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| Enhanced differentiation of human embryonic stem cells to mesenchymal progenitors by inhibition of TGF-beta/Activin/Nodal signaling using SB-431542.
March 5, 2010 at 8:26 AM
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| | Enhanced differentiation of human embryonic stem cells to mesenchymal progenitors by inhibition of TGF-beta/Activin/Nodal signaling using SB-431542. J Bone Miner Res. 2010 Jan 29; Authors: Mahmood A, Harkness L, Schrøder HD, Abdallah BM, Kassem M Directing differentiation of human embryonic stem cells (hESC) into specific cell types using an easy and reproducible protocol is a prerequisite for the clinical use of hESC in regenerative medicine procedures. Here, we report a protocol for directing the differentiation of hESC into mesenchymal progenitor cells. We demonstrate that inhibition of TGF-beta/Activin/Nodal signaling during embryoid bodies (EB) formation using SB-431542 (SB) in serum free medium, markedly up-regulated paraxial mesodermal markers (TBX6, TBX5), and several myogenic developmental markers including early myogenic transcriptional factors (Myf5, Pax7) as well as myocyte committed markers (NCAM, CD34, Desmin, MHC (fast), alpha-smooth muscle actin, Nkx2.5, cTNT). Continuous inhibition of TGF-beta signaling in EB outgrowth cultures (SB-OG) enriched for myocyte progenitor cells that were PAX7(+) (25%), MYOD1(+) (52%) and NCAM(+) (CD56) (73%). DNA microarray analysis revealed differential up-reg! ulation of 117 genes (>2-fold compared to control cells) annotated to myogenic development and function. Moreover, these cells showed the ability to contract (80% of the population) and formed myofibres when implanted intramuscularly in vivo. Interestingly, SB-OG cells cultured in 10% fetal bovine serum (FBS) developed into a homogeneous population of mesenchymal progenitors that expressed CD markers characteristic of mesenchymal stem cells (MSC): CD44(+) (100%), CD73(+) (98%), CD146(+) (96%) and CD166(+) (88%) with the ability to differentiate into osteoblasts, adipocytes and chondrocytes in vitro and in vivo. Furthermore, microarray analysis of these cells revealed down-regulation of genes related to myogenesis: MYH3 (-167.9 fold), ACTA1 (-161), MYBPH (-139 fold), ACTC (-100.3), MYH8 (-45.5 fold) and MYOT (-41.8 fold) and marked up-regulation of genes related to mesoderm-derived cell lineages. In conclusion, our data provides a simple and versatile protocol for directi! ng the differentiation of hESC into a myogenic lineage and the! n furthe r into mesenchymal progenitors by blocking the TGF-beta signaling pathway. (c) 2010 American Society for Bone and Mineral Research. PMID: 20200949 [PubMed - as supplied by publisher] | | |
| Dramatic increase in cortical thickness induced by femoral marrow ablation followed by a three-month treatment with PTH in rats.
March 5, 2010 at 8:26 AM
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| | Dramatic increase in cortical thickness induced by femoral marrow ablation followed by a three-month treatment with PTH in rats. J Bone Miner Res. 2010 Feb 2; Authors: Zhang Q, Carlson J, Ke HZ, Li J, Kim M, Murphy K, Mehta N, Gilligan J, Vignery A We previously reported that following mechanical ablation of the marrow from the mid-shaft of rat femurs, there is a rapid and abundant, but transient growth of bone, and this growth is enhanced and maintained over a three-week period by the bone anabolic hormone PTH. Here we ask whether further treatment with PTH or bisphosphonates can extend the half-life of the new bone formed in lieu of marrow. We subjected the left femur of rats to mechanical marrow ablation and treated the animals five days a week with PTH for three weeks to replace the marrow by bone, or with vehicle as a control. Some rats were sacrificed and used as positive controls or treated with vehicle, PTH or the bisphosphonate alendronate for a further nine weeks. We subjected both femurs from each rat to soft X-ray, pQCT, microCT, dynamic histomorphometry analysis and biomechanical testing. We also determined the concentrations of serum osteocalcin to confirm the efficacy of PTH. Treatment with PT! H for three months dramatically enhanced endosteal and periosteal bone formation leading to a 30% increase in cortical thickness. In contrast, alendronate protected the bone that had formed in the femoral marrow cavity after marrow ablation and three weeks treatment with PTH, but failed to promote endosteal bone growth and to improve the biomechanical properties of ablated femurs. We further asked whether calcium-phosphate cements could potentiate the formation of bone after marrow ablation. Marrow cavities from ablated femurs were filled with one of two calcium-phosphate cements and rats were treated with PTH or PBS for 84 days. Both cements helped to protect the new bone formed after ablation. To some extent, they promoted the formation of bone after ablation even in the absence of any anabolic hormone. Our data therefore expand the role of PTH in bone engineering and open new avenues of investigations to the field of regenerative medicine and tissue engineering. Local bo! ne marrow removal in conjunction with an anabolic agent, a bis! phosphon ate or a calcium-phosphate cement might provide a new platform for rapid preferential site-directed bone growth in areas of high bone loss. (c) 2010 American Society for Bone and Mineral Research. PMID: 20200940 [PubMed - as supplied by publisher] | | |
| Isolation of amniotic mesenchymal stem cells.
March 5, 2010 at 8:26 AM
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| | Isolation of amniotic mesenchymal stem cells. Curr Protoc Stem Cell Biol. 2010 Mar;Chapter 1:Unit1E.5 Authors: Marongiu F, Gramignoli R, Sun Q, Tahan V, Miki T, Dorko K, Ellis E, Strom SC Mesenchymal stem cells (MSCs) have the ability to differentiate into osteocytes, chondrocytes, and adipocytes and possess immunomodulatory properties. Amniotic membrane from human term placenta is a potential source of multipotent MSCs that could be useful for regenerative medicine. This unit describes a detailed and simple protocol for the isolation of amniotic mesenchymal cells. We also introduce a simple density separation technique for the purification of this cell type from possible contamination with amniotic epithelial cells. Curr. Protoc. Stem Cell Biol. 12:1E.5.1-1E.5.11. (c) 2010 by John Wiley & Sons, Inc. PMID: 20200854 [PubMed - in process] | | |
| Progressive maturation in contracting cardiomyocytes derived from human embryonic stem cells: Qualitative effects on electrophysiological responses to drugs.
March 5, 2010 at 8:26 AM
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| | Progressive maturation in contracting cardiomyocytes derived from human embryonic stem cells: Qualitative effects on electrophysiological responses to drugs. Stem Cell Res. 2010 Feb 6; Authors: Otsuji TG, Minami I, Kurose Y, Yamauchi K, Tada M, Nakatsuji N The field of drug testing currently needs a new integrated assay system, as accurate as systems using native tissues, that will allow us to predict arrhythmia risks of candidate drugs and the relationship between genetic mutations and acquired electrophysiological phenotypes. This could be accomplished by combining the microelectrode array (MEA) system with cardiomyocytes (CMs) derived from human embryonic stem cells (hESC) and induced pluripotential stem cells. CMs have been successfully induced from both types, but their maturation process is not systematically controlled; this results in loss of beating potency and insufficient ion channel function. We generated a transgenic hESC line that facilitates maintenance of hESC-CM clusters every 2 weeks by expressing GFP driven by a cardiac-specific alphaMHC promoter, thereby producing a compact pacemaker lineage within a ventricular population over a year. Further analyses, including quantitative RT-PCR, patch-clamp,! and MEA-mediated QT tests, demonstrated that replating culturing continuously enhanced gene expression, ionic current amplitudes, and resistance to K(+) channel blockades in hESC-CMs. Moreover, temporal three-dimensional (3D) culturing accelerated maturation by restoring the global gene repressive status established in the adhesive status. Replating/3D culturing thus produces hESC-CMs that act as functional syncytia suitable for use in regenerative medicine and accurate drug tests. PMID: 20199896 [PubMed - as supplied by publisher] | | |
| Decontamination of chemical and biological warfare agents with a single multi-functional material.
March 5, 2010 at 8:26 AM
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| | Decontamination of chemical and biological warfare agents with a single multi-functional material. Biomaterials. 2010 Mar 1; Authors: Amitai G, Murata H, D Andersen J, R Koepsel R, J Russell A We report the synthesis of new polymers based on a dimethylacrylamide-methacrylate (DMAA-MA) co-polymer backbone that support both chemical and biological agent decontamination. Polyurethanes containing the redox enzymes glucose oxidase and horseradish peroxidase can convert halide ions into active halogens and exert striking bactericidal activity against gram positive and gram negative bacteria. New materials combining those biopolymers with a family of N-alkyl 4-pyridinium aldoxime (4-PAM) halide-acrylate co-polymers offer both nucleophilic activity for the detoxification of organophosphorus nerve agents and internal sources of halide ions for generation of biocidal activity. Generation of free bromine and iodine was observed in the combined material resulting in bactericidal activity of the enzymatically formed free halogens that caused complete kill of E. coli (>6 log units reduction) within 1 h at 37 degrees C. Detoxification of diisopropylfluorophosphate ! (DFP) by the polyDMAA MA-4-PAM iodide component was dose-dependent reaching 85% within 30 min. A subset of 4-PAM-halide co-polymers was designed to serve as a controlled release reservoir for N-hydroxyethyl 4-PAM (HE 4-PAM) molecules that reactivate nerve agent-inhibited acetylcholinesterase (AChE). Release rates for HE 4-PAM were consistent with hydrolysis of the HE 4-PAM from the polymer backbone. The HE 4-PAM that was released from the polymer reactivated DFP-inhibited AChE at a similar rate to the oxime antidote 4-PAM. PMID: 20199807 [PubMed - as supplied by publisher] | | |
| PHBV microspheres - PLGA matrix composite scaffold for bone tissue engineering.
March 5, 2010 at 8:26 AM
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| | PHBV microspheres - PLGA matrix composite scaffold for bone tissue engineering. Biomaterials. 2010 Mar 1; Authors: Huang W, Shi X, Ren L, Du C, Wang Y Polymer scaffolds, particularly in the form of microspheres, have been employed to support cells growth and deliver drugs or growth factors in tissue engineering. In this study, we have established a scaffold by embedding poly (beta-hydroxybutyrate-co-beta-hydroxyvalerate) (PHBV) microspheres into poly (l-lactic-co-glycolic acid) (PLGA) matrix, according to their different solubility in acetone, with the aim of repairing bone defects. PLGA/PHBV scaffolds had good pore parameters, for example, the porosity of PLGA/30% PHBV scaffold can reach to 81.273 +/- 2.192%. Besides, the pore size distribution of the model was evaluated and the results revealed that the pore size mainly distributed between 50 mum and 200 mum. With increasing the amount of PHBV microspheres, the compressive strength of the PLGA/PHBV scaffold enhanced. The morphology of the hybrid scaffold was rougher than that of pure PLGA scaffold, which had no significant effect on the cell behavior. The in v! itro evaluation suggested that the model is suitable as a scaffold for engineering bone tissue, and has the potential for further applications in drug delivery system. PMID: 20199806 [PubMed - as supplied by publisher] | | |
| Preliminary results of the application of a silk fibroin scaffold to otology.
March 5, 2010 at 8:26 AM
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| | Preliminary results of the application of a silk fibroin scaffold to otology. Otolaryngol Head Neck Surg. 2010 Mar;142(3 Suppl 1):S33-5 Authors: Levin B, Redmond SL, Rajkhowa R, Eikelboom RH, Marano RJ, Atlas MD The surgical treatment to repair chronic tympanic membrane perforations is myringoplasty. Although multiple autologous grafts, allografts, and synthetic graft materials have been used over the years, no single graft material is superior for repairing all perforation types. Recently, the remarkable properties of silk fibroin protein have been studied, with biomedical and tissue engineering applications in mind, across a number of medical and surgical disciplines. The present study examines the use of silk fibroin for its potential suitability as an alternative graft in myringoplasty surgery by investigating the growth and proliferation of human tympanic membrane keratinocytes on a silk fibroin scaffold in vitro. Light microscopy, immunofluorescent staining, and confocal imaging all reveal promising preliminary results. The biocompatibility, transparency, stability, high tensile strength, and biodegradability of fibroin make this biomaterial an attractive option to ! study for this utility. PMID: 20176279 [PubMed - in process] | | |
| Invited commentary.
March 5, 2010 at 8:26 AM
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| | Invited commentary. Ann Thorac Surg. 2010 Jan;89(1):264 Authors: Richenbacher WE PMID: 20103249 [PubMed - indexed for MEDLINE] | | |
| Three-dimensional characterization of osteoclast bone-resorbing activity in the resorption lacunae.
March 5, 2010 at 8:26 AM
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| | Three-dimensional characterization of osteoclast bone-resorbing activity in the resorption lacunae. J Med Dent Sci. 2009 Sep;56(3):107-12 Authors: Soysa NS, Alles N, Aoki K, Ohya K Confocal laser microscopy is a well-recognized research tool in the fields of biological and material science which enables high-resolution images of samples with minimum requirements for specimen preparation. Here we introduce an innovative technique for the 3-D description and measurement of resorption pits using Super Depth Surface Profile Measurement Microscope based on the principle of confocal microscope. We show one example of culturing for 48 h with an established NF-kappaB inhibitor named NBD-peptide after plating mature osteoclasts on dentine slices with osteoblasts. The activity of osteoclasts is measured by determining the volume of resorbed portion of dentine by osteoclasts in vitro. The 3-D surface profile could be obtained by detecting the position at which the reflected laser intensity from the target becomes the maximum on z-axis. The volume and depth of resorption lacunae by stimulated osteoclasts is significantly increased compared to the un-sti! mulated group without changing of resorption area. The increase in volume and depth are dose-dependently inhibited by the NBD-peptide. Comparing to the classical method by measuring 2-D area of pits, analysis based on this technique could provide reliable quantitative assessment reflecting the osteoclast activity. PMID: 20099473 [PubMed - indexed for MEDLINE] | | |
| Augmentation of degenerated human cartilage in vitro using magnetically labeled mesenchymal stem cells and an external magnetic device.
March 5, 2010 at 8:26 AM
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| | Augmentation of degenerated human cartilage in vitro using magnetically labeled mesenchymal stem cells and an external magnetic device. Arthroscopy. 2009 Dec;25(12):1435-41 Authors: Kobayashi T, Ochi M, Yanada S, Ishikawa M, Adachi N, Deie M, Arihiro K PURPOSE: The purpose of this study was to investigate whether it is possible to regenerate degenerated human cartilage in vitro by use of magnetically labeled mesenchymal stem cells (MSCs) and an external magnetic device. METHODS: MSCs from human bone marrow were cultured and magnetically labeled. Degenerated human cartilage was obtained during total knee arthroplasty. The osteochondral fragments were attached to the sidewall of tissue culture flasks, and magnetically labeled MSCs were injected into the flasks. By use of an external magnetic device, a magnetic force was applied for 6 hours to the direction of the cartilage, and then the degenerated cartilage was cultured in chondrogenic differentiation medium for 3 weeks. In the control group a magnetic force was not applied. The specimens were evaluated histologically. RESULTS: A cell layer was formed on the degenerated cartilage as shown by H&E staining. The cell layer was also stained in toluidine blue and ! safranin O and with anti-collagen type II immunostaining, indicating that the cell layer contained an extracellular matrix. In the control group a cell layer was not observed on the cartilage. CONCLUSIONS: We were able to show that our system could deliver MSCs onto degenerated human cartilage and then form an extracellular matrix on the degenerated cartilage in vitro. CLINICAL RELEVANCE: Our novel cell delivery system using magnetic force may lead toward a new treatment option for osteoarthritis. PMID: 19962071 [PubMed - indexed for MEDLINE] | | |
| Bidirectional extracellular matrix signaling during tissue morphogenesis.
March 5, 2010 at 8:26 AM
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| | Bidirectional extracellular matrix signaling during tissue morphogenesis. Cytokine Growth Factor Rev. 2009 Oct-Dec;20(5-6):459-65 Authors: Gjorevski N, Nelson CM Normal tissue development and function are regulated by the interplay between cells and their surrounding extracellular matrix (ECM). The ECM provides biochemical and mechanical contextual information that is conveyed from the cell membrane through the cytoskeleton to the nucleus to direct cell phenotype. Cells, in turn, remodel the ECM and thereby sculpt their local microenvironment. Here we review the mechanisms by which cells interact with, respond to, and influence the ECM, with particular emphasis placed on the role of this bidirectional communication during tissue morphogenesis. We also discuss the implications for successful engineering of functional tissues ex vivo. PMID: 19896886 [PubMed - indexed for MEDLINE] | | |
| Vibration stimulates vocal mucosa-like matrix expression by hydrogel-encapsulated fibroblasts.
March 5, 2010 at 8:26 AM
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| | Vibration stimulates vocal mucosa-like matrix expression by hydrogel-encapsulated fibroblasts. J Tissue Eng Regen Med. 2010 Jan;4(1):62-72 Authors: Kutty JK, Webb K The composition and organization of the vocal fold extracellular matrix (ECM) provide the viscoelastic mechanical properties that are required to sustain high-frequency vibration during voice production. Although vocal injury and pathology are known to produce alterations in matrix physiology, the mechanisms responsible for the development and maintenance of vocal fold ECM are poorly understood. The objective of this study was to investigate the effect of physiologically relevant vibratory stimulation on ECM gene expression and synthesis by fibroblasts encapsulated within hyaluronic acid hydrogels that approximate the viscoelastic properties of vocal mucosa. Relative to static controls, samples exposed to vibration exhibited significant increases in mRNA expression levels of HA synthase 2, decorin, fibromodulin and MMP-1, while collagen and elastin expression were relatively unchanged. Expression levels exhibited a temporal response, with maximum increases observe! d after 3 and 5 days of vibratory stimulation and significant downregulation observed at 10 days. Quantitative assays of matrix accumulation confirmed significant increases in sulphated glycosaminoglycans and significant decreases in collagen after 5 and 10 days of vibratory culture, relative to static controls. Cellular remodelling and hydrogel viscosity were affected by vibratory stimulation and were influenced by varying the encapsulated cell density. These results indicate that vibration is a critical epigenetic factor regulating vocal fold ECM and suggest that rapid restoration of the phonatory microenvironment may provide a basis for reducing vocal scarring, restoring native matrix composition and improving vocal quality. PMID: 19842110 [PubMed - indexed for MEDLINE] | | |
| Donor-matched comparison of dental pulp stem cells and bone marrow-derived mesenchymal stem cells in a rat model.
March 5, 2010 at 8:26 AM
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| | Donor-matched comparison of dental pulp stem cells and bone marrow-derived mesenchymal stem cells in a rat model. J Tissue Eng Regen Med. 2010 Jan;4(1):73-81 Authors: Alge DL, Zhou D, Adams LL, Wyss BK, Shadday MD, Woods EJ, Gabriel Chu TM, Goebel WS Dental pulp stem cells (DPSCs) have drawn much interest for the regeneration of mineralized tissues, and several studies have compared DPSCs to bone marrow-derived mesenchymal stem cells (BMMSCs). However, conflicting results, possibly due to donor-associated variability, have been published and the regenerative potential of DPSCs is currently unclear. In the present study we have sought to address this problem using a donor-matched experimental design to robustly compare the biological properties of DPSCs and BMMSCs. All experiments were performed using cells isolated from a single adult Sprague-Dawley rat. Our results show that DPSCs and BMMSCs had similar morphologies and flow cytometry profiles, were capable of forming colonies in vitro and were capable of osteogenic, chondrogenic and adipogenic differentiation. However, quantitative comparisons revealed that DPSCs had a faster population doubling time and a higher percentage of stem/progenitor cells in the po! pulation, as determined by clonogenic assays. Furthermore, while both cell populations formed mineral in vitro, DPSCs had significantly higher alkaline phosphatase activity than BMMSCs after 3 weeks in osteogenic medium. These data show several key differences between DPSCs and BMMSCs and support the possibility of using DPSCs for mineralized tissue regeneration. PMID: 19842108 [PubMed - indexed for MEDLINE] | | |
| A nanofibrous cell-seeded hydrogel promotes integration in a cartilage gap model.
March 5, 2010 at 8:26 AM
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| | A nanofibrous cell-seeded hydrogel promotes integration in a cartilage gap model. J Tissue Eng Regen Med. 2010 Jan;4(1):25-9 Authors: Maher SA, Mauck RL, Rackwitz L, Tuan RS The presence of a defect in mature articular cartilage can lead to degenerative changes of the joint. This is in part caused by the inability of cartilage to regenerate tissue that is capable of spanning a fissure or crack. In this study, we hypothesized that introduction of a biodegradable cell-seeded nanofibrous hydrogel, Puramatrix(), into a cartilage gap would facilitate the generation of a mechanically stable interface. The effects of chondrocyte incorporation within the hydrogel and supplementation with transforming growth factor-beta3 (TGFbeta3), a known regulator of cell growth and differentiation, on cartilage integration were examined mechanically and histologically as a function of cell density and incubation time. When supplemented with TGFbeta3, the cell-seeded hydrogel exhibited abundant matrix generation within the hydrogel and a corresponding increase in maximum push-out stress as compared to all other groups. Furthermore, initial cell seeding dens! ity affected interfacial strength in a time-dependent manner. This study suggests that a cell-seeded TGFbeta3-supplemented hydrogel can encourage integration between two opposing pieces of articular cartilage. PMID: 19834956 [PubMed - indexed for MEDLINE] | | |
| Human tissue-engineered bone produced in clinically relevant amounts using a semi-automated perfusion bioreactor system: a preliminary study.
March 5, 2010 at 8:26 AM
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| | Human tissue-engineered bone produced in clinically relevant amounts using a semi-automated perfusion bioreactor system: a preliminary study. J Tissue Eng Regen Med. 2010 Jan;4(1):12-24 Authors: Janssen FW, van Dijkhuizen-Radersma R, Van Oorschot A, Oostra J, de Bruijn JD, Van Blitterswijk CA The aim of this study was to evaluate a semi-automated perfusion bioreactor system for the production of clinically relevant amounts of human tissue-engineered bone. Human bone marrow stromal cells (hBMSCs) of eight donors were dynamically seeded and proliferated in a perfusion bioreactor system in clinically relevant volumes (10 cm(3)) of macroporous biphasic calcium phosphate scaffolds (BCP particles, 2-6 mm). Cell load and distribution were shown using methylene blue staining. MTT staining was used to demonstrate viability of the present cells. After 20 days of cultivation, the particles were covered with a homogeneous layer of viable cells. Online oxygen measurements confirmed the proliferation of hBMSCs in the bioreactor. After 20 days of cultivation, the hybrid constructs became interconnected and a dense layer of extracellular matrix was present, as visualized by scanning electron microscopy (SEM). Furthermore, the hBMSCs showed differentiation towards the ! osteogenic lineage as was indicated by collagen type I production and alkaline phosphatase (ALP) expression. We observed no significant differences in osteogenic gene expression profiles between static and dynamic conditions like ALP, BMP2, Id1, Id2, Smad6, collagen type I, osteocalcin, osteonectin and S100A4. For the donors that showed bone formation, dynamically cultured hybrid constructs showed the same amount of bone as the statically cultured hybrid constructs. Based on these results, we conclude that a semi-automated perfusion bioreactor system is capable of producing clinically relevant and viable amounts of human tissue-engineered bone that exhibit bone-forming potential after implantation in nude mice. PMID: 19834955 [PubMed - indexed for MEDLINE] | | |
| In situ adipogenesis in fat tissue augmented by collagen scaffold with gelatin microspheres containing basic fibroblast growth factor.
March 5, 2010 at 8:26 AM
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| | In situ adipogenesis in fat tissue augmented by collagen scaffold with gelatin microspheres containing basic fibroblast growth factor. J Tissue Eng Regen Med. 2010 Jan;4(1):55-61 Authors: Kimura Y, Tsuji W, Yamashiro H, Toi M, Inamoto T, Tabata Y In situ adipose tissue regeneration in fat tissue by collagen sponges and gelatin microspheres containing basic fibroblast growth factor (bFGF) was investigated. A minced collagen sponge scaffold (1 ml) was incorporated with microspheres containing 10 microg bFGF and administered into a defect of rabbit fat tissues. Adipogenesis at the administered site was evaluated histologically. The adipose tissue regeneration induced by the administration of mixed collagen scaffold and microspheres containing bFGF was significantly stronger than that of either collagen scaffold alone or microspheres containing bFGF alone. The histological area of in situ adipogenesis by the mixed collagen scaffold and microspheres containing bFGF was enhanced over time by repeated administration. It is concluded that the repeated administration of collagen scaffold and microspheres containing bFGF is a promising way to achieve adipose tissue regeneration inside inherent fat tissue. This techn! ique might be applicable for the reconstruction of volume contour deformities by trauma or surgical interventions of adipose tissue in a minimally invasive manner. PMID: 19830791 [PubMed - indexed for MEDLINE] | | |
| Utilization of human limbal mesenchymal cells as feeder layers for human limbal stem cells cultured on amniotic membrane.
March 5, 2010 at 8:26 AM
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| | Utilization of human limbal mesenchymal cells as feeder layers for human limbal stem cells cultured on amniotic membrane. J Tissue Eng Regen Med. 2010 Jan;4(1):38-44 Authors: Zhang X, Sun H, Li X, Yuan X, Zhang L, Zhao S Various cell culture techniques for limbal epithelial cells are currently being used for the transplantation of cultured limbal stem cells. In this study, we explored the possibility of using human limbal mesenchymal cells (HLMCs) as feeder layer for the human limbal epithelial cells (HLECs). Single cell suspension of HLECs was seeded onto denuded amniotic membranes with inactivated 3T3 fibroblasts or HLMCs as feeder layer. Expressions of Cytokeratin 3, Np63 and connexin 43 (Cx43) of the cultured epithelial cells were determined at 28 days and the ultrastructure of the epithelium was examined by transmission electron microscope after 14 days and 28 days of cultivation. In both groups, cells were differentiated into multilayer epithelium at 28 days. Basal cells of the cultured epithelium showed a strong nuclear labeling of Np63, but lacked CK3 and Cx43 expression. Transmission electron microscopy examination showed that there were abundant desmosomal contacts betwe! en the cells. The key feature the cultured epithelium was occurrence of a typical basement membrane. These results suggested that HLMCs can be used as an alternative feeder layer for HLECs, which makes the bioengineering product biologically safer for the clinical applications. PMID: 19813216 [PubMed - indexed for MEDLINE] | | |
| Fibulin-1 and fibrinogen in human atherosclerotic lesions.
March 5, 2010 at 8:26 AM
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| | Fibulin-1 and fibrinogen in human atherosclerotic lesions. Histochem Cell Biol. 2009 Nov;132(5):559-65 Authors: Argraves WS, Tanaka A, Smith EP, Twal WO, Argraves KM, Fan D, Haudenschild CC Fibulin-1 is a fibrinogen-binding blood protein and a component of many extracellular matrices (ECM) including those of blood vessels. In this study, the deposition patterns of fibulin-1 and fibrinogen were examined in human coronary artery atherosclerotic lesions excised by atherectomy from 20 patients. Fibulin-1 deposition was found to be closely overlapping with fibrinogen located within the atherosclerotic lesions and in regions containing fresh thrombi. Pronounced intracellular fibulin-1 immunostaining was apparent in lesion areas rich in macrophages and foam cells, although THP-1 macrophages and foam cells were found not to express fibulin-1. Strong ECM deposition of fibulin-1 was observed in acellular atheromatous and myxomatous regions. By contrast, fibulin-1 was present at relatively low levels in the ECM associated with smooth muscle cells within and outside of lesions and was not detected in sclerotic regions. These results reveal the pattern of fibulin! -1 within human atherosclerotic lesions and highlight the potential for fibulin-1, perhaps derived from the blood and acting in conjunction with fibrinogen, to play a role in the etiology and cardiovascular disease progression, particularly with respect to thrombotic aspects of atherosclerosis. PMID: 19693531 [PubMed - indexed for MEDLINE] | | |
| [Cell sheet fabrication of hepatocyte-like cells differentiated from adipose tissue mesenchymal stem cells]
March 5, 2010 at 8:26 AM
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| | [Cell sheet fabrication of hepatocyte-like cells differentiated from adipose tissue mesenchymal stem cells] Sheng Wu Gong Cheng Xue Bao. 2009 Apr;25(4):599-604 Authors: Lu Y, Qiu F, Chen Y, Zhao X Adult pluripotent stem cells, such as mesenchymal stem cells derived from bone marrow and adipose tissue are capable of multilineage differentiation. Although autologous stem cell transplantation is an effective alternative to organ transplantation, the loss of cell viability and differentiation confinement of implanted cells has largely impaired the therapeutic efficacy. To produce biomaterial-free liver construct to integrate into living tissue, we isolated adipose mesenchymal stem cells and subjected them to a delicate culture configuration to mediate the hepatocyte differentiation. The differentiated hepatocyte-like cells were then inoculated onto poly (N-isopropylacrylamide) (PNIPAAm) grafted cell culture dish. By lowering the culture temperature to 20 degrees C, cells detached from the dish surface into a complete cell sheet. Hematoxylin and eosin staining and immunohistochemistry results showed that cell sheet was composed of 2-3 layers of cells and extrace! llular matrix was maintained intact. As compared with traditional cell harvest using trypsin digestion, cell sheet fabrication causes no damage to cell membrane and extracellular matrix. Hence, cell sheets would form a better interaction with tissues in situ, and a higher cell viability and therapeutic efficiency would be expected. PMID: 19637638 [PubMed - indexed for MEDLINE] | | |
| [Characterization and biocompatibility of human-like collagen-hyaluronic acid scaffold for blood vessel]
March 5, 2010 at 8:26 AM
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| | [Characterization and biocompatibility of human-like collagen-hyaluronic acid scaffold for blood vessel] Sheng Wu Gong Cheng Xue Bao. 2009 Apr;25(4):591-8 Authors: Sun X, Fan D, Zhu C, Ma X, Luo Y, Chen L, Guo J Human-like collagen (HLC) was cross-linked with hyaluronic acid by genipin in different ratio. The concentrations of hyaluronic acid in the mixture were 0, 0.01%, 0.05% and 0.1%. The blood vessel tubular grafts were then fabricated by freeze-drying. Microstructure, element composite, mechanical properties, cytotoxicity grade, and biocompatibility of different vascular scaffold groups were studied by scanning electron microscope (SEM), X-ray photoelectron spectroscopy (XPS), tensile test, burst pressure experiment, cytotoxicity experiment, endothelial cells planted in blood vessel scaffolds and hypodermic embedding of mice. The results showed that HLC-HA (0.05%) tubular scaffold exhibited interconnected well-distributed and porous structure and porosity of 94.38%; achieved the desirable mechanical property with stress of (1000.8 +/- 7.9) kPa and burst pressure of (1058.6 +/- 8.2) kPa, hypocytotoxicity, favourable cytocompatibility, hisocompatibility and disposition! of degradation. PMID: 19637637 [PubMed - indexed for MEDLINE] | | |
| Liver-derived extracellular matrix as a biologic scaffold for acute vocal fold repair in a canine model.
March 5, 2010 at 8:26 AM
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| | Liver-derived extracellular matrix as a biologic scaffold for acute vocal fold repair in a canine model. Laryngoscope. 2009 Sep;119(9):1856-63 Authors: Gilbert TW, Agrawal V, Gilbert MR, Povirk KM, Badylak SF, Rosen CA OBJECTIVES/HYPOTHESIS: The objective of the study was to evaluate a naturally derived liver extracellular matrix (L-ECM) scaffold for repair of an acute injury to the vocal fold lamina propria in a canine model. METHODS: The vocal fold lamina propria was removed bilaterally in four dogs. One vocal fold in each dog was repaired with a porcine L-ECM scaffold, which was chosen because it contains hepatocyte growth factor, an antifibrotic growth factor that aids the healing of vocal folds. The other vocal fold was left untreated. At 3 months after surgery, morphologic and histologic analysis was performed to assess the vocal fold shape, collagen density, collagen composition, elastic fiber content, and glycosaminoglycan content. RESULTS: The L-ECM-treated vocal fold showed increased collagen density in the superficial aspect of the vocal fold (P < .05). The L-ECM-treated vocal fold also showed an increased collagen III/I ratio as compared to the nontreated group (P! < .05). However, the elastic fiber content was found to be increased in both groups, and the glycosaminoglycan content was decreased in both groups as compared to the normal vocal fold (P < .05) with no differences detected between groups. CONCLUSIONS: The L-ECM scaffold did not restore the biochemical composition or histologic appearance of the injured vocal fold as compared to normal. However, the increased ratio of collagen III/I and elastic fiber content suggests that L-ECM leads to formation of connective tissue that may be more pliable as compared to no treatment. Additional investigation, including functional assessment, is warranted. PMID: 19572393 [PubMed - indexed for MEDLINE] | | |
| Proteomic analysis reveals significant elevation of heat shock protein 70 in patients with chronic heart failure due to arrhythmogenic right ventricular cardiomyopathy.
March 5, 2010 at 8:26 AM
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| | Proteomic analysis reveals significant elevation of heat shock protein 70 in patients with chronic heart failure due to arrhythmogenic right ventricular cardiomyopathy. Mol Cell Biochem. 2009 Dec;332(1-2):103-11 Authors: Wei YJ, Huang YX, Shen Y, Cui CJ, Zhang XL, Zhang H, Hu SS As proteins are the ultimate biological determinants of phenotype of disease, we screened altered proteins associated with heart failure due to arrhythmogenic right ventricular cardiomyopathy (ARVC) to identify biomarkers potential for rapid diagnosis of heart failure. By 2-dimensional gel electrophoresis and mass spectrometry, we identified five commonly altered proteins with more than 1.5 fold changes in eight ARVC failing hearts using eight non-failing hearts as reference. Noticeably, one of the altered proteins, heat shock protein 70 (HSP70), was increased by 1.64 fold in ARVC failing hearts compared with non-failing hearts. The increase of cardiac HSP70 was further validated by Western blot, immunochemistry, and enzyme-linked immunosorbent assay (ELISA) in failing hearts due to not only ARVC, but also dilated (DCM, n = 18) and ischemic cardiomyopathy (ICM, n = 8). Serum HSP70 was also observed to be significantly increased in heart failure patients derived fr! om the three forms of cardiomyopathies. In addition, we observed hypoxia/serum depletion stimulation induced significantly elevation of intracellular and extracellular HSP70 in cultured neonatal rat cardiomyocytes. For the first time to our knowledge, we revealed and clearly demonstrated significant up-regulation of cardiac and serum HSP70 in ARVC heart failure patients. Our results indicate that elevated HSP70 is the common feature of heart failure due to ARVC, DCM, and ICM, which suggests that HSP70 may be used as a biomarker for the presence of heart failure due to cardiomyopathies of different etiologies and may hold diagnostic/prognostic potential in clinical practice. PMID: 19543852 [PubMed - indexed for MEDLINE] | | |
| Effects of high doses of ionising radiation on bone in rats: a new model for evaluation of bone engineering.
March 5, 2010 at 8:26 AM
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| | Effects of high doses of ionising radiation on bone in rats: a new model for evaluation of bone engineering. Br J Oral Maxillofac Surg. 2009 Dec;47(8):602-7 Authors: Lerouxel E, Moreau A, Bouler JM, Giumelli B, Daculsi G, Weiss P, Malard O The purpose of this study was to assess the effects of high doses of ionising radiation on the histology and healing of bone in an experimental model of 12 inbred rats. Ten of the rats had external irradiation of a single dose of 30 or 45 Gy on the hind limbs, which is equivalent to 2 or 3 times the routine doses used for treatment in humans. Three weeks later, two bony defects were created on their left sides, and the animals were killed 12 or 18 weeks after irradiation. Decalcified bony specimens were studied with light microscopy for qualitative analysis. Thirty Gy irradiation induced medullar oedema or fibro-oedema and normal or fibrous healing of the defects. Forty-five Gy induced medullar oedema or fibro-oedema and depletion in bone marrow. In addition, pathological healing of the defects was obvious and characterised by oedema, fibrosis, and necrosis. In this study high doses of ionising radiation modified the histology of bone, particularly into fibro-oede! ma, and delayed healing. This new animal model could be used to evaluate the capacities of tissue-engineered materials to repair bony defects after irradiation and osteoradionecrosis. PMID: 19200627 [PubMed - indexed for MEDLINE] | | |
| Stem cell transplantation for autoimmune diseases: what can we learn from experimental models?
March 5, 2010 at 8:26 AM
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| | Stem cell transplantation for autoimmune diseases: what can we learn from experimental models? Autoimmunity. 2008 Dec;41(8):563-9 Authors: Ikehara S Using animal models for autoimmune diseases, we have previously shown that allogeneic bone marrow transplantation (allo BMT) can be used to treat autoimmune diseases. Using cynomolgus monkeys, we have recently developed new BMT methods for the treatment of autoimmune diseases. The methods include the perfusion method (PM) for the collection of bone marrow cells (BMCs), and intra-bone marrow (IBM)-BMT for the direct injection of collected whole BMCs into the bone marrow cavity. The PM, in comparison with the conventional aspiration method, can minimize the contamination of BMCs with T cells from the peripheral blood. Therefore, without removing T cells, no graft-versus-host disease (GvHD) develops in the case of the PM. Since BMCs collected by the PM contain not only hemopoietic stem cells (HSCs) but also mesenchymal stem cells (MSCs), the injection of both cells directly into the bone marrow cavity (IBM-BMT) facilitates the engraftment of donor hemopoietic cells. ! In organ allografts with IBM-BMT, no graft failure occurs even if the radiation dose is reduced. In addition, IBM-BMT is applicable to regeneration therapy and various age-associated diseases such as osteoporosis, since it can efficiently recruit donor-derived normal MSCs. We have also found that IBM-BMT in conjunction with donor lymphocyte infusion can prevent GvHD, but suppress tumor growth. We believe that this strategy heralds a revolution in the field of transplantation (BMT and organ allografts) and regeneration therapy. PMID: 18958757 [PubMed - indexed for MEDLINE] | | | | 
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