| 1. |
- Ahmed, Tamer A. E., et al.
(författare)
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Characterization and inhibition of fibrin hydrogel-degrading enzymes during development of tissue engineering scaffolds
- 2007
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Ingår i: Tissue engineering. - : Mary Ann Liebert. - 1076-3279 .- 1557-8690. ; 13:7, s. 1469-1477
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Tidskriftsartikel (refereegranskat)abstract
- The goal of articular cartilage tissue engineering is to provide cartilaginous constructs to replace abnormal cartilage. We have evaluated the chondroprogenitor clonal cell line RCJ3.1C5.18 (C5.18) as a model to guide the development of appropriate scaffolds for tissue engineering. Rapid degradation of fibrin hydrogels was observed after encapsulation of C5.18 cells. The enzymes responsible for this fibrin gel breakdown were characterized to control their activity and regulate gel stability. Western blotting, confirming zymography, revealed bands due to matrix metalloproteinases (MMP-2, MMP-3) that are secreted concomitantly with fibrin hydrogels breakdown. High plasmin activity was detected in conditioned media during hydrogel breakdown but not in the confluent cells before encapsulation. Reverse transcriptase polymerase chain reaction indicated the expression of MMP-2, -3, and -9 and plasminogen in the cells. MMP-9 was 100 times higher at day 1, whereas MMP-2 started to increase and reached its maximum level by day 7. Aprotinin, a known serine protease inhibitor, and galardin (GM6001), a potent MMP inhibitor, in combination or separately, prevented the breakdown of fibrin-C5.18 hydrogels, whereas only the combination of both promoted the accumulation of extracellular matrix. These findings suggest that plasmin and MMPs contribute independently to fibrin hydrogel breakdown, but that either enzyme can achieve extracellular matrix breakdown.
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| 2. |
- Ananta, M., et al.
(författare)
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A Novel Poly(L-Lactide-co-e-Caprolactone)-Collagen Hybrid Construct for Application in Tissue Engineering
- 2007
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Ingår i: Termis-EU Meeting Abstracts, London, UK September 4-7 2007. - : Mary Ann Liebert Inc.. ; , s. 1637-1637
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- A biodegradable hybrid construct consisting of a slow degrading poly(L-lactide-co-e-caprolactone) (PLA-e-CL) knitted mesh, plastically compressed (1) between two collagen gels was fabricated and tested in vitro for tissue engineering applications. The polymer mesh was incorporated to give greater mechanical stability to the compressed collagen scaffolds. The hybrid construct was characterized for fluid (weight) loss and cell viability during compression and mechanical properties. Hybrid constructs embedded and surface layered with human dermal fibroblasts (2, Eþ5 per 5 ml) were cultured for up to one week in static culture. Quantitative and qualitative data on cell viability and proliferation were obtained. It was found that the fluid (weight) loss in plastic compression of the hybrid construct was time dependent and not weight dependent at an applied load of 240 grams. No significant cell death was observed during the plastic compression process and a homogenous cell distribution was achieved. One week of static culture showed that the cultivated hybrid construct retained its mechanical properties with no evidence of degradation, and cells inside the constructs as well as layered on top of the constructs proliferated. We found the PLA-e-CL-Collagen hybrid construct a useful three-dimensional scaffold for tissue engineering of stratified tissues and potential applications in bladder wall, blood vessels and skin are currently being explored.
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| 3. |
- Armstrong, Stephanie J., et al.
(författare)
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ECM molecules mediate both Schwann cell proliferation and activation to enhance neurite outgrowth
- 2007
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Ingår i: Tissue engineering. - : Mary Ann Liebert. - 1076-3279 .- 1557-8690. ; 13:12, s. 2863-2870
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Tidskriftsartikel (refereegranskat)abstract
- Tissue engineering using a combination of biomaterials and cells represents a new approach to nerve repair. We have investigated the effect that extracellular matrix (ECM) molecules have on Schwann cell (SC) attachment and proliferation on the nerve conduit material poly-3-hydroxybutyrate (PHB), and SC influence on neurite outgrowth in vitro. Initial SC attachment to PHB mats was unaffected by ECM molecules but proliferation increased (laminin > fibronectin > collagen). SCs seeded onto ECM-coated culture inserts suspended above a monolayer of NG108-15 cells determined the effect of released diffusible factors. The effect of direct contact between the two cell types on ECM molecules was also investigated. In both systems SCs enhanced neurite number per cell and percentage of NG108-15 cells sprouting neurites. NG108-15 cells grown in direct contact with SCs had significantly longer neurites than those exposed to diffusible factors when seeded on laminin or fibronectin. Diffusible factors released from SCs cultured on ECM molecules appear to initiate neurite outgrowth, whereas SC-neuron contact promotes neurite elongation. SC proliferation was maximal on poly-D-lysine-coated surfaces, but these cells did not influence neurite outgrowth to the levels of laminin or fibronectin. This suggests that ECM molecules enhance cell number and activate SCs to release neurite promoting factors. Addition of ECM molecules to PHB nerve conduits containing SCs is likely to provide benefits for the treatment of nerve injuries.
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| 4. |
- Aulin, Cecilia, et al.
(författare)
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Designing Extracellular Matrix Scaffolds by Dynamic culture of fibroblasts
- 2007
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Ingår i: TERMIS-EU Meeting Abstracts London, UK September 4–7, 2007. - : Mary Ann Liebert. ; , s. 1667-1667
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Our bodies are constantly exposed to different sorts of mechanical forces, from muscle tension to wound healing. Connective tissue adapts its extracellular matrix (ECM) to changes in mechanical load and the influence of mechanical stimulation on fibroblasts has been studied for a long time [1, 2]. When exposed to forces, fibroblasts are known to respond with expression and remodeling of ECM proteins, in particular collagen type I [3]. In this study the effect of dynamic culture conditions on human dermal fibroblasts was evaluated in terms of deposition and remodeling of ECM, with the aim of producing an ECM based scaffold. The fibroblasts were grown on compliant polymer supports either in a bioreactor with a pulsating flow or under static conditions. By applying dynamic culture conditions, the collagen deposition on the polymer supports increased fivefold. Scanning electron microscopy showed that polymer fibers were well integrated with cells and ECM and alignment along the polymer fibers was observed. Scaffold design should aim at creating structures that can help guiding the cells to form new, functional tissue. The presented system may present a new way of producing designed extracellular matrix based scaffolds for tissue engineering.
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| 5. |
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| 6. |
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| 7. |
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| 8. |
- Heydarkhan-Hagvall, Sepideh, 1969, et al.
(författare)
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Production of extracellular matrix components in tissue-engineered blood vessels
- 2006
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Ingår i: Tissue engineering. - : Mary Ann Liebert Inc. - 1076-3279 .- 1557-8690. ; 12:4, s. 831-42
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Tidskriftsartikel (refereegranskat)abstract
- Morphology and compliance of tissue-engineered blood vessels (TEBV) are dependent on the culture period and production of extracellular matrix (ECM) components in order to increase the strength of the developing tissue. The aim of the present study was to evaluate the potential of TEBVs to produce an ECM similar to native arteries and veins. Human smooth muscle cells (SMC) were seeded onto the poly(glycolic acid) (PGA) scaffold and placed in bioreactors filled with DMEM supplemented with growth factors. After 6 weeks, the vessels were harvested from the bioreactors and seeded with human endothelial cells at the lumen for another 3 days. Then, the TEBVs were harvested for RNA and protein isolation for further RT-PCR and Western blot. TEBVs had a similar macroscopic appearance to that of native vessels with no visible evidence of the original PGA. Histological and immunohistochemical analyses indicated the presence of high cell density and development of a highly organized structure of ECM. After 6 weeks of culture, there were significantly lower gene expression of SMC-specific markers, such as alpha-actin, caldesmon, and vimentin, and proteoglycans, such as biglycan, decorin, and versican, and other ECM components, such as collagen I and elastin, in TEBVs, with and without pulsatile conditions, compared to that of native arteries. Gene expression of fibronectin was significantly lower in TEBVs grown during pulsatile conditions compared to that of native arteries. No difference was observed in TEBVs grown during non-pulsatile conditions. The presence of alpha-actin, collagen I, decorin, and fibronectin at protein level was demonstrated in TEBVs with and without pulsatile conditions after 6 weeks and in native veins and arteries as well. How this deviation translates into mechanical properties remains to be explored.
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| 9. |
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| 10. |
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| 11. |
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| 12. |
- Pulliainen, Outi, et al.
(författare)
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Poly-L-D-lactic acid scaffold in the repair of porcine knee cartilage lesions.
- 2007
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Ingår i: Tissue engineering. - : Mary Ann Liebert Inc. - 1076-3279 .- 1557-8690. ; 13:6, s. 1347-55
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Tidskriftsartikel (refereegranskat)abstract
- Articular cartilage injuries cause a major clinical problem because of the negligible repair capacity of cartilage. Autologous chondrocyte transplantation is a surgical method developed to repair cartilage lesions. In the operation, cartilage defect is covered with a periosteal patch and the suspension of cultured autologous chondrocytes is injected into the lesion site. The method can form good repair tissue, but new techniques are needed to make the operation easier and to increase the postoperative biomechanical properties of the repair tissue. In this study, we investigated poly-L,D-lactic acid (PLDLA) scaffolds alone or seeded with autologous chondrocytes in the repair of circular 6-mm cartilage lesions in immature porcine knee joints. Spontaneous repair was used as a reference. Histologic evaluation of the repair tissue showed that spontaneous repair exhibited higher scores than either PLDLA scaffold group (with or without seeded chondrocytes). The scaffold material was most often seen embedded in the subchondral bone underneath the defect area, probably because of the hardness of the PLDLA material. However, some of the cell-seeded and nonseeded scaffolds contained cartilaginous tissue, suggesting that invasion of mesenchymal cells inside nonseeded scaffolds had occurred. Hyaluronan deposited in the scaffold had possibly acted as a chemoattractant for the cell recruitment. In conclusion, the PLDLA scaffold material used in this study was obviously mechanically too hard to be used for cartilage repair in immature animals.
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| 13. |
- Scherman, Peter, et al.
(författare)
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Local effects on triiodothyronine-treated polyglactin sutures on regeneration across peripheral nerve defects.
- 2004
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Ingår i: Tissue Engineering. - : Mary Ann Liebert Inc. - 1076-3279 .- 1557-8690. ; 10:3-4, s. 455-464
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Tidskriftsartikel (refereegranskat)abstract
- We have previously described a new and simple method for nerve repair in which continuous longitudinal polyglactin sutures alone are used to bridge limited nerve defects in rats. Here we examined whether such sutures could be used to deliver a growth-promoting substance, triiodothyronine (T3), and enhance regeneration of the rat sciatic nerve. Sutures were pretreated in highly concentrated solutions of T3 for 24 h. In vitro measurements showed that such sutures released T3 with an initial rapid phase followed by a slow-release phase lasting at least 3 weeks. Bilateral sciatic nerve defects (7 mm) in rats were bridged by either T3- or saline-incubated sutures. Immunocytochemistry for Schwann cells and axons at 2 weeks showed no differences in Schwann cell distribution or axonal outgrowth length. Morphometric analysis 4 and 12 weeks after the repair revealed a slight but significant (p < 0.05) increase in the mean myelin area in T3-treated nerves. No differences were seen in the number of axons or return of force in the gastrocnemius muscle at 12 weeks. The results show that sutures can be used both for the bridging of defects in peripheral nerves and for the delivery of a growth-promoting substance to regenerating nerve structures.
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| 14. |
- Stading, Mats, et al.
(författare)
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Mechanical shear properties of cell-polymer cartilage constructs
- 1999
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Ingår i: Tissue engineering. - : Mary Ann Liebert Inc. - 1076-3279 .- 1557-8690. ; 5:3, s. 241-250
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Tidskriftsartikel (refereegranskat)abstract
- Cartilaginous constructs were created by using bovine chondrocytes on synthetic, biodegradable scaffolds made of fibrous polyglycolic acid (PGA). The constructs have previously been shown to resemble natural articular cartilage biochemically and histologically. The mechanical properties of articular cartilage mainly depend on the swollen extracellular matrix (ECM), which is a gel consisting of collagen fibers and proteoglycans in a fluid phase of water and electrolytes. The biomechanical properties of the constructs and the build-up of the ECM were studied using dynamic, nondestructive measurements in shear. A small, harmonic strain, ? ? 5 x 10-4, was applied to the sample, and the resulting stress was recorded and used for calculating the complex shear modulus G*. The applied strain was much smaller than that used in confined compression. The shear modulus G* correlated well with both the collagen and glycosaminoglycan content of the constructs but did not reach the same level as in natural cartilage. Collagen is the dominant component contributing to the shear strength of cartilage, and G* was shown to depend approximately quadratically on the collagen content of the constructs. The difference in G* between the constructs and natural cartilage was shown to depend on both the biochemical composition and the microstructure of the constructs.
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| 15. |
- Tohill, Mel P, et al.
(författare)
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Green fluorescent protein is a stable morphological marker for schwann cell transplants in bioengineered nerve conduits
- 2004
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Ingår i: Tissue engineering. - Larchmont, NY : Mary Ann Liebert, Inc.. - 1076-3279 .- 1557-8690. ; 10:9-10, s. 1359-1367
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Tidskriftsartikel (refereegranskat)abstract
- Bioengineered systems incorporate cultured cells to mimic the substituted tissue. A labeling method is necessary to monitor the survival of transplanted cells within the host. This labeling method must be compatible with the histochemical methods used for morphological analysis. This study assessed (1) The in vitro characteristics of Schwann cells (SCs) labeled with green fluorescent protein (GFP), (2) the in vivo effect of transplanted GFP-SCs in a model of peripheral nerve injury, and (3) the compatibility of GFP-SCs with immunofluorescence histochemical techniques. SCs were retrovirally labeled with GFP and their growth characteristics were compared with those of nontransduced SCs (ntSCs). GFP-SCs were seeded in a resorbable nerve conduit for grafting into a 1-cm gap in rat sciatic nerve. Grafts were harvested after 2 weeks and immunofluorescent staining was performed to measure axonal and SC regeneration distances and to identify GFP-SCs. Results of GFP-SC vitality assays did not vary significantly from those of ntSC assays. GFP-SCs were readily located ex vivo and stimulated significantly better axonal and SC regeneration distances in comparison with empty conduits. These findings show that GFP labeling does not have a deleterious effect on SCs and that it is a useful labeling method for the study of bioengineered systems.
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| 16. |
- Helenius, Gisela, 1973, et al.
(författare)
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Expression of fibrinolytic and coagulation factors in cocultured human endothelial and smooth muscle cells
- 2004
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Ingår i: Tissue engineering. - 1076-3279. ; 10:3-4, s. 353-60
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Tidskriftsartikel (refereegranskat)abstract
- Interactions between endothelial cells and smooth muscle cells are interesting from a tissue-engineering point of view. We have developed a coculture system that allows direct contact between these two cell types. The fibrinolytic factors PAI-1, tPA, and uPA and the coagulation factor TF, were studied at the gene level by RT-PCR and at the protein level by ELISA. Significant changes of all studied factors were seen at the gene level in cocultured endothelial cells. tPA and TF were upregulated 4- and 7-fold, respectively, and PAI-1 and uPA were downregulated 4- and 1.5-fold, respectively, compared with single-cultured controls. In cocultured smooth muscle cells alterations of PAI-1 and TF were significant, with a 1.5-fold upregulation of PAI-1 and a 2.5-fold downregulation of TF. Results at the protein level mirrored the gene expression results. These findings indicate that cocultured endothelial cells are rendered both hypercoagulative and hyperfibrinolytic.
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| 17. |
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| 18. |
- Nishiura, Y, et al.
(författare)
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Addition of cultured Schwann cells to tendon autografts and freeze-thawed muscle grafts improves peripheral nerve regeneration
- 2004
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Ingår i: Tissue Engineering. - 1076-3279. ; 10:1-2, s. 157-164
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Tidskriftsartikel (refereegranskat)abstract
- The effects of addition of Schwann cells on peripheral nerve regeneration through a novel graft material-the tendon autograft-and a conventional freeze-thawed muscle graft, were studied in the rat sciatic nerve. Adult Schwann cell cultures were established from predegenerated nerves. The Schwann cells were added to the autologous grafts by coculture (tendon autograft) or injection (freeze-thawed muscle graft). Both graft types supported adherence of the added Schwann cells. Addition of cultured Schwann cells to the two different graft models improved regeneration by increasing the rate of axonal outgrowth as compared with similar grafts without added cells.
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| 19. |
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