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| A survey on cell and engineered tissue therapies in Europe in 2008.
February 27, 2010 at 2:08 PM
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| | A survey on cell and engineered tissue therapies in Europe in 2008. Tissue Eng Part A. 2010 Feb 25; Authors: Martin I, Baldomero H, Tyndall A, Niederwieser D, Gratwohl A Cellular therapy is an evolving investigational treatment modality in regenerative medicine but little published information is available on its current use. Starting from the established European group for Blood and Marrow Transplantation (EBMT) activity survey on hematopoietic stem cell (HSC) transplantation, a joint committee of four major scientific organizations made a coordinated attempt to collect detailed information in Europe for the year 2008. Thirty-three teams from 16 countries reported data on 656 patients to a "novel cellular therapy" survey, which were combined to additional 384 records reported to the standard EBMT survey. Indications were cardiovascular (29%; 100% autologous), musculoskeletal (18%; 97% autologous), neurological (9%; 39% autologous), epithelial/parenchymal (9%; 18% autologous), autoimmune diseases (12%; 77% autologous) or graft-vs-host-disease (23%; 13% autologous). Reported cell types were HSC (39%), mesenchymal stromal cells (47%! ), chondrocytes (5%), keratinocytes (7%), myoblasts (2%) and others (1%). In 51% of the grafts, cells were delivered following expansion, in 4% of the cases cells were transduced. Cells were delivered intravenously (31%), intraorgan (45%), on a membrane or gel (14%) or using 3D scaffolds (10%). This data collection platform is expected to capture and foresee trends for novel cellular therapies in Europe, and warrants further consolidation and extension. PMID: 20184422 [PubMed - as supplied by publisher] | | |
| Injectable gellan gum hydrogels with autologous cells for the treatment of rabbit articular cartilage defects.
February 27, 2010 at 7:09 AM
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| | Injectable gellan gum hydrogels with autologous cells for the treatment of rabbit articular cartilage defects. J Orthop Res. 2010 Feb 24; Authors: Oliveira JT, Gardel LS, Rada T, Martins L, Gomes ME, Reis RL In this work, the ability of gellan gum hydrogels coupled with autologous cells to regenerate rabbit full-thickness articular cartilage defects was tested. Five study groups were defined: (a) gellan gum with encapsulated chondrogenic predifferentiated rabbit adipose stem cells (ASC + GF); (b) gellan gum with encapsulated nonchondrogenic predifferentiated rabbit adipose stem cells (ASC); (c) gellan gum with encapsulated rabbit articular chondrocytes (AC) (standard control); (d) gellan gum alone (control); (e) empty defect (control). Full-thickness articular cartilage defects were created and the gellan gum constructs were injected and left for 8 weeks. The macroscopic aspect of the explants showed a progressive increase of similarity with the lateral native cartilage, stable integration at the defect site, more pronouncedly in the cell-loaded constructs. Tissue scoring showed that ASC + GF exhibited the best results regarding tissue quality progression. Alcian blue! retrieved similar results with a better outcome for the cell-loaded constructs. Regarding real-time PCR analyses, ASC + GF had the best progression with an upregulation of collagen type II and aggrecan, and a downregulation of collagen type I. Gellan gum hydrogels combined with autologous cells constitute a promising approach for the treatment of articular cartilage defects, and adipose derived cells may constitute a valid alternative to currently used articular chondrocytes. (c) 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res. PMID: 20187118 [PubMed - as supplied by publisher] | | |
| Extracellular matrix expression of human tenocytes in three-dimensional air-liquid and PLGA cultures compared with tendon tissue: Implications for tendon tissue engineering.
February 27, 2010 at 7:09 AM
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| | Extracellular matrix expression of human tenocytes in three-dimensional air-liquid and PLGA cultures compared with tendon tissue: Implications for tendon tissue engineering. J Orthop Res. 2010 Feb 24; Authors: Stoll C, John T, Endres M, Rosen C, Kaps C, Kohl B, Sittinger M, Ertel W, Schulze-Tanzil G Tenocyte transplantation may prove to be an approach to support healing of tendon defects. Cell-cell and cell-matrix contacts within three-dimensional (3D) cultures may prevent tenocyte dedifferentiation observed in monolayer (2D) culture. The present study compares both neotissue formation and tenocyte extracellular matrix (ECM) expression in 2D and 3D cultures directly with that of native tendon, in order to determine optimal conditions for tendon tissue engineering. Primary human tenocytes were embedded in poly[lactic-co-glycolic-acid] (PLGA)-scaffolds and high-density cultures. Neotissue formation was examined by hematoxyline-eosine (H&E) and immunofluorescence staining. Gene expression of ECM proteins and vascular endothelial growth factor (VEGF) was compared at days 0 (2D), 14, and 28 in 3D cultures and tendon. Histomorphology of 3D culture showed tendon-like tissue as tenocyte cell nuclei became more elongated and ECM accumulated. Type I collagen gene e! xpression was higher in 2D culture than in tendon and decreased in 4-week-old 3D cultures, whereas type III collagen was only elevated in high-density culture compared with tendon. Decorin and COMP were reduced in 2D and increased in 3D culture almost to ex vivo level. These results suggest that the 3D high-density or biodegradable scaffolds cultures encourage the differentiation of expanded monolayer tenocytes in vitro to tendon-like tissue. (c) 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res. PMID: 20187116 [PubMed - as supplied by publisher] | | |
| Age dependence of expression of growth factor receptors in porcine ACL fibroblasts.
February 27, 2010 at 7:09 AM
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| | Age dependence of expression of growth factor receptors in porcine ACL fibroblasts. J Orthop Res. 2010 Feb 22; Authors: Vavken P, Saad FA, Murray MM Tissue engineering approaches that harness the stimulatory power of platelet-rich plasma have produced encouraging results in anterior cruciate ligament (ACL) repair. However, a number of recent studies have demonstrated age-dependent differences in cellular responses to such an approach. Identifying the reasons for these differences would allow counteracting them and consequently improve outcomes. In this study we hypothesized that these age-related effects are caused by differences in the expression of the receptors for growth factors released from platelet-rich plasma (PRP). Porcine ACL fibroblasts from a predetermined number of animals of different ages were obtained, and mRNA levels of the receptors of platelet-derived growth factor (PDGF), transforming growth factor beta (TGF-beta), fibroblast growth factor (FGF), and vascular endothelial growth factor (VEGF) were determined. Expression levels were compared across age groups (young and adolescent) and regres! sed on age in days. While no significant difference was seen across groups, the regression analysis showed decreases in receptor expression with increasing age. These differences were statistically significant for TGF-beta receptor 1, FGF receptor, and VEGF receptor 2; and borderline significant for TGF-beta receptor 3 and PDGF receptor. The only receptor that was not associated with age was VEGF receptor 1, a regulator of VEGF receptor 2. These findings suggest that the decrease in growth factor receptor expression as a likely reason for reduced PRP action with increasing age. (c) 2010 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res. PMID: 20186834 [PubMed - as supplied by publisher] | | |
| Calcium phosphate cement reinforcement by polymer infiltration and in situ curing: A method for 3D scaffold reinforcement.
February 27, 2010 at 7:09 AM
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| | Calcium phosphate cement reinforcement by polymer infiltration and in situ curing: A method for 3D scaffold reinforcement. J Biomed Mater Res A. 2010 Feb 22; Authors: Alge DL, Chu TM This study describes a novel method of calcium phosphate cement reinforcement based on infiltrating a pre-set cement with a reactive polymer and then cross-linking the polymer in situ. This method can be used to reinforce 3D calcium phosphate cement scaffolds, which we demonstrate using poly(ethylene glycol) diacrylate (PEGDA) as a model reinforcing polymer. The compressive strength of a 3D scaffold comprised of orthogonally intersecting beams was increased from 0.31 +/- 0.06 MPa to 1.65 +/- 0.13 MPa using PEGDA 600. In addition, the mechanical properties of reinforced cement were characterized using three PEGDA molecular weights (200, 400, and 600 Da) and three cement powder to liquid (P/L) ratios (0.8, 1.0, and 1.43). Higher molecular weight increased reinforcement efficacy, and P/L controlled cement porosity and determined the extent of polymer incorporation. Although increasing polymer incorporation resulted in a transition from brittle, cement-like behavior t! o ductile, polymer-like behavior, maximizing polymer incorporation was not advantageous. Polymerization shrinkage produced microcracks in the cement, which reduced the mechanical properties. The most effective reinforcement was achieved with P/L of 1.43 and PEGDA 600. In this group, flexural strength increased from 0.44 +/- 0.12 MPa to 7.04 +/- 0.51 MPa, maximum displacement from 0.05 +/- 0.01 mm to 1.44 +/- 0.17 mm, and work of fracture from 0.64 +/- 0.10 J/m(2) to 677.96 +/- 70.88 J/m(2) compared to non-reinforced controls. These results demonstrate the effectiveness of our novel reinforcement method, as well as its potential for fabricating reinforced 3D calcium phosphate cement scaffolds useful for bone tissue engineering. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20186776 [PubMed - as supplied by publisher] | | |
| Endothelial and stem cell interactions on dielectrophoretically aligned fibrous silk fibroin-chitosan scaffolds.
February 27, 2010 at 7:09 AM
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| | Endothelial and stem cell interactions on dielectrophoretically aligned fibrous silk fibroin-chitosan scaffolds. J Biomed Mater Res A. 2010 Feb 22; Authors: Gupta V, Davis G, Gordon A, Altman AM, Reece GP, Gascoyne PR, Mathur AB Regenerative tissue engineering requires biomaterials that would mimic structure and composition of the extracellular matrix to facilitate cell infiltration, differentiation, and vascularization. Engineered scaffolds composed of natural biomaterials silk fibroin (SF) and chitosan (CS) blend were fabricated to achieve fibrillar nano-structures aligned in three-dimensions using the technique of dielectrophoresis. The effect of scaffold properties on adhesion and migration of human adipose-derived stem cells (hASC) and endothelial cells (HUVEC) was studied on SFCS (micro-structure, unaligned) and engineered SFCS (E-SFCS; nano-structure, aligned). E-SFCS constituted of a nano-featured fibrillar sheets, whereas SFCS sheets had a smooth morphology with unaligned micro-fibrillar extensions at the ends. Adhesion of hASC to either scaffolds occurred within 30 min and was higher than HUVEC adhesion. The percentage of moving cells and average speed was highest for hASC on SF! CS scaffold as compared to hASC cocultured with HUVEC. HUVEC interactions with hASC appeared to slow the speed of hASC migration (in coculture) on both scaffolds. It is concluded that the guidance of cells for regenerative tissue engineering using SFCS scaffolds requires a fine balance between cell-cell interactions that affect the migration speed of cells and the surface characteristics that affects the overall adhesion and direction of migration. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2010. PMID: 20186770 [PubMed - as supplied by publisher] | | |
| Characterization of cyclic acetal hydroxyapatite nanocomposites for craniofacial tissue engineering.
February 27, 2010 at 7:09 AM
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| | Characterization of cyclic acetal hydroxyapatite nanocomposites for craniofacial tissue engineering. J Biomed Mater Res A. 2010 Feb 22; Authors: Patel M, Patel KJ, Caccamese JF, Coletti DP, Sauk JJ, Fisher JP Cyclic acetal hydrogels are a novel group of biomaterials which may facilitate osteogenic differentiation of encapsulated bone marrow stromal cells (BMSCs) because of their neutral degradation products. Here, we have incorporated hydroxyapatite nanoparticles within cyclic acetal hydrogels to create cyclic acetal nanocomposites for craniofacial tissue engineering applications. We hypothesized that inclusion of nanosized hydroxyapatite particles within cyclic acetal hydrogels would upregulate osteogenic signal expression of encapsulated BMSCs, due to enhanced cell adhesion, and therefore promote osteodifferentiation. Experimental nanocomposite groups consisted of lower (5 ng/mL) and higher (50 ng/mL) concentrations of nanoparticles. The nanocomposites were characterized by scanning electron microscopy, transmission electron microscopy, and energy dispersive spectroscopy. Swelling parameters of hydrogels in the presence of nanoparticles was studied. Osteoblastic diff! erentiation was characterized by alkaline phosphatase (ALP) and osteocalcin (OC) expression, whereas endogenous osteogenic signal expression was characterized by morphogenetic protein-2 (BMP-2) expression. Finally, immunohistochemistry was performed to detect the presence of OC at the protein level. Results indicated that hydroxyapatite nanoparticles were uniformly distributed throughout the hydrogels and did not affect material properties of the gels. Viability of cells was not affected by nanoparticle concentration, and BMP-2 and OC mRNA expression was enhanced in the presence of nanoparticles. However, a difference in BMP-2, ALP, and OC mRNA expression was not noted between the lower and higher concentrations of nanoparticles. This work demonstrates that inclusion of hydroxyapatite nanoparticles within a cyclic acetal nanocomposite hydrogel may enhance BMSC differentiation by promoting endogenous osteogenic signal expression. (c) 2010 Wiley Periodicals, Inc. J Biomed Mat! er Res 2010. PMID: 20186741 [PubMed - as supplied by publisher] | | |
| An improved collagen scaffold for skeletal regeneration.
February 27, 2010 at 7:09 AM
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| | An improved collagen scaffold for skeletal regeneration. J Biomed Mater Res A. 2010 Feb 22; Authors: Oliveira SM, Ringshia RA, Legeros RZ, Clark E, Yost MJ, Terracio L, Teixeira CC Bone repair and regeneration is one of the most extensively studied areas in the field of tissue engineering. All of the current tissue engineering approaches to create bone focus on intramembranous ossification, ignoring the other mechanism of bone formation, endochondral ossification. We propose to create a transient cartilage template in vitro, which could serve as an intermediate for bone formation by the endochondral mechanism once implanted in vivo. The goals of the study are (1) to prepare and characterize type I collagen sponges as a scaffold for the cartilage template, and (2) to establish a method of culturing chondrocytes in type I collagen sponges and induce cell maturation. Collagen sponges were generated from a 1% solution of type I collagen using a freeze/dry technique followed by UV light crosslinking. Chondrocytes isolated from two locations in chick embryo sterna were cultured in these sponges and treated with retinoic acid to induce chondrocyte ! maturation and extracellular matrix deposition. Material strength testing as well as microscopic and biochemical analyzes were conducted to evaluate the properties of sponges and cell behavior during the culture period. We found that our collagen sponges presented improved stiffness and supported chondrocyte attachment and proliferation. Cells underwent maturation, depositing an abundant extracellular matrix throughout the scaffold, expressing high levels of type X collagen, type I collagen and alkaline phosphatase. These results demonstrate that we have created a transient cartilage template with potential to direct endochondral bone formation after implantation. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res 2010. PMID: 20186736 [PubMed - as supplied by publisher] | | |
| Collagen type I hydrogel allows migration, proliferation, and osteogenic differentiation of rat bone marrow stromal cells.
February 27, 2010 at 7:09 AM
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| | Collagen type I hydrogel allows migration, proliferation, and osteogenic differentiation of rat bone marrow stromal cells. J Biomed Mater Res A. 2010 Feb 22; Authors: Hesse E, Hefferan TE, Tarara JE, Haasper C, Meller R, Krettek C, Lu L, Yaszemski MJ Hydrogels are potentially useful for many purposes in regenerative medicine including drug and growth factor delivery, as single scaffold for bone repair or as a filler of pores of another biomaterial in which host mesenchymal progenitor cells can migrate in and differentiate into matrix-producing osteoblasts. Collagen type I is of special interest as it is a very important and abundant natural matrix component. The purpose of this study was to investigate whether rat bone marrow stromal cells (rBMSCs) are able to adhere to, to survive, to proliferate and to migrate in collagen type I hydrogels and whether they can adopt an osteoblastic fate. rBMSCs were obtained from rat femora and plated on collagen type I hydrogels. Before harvest by day 7, 14, and 21, hydrogels were fluorescently labeled, cryo-cut and analyzed by fluorescent-based and laser scanning confocal microscopy to determine cell proliferation, migration, and viability. Osteogenic differentiation was de! termined by alkaline phosphatase activity. Collagen type I hydrogels allowed the attachment of rBMSCs to the hydrogel, their proliferation, and migration towards the inner part of the gel. rBMSCs started to differentiate into osteoblasts as determined by an increase in alkaline phosphatase activity after two weeks in culture. This study therefore suggests that collagen type I hydrogels could be useful for musculoskeletal regenerative therapies. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res 2010. PMID: 20186733 [PubMed - as supplied by publisher] | | |
| Fabrication and characterization of a biomimetic composite scaffold for bone defect repair.
February 27, 2010 at 7:09 AM
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| | Fabrication and characterization of a biomimetic composite scaffold for bone defect repair. J Biomed Mater Res A. 2010 Feb 22; Authors: Nitzsche H, Lochmann A, Metz H, Hauser A, Syrowatka F, Hempel E, Müller T, Thurn-Albrecht T, Mäder K For successful bone tissue engineering, scaffolds with tailored properties are a basic requirement. The combination of different available materials not only appears to be desirable but also very challenging. In this study, a composite material consisting of hydroxyapatite and collagen was produced by a biomimetic precipitation method and characterized by X-ray diffraction (XRD) and thermogravimetry (TGA). Subsequently, a suspension-quick-freezing and lyophilization method was used to incorporate the hydroxyapatite into a polymeric matrix consisting of collagen and chitosan. Before physicochemical characterization, the highly porous scaffolds were consolidated by a dehydrothermal treatment (DHT). The main attention was focused on the particle size of hydroxyapatite, which should be in the nanometer range. This is relevant to achieve a homogeneous resorption of the material by osteoclasts. Small-angle X-ray scattering (SAXS), atomic force microscopy (AFM), and envi! ronmental scanning electron microscopy (ESEM) were used to evaluate the outcome. The results suggest a successful polymeric embedding of nanoscaled hydroxyapatite particles into the matrix of the spongy construct. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res, 2010. PMID: 20186731 [PubMed - as supplied by publisher] | | |
| In vitro osteoclastogenesis on textile chitosan scaffold.
February 27, 2010 at 7:09 AM
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| | In vitro osteoclastogenesis on textile chitosan scaffold. Eur Cell Mater. 2010;19:96-106 Authors: Heinemann C, Heinemann S, Bernhardt A, Lode A, Worch H, Hanke T Textile chitosan fibre scaffolds were evaluated in terms of interaction with osteoclast-like cells, derived from human primary monocytes. Part of the scaffolds was further modified by coating with fibrillar collagen type I in order to make the surface biocompatible. Monocytes were cultured directly on the scaffolds in the presence of macrophage colony stimulating factor (M-CSF) and receptor activator of nuclear factor kappaB ligand (RANKL) for up to 18 days. Confocal laser scanning microscopy (CLSM) as well as scanning electron microscopy (SEM) revealed the formation of multinuclear osteoclast-like cells on both the raw chitosan fibres and the collagen-coated scaffolds. The modified surface supported the osteoclastogenesis. Differentiation towards the osteoclastic lineage was confirmed by the microscopic detection of cathepsin K, tartrate resistant acid phosphatase (TRAP), acidic compartments using 3-(2,4-dinitroanillino)-3'-amino-N-methyldipropylamine (DAMP), imm! unological detection of TRAP isoform 5b, and analysis of gene expression of the osteoclastic markers TRAP, cathepsin K, vitronectin receptor, and calcitonin receptor using reverse transcription-polymerase chain reaction (RT-PCR). The feature of the collagen-coated but also of the raw chitosan fibre scaffolds to support attachment and differentiation of human monocytes facilitates cell-induced material resorption - one main requirement for successful bone tissue engineering. PMID: 20186669 [PubMed - in process] | | |
| The composition of hydrogels for cartilage tissue engineering can influence glycosaminoglycan profile.
February 27, 2010 at 7:09 AM
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| | The composition of hydrogels for cartilage tissue engineering can influence glycosaminoglycan profile. Eur Cell Mater. 2010;19:86-95 Authors: Wang QG, Hughes N, Cartmell SH, Kuiper NJ The injectable and hydrophilic nature of hydrogels makes them suitable candidates for cartilage tissue engineering. To date, a wide range of hydrogels have been proposed for articular cartilage regeneration but few studies have quantitatively compared chondrocyte behaviour and extracellular matrix (ECM) synthesis within the hydrogels. Herein we have examined the nature of ECM synthesis by chondrocytes seeded into four hydrogels formed by either temperature change, self-assembly or chemical cross-linking. Bovine articular cartilage chondrocytes were cultured for 14 days in Extracel(R), Pluronic F127 blended with Type II collagen, Puramatrix(R) and Matrixhyal(R). The discriminatory and sensitive technique of fluorophore-assisted carbohydrate electrophoresis (FACE) was used to determine the fine detail of the glycosaminoglycans (GAG); hyaluronan and chondroitin sulphate. FACE analysis for chondroitin sulphate and hyaluronan profiles in Puramatrix(R) closely matched t! hat of native cartilage. For each hydrogel, DNA content, viability and morphology were assessed. Total collagen and total sulphated GAG production were measured and normalised to DNA content. Significant differences were found in total collagen synthesis. By day 14, Extracel(R) and Puramatrix(R) had significantly more total collagen than Matrixhyal(R) (1.77+/-0.26 mug and 1.97+/-0.26 mug vs. 0.60+/-0.26 mug; p<0.05). sGAG synthesis occurred in all hydrogels but a significantly higher amount of sGAG was retained within Extracel(R) at days 7 and 14 (p<0.05). In summary, we have shown that the biochemical and biophysical characteristics of each hydrogel directly or indirectly influenced ECM formation. A detailed understanding of the ECM in the development of engineered constructs is an important step in monitoring the success of cartilage regeneration strategies. PMID: 20186668 [PubMed - in process] | | |
| Long-term dynamic loading improves the mechanical properties of chondrogenic mesenchymal stem cell-laden hydrogel.
February 27, 2010 at 7:09 AM
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| | Long-term dynamic loading improves the mechanical properties of chondrogenic mesenchymal stem cell-laden hydrogel. Eur Cell Mater. 2010;19:72-85 Authors: Huang AH, Farrell MJ, Kim M, Mauck RL Mesenchymal stem cells (MSCs) are an attractive cell source for cartilage tissue engineering given their ability to undergo chondrogenesis in 3D culture systems. Mechanical forces play an important role in regulating both cartilage development and MSC chondrogenic gene expression, however, mechanical stimulation has yet to enhance the mechanical properties of engineered constructs. In this study, we applied long-term dynamic compression to MSC-seeded constructs and assessed whether varying pre-culture duration, loading regimens and inclusion of TGF-beta3 during loading would influence functional outcomes and these phenotypic transitions. Loading initiated before chondrogenesis decreased functional maturation, although chondrogenic gene expression increased. In contrast, loading initiated after chondrogenesis and matrix elaboration further improved the mechanical properties of MSC-based constructs, but only when TGF-beta3 levels were maintained and under specific l! oading parameters. Although matrix quantity was not affected by dynamic compression, matrix distribution, assessed histologically and by FT-IRIS analysis, was significantly improved on the micro- (pericellular) and macro- (construct expanse) scales. Further, whole genome expression profiling revealed marked shifts in the molecular topography with dynamic loading. These results demonstrate, for the first time, that dynamic compressive loading initiated after a sufficient period of chondro-induction and with sustained TGF-beta exposure enhances matrix distribution and the mechanical properties of MSC-seeded constructs. PMID: 20186667 [PubMed - in process] | | |
| A new fish scale-derived scaffold for corneal regeneration.
February 27, 2010 at 7:09 AM
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| | A new fish scale-derived scaffold for corneal regeneration. Eur Cell Mater. 2010;19:50-7 Authors: Lin CC, Ritch R, Lin SM, Ni MH, Chang YC, Lu YL, Lai HJ, Lin FH The purpose of this study is to develop a novel scaffold, derived from fish scales, as an alternative functional material with sufficient mechanical strength for corneal regenerative applications. Fish scales, which are usually considered as marine wastes, were acellularized, decalcified and fabricated into collagen scaffolds. The microstructure of the acellularized scaffold was imaged by scanning electron microscopy (SEM). The acellularization and decalcification treatments did not affect the naturally 3-dimentional, highly centrally-oriented micropatterned structure of the material. To assess the cytocompatibility of the scaffold with corneal cells, rabbit corneal cells were cultured on the scaffold and examined under SEM and confocal microscopy at different time periods. Rapid cell proliferation and migration on the scaffold were observed under SEM and confocal microscopy. The highly centrally-oriented micropatterned structure of the scaffold was beneficial for! efficient nutrient and oxygen supply to the cells cultured in the three-dimensional matrices, and therefore it is useful for high-density cell seeding and spreading. Collectively, we demonstrate the superior cellular conductivity of the newly developed material. We provide evidences for the feasibility of the scaffold as a template for corneal cells growth and migration, and thus the fish scale-derived scaffold can be developed as a promising material for tissue-engineering of cornea. PMID: 20186665 [PubMed - in process] | | |
| The Vastus Intermedius Periosteal (VIP) Flap: A Novel Flap for Osteoinduction.
February 27, 2010 at 7:09 AM
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| | The Vastus Intermedius Periosteal (VIP) Flap: A Novel Flap for Osteoinduction. J Reconstr Microsurg. 2010 Feb 25; Authors: Maercks RA, Runyan CM, Jones DC, Taylor JA Periosteum's role in fracture healing is widely recognized, and its function in bone tissue engineering shows great potential. Here we introduce a novel periosteal free flap to be used as an abundant source of periosteum in the engineering and repair of bone. The descending branch of the lateral femoral circumflex vessels were isolated on 11 fresh human cadavers, preserving perforators to the vastus intermedius muscle. A cuff of vastus intermedius and ~75% of the circumference of the femoral periosteum were harvested from 6 cm proximal to the knee to 8 cm distal to the greater trochanter. Flap pedicle length and periosteal dimensions were measured. The pedicle arteries were injected with radiopaque dye, and radiographs were taken. A musculoperiosteal flap was elevated with visible descending perforators in each case. Mean flap surface area was 128 cm (2) (+/-99-143 cm (2)). Average pedicle length was 8 cm (+/-6-11 cm). Dye injection confirmed that the flaps blood ! supply was the descending branch of the lateral femoral circumflex artery. This anatomical study confirms the vascular supply of this large musculoperiosteal flap. Future work will test its efficacy as an osteoinductive agent in bone repair and tissue engineering in humans. PMID: 20186660 [PubMed - as supplied by publisher] | | |
| Low-level Er:YAG laser irradiation enhances osteoblast proliferation through activation of MAPK/ERK.
February 27, 2010 at 7:09 AM
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| | Low-level Er:YAG laser irradiation enhances osteoblast proliferation through activation of MAPK/ERK. Lasers Med Sci. 2010 Feb 26; Authors: Aleksic V, Aoki A, Iwasaki K, Takasaki AA, Wang CY, Abiko Y, Ishikawa I, Izumi Y Although the use of high-level Er:YAG laser irradiation has been increasing in periodontal and peri-implant therapy, the effects of low-level Er:YAG laser on surrounding tissues and cells remain unclear. In the present study, the effects of low-level Er:YAG laser irradiation on osteoblast proliferation were investigated. Cells of the osteoblastic cell line MC3T3-E1 were treated with low-level Er:YAG laser irradiation with various combinations of laser settings (fluence 0.7-17.2 J/cm(2)) and in the absence or presence of culture medium during irradiation. On day 1 and/or day 3, cell proliferation and death were determined by cell counting and by measurement of lactate dehydrogenase (LDH) levels. Further, the role of mitogen-activated protein kinase (MAPK) pathways in laser-enhanced cell proliferation was investigated by inhibiting the MAPK pathways and then measuring MAPK phosphorylation by Western blotting. Higher proliferation rates were found with various combin! ations of irradiation parameters on days 1 and 3. Significantly higher proliferation was also observed in laser-irradiated MC3T3-E1 cells at a fluence of approximately 1.0-15.1 J/cm(2), whereas no increase in LDH activity was observed. Further, low-level Er:YAG irradiation induced the phosphorylation of extracellular signal-regulated protein kinase (MAPK/ERK) 5 to 30 min after irradiation. Although MAPK/ERK 1/2 inhibitor U0126 significantly inhibited laser-enhanced cell proliferation, activation of stress-activated protein kinases/Jun N-terminal kinase (SAPK/JNK) and p38 MAPK was not clearly detected. These results suggest that low-level Er:YAG laser irradiation increases osteoblast proliferation mainly by activation of MAPK/ERK, suggesting that the Er:YAG laser may be able to promote bone healing following periodontal and peri-implant therapy. PMID: 20186556 [PubMed - as supplied by publisher] | | |
| Derivation, characterization, differentiation, and registration of seven human embryonic stem cell lines (VAL-3, -4, -5, -6M, -7, -8, and -9) on human feeder.
February 27, 2010 at 7:09 AM
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| | Derivation, characterization, differentiation, and registration of seven human embryonic stem cell lines (VAL-3, -4, -5, -6M, -7, -8, and -9) on human feeder. In Vitro Cell Dev Biol Anim. 2010 Feb 26; Authors: Aguilar-Gallardo C, Poo M, Gomez E, Galan A, Sanchez E, Marques-Mari A, Ruiz V, Medrano J, Riboldi M, Valbuena D, Simon C Derivation of human embryonic stem cell lines has been a remarkable scientific achievement during the last decade. Human embryonic stem cells are regarded as an unlimited cell source for replacement therapy in regenerative medicine. Clearly, the scientific community requires proper derivation, characterization, and registration with the purpose of making them available for research and future medical applications worldwide. In this paper, we report our derivation work as the Valencian Node of the Spanish Stem Cell Bank in the generation, characterization, and registration of VAL-3, -4, -5, -6M, -7, -8, and 9 ( www.isciii/htdocs/terapia/terapia_bancocelular.jsp ). The derivation process was performed on microbiologically tested and irradiated human foreskin fibroblasts and designed to minimize contact with xeno-components in knockout Dulbecco's modified Eagle's medium supplemented with knockout serum replacement and basic fibroblast growth factor. Fingerprinting of! the cell lines was performed to allow their identification and traceability. All lines were expressed at the mRNA and specific protein markers for undifferentiation and were found to be negative for classical differentiation markers such as neurofilament heavy chain (ectoderm), renin (mesoderm), and amylase (endoderm). All lines displayed high levels of telomerase activity and were shown to successfully overcome cryopreservation and thawing. Finally, we demonstrated the potential to differentiate in vitro (embryoid body formation) and in vivo (teratoma formation) into cell types from all three germ layers. Teratoma derived from all human embryonic stem cell lines present similar morphological features except VAL-8 that display more aggressive tumor behavior with a larger proportion of solid tissues, as opposed to cyst formation in the other cell lines. PMID: 20186513 [PubMed - as supplied by publisher] | | |
| Targeting a20 decreases glioma stem cell survival and tumor growth.
February 27, 2010 at 7:09 AM
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| | Targeting a20 decreases glioma stem cell survival and tumor growth. PLoS Biol. 2010 Feb;8(2):e1000319 Authors: Hjelmeland AB, Wu Q, Wickman S, Eyler C, Heddleston J, Shi Q, Lathia JD, Macswords J, Lee J, McLendon RE, Rich JN Glioblastomas are deadly cancers that display a functional cellular hierarchy maintained by self-renewing glioblastoma stem cells (GSCs). GSCs are regulated by molecular pathways distinct from the bulk tumor that may be useful therapeutic targets. We determined that A20 (TNFAIP3), a regulator of cell survival and the NF-kappaB pathway, is overexpressed in GSCs relative to non-stem glioblastoma cells at both the mRNA and protein levels. To determine the functional significance of A20 in GSCs, we targeted A20 expression with lentiviral-mediated delivery of short hairpin RNA (shRNA). Inhibiting A20 expression decreased GSC growth and survival through mechanisms associated with decreased cell-cycle progression and decreased phosphorylation of p65/RelA. Elevated levels of A20 in GSCs contributed to apoptotic resistance: GSCs were less susceptible to TNFalpha-induced cell death than matched non-stem glioma cells, but A20 knockdown sensitized GSCs to TNFalpha-mediated ap! optosis. The decreased survival of GSCs upon A20 knockdown contributed to the reduced ability of these cells to self-renew in primary and secondary neurosphere formation assays. The tumorigenic potential of GSCs was decreased with A20 targeting, resulting in increased survival of mice bearing human glioma xenografts. In silico analysis of a glioma patient genomic database indicates that A20 overexpression and amplification is inversely correlated with survival. Together these data indicate that A20 contributes to glioma maintenance through effects on the glioma stem cell subpopulation. Although inactivating mutations in A20 in lymphoma suggest A20 can act as a tumor suppressor, similar point mutations have not been identified through glioma genomic sequencing: in fact, our data suggest A20 may function as a tumor enhancer in glioma through promotion of GSC survival. A20 anticancer therapies should therefore be viewed with caution as effects will likely differ depending on t! he tumor type. PMID: 20186265 [PubMed - in process] | | |
| A Simple Modification of the Separation Method Reduces Heterogeneity of Adipose-Derived Stem Cells.
February 27, 2010 at 7:09 AM
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| | A Simple Modification of the Separation Method Reduces Heterogeneity of Adipose-Derived Stem Cells. Cells Tissues Organs. 2010 Feb 24; Authors: Griesche N, Luttmann W, Luttmann A, Stammermann T, Geiger H, Baer PC High hopes are put into the use of mesenchymal stem cells (MSCs) in various approaches for tissue engineering and regenerative medicine. MSCs are derived from different tissues with only small differences in their phenotype or their differentiation potential, but higher differences in the cell yield. Since fat is easily accessible and contains a high amount of MSCs to be isolated, adipose-derived stem cells (ASCs) are very promising for clinical approaches. ASCs are not a completely homogeneous cell population. Our study was initiated to explore an easy and convenient method to reduce heterogeneity. We tested different isolation methods: (1) the standard isolation method for ASCs based on plastic attachment, (2) the standard method with an initial washing step after 60 min of adherence and (3) immunomagnetic isolation by 4 typical markers (CD49a, CD90, CD105 and CD271). Cells isolated by these methods were evaluated using quantitative PCR and flow cytometry as wel! l as by their differentiation potential. Washing led to a significantly lower expression of desmin, smA and six2, and a higher expression of the stem cell markers nestin, oct-4 and sall-1, compared to standard isolated cells, while the immunomagnetically isolated cells showed no significant changes. All cells independent of the isolation method could be induced to differentiate into adipocytes and osteoblasts. Our study demonstrates that a simple washing step reduces heterogeneity of cultured ASCs according to PCR analysis, whereas the immunomagnetic isolation only showed minor advantages compared to the standard method, but the disadvantage of significantly lower cell yields in the primary isolates. PMID: 20185896 [PubMed - as supplied by publisher] | | |
| Cortical plasticity induced by inhibitory neuron transplantation.
February 27, 2010 at 7:09 AM
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| | Cortical plasticity induced by inhibitory neuron transplantation. Science. 2010 Feb 26;327(5969):1145-8 Authors: Southwell DG, Froemke RC, Alvarez-Buylla A, Stryker MP, Gandhi SP Critical periods are times of pronounced brain plasticity. During a critical period in the postnatal development of the visual cortex, the occlusion of one eye triggers a rapid reorganization of neuronal responses, a process known as ocular dominance plasticity. We have shown that the transplantation of inhibitory neurons induces ocular dominance plasticity after the critical period. Transplanted inhibitory neurons receive excitatory synapses, make inhibitory synapses onto host cortical neurons, and promote plasticity when they reach a cellular age equivalent to that of endogenous inhibitory neurons during the normal critical period. These findings suggest that ocular dominance plasticity is regulated by the execution of a maturational program intrinsic to inhibitory neurons. By inducing plasticity, inhibitory neuron transplantation may facilitate brain repair. PMID: 20185728 [PubMed - in process] | | |
| Effect of nanoscale topography on fibronectin adsorption, focal adhesion size and matrix organisation.
February 27, 2010 at 7:09 AM
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| | Effect of nanoscale topography on fibronectin adsorption, focal adhesion size and matrix organisation. Colloids Surf B Biointerfaces. 2010 Feb 4; Authors: González-García C, Sousa SR, Moratal D, Rico P, Salmerón-Sánchez M Phase separation of PLLA/PS (50/50, w/w) solutions during a spin-casting process gives rise to well-defined nanotopographies of 14, 29 and 45nm deep pits depending on the concentration of the solution. Their influence on the biological activity of fibronectin (FN) was investigated. FN adsorption was quantified by radiolabelling the protein. The amount of adsorbed FN was higher on the 14nm deep pit nanotopography than on the other two surfaces. FN distribution between valleys and peaks was investigated by AFM combined with image analysis. FN tends to adsorb preferentially on the valleys of the nanotopography only for the 14nm system and when adsorbed from solutions of concentration lower than 10mug/ml. Higher concentration of the FN solution leads to evenly distribution of the protein throughout the surface; moreover, there is no difference in the distribution of the protein between valleys and peaks for the other two systems (29 and 45nm) irrespective of the conce! ntration of the FN solution. The biological activity of the adsorbed protein layer was assessed by investigating MC3T3 osteoblast-like cells adhesion, FN reorganisation and late matrix formation on the different substrates. Even if initial cell adhesion is excellent for every substrate, the size of the focal adhesion plaques increases as the size of the pits in the nanotopography does. This is correlated to FN reorganisation, which only takes places on the 29 and 45nm deep pits surfaces, where enhanced late matrix production was also found. PMID: 20185279 [PubMed - as supplied by publisher] | | |
| Biomimetic hydrogels with pro-angiogenic properties.
February 27, 2010 at 7:09 AM
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| | Biomimetic hydrogels with pro-angiogenic properties. Biomaterials. 2010 Feb 23; Authors: Moon JJ, Saik JE, Poché RA, Leslie-Barbick JE, Lee SH, Smith AA, Dickinson ME, West JL To achieve the task of fabricating functional tissues, scaffold materials that can be sufficiently vascularized to mimic functionality and complexity of native tissues are yet to be developed. Here, we report development of synthetic, biomimetic hydrogels that allow the rapid formation of a stable and mature vascular network both in vitro and in vivo. Hydrogels were fabricated with integrin binding sites and protease-sensitive substrates to mimic the natural provisional extracellular matrices, and endothelial cells cultured in these hydrogels organized into stable, intricate networks of capillary-like structures. The resulting structures were further stabilized by recruitment of mesenchymal progenitor cells that differentiated into a smooth muscle cell lineage and deposited collagen IV and laminin in vitro. In addition, hydrogels transplanted into mouse corneas were infiltrated with host vasculature, resulting in extensive vascularization with functional blood ves! sels. These results indicate that these hydrogels may be useful for applications in basic biological research, tissue engineering, and regenerative medicine. PMID: 20185173 [PubMed - as supplied by publisher] | | |
| The effect of incorporation of SDF-1alpha into PLGA scaffolds on stem cell recruitment and the inflammatory response.
February 27, 2010 at 7:09 AM
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| | The effect of incorporation of SDF-1alpha into PLGA scaffolds on stem cell recruitment and the inflammatory response. Biomaterials. 2010 Feb 23; Authors: Thevenot PT, Nair AM, Shen J, Lotfi P, Ko CY, Tang L Despite significant advances in the understanding of tissue responses to biomaterials, most implants are still plagued by inflammatory responses which can lead to fibrotic encapsulation. This is of dire consequence in tissue engineering, where seeded cells and bioactive components are separated from the native tissue, limiting the regenerative potential of the design. Additionally, these interactions prevent desired tissue integration and angiogenesis, preventing functionality of the design. Recent evidence supports that mesenchymal stem cells (MSC) and hematopoietic stem cells (HSC) can have beneficial effects which alter the inflammatory responses and improve healing. The purpose of this study was to examine whether stem cells could be targeted to the site of biomaterial implantation and whether increasing local stem cell responses could improve the tissue response to PLGA scaffold implants. Through incorporation of SDF-1alpha through factor adsorption and mini-! osmotic pump delivery, the host-derived stem cell response can be improved resulting in 3X increase in stem cell populations at the interface for up to 2 weeks. These interactions were found to significantly alter the acute mast cell responses, reducing the number of mast cells and degranulated mast cells near the scaffold implants. This led to subsequent downstream reduction in the inflammatory cell responses, and through altered mast cell activation and stem cell participation, increased angiogenesis and decreased fibrotic responses to the scaffold implants. These results support that enhanced recruitment of autologous stem cells can improve the tissue responses to biomaterial implants through modifying/bypassing inflammatory cell responses and jumpstarting stem cell participation in healing at the implant interface. PMID: 20185171 [PubMed - as supplied by publisher] | | |
| Computational modeling of flow-induced shear stresses within 3D salt-leached porous scaffolds imaged via micro-CT.
February 27, 2010 at 7:09 AM
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| | Computational modeling of flow-induced shear stresses within 3D salt-leached porous scaffolds imaged via micro-CT. J Biomech. 2010 Feb 23; Authors: Voronov R, Vangordon S, Sikavitsas VI, Papavassiliou DV Flow-induced shear stresses have been found to be a stimulatory factor in pre-osteoblastic cells seeded in 3D porous scaffolds and cultured under continuous flow perfusion. However, due to the complex internal structure of porous scaffolds, analytical estimation of the local shear forces is impractical. The primary goal of this work is to investigate the shear stress distributions within Poly(l-lactic acid) scaffolds via computation. Scaffolds used in this study are prepared via salt leeching with various geometric characteristics (80-95% porosity and 215-402.5mum average pore size). High resolution micro-computed tomography is used to obtain their 3D structure. Flow of osteogenic media through the scaffolds is modeled via lattice Boltzmann method. It is found that the surface stress distributions within the scaffolds are characterized by long tails to the right (a positive skewness). Their shape is not strongly dependent on the scaffold manufacturing parameters, ! but the magnitudes of the stresses are. Correlations are prepared for the estimation of the average surface shear stress experienced by the cells within the scaffolds and of the probability density function of the surface stresses. Though the manufacturing technique does not appear to affect the shape of the shear stress distributions, presence of manufacturing defects is found to be significant: defects create areas of high flow and high stress along their periphery. The results of this study are applicable to other polymer systems provided that they are manufactured by a similar salt leeching technique, while the imaging/modeling approach is applicable to all scaffolds relevant to tissue engineering. PMID: 20185132 [PubMed - as supplied by publisher] | | |
| Conditional TGF-beta1 treatment increases stem cell-like cell population in myoblasts.
February 27, 2010 at 7:09 AM
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| | Conditional TGF-beta1 treatment increases stem cell-like cell population in myoblasts. J Cell Mol Med. 2010 Feb 23; Authors: Mu X, Li Y The limitation in successfully acquiring large populations of stem cell has impeded their application. A new method based on the dedifferentiation of adult somatic cells to generate induced multipotent stem cells would allow us to obtain a large amount of autologous stem cells for regenerative medicine. The current work was proposed to induce a sub-population of cells with characteristics of muscle stem cells from myoblasts through conditional treatment of Transforming Growth Factor (TGF)-beta1. Our results show that a lower concentration of TGF-beta1 is able to promote C2C12 myoblasts to express stem cell markers as well as to repress myogenic proteins, which involves a mechanism of dedifferentiation. Moreover, TGF-beta1 treatment promoted the proliferation-arrested C2C12 myoblasts to re-enter the S-phase. We also investigated the multi-differentiation potentials of the dedifferentiated cells. TGF-beta1 pretreated C2C12 myoblasts were implanted into mice to repai! r dystrophic skeletal muscle or injured bone. In addition to the C2C12 myoblasts, similar effects of TGF-beta1 were also observed in the primary myoblasts of mice. Our results suggest that TGF-beta1 is effective as a molecular trigger for the dedifferentiation of skeletal muscle myoblasts and could be used to generate a large pool of progenitor cells that collectively behave as multipotent stem cell-like cells for regenerative medicine applications. PMID: 20184662 [PubMed - as supplied by publisher] | | |
| The first COL7A1 mutation survey in a large Spanish Dystrophic Epidermolysis Bullosa cohort: c.6527insC disclosed as an unusually recurrent mutation.
February 27, 2010 at 7:09 AM
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| | The first COL7A1 mutation survey in a large Spanish Dystrophic Epidermolysis Bullosa cohort: c.6527insC disclosed as an unusually recurrent mutation. Br J Dermatol. 2010 Feb 22; Authors: Escámez MJ, García M, Cuadrado-Corrales N, Llames SG, Charlesworth A, De Luca N, Illera N, Sánchez-Jimeno C, Holguín A, Duarte B, Trujillo-Tiebas MJ, Vicario JL, Santiago JL, Hernández-Martín A, Torrelo A, Castiglia D, Ayuso C, Larcher F, Jorcano JL, Meana A, Meneguzzi G, Zambruno G, Del Rio M Summary Background: Dystrophic epidermolysis bullosa (DEB) is a genodermatosis caused by mutations in COL7A1. The clinical manifestations are highly variable from nail dystrophy to life-threatening blistering, making early molecular diagnosis and prognosis of utmost importance for the affected families. Mutation identification is mandatory for prenatal testing. Objective: To conduct the first mutational analysis of COL7A1 in a Spanish cohort. To assess mutation consequences at protein/mRNA level and to establish genotype-phenotype correlations. Patients/Methods: Forty-nine Spanish DEB patients were studied. Antigen mapping was performed on patient skin biopsies. COL7A1 mutation screening in genomic DNA was performed by touchdown-PCR and direct sequencing. Mutation consequences were determined by reverse transcription polymerase chain reaction (RT-PCR). Results: Eight patients belonged to three unrelated families with Dominant DEB (DDEB). Forty-one were affected wi! th Recessive DEB (RDEB). Specifically, 27 displayed the severe generalized subtype, 8 the other generalized subtype and 6 a localized phenotype (2 pretibial, 3 acral and 1 inversa). Thirty-five mutations were identified, 20 of which are novel. The pathogenic mutation c.6527insC accounted for 46.3% of RDEB Spanish alleles. A consistent genotype-phenotype correlation was established. Conclusions: Although COL7A1 database indicates that most of the DEB mutations are family specific, the pathogenic mutation c.6527insC was highly recurrent in our cohort. This level of recurrence for a single genetic defect was never reported for COL7A1. Our findings are essential to the clinicians caring for DEB patients in Spain and in the large population of Spanish descendants in Latin America. They also provide geneticists a molecular clue for a priority mutation screening strategy. PMID: 20184583 [PubMed - as supplied by publisher] | | |
| The Sequential Production Profiles of Growth Factors and Their Relations to Bone Volume in Ossifying Bone Marrow Explants.
February 27, 2010 at 7:09 AM
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| | The Sequential Production Profiles of Growth Factors and Their Relations to Bone Volume in Ossifying Bone Marrow Explants. Tissue Eng Part A. 2010 Feb 25; Authors: Gurkan UA, Gargac J, Akkus O Osteogenesis is a complex process that involves the synergistic contribution of multiple cell types and numerous growth factors (GFs). In order to develop effective bone tissue engineering strategies employing GFs, it is essential to delineate the complex and interconnected role of GFs in osteogenesis. The studies investigating the temporal involvement of GFs in osteogenesis are limited to in vitro studies with single cell types or complex in vivo studies. There is a need for platforms that embody the physiological characteristics and the multicellular environment of natural osteogenesis. Marrow tissue houses various cell types that are known to be involved in osteogenesis and in vitro cultures of marrow inherently undergo osteogenesis process. Self-inductive ossification of marrow explants in vitro can be employed as a representative multicellular and 3-dimensional model of osteogenesis. Therefore, the aims of this study were to employ the rat bone marrow explant! ossification model to determine: 1) the temporal production profiles of key GFs involved in osteogenesis, 2) the relation between GF production and ossification, and, 3) the relations between the GF levels throughout ossification. Temporal production profiles of transforming growth factor beta-1 (TGF-beta1), bone morphogenetic protein-2 (BMP-2), vascular endothelial growth factor (VEGF) and insulin-like growth factor-1 (IGF-1); and the bone-related proteins alkaline phosphatase (AP), osteocalcin (OC) were obtained by enzyme-linked immunosorbent assay (ELISA) conducted at days 2, 7, 12, 14, 19 and 21. The final amount of ossification (ossified volume, OV) was measured by micro computed tomography (muCT) at day 21. TGF-beta1, BMP-2, VEGF, IGF-1, AP and OC were produced by the ossifying marrow explants differentially over time. The early production of IGF-1 (day 2) correlated positively (r=0.868) with OV; however, latent production of IGF-1 correlated negatively (day14: r=-0.! 813; and day-19: r=-0.865) with OV. OV also correlated with VE! GF level s at day-12 (r=0.988) and at day-14 (r=0.970). Production of GFs also correlated to each other across time points which indicate the complex and interconnected contribution of various GFs in osteogenesis. Therefore tissue engineering strategies towards bone regeneration should consider the richness of GFs involved in osteogenesis as well as their dynamically varying participation over time. PMID: 20184436 [PubMed - as supplied by publisher] | | |
| Differential gene expression in adipose stem cells cultured in allogeneic human serum versus fetal bovine serum.
February 27, 2010 at 7:09 AM
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| | Differential gene expression in adipose stem cells cultured in allogeneic human serum versus fetal bovine serum. Tissue Eng Part A. 2010 Feb 25; Authors: Lindroos B, Aho KL, Kuokkanen H, Räty S, Huhtala H, Lemponen R, Yli-Harja O, Miettinen S, Suuronen R In pre-clinical studies, human adipose stem cells (ASCs) have been shown to have therapeutic applicability, but standard expansion methods for clinical applications remain yet to be established. ASCs are typically expanded in medium containing fetal bovine serum (FBS). However, sera and other animal-derived culture reagents stage safety issues in clinical therapy, including possible infections and severe immune reactions. By expanding ASCs in medium containing human serum, the problem can be eliminated. To define how allogeneic human serum (alloHS) performs in ASC expansion compared to FBS, a comparative in vitro study in both serum supplements was performed. The choice of serum had a significant effect on ASCs. Firstly, to reach cell proliferation levels comparable with 10% FBS, at least 15% alloHS was required. Secondly, while genes of the cell cycle pathway were overexpressed in alloHS, genes of the BMP receptor mediated signaling on the TGF-beta signaling path! way, regulating e.g. osteoblast differentiation, were overexpressed in FBS. The result was further supported by differentiation analysis, where early osteogenic differentiation was significantly enhanced in FBS. The data presented here underscore the importance of thorough investigation of ASCs for utilization in cell therapies. This study is a step forward in the understanding of these potential cells. PMID: 20184435 [PubMed - as supplied by publisher] | | |
| Accelerated angiogenic host tissue response to PLGA- scaffolds by vitalization with osteoblast like cells.
February 27, 2010 at 7:09 AM
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| | Accelerated angiogenic host tissue response to PLGA- scaffolds by vitalization with osteoblast like cells. Tissue Eng Part A. 2010 Feb 25; Authors: Tavassol F, Schumann P, Lindhorst D, Sinikovic B, Voss A, Stühmer C, Kampmann A, Bormann KH, Carvalho C, Mülhaupt R, Harder Y, Laschke MW, Menger M, Gellrich NC, Rücker M Bone substitutes should ideally promote rapid vascularization, which could be accelerated if these substitutes were vitalized by autologous cells. Although adequate engraftment of porous poly(L-lactide-co-glycolide) (PLGA) scaffolds has been demonstrated in the past, it has not yet been investigated how vascularization is influenced by vitalization or, more precisely, by seeding PLGA scaffolds with osteoblast-like cells (OLCs). For this reason, we conducted an in vivo study to assess host angiogenic and inflammatory responses after the implantation of PLGA scaffolds vitalized with isogeneic OLCs. OLCs were seeded on collagen-coated PLGA scaffolds that were implanted into dorsal skinfold chambers in BALB/c mice (n=8). Two further groups of animals received either collagen-coated (n=8) or uncoated PLGA scaffolds (n=8). Animals that received chambers without implants served as controls (n=8). Angiogenesis, neovascularization and leukocyte-endothelial cell interaction! were analyzed during a 14-day period using intravital fluorescence microscopy. PLGA scaffolds with and without OLCs showed a temporary increase in leukocyte recruitment. At day 3 after implantation, a marked angiogenic host tissue response was observed in close vicinity of all scaffolds studied. At days 6 and 10, the angiogenic response was significantly higher (p<0.05) in PLGA scaffolds vitalized with OLCs than in uncoated or collagen-coated PLGA scaffolds. The majority of OLCs, however, died within 14 days after implantation. Our study demonstrates that PLGA scaffold vitalization with OLCs accelerates the angiogenic response in the surrounding host tissue. Bone substitutes created by tissue engineering may thus be superior to non-vitalized substitutes although the seeded cells do not survive for long periods of time. PMID: 20184434 [PubMed - as supplied by publisher] | | |
| Induction of haematopoietic differentiation of mouse embryonic stem cells by an AGM-derived stromal cell line is not further enhanced by over-expression of HOXB4.
February 27, 2010 at 7:09 AM
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| | Induction of haematopoietic differentiation of mouse embryonic stem cells by an AGM-derived stromal cell line is not further enhanced by over-expression of HOXB4. Stem Cells Dev. 2010 Feb 25; Authors: Gordon-Keylock SA, Jackson M, Huang C, Samuel K, Axton RA, Oostendorp R, Taylor AH, Wilson J, Forrester L Haematopoietic differentiation of embryonic stem (ES) cells can be enhanced by co-culture with stromal cells derived from haematopoietic tissues and by over-expression of the transcription factor HOXB4. In this study we compare the haematopoietic inductive effects of stromal cell lines derived from different sub-regions of the embryonic aorta-gonad-mesonephros tissue with the commonly used OP9 stromal cell line and with HOXB4 activation. We show that stromal cell lines derived from the aorta and surrounding mesenchyme (AM) act at an earlier stage of the differentiation process compared to bone marrow derived OP9 stromal cells. AM stromal cells were able to promote the further differentiation of isolated brachyury-GFP+ mesodermal cells into haematopoietic progenitors, whereas the OP9 stromal cells could not support the differentiation of these cells. Co-culture and analyses of individual embryoid bodies support the hypothesis that the AM stromal cell lines could en! hance the de novo production of haematopoietic progenitors, lending support to the idea that AM stromal cells might act on pre-haematopoietic mesoderm. The induction level observed for AM stromal cells was comparable to HOXB4 activation, but no additive effect was observed when these two inductive strategies were combined. Addition of a g-secretase inhibitor reduced the inductive effects of both the stromal cell line and HOXB4, providing clues to possible shared molecular mechanisms. PMID: 20184433 [PubMed - as supplied by publisher] | | |
| Self-assembled Rosette Nanotube/Hydrogel Composites for Cartilage Tissue Engineering.
February 27, 2010 at 7:09 AM
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| | Self-assembled Rosette Nanotube/Hydrogel Composites for Cartilage Tissue Engineering. Tissue Eng Part C Methods. 2010 Feb 25; Authors: Chen Y, Bilgen B, Aaron R, Ciombor D, Fenniri H, Webster TJ Recently, hydrogels (such as alginate, agarose, polyethylene glycol (PEG), etc.) have been investigated as promising cartilage healing materials. To further improve cell-material interactions or mechanical properties of such hydrogel scaffolds, many materials (such as ceramics or carbon nanotubes) have been added to produce composites with tailored properties. In this study, novel rosette nanotubes (RNTs, self-assembled nanotubes built from DNA base pairs), hydrogels and cartilage cells (specifically, fibroblast-like type-B synoviocytes (SFB cells) and chondrocytes) were combined via a novel electrospinning technique to generate three-dimensional, implantable scaffolds for cartilage repair. Importantly, results of this study showed that electrospun RNT/hydrogel composites improved both SFB cell and chondrocyte functions. Especially, RNT/hydrogel composites promoted SFB cell chondrogenic differentiation in two week culture experiments. Furthermore, studies demonstr! ated that RNTs enhanced hydrogel adhesive strength to severed collagen. Results of this study, thus, provided a nanostructured scaffold that enhanced SFB cell adhesion, viability and chondrogenic differentiation compared to nano-smooth hydrogels without RNTs. This study provided an alternative cartilage regenerative material derived from RNTs that could be directly electrospun into cartilage defects (with SFB cells and/or chondrocytes) to bond to severed collagen and promote cartilage cell adhesion, viability, and subsequent functions. PMID: 20184414 [PubMed - as supplied by publisher] | | |
| Engineering the cell wall by reducing de-methyl-esterified homogalacturonan improves saccharification of plant tissues for bioconversion.
February 27, 2010 at 7:09 AM
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| | Engineering the cell wall by reducing de-methyl-esterified homogalacturonan improves saccharification of plant tissues for bioconversion. Proc Natl Acad Sci U S A. 2010 Jan 12;107(2):616-21 Authors: Lionetti V, Francocci F, Ferrari S, Volpi C, Bellincampi D, Galletti R, D'Ovidio R, De Lorenzo G, Cervone F Plant cell walls represent an abundant, renewable source of biofuel and other useful products. The major bottleneck for the industrial scale-up of their conversion to simple sugars (saccharification), to be subsequently converted by microorganisms into ethanol or other products, is their recalcitrance to enzymatic saccharification. We investigated whether the structure of pectin that embeds the cellulose-hemicellulose network affects the exposure of cellulose to enzymes and consequently the process of saccharification. Reduction of de-methyl-esterified homogalacturonan (HGA) in Arabidopsis plants through the expression of a fungal polygalacturonase (PG) or an inhibitor of pectin methylesterase (PMEI) increased the efficiency of enzymatic saccharification. The improved enzymatic saccharification efficiency observed in transformed plants could also reduce the need for acid pretreatment. Similar results were obtained in PG-expressing tobacco plants and in PMEI-expres! sing wheat plants, indicating that reduction of de-methyl-esterified HGA may be used in crop species to facilitate the process of biomass saccharification. PMID: 20080727 [PubMed - indexed for MEDLINE] | | |
| Cytoskeleton-based forecasting of stem cell lineage fates.
February 27, 2010 at 7:09 AM
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| | Cytoskeleton-based forecasting of stem cell lineage fates. Proc Natl Acad Sci U S A. 2010 Jan 12;107(2):610-5 Authors: Treiser MD, Yang EH, Gordonov S, Cohen DM, Androulakis IP, Kohn J, Chen CS, Moghe PV Stem cells that adopt distinct lineages cannot be distinguished based on traditional cell shape. This study reports that higher-order variations in cell shape and cytoskeletal organization that occur within hours of stimulation forecast the lineage commitment fates of human mesenchymal stem cells (hMSCs). The unique approach captures numerous early (24 h), quantitative features of actin fluororeporter shapes, intensities, textures, and spatial distributions (collectively termed morphometric descriptors). The large number of descriptors are reduced into "combinations" through which distinct subpopulations of cells featuring unique combinations are identified. We demonstrate that hMSCs cultured on fibronectin-treated glass substrates under environments permissive to bone lineage induction could be readily discerned within the first 24 h from those cultured in basal- or fat-inductive conditions by such cytoskeletal feature groupings. We extend the utility of this app! roach to forecast osteogenic stem cell lineage fates across a series of synthetic polymeric materials of diverse physicochemical properties. Within the first 24 h following stem cell seeding, we could successfully "profile" the substrate responsiveness prospectively in terms of the degree of bone versus nonbone predisposition. The morphometric methodology also provided insights into how substrates may modulate the pace of osteogenic lineage specification. Cells on glass substrates deficient in fibronectin showed a similar divergence of lineage fates, but delayed beyond 48 h. In summary, this high-content imaging and single cell modeling approach offers a framework to elucidate and manipulate determinants of stem cell behaviors, as well as to screen stem cell lineage modulating materials and environments. PMID: 20080726 [PubMed - indexed for MEDLINE] | | |
| Nanoscale cues regulate the structure and function of macroscopic cardiac tissue constructs.
February 27, 2010 at 7:09 AM
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| | Nanoscale cues regulate the structure and function of macroscopic cardiac tissue constructs. Proc Natl Acad Sci U S A. 2010 Jan 12;107(2):565-70 Authors: Kim DH, Lipke EA, Kim P, Cheong R, Thompson S, Delannoy M, Suh KY, Tung L, Levchenko A Heart tissue possesses complex structural organization on multiple scales, from macro- to nano-, but nanoscale control of cardiac function has not been extensively analyzed. Inspired by ultrastructural analysis of the native tissue, we constructed a scalable, nanotopographically controlled model of myocardium mimicking the in vivo ventricular organization. Guided by nanoscale mechanical cues provided by the underlying hydrogel, the tissue constructs displayed anisotropic action potential propagation and contractility characteristic of the native tissue. Surprisingly, cell geometry, action potential conduction velocity, and the expression of a cell-cell coupling protein were exquisitely sensitive to differences in the substratum nanoscale features of the surrounding extracellular matrix. We propose that controlling cell-material interactions on the nanoscale can stipulate structure and function on the tissue level and yield novel insights into in vivo tissue physio! logy, while providing materials for tissue repair. PMID: 20018748 [PubMed - indexed for MEDLINE] | | |
| Dynamic distribution and stem cell characteristics of Sox1-expressing cells in the cerebellar cortex.
February 27, 2010 at 7:09 AM
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| | Dynamic distribution and stem cell characteristics of Sox1-expressing cells in the cerebellar cortex. Cell Res. 2009 Dec;19(12):1324-33 Authors: Alcock J, Sottile V Bergmann glia cells are a discrete radial glia population surrounding Purkinje cells in the cerebellar cortex. Although Bergmann glia are essential for the development and correct arborization of Purkinje cells, little is known about the regulation of this cell population after the developmental phase. In an effort to characterize this population at the molecular level, we have analyzed marker expression and established that adult Bergmann glia express Sox1, Sox2 and Sox9, a feature otherwise associated with neural stem cells (NSCs). In the present study, we have further analyzed the developmental pattern of Sox1-expressing cells in the developing cerebellum. We report that before becoming restricted to the Purkinje cell layer, Sox1-positive cells are present throughout the immature tissue, and that these cells show characteristics of Bergmann glia progenitors. Our study shows that these progenitors express Sox1, Sox2 and Sox9, a signature maintained throughout ce! rebellar maturation into adulthood. When isolated in culture, the Sox1-expressing cerebellar population exhibited neurosphere-forming ability, NSC-marker characteristics, and demonstrated multipotency at the clonal level. Our results show that the Bergmann glia population expresses Sox1 during cerebellar development, and that these cells can be isolated and show stem cell characteristics in vitro, suggesting that they could hold a broader potential than previously thought. PMID: 19823196 [PubMed - indexed for MEDLINE] | | |
| Effective treatment of intractable skin ulcers using allogeneic cultured dermal substitutes in patients with systemic lupus erythematosus.
February 27, 2010 at 7:09 AM
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| | Effective treatment of intractable skin ulcers using allogeneic cultured dermal substitutes in patients with systemic lupus erythematosus. Eur J Dermatol. 2009 Nov-Dec;19(6):594-6 Authors: Toyozawa S, Yamamoto Y, Kishioka A, Yonei N, Kanazawa N, Matsumoto Y, Kuroyanagi Y, Furukawa F Skin ulcers in systemic lupus erythematosus (SLE) patients are non-healing or intractable, because various factors or complications, including vasculitis and immunosuppressants, impair wound healing. In the present study, we applied cultured dermal substitutes (CDSs) to 3 cases of SLE skin ulcers because various systemic or topical therapies were ineffective. CDSs are prepared by culturing human fibroblasts on two-layered spongy matrices of hyaluronic acid and atelo-collagen, and they effectively promote the healing of severe skin defects. After using CDSs in the 3 cases, healthy granulation tissues formed within 6 weeks, and skin grafts were successfully performed. These results indicate that allogeneic CDSs provide new therapeutic alternatives as topical therapies for intractable skin ulcers in SLE. PMID: 19666399 [PubMed - indexed for MEDLINE] | | |
| A model for studying human articular cartilage integration in vitro.
February 27, 2010 at 1:09 AM
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| | A model for studying human articular cartilage integration in vitro. J Biomed Mater Res A. 2010 Feb 22; Authors: Enders JT, Otto TJ, Peters HC, Wu J, Hardouin S, Moed BR, Zhang Z One of the major obstacles hindering cartilage repair is the integration of the reparative cartilage with the recipient cartilage. The purpose of this study was to develop an in vitro model that can be conveniently applied to simulate and improve the integration of tissue engineered cartilage with native articular cartilage. This model, a cartilage integration construct, consists of a cartilage explant and isolated chondrocytes. The explant was anchored to agarose gel on a culture plate as agarose gelation at 4 degrees C to seal the gap between the bottom of the explant and culture plate surface. Isolated chondrocytes were added and confined in the defect created in the center of the explant. After 4 weeks of culture, neocartilage containing proteoglycans and type II collagen was formed. Minimal integration occurred between the neocartilage and the cartilage explant, resembling the failure of cartilage integration manifested in experimental and clinical cartilage ! repair. In this model, agarose gel anchors the explant onto culture plate by altering temperatures and effectively prevents "leakage" of the isolated chondrocytes from the defect of the explant. This model provides a convenient simulation of the cartilage integration process in vitro and has applications in studies of cartilage integration and cartilage tissue engineering. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res part A, 2010. PMID: 20186769 [PubMed - as supplied by publisher] | | | | 
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