm from the AV loop endothelium (7-28 days chambers) did not exhibit pimonidazole binding due to a significant increase in the number of new blood vessels and was, therefore, adequately oxygenated.

Animals↗

PubMed1999

[Tissue engineering].

The new field of tissue engineering, applies the principles of biology and engineering to the development of functional substitutes for the loss or failure of an organ or tissue. This article describes the previous challenges and present status of this interdisciplinary field.

Animals↗

PubMed2007

Electrospun fibrinogen: feasibility as a tissue engineering scaffold in a rat cell culture model.

Fibrinogen has a well-established tissue engineering track record because of its ability to induce improved cellular interaction and scaffold remodeling compared to synthetic scaffolds. While the feasibility of electrospinning fibrinogen scaffolds of submicron diameter fibers and their mechanical properties have been demonstrated, in vitro cellular interaction has not yet been evaluated. The goal of this study was to demonstrate, based on cellular interaction and scaffold remodeling, that electrospun fibrinogen can be used successfully as a tissue engineering scaffold. Electrospun fibrinogen scaffolds were disinfected, seeded with neonatal rat cardiac fibroblasts, and cultured for 2, 7, and 14 days. Cultures were treated to regulate scaffold degradation by either supplementing serum-containing media with aprotinin or crosslinking the scaffolds with glutaraldehyde vapor. Biocompatibility was assessed through a WST-1 cell proliferation assay. Postculture scaffolds were evaluated by scanning electron microscopy and histology. Cell culture demonstrated that fibroblasts readily migrate into and remodel electrospun fibrinogen scaffolds with deposition of native collagen. Supplementation of culture media with different concentrations of aprotinin-modulated scaffold degradation in a predictable fashion, but glutaraldehyde vapor fixation was less reliable. Based on the observed cellular interactions, there is tremendous potential for electrospun fibrinogen as a tissue engineering scaffold.

Animals↗

PubMed2000

[Huge thoracic wall defect repaired by tissue engineered bone transplantation].

OBJECTIVE: A rare huge desmoplastic fibroma on thoracic wall in 1 female case of 25 years old was resected, and the accompanying huge thoracic wall defect, ribs and soft tissues were repaired by tissue engineered bone and pedicled flap. The paper aims to explore the clinical results of early stage after operation. METHODS: Autogeneic bone marrow stromal cells (MSC) were obtained from bone marrow puncture of iliac bone and isolated and cultured according to the Houghton's methods, MSC were directively induced and differentiated to osteoblasts. Allogeneic ribs were made to the bio-derived bone scaffold materials after treatment of decell, deantigen, decalcification and dry freezing. 5 x 10(6)/ml MSC were cocultured with the bio-derived bone for 6 days in vitro. After intact resection of tumor, the diaphragm flap was applied to repair pleural cavity, the three defect ribs were repaired by tissue engineered bone and the soft tissue defect was repaired by transfer of pedicled ipsilateral abdominal flaps. RESULTS: The patient recovered well with first intention. Followed up for 3 months, tissue engineered ribs were matured in vitro and the heart and pulmonary functions were improved markedly. CONCLUSION: The tissue engineered bone constructed by autogeneic MSC is advantageous in individual treatment.

Adult↗

PubMed2006

Enhancement of in vivo endothelialization of tissue-engineered vascular grafts by granulocyte colony-stimulating factor.

Successful reconstruction of large-diameter blood vessel in humans has been demonstrated using the tissue engineering technique, but improvement in patency of small-diameter bioartificial vascular graft remains a great challenge. This study reports that granulocyte colony-stimulating factor (G-CSF) can enhance in vivo endothelialization of tissue-engineered vascular grafts, which could be used to improve patency of small-diameter vascular graft. Vascular grafts were tissue engineered with decellularized canine abdominal aortas and canine autologous bone marrow-derived cells. Prior to cell seeding onto decellularized graft matrices, bone marrow-derived cells were induced to differentiate into endothelial cells and smooth muscle cells. The cell-seeded vascular grafts were implanted into the abdominal aortas of bone marrow donor dogs. Before and after graft implantation, G-CSF was administered subcutaneously to the dogs (n = 3). The grafts implanted into the dogs not receiving G-CSF were used as controls (n = 3). Eight weeks after implantation, grafts in both groups showed regeneration of vascular tissues including endothelium and smooth muscle. Importantly, endothelium formation was more extensive in the G-CSF-treated grafts than in the control grafts, as assessed with reverse transcription polymerase chain reaction, western blot, and immunohistochemistry. In addition, intimal hyperplasia was significantly reduced in the G-CSF-treated grafts compared to the control grafts. This study suggests that G-CSF administration could be applied to improve patency of small-diameter tissue-engineered vascular grafts.

Animals↗

PubMed2005

In vitro cartilage tissue engineering with 3D porous aqueous-derived silk scaffolds and mesenchymal stem cells.

Adult cartilage tissue has limited self-repair capacity, especially in the case of severe damages caused by developmental abnormalities, trauma, or aging-related degeneration like osteoarthritis. Adult mesenchymal stem cells (MSCs) have the potential to differentiate into cells of different lineages including bone, cartilage, and fat. In vitro cartilage tissue engineering using autologous MSCs and three-dimensional (3-D) porous scaffolds has the potential for the successful repair of severe cartilage damage. Ideally, scaffolds designed for cartilage tissue engineering should have optimal structural and mechanical properties, excellent biocompatibility, controlled degradation rate, and good handling characteristics. In the present work, a novel, highly porous silk scaffold was developed by an aqueous process according to these criteria and subsequently combined with MSCs for in vitro cartilage tissue engineering. Chondrogenesis of MSCs in the silk scaffold was evident by real-time RT-PCR analysis for cartilage-specific ECM gene markers, histological and immunohistochemical evaluations of cartilage-specific ECM components. Dexamethasone and TGF-beta3 were essential for the survival, proliferation and chondrogenesis of MSCs in the silk scaffolds. The attachment, proliferation, and differentiation of MSCs in the silk scaffold showed unique characteristics. After 3 weeks of cultivation, the spatial cell arrangement and the collagen type-II distribution in the MSCs-silk scaffold constructs resembles those in native articular cartilage tissue, suggesting promise for these novel 3-D degradable silk-based scaffolds in MSC-based cartilage repair. Further in vivo evaluation is necessary to fully recognize the clinical relevance of these observations.

Adult↗

PubMed2003

Current state of cartilage tissue engineering.

Damage to cartilage is of great clinical consequence given the tissue's limited intrinsic potential for healing. Current treatments for cartilage repair are less than satisfactory, and rarely restore full function or return the tissue to its native normal state. The rapidly emerging field of tissue engineering holds great promise for the generation of functional cartilage tissue substitutes. The general approach involves a biocompatible, structurally and mechanically sound scaffold, with an appropriate cell source, which is loaded with bioactive molecules that promote cellular differentiation and/or maturation. This review highlights aspects of current progress in cartilage tissue engineering.

Animals↗