|  |  |  | | | | | TE-RegenMed-StemCell feed | | | | | | | | | | | | | | | | | | | Immunohistochemical parcellation of the ferret (Mustela putorius) visual cortex reveals substantial homology with the cat (Felis catus). J Comp Neurol. 2010 Nov 1;518(21):4439-62 Authors: Homman-Ludiye J, Manger PR, Bourne JA Electrophysiological mapping of the adult ferret visual cortex has until now determined the existence of 12 retinotopically distinct areas; however, in the cat, another member of the Carnivora, 20 distinct visual areas have been identified by using retinotopic mapping and immunolabeling. In the present study, the immunohistochemical approach to demarcate the areal boundaries of the adult ferret visual cortex was applied in order to overcome the difficulties in accessing the sulcal surfaces of a small, gyrencephalic brain. Nonphosphorylated neurofilament (NNF) expression profiles were compared with another classical immunostain of cortical nuclei, Cat-301 chondroitin sulfate proteoglycan (CSPG). Together, these two markers reliably demarcated the borders of the 12 previously defined areas and revealed further arealization beyond those borders to a total of 19 areas: 21a and 21b; the anterolateral, posterolateral, dorsal, and ventral lateral suprasylvian areas (ALLS, PLLS, DLS, and VLS, respectively); and the splenial and cingulate visual areas (SVA and CVA). NNF expression profile and location of the newly defined areas correlate with previously defined areas in the cat. Moreover, NNF and Cat-301 together revealed discrete expression domains in the posteroparietal (PP) cortex, demarcating four subdivisions in the caudal lateral and medial domains (PPcL and PPcM) and rostral lateral and medial domains (PPrL and PPrM), where only two retinotopic maps have been previously identified (PPc and PPr). Taken together, these studies suggest that NNF and Cat-301 can illustrate the homology between cortical areas in different species and draw out the principles that have driven evolution of the visual cortex. J. Comp. Neurol. 518:4439-4462, 2010. © 2010 Wiley-Liss, Inc. PMID: 20853515 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | The Potential of Adipose Stem Cells in Regenerative Medicine. Stem Cell Rev. 2010 Sep 18; Authors: Lindroos B, Suuronen R, Miettinen S Adipose stem cells (ASCs) are an attractive and abundant stem cell source with therapeutic applicability in diverse fields for the repair and regeneration of acute and chronically damaged tissues. Importantly, unlike the human bone marrow stromal/stem stem cells (BMSCs) that are present at low frequency in the bone marrow, ASCs can be retrieved in high number from either liposuction aspirates or subcutaneous adipose tissue fragments and can easily be expanded in vitro. ASCs display properties similar to that observed in BMSCs and, upon induction, undergo at least osteogenic, chondrogenic, adipogenic and neurogenic, differentiation in vitro. Furthermore, ASCs have been shown to be immunoprivileged, prevent severe graft-versus-host disease in vitro and in vivo and to be genetically stable in long-term culture. They have also proven applicability in other functions, such as providing hematopoietic support and gene transfer. Due to these characteristics, ASCs have rapidly advanced into clinical trials for treatment of a broad range of conditions. As cell therapies are becoming more frequent, clinical laboratories following good manufacturing practices are needed. At the same time as laboratory processes become more extensive, the need for control in the processing laboratory grows consequently involving a greater risk of complications and possibly adverse events for the recipient. Therefore, the safety, reproducibility and quality of the stem cells must thoroughly be examined prior to extensive use in clinical applications. In this review, some of the aspects of examination on ASCs in vitro and the utilization of ASCs in clinical studies are discussed. PMID: 20853072 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Three-Dimensional Quantitative Micromorphology of Pre- and Post-Implanted Engineered Heart Valve Tissues. Ann Biomed Eng. 2010 Sep 18; Authors: Eckert CE, Mikulis BT, Gottlieb D, Gerneke D, Legrice I, Padera RF, Mayer JE, Schoen FJ, Sacks MS There is a significant gap in our knowledge of engineered heart valve tissue (EHVT) development regarding detailed three-dimensional (3D) tissue formation and remodeling from the point of in vitro culturing to full in vivo function. As a step toward understanding the complexities of EHVT formation and remodeling, a novel serial confocal microscopy technique was employed to obtain 3D microstructural information of pre-implant (PRI) and post-implant for 12 weeks (POI) EHVT fabricated from PGA:PLLA scaffolds and seeded with ovine bone-marrow-derived mesenchymal stem cells. Custom scaffold fiber tracking software was developed to quantify scaffold fiber architectural features such as length, tortuosity, and minimum scaffold fiber-fiber separation distance and scaffold fiber orientation was quantified utilizing a 3D fabric tensor. In addition, collagen and cellular density of ovine pulmonary valve leaflet tissue were also analyzed for baseline comparisons. Results indicated that in the unseeded state, scaffold fibers formed a continuous, oriented network. In the PRI state, the scaffold showed some fragmentation with a scaffold volume fraction of 7.79%. In the POI specimen, the scaffold became highly fragmented, forming a randomly distributed short fibrous network (volume fraction of 2.03%) within a contiguous, dense collagenous matrix. Both PGA and PLLA scaffold fibers were observed in the PRI and POI specimens. Collagen density remained similar in both PRI and POI specimens (74.2 and 71.5%, respectively), though the distributions in the transmural direction appeared slightly more homogenous in the POI specimen. Finally, to guide future 2D histological studies for large-scale studies (since acquisition of high-resolution volumetric data is not practical for all specimens), we investigated changes in relevant collagen and scaffold metrics (collagen density and scaffold fiber orientation) with varying section spacing. It was found that a sectioning spacing up to 25 μm (for scaffold morphology) and 50 μm (for collagen density) in both PRI and POI tissues did not result in loss of information fidelity, and that sectioning in the circumferential or radial direction provides the greatest preservation of information. This is the first known work to investigate EHVT microstructure over a large volume with high resolution and to investigate time evolving in vivo EHVT morphology. The important scaffold fiber structural changes observed provide morphological information crucial for guiding future structurally based constitutive modeling efforts focused on better understanding EHVT tissue formation and remodeling. PMID: 20853027 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Microsatellite Instability Detection by High-Resolution Melting Analysis. Clin Chem. 2010 Sep 17; Authors: Janavicius R, Matiukaite D, Jakubauskas A, Griskevicius L BACKGROUND: Microsatellite instability (MSI) is an important marker for screening for hereditary nonpolyposis colorectal cancer (Lynch syndrome) as well as a prognostic and predictive marker for sporadic colorectal cancer (CRC). The mononucleotide microsatellite marker panel is a well-established and superior alternative to the traditional Bethesda MSI analysis panel, and does not require testing for corresponding normal DNA. The most common MSI detection techniques-fluorescent capillary electrophoresis and denaturing HPLC (DHPLC)-both have advantages and drawbacks. A new high-resolution melting (HRM) analysis method enables rapid identification of heteroduplexes in amplicons by their lower thermal stability, a technique that overcomes the main shortcomings of capillary electrophoresis and DHPLC. METHODS: We investigated the straightforward application of HRM for the detection of MSI in 70 archival CRC samples. HRM analysis for 2 MSI markers (BAT25 and BAT26) was evaluated, and 2 different HRM-enabled instruments were compared-the LightCycler® 480 (Roche Diagnostics) and the LightScanner(TM) (Idaho Technology). We also determined the analytical sensitivity and specificity of the HRM assay on both instruments using 11 known MSI-positive and 54 microsatellite-stable CRC samples. RESULTS: All MSI-positive samples were detected on both instruments (100% analytical sensitivity). The LightScanner performed better for analytical specificity, giving a combined specificity value of 99.1% compared with 92.3% on the LightCycler 480. CONCLUSIONS: We expanded the application of the HRM analysis method as an effective MSI detection technique for clinical samples. PMID: 20852132 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Increased mechanosensitivity of cells cultured on nanotopographies. J Biomech. 2010 Sep 17; Authors: Salvi JD, Yul Lim J, Donahue HJ Enhancing cellular mechanosensitivity is recognized as a novel tool for successful musculoskeletal tissue engineering. We examined the hypothesis that mechanosensitivity of human mesenchymal stem cells (hMSCs) is enhanced on nanotopographic substrates relative to flat surfaces. hMSCs were cultured on polymer-demixed, randomly distributed nanoisland surfaces with varying island heights and changes in intracellular calcium concentration, [Ca(2+)](i), in response to fluid flow induced shear stress were quantifide. Stem cells cultured on specific scale nanotopographies displayed greater intracellular calcium responses to fluid flow. hMSCs cultured on 10-20nm high nanoislands displayed a greater percentage of cells responding in calcium relative to cells cultured on flat control, and showed greater average [Ca(2+)](i) increase relative to cells cultured on other nanoislands (45-80nm high nanoislands). As [Ca(2+)](i) is an important regulator of downstream signaling, as well as proliferation and differentiation of hMSCs, this observation suggests that specific scale nanotopographies provide an optimal milieu for promoting stem cell mechanotransduction activity. That mechanical signals and substrate nanotopography may synergistically regulate cell behavior is of significant interest in the development of regenerative medicine protocols. PMID: 20851397 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Dystrophin-deficient zebrafish feature aspects of the Duchenne muscular dystrophy pathology. Neuromuscul Disord. 2010 Sep 15; Authors: Berger J, Berger S, Hall TE, Lieschke GJ, Currie PD Duchenne muscular dystrophy is caused by mutations in the dystrophin gene. As in humans, zebrafish dystrophin is initially expressed at the peripheral ends of the myofibres adjacent to the myotendinous junction and gradually shifts to non-junctional sites. Dystrophin-deficient zebrafish larvae are characterised by abundant necrotic fibres being replaced by mono-nucleated infiltrates, extensive fibrosis accompanied by inflammation, and a broader variation in muscle fibre cross-sectional areas. Muscle progenitor proliferation cannot compensate for the extensive skeletal muscle loss. Live imaging of dystrophin-deficient zebrafish larvae documents detaching myofibres elicited by muscle contraction. Correspondingly, the progressive phenotype of dystrophin-deficient zebrafish resembles many aspects of the human disease, suggesting that specific advantages of the zebrafish model system, such as the ability to undertake in vivo drug screens and real time analysis of muscle fibre loss, could be used to make novel insights relevant to understanding and treating the pathological basis of dystrophin-deficient muscular dystrophy. PMID: 20850317 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Gene profiling of bone marrow- and adipose tissue-derived stromal cells: a key role of Kruppel-like factor 4 in cell fate regulation. Cytotherapy. 2010 Sep 20; Authors: Saulnier N, Puglisi MA, Lattanzi W, Castellini L, Pani G, Leone G, Alfieri S, Michetti F, Piscaglia AC, Gasbarrini A Abstract Background aims. Bone marrow- and adipose tissue-derived mesenchymal stromal cells (MSC) represent promising sources for regenerative medicine. However, the precise molecular mechanisms underlying MSC stemness maintenance versus differentiation are not fully understood. The aim of this study was to compare the genome-wide expression profiles of bone marrow- and adipose tissue-derived MSC, in order to identify a common molecular stemness core. Methods. Molecular profiling was carried out using Affymetrix microarray and relevant genes were further validated by Q-PCR. Results. We identified an overlapping dataset of 190 transcripts commonly regulated in both cell populations, which included several genes involved in stemness regulation (i.e. self-renewal potential and the ability to generate differentiated cells), various signaling pathways and transcription factors. In particular, we identified a central role of the Kruppel-like factor 4 (KLF4) DNA-binding protein in regulating MSC transcriptional activity. Conclusions. Our results provide new insights toward understanding the molecular basis of MSC stemness maintenance and underline the ability of KLF4 to maintain cells in an undifferentiated state. PMID: 20849362 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | California jockey Michael Martinez, whose spine was severely damaged earlier this month, will not be enrolled in Geron's clincal trials for hESC treatment, the only such in the nation.
Reporter Matt Hegarty of the Daily Racing Form wrote this afternoon,
"Specialists at Northwestern University near Chicago have decided that the rider Michael Martinez is not a candidate for a clinical trial using | | | | | | | | | | | | | | | | | | | | | | | | Benoit & AssociatesJockey Michael Martinez (left), with cousin Alex Solis.
More details are emerging on the possible use of experimental hESC treatments within the next two weeks on a California jockey with a severed spinal cord.
According to an article yesterday by Chuck Dybdal in the Racing Form, which appears to have carried the first account of the possible treatment, it involves | | | | | | | | | | | | | | | | | | | | | | | | The sports section of the San Francisco Chronicle carried an intriguing but brief story this morning dealing with use of embryonic stem cells to "repair" the severed spinal cord of an injured jockey.
The story by Larry Stumes said Michael Martinez is in the UCSF medical center for an MRI exam to determine whether he will be flown to Chicago this week to determine whether he is a candidate for | | | | | | | | | | | | | | | | | | | | | | | | | Adipose tissue engineering from adult human stem cells: a new concept in biosurgery. Facial Plast Surg. 2010 Oct;26(5):413-20 Authors: Ahn JM, Mao JJ Current autologous fat grafting technique suffers from the drawbacks of donor site morbidity and, more importantly, significant resorption of the grafted fat. Adipose tissue engineering using adult human stem cells has been found to overcome the shortcomings of autologous fat grafting in reconstructing facial defects. Mesenchymal stem cells that can self-renew and differentiate into mature adipocytes have been used to generate adipose tissue, in both in vitro and in vivo cell transplantation studies. However, long-term maintenance of the shape and dimension of the produced adipose tissue remains a challenge, even in tissue engineering with cell transplantation. The choice of appropriate scaffolds to promote stem cell adhesion, proliferation, and differentiation is essential for successful adipogenesis. Recent advances in nanotechnology allow the development of nanostructured scaffolds with a cellular environment that maximally enhances not only cell expansion but also the neovascularization that is crucial for long-term maintenance of cell volume. Cell homing is a technique that actively recruits endogenous host stem cells into a predefined anatomic location for the desired tissue generation. Bypassing ex vivo cell manipulation, the cell homing technique eliminates donor site morbidity and rejection, reducing the regulation issue in clinical translation. Mao et al. introduced the concept of biosurgery, which combined nanostructured scaffolds and growth factor biocues, with or without cell transplantation, for successful de novo adipogenesis in restoring facial defects. Important questions, such as the necessity of cell transplantation in scaling up the size of engineered adipose tissue, need to be answered with further studies. However, the era of biosurgery replacing conventional treatments such as biologically inactive filler injections and alloplastic implants appears to be in the near future. PMID: 20853233 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | The Potential of Adipose Stem Cells in Regenerative Medicine. Stem Cell Rev. 2010 Sep 18; Authors: Lindroos B, Suuronen R, Miettinen S Adipose stem cells (ASCs) are an attractive and abundant stem cell source with therapeutic applicability in diverse fields for the repair and regeneration of acute and chronically damaged tissues. Importantly, unlike the human bone marrow stromal/stem stem cells (BMSCs) that are present at low frequency in the bone marrow, ASCs can be retrieved in high number from either liposuction aspirates or subcutaneous adipose tissue fragments and can easily be expanded in vitro. ASCs display properties similar to that observed in BMSCs and, upon induction, undergo at least osteogenic, chondrogenic, adipogenic and neurogenic, differentiation in vitro. Furthermore, ASCs have been shown to be immunoprivileged, prevent severe graft-versus-host disease in vitro and in vivo and to be genetically stable in long-term culture. They have also proven applicability in other functions, such as providing hematopoietic support and gene transfer. Due to these characteristics, ASCs have rapidly advanced into clinical trials for treatment of a broad range of conditions. As cell therapies are becoming more frequent, clinical laboratories following good manufacturing practices are needed. At the same time as laboratory processes become more extensive, the need for control in the processing laboratory grows consequently involving a greater risk of complications and possibly adverse events for the recipient. Therefore, the safety, reproducibility and quality of the stem cells must thoroughly be examined prior to extensive use in clinical applications. In this review, some of the aspects of examination on ASCs in vitro and the utilization of ASCs in clinical studies are discussed. PMID: 20853072 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Composite cell sheets: a further step toward safe and effective myocardial regeneration by cardiac progenitors derived from embryonic stem cells. Circulation. 2010 Sep 14;122(11 Suppl):S118-23 Authors: Bel A, Planat-Bernard V, Saito A, Bonnevie L, Bellamy V, Sabbah L, Bellabas L, Brinon B, Vanneaux V, Pradeau P, Peyrard S, Larghero J, Pouly J, Binder P, Garcia S, Shimizu T, Sawa Y, Okano T, Bruneval P, Desnos M, Hagège AA, Casteilla L, Pucéat M, Menasché P BACKGROUND: The safety and efficacy of myocardial regeneration using embryonic stem cells are limited by the risk of teratoma and the high rate of cell death. METHODS AND RESULTS: To address these issues, we developed a composite construct made of a sheet of adipose tissue-derived stroma cells and embryonic stem cell-derived cardiac progenitors. Ten Rhesus monkeys underwent a transient coronary artery occlusion followed, 2 weeks later, by the open-chest delivery of the composite cell sheet over the infarcted area or a sham operation. The sheet was made of adipose tissue-derived stroma cells grown from a biopsy of autologous adipose tissue and cultured onto temperature-responsive dishes. Allogeneic Rhesus embryonic stem cells were committed to a cardiac lineage and immunomagnetically sorted to yield SSEA-1(+) cardiac progenitors, which were then deposited onto the cell sheet. Cyclosporine was given for 2 months until the animals were euthanized. Preimplantation studies showed that the SSEA-1(+) progenitors expressed cardiac markers and had lost pluripotency. After 2 months, there was no teratoma in any of the 5 cell-treated monkeys. Analysis of >1500 histological sections showed that the SSEA-1(+) cardiac progenitors had differentiated into cardiomyocytes, as evidenced by immunofluorescence and real-time polymerase chain reaction. There were also a robust engraftment of autologous adipose tissue-derived stroma cells and increased angiogenesis compared with the sham animals. CONCLUSIONS: These data collected in a clinically relevant nonhuman primate model show that developmentally restricted SSEA-1(+) cardiac progenitors appear to be safe and highlight the benefit of the epicardial delivery of a construct harboring cells with a cardiomyogenic differentiation potential and cells providing them the necessary trophic support. PMID: 20837902 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Promoter DNA methylation patterns of differentiated cells are largely programmed at the progenitor stage. Mol Biol Cell. 2010 Jun 15;21(12):2066-77 Authors: Sørensen AL, Jacobsen BM, Reiner AH, Andersen IS, Collas P Mesenchymal stem cells (MSCs) isolated from various tissues share common phenotypic and functional properties. However, intrinsic molecular evidence supporting these observations has been lacking. Here, we unravel overlapping genome-wide promoter DNA methylation patterns between MSCs from adipose tissue, bone marrow, and skeletal muscle, whereas hematopoietic progenitors are more epigenetically distant from MSCs as a whole. Commonly hypermethylated genes are enriched in signaling, metabolic, and developmental functions, whereas genes hypermethylated only in MSCs are associated with early development functions. We find that most lineage-specification promoters are DNA hypomethylated and harbor a combination of trimethylated H3K4 and H3K27, whereas early developmental genes are DNA hypermethylated with or without H3K27 methylation. Promoter DNA methylation patterns of differentiated cells are largely established at the progenitor stage; yet, differentiation segregates a minor fraction of the commonly hypermethylated promoters, generating greater epigenetic divergence between differentiated cell types than between their undifferentiated counterparts. We also show an effect of promoter CpG content on methylation dynamics upon differentiation and distinct methylation profiles on transcriptionally active and inactive promoters. We infer that methylation state of lineage-specific promoters in MSCs is not a primary determinant of differentiation capacity. Our results support the view of a common origin of mesenchymal progenitors. PMID: 20410135 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Adipose tissue engineering from adult human stem cells: a new concept in biosurgery. Facial Plast Surg. 2010 Oct;26(5):413-20 Authors: Ahn JM, Mao JJ Current autologous fat grafting technique suffers from the drawbacks of donor site morbidity and, more importantly, significant resorption of the grafted fat. Adipose tissue engineering using adult human stem cells has been found to overcome the shortcomings of autologous fat grafting in reconstructing facial defects. Mesenchymal stem cells that can self-renew and differentiate into mature adipocytes have been used to generate adipose tissue, in both in vitro and in vivo cell transplantation studies. However, long-term maintenance of the shape and dimension of the produced adipose tissue remains a challenge, even in tissue engineering with cell transplantation. The choice of appropriate scaffolds to promote stem cell adhesion, proliferation, and differentiation is essential for successful adipogenesis. Recent advances in nanotechnology allow the development of nanostructured scaffolds with a cellular environment that maximally enhances not only cell expansion but also the neovascularization that is crucial for long-term maintenance of cell volume. Cell homing is a technique that actively recruits endogenous host stem cells into a predefined anatomic location for the desired tissue generation. Bypassing ex vivo cell manipulation, the cell homing technique eliminates donor site morbidity and rejection, reducing the regulation issue in clinical translation. Mao et al. introduced the concept of biosurgery, which combined nanostructured scaffolds and growth factor biocues, with or without cell transplantation, for successful de novo adipogenesis in restoring facial defects. Important questions, such as the necessity of cell transplantation in scaling up the size of engineered adipose tissue, need to be answered with further studies. However, the era of biosurgery replacing conventional treatments such as biologically inactive filler injections and alloplastic implants appears to be in the near future. PMID: 20853233 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Embryonic progenitor cells in adipose tissue engineering. Facial Plast Surg. 2010 Oct;26(5):405-12 Authors: Hillel AT, Elisseeff JH Adipose tissue is extensively used in facial plastic surgery as a soft tissue filler for small-to-large facial defects. Variable results with autologous fat grafting and lipoinjection has led to interest in alternative forms of adipose tissue, including tissue engineered adipose tissue. Tissue engineering combines cells, scaffolds, and bioactive signals to regenerate organs or tissue. Cell sources include preadipocytes, adult stem cells, and embryonic stem cells. Although adult cells may be easily accessible from a patient, embryonic progenitor cells have comparative advantages over adult stem cells including indefinite self-renewal (high proliferative and expansion capacity) and strong tissue-forming capacity. This article will describe the types of embryonic progenitor cells and the cell culture conditions, common biomaterials, signaling factors, and biomechanical forces used in adipose tissue engineering. We will identify optimal conditions to generate functional, long-lasting adipose-like tissue. Lastly, we will propose potential future directions for the rapidly expanding field of adipose tissue engineering. PMID: 20853232 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Septal cartilage tissue engineering: new horizons. Facial Plast Surg. 2010 Oct;26(5):396-404 Authors: Greene JJ, Watson D Cartilage tissue engineering is a dynamically changing field that has the potential to address some of the tissue repair challenges seen in nasal and craniofacial reconstructive surgeries. The scope of the problem includes limited autologous tissue availability, donor site morbidity associated with the harvesting of these tissue grafts, and the risk of an immune reaction to allogenic or synthetic implants that might be used as alternatives. Current tissue engineering strategies involve harvesting a small biopsy specimen from a patient and then isolating chondrocytes through enzymatic digestion of the extracellular matrix. These isolated chondrocytes can be expanded in monolayer and reseeded into a three-dimensional scaffold that could potentially be used as autologous surgical grafts. Using cell-expansion techniques, it would be feasible to generate abundant amounts of cartilage in defined shapes and sizes. The ideal tissue-engineered cartilage would resemble native tissue in terms of its biochemical, structural, and metabolic properties so that it could restore stability, function, and contour to the damaged or defective facial region. In this article, emerging technology and major challenges are described to highlight recent advances and overall trends within septal cartilage tissue engineering. PMID: 20853231 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Clinical applications of stem cells in craniofacial surgery. Facial Plast Surg. 2010 Oct;26(5):385-95 Authors: Runyan CM, Taylor JA Few areas of translational medicine carry as much excitement and hope as stem cell therapies. Because of recent advances in material science and stem cell and developmental biology that help to target molecules and pathways to restore the body's regenerative capacity, the "engineering" of missing tissue is quickly becoming a reality. Classically, tissue engineering has been thought to require external regenerative resources including a scaffold, cells, and growth factors. The allure of providing an exact replica of a missing bone that incorporates to become indistinguishable from self, has the capacity to heal and grow, is resistant to infection, and has minimal morbidity is a "holy grail" to all surgeons who work with bone. This article attempts to shed light on the use of stem cells for craniofacial reconstruction, including important principles learned from other scientific disciplines, relevant animal models for tissue engineering, early clinical reports from our experience and that of others, and future directions. PMID: 20853230 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Tissue engineered biological augmentation for tendon healing: a systematic review. Br Med Bull. 2010 Sep 17; Authors: Longo UG, Lamberti A, Maffulli N, Denaro V Tendon injuries give rise to significant morbidity. In the last few decades, several techniques have been increasingly used to optimize tendon healing. We performed a comprehensive search of PubMed, Medline, Cochrane, CINAHL and Embase databases using various combinations of the commercial names of each scaffold and the keywords 'tendon', 'rotator cuff', 'supraspinatus tendon', 'Achilles tendon', 'growth factors', 'cytokines', 'gene therapy', 'tissue engineering', 'mesenchymal' and 'stem cells' over the years 1966-2009. All articles relevant to the subject were retrieved, and their bibliographies were hand searched for further references in the context to tissue-engineered biological augmentation for tendon healing. Several new techniques are available for tissue-engineered biological augmentation for tendon healing, growth factors, gene therapy and mesenchimal stem cells. Data are lacking to allow definitive conclusions on the use of these techniques for routine management of tendon ailments. The emerging field of tissue engineering holds the promise to use new techniques for tendon augmentation and repair. Preliminary studies support the idea that these techniques can provide an alternative for tendon augmentation with great therapeutic potential. The optimization strategies discussed in this article are currently at an early stage of development. Although these emerging technologies may develop into substantial clinical treatment options, their full impact needs to be critically evaluated in a scientific fashion. PMID: 20851817 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Increased mechanosensitivity of cells cultured on nanotopographies. J Biomech. 2010 Sep 17; Authors: Salvi JD, Yul Lim J, Donahue HJ Enhancing cellular mechanosensitivity is recognized as a novel tool for successful musculoskeletal tissue engineering. We examined the hypothesis that mechanosensitivity of human mesenchymal stem cells (hMSCs) is enhanced on nanotopographic substrates relative to flat surfaces. hMSCs were cultured on polymer-demixed, randomly distributed nanoisland surfaces with varying island heights and changes in intracellular calcium concentration, [Ca(2+)](i), in response to fluid flow induced shear stress were quantifide. Stem cells cultured on specific scale nanotopographies displayed greater intracellular calcium responses to fluid flow. hMSCs cultured on 10-20nm high nanoislands displayed a greater percentage of cells responding in calcium relative to cells cultured on flat control, and showed greater average [Ca(2+)](i) increase relative to cells cultured on other nanoislands (45-80nm high nanoislands). As [Ca(2+)](i) is an important regulator of downstream signaling, as well as proliferation and differentiation of hMSCs, this observation suggests that specific scale nanotopographies provide an optimal milieu for promoting stem cell mechanotransduction activity. That mechanical signals and substrate nanotopography may synergistically regulate cell behavior is of significant interest in the development of regenerative medicine protocols. PMID: 20851397 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Development of an Acellular Bioengineered Matrix with a Dominant Vascular Pedicle. J Surg Res. 2010 Aug 6; Authors: Henderson PW, Nagineni VV, Harper A, Bavinck N, Sohn AM, Krijgh DD, Jimenez N, Weinstein AL, Spector JA BACKGROUND: This study assessed the feasibility of creating a tissue engineering platform by decellularization of fasciocutaneous tissue. MATERIALS AND METHODS: A fasciocutaneous flap based upon the superficial inferior epigastric artery was harvested from the abdominal wall of 8-wk-old male Sprague-Dawley rats. All cellular components were removed by sequential treatment with sodium azide, DNAse, and sodium deoxycholate. The degree of decellularization was qualitatively assessed by histology and quantitatively assessed by spectrophotometry. Persistence of relevant extracellular matrix proteins was shown following staining with orcein and hematoxylin. The duration of circuit patency was determined by continuous perfusion with a peristaltic perfusion pump. RESULTS: Gross and histologic examination demonstrated removal of cellular constituents with preservation of tissue matrix architecture, including macrochannels and microchannels. This was confirmed by the application of spectrophotometry to DNA isolates, which showed that the decellularized flap retained 4.04 ng/μL DNA, compared with the non-processed control, which retained 37.03 ng/μL DNA, and the acellular control, which was read as having 0.65 ng/μL DNA. The extracellular matrix of vessel walls was shown to remain intact. Peristaltic perfusion of the cannulated pedicle inflow channel with phosphate buffered saline at a rate of 200 μL/min confirmed circuit patency for 6 h. CONCLUSION: Fasciocutaneous flaps harvested with an intact vascular pedicle and associated tissue vascular network can be successfully decellularized and perfused ex vivo. This methodology, which is scalable to human size tissues, provides promise as a technique for the production of customizable engineered tissues. PMID: 20850792 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | A neurospheroid network-stamping method for neural transplantation to the brain. Biomaterials. 2010 Sep 15; Authors: Kato-Negishi M, Tsuda Y, Onoe H, Takeuchi S Neural transplantation therapy using neural stem cells has received as potential treatments for neurodegenerative diseases. Indeed, this therapy is thought to be effective for replacement of degenerating neurons in restricted anatomical region. However, because injected neural stem cells integrate randomly into the host neural network, another approach is needed to establish a neural pathway between selective areas of the brain or treat widespread degeneration across multiple brain regions. One of the promising approaches might be a therapy using pre-made neural network in vitro by the tissue engineering technique. In this study, we engineered a three-dimensional (3D) tissue with a neuronal network that can be easily manipulated and transplanted onto the host brain tissue in vivo. A polydimethylsiloxane microchamber array facilitated the formation of multiple neurospheroids, which in turn interconnected via neuronal processes to form a centimeter-sized neurospheroid network (NSN). The NSN was transferable onto the cortical surface of the brain without damage of the neuronal network. After transfer onto the cortical tissue, the NSN showed neural activity for more than 8 days. Moreover, neurons of the transplanted NSN extended their axons into the host cortical tissue and established synaptic connections with host neurons. Our findings suggest that this method could lay the foundation for treating severe degenerative brain disease. PMID: 20850180 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Long-term in vitro degradation of PDLLA/Bioglass® bone scaffolds in acellular simulated body fluid. Acta Biomater. 2010 Sep 14; Authors: Blaker JJ, Nazhat SN, Maquet V, Boccaccini AR The long-term (600 days) in vitro degradation of highly porous PDLLA/Bioglass®-filled composite foams developed for bone tissue engineering scaffolds has been investigated in simulated body fluid (SBF). Foams of circa 93% porosity were produced by thermally induced phase separation (TIPS). The degradation profile for foams of neat PDLLA and the influence of Bioglass® addition were comprehensively assessed in terms of changes in dimensional stability, pore morphology, weight loss, molecular weight and mechanical properties (dry and wet states). It is shown that the degradation process proceeded in several stages: a) the quasi-stable stage, where water absorption and plasticization occur together with weight loss due to Bioglass® particle loss and dissolution, resulting in decreased wet mechanical properties, b) a stage of slight increase in the wet mechanical properties and moderate decrease in dimensions, with properties remaining moderately constant until the onset of significant weight loss, whilst molecular weight continues to decrease, and c) an end-stage of massive weight loss, disruption of pore structure and the formation of blisters and embrittlement of the scaffold (evident on handling). The findings from this long-term in vitro degradation investigation underpin studies that have been and continue to be performed on highly porous bioactive glass filled poly(α-hydroxyesters) for bone tissue engineering applications. PMID: 20849987 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Through-thickness control of polymer bioresorption via electron beam irradiation. Acta Biomater. 2010 Sep 14; Authors: Cairns ML, Sykes A, Dickson GR, Orr JF, Farrar D, Dumba A, Buchanan FJ Predicable and controlled degradation is not only central to the accurate delivery of bioactive agents and drugs, it also plays a vital role in key aspects of bone tissue engineering. The work addressed in this paper investigates the utilization of e-beam irradiation in order to achieve a controlled (surface) degradation profile. This study focuses on the modification of commercially and clinically relevant materials, namely PLLA, PLLA-HA PLG, and PLDL. Samples were subjected to irradiation treatments using a 0.5 MeV electron beam with delivered surface doses of 150 kGy and 500 kGy. In addition, an acrylic attenuation shield was used for selected samples to control the penetration of the e-beam. E-beam irradiation induced chain scission in all polymers as characterized by reduced molecular weights and glass transition temperatures (Tg). Irradiation not only produced changes in the physical properties of the polymers but also had associated effects on the surface erosion of the materials during hydrolytic degradation. Moreover, the extent to which both mechanical and hydrolytic degradation was observed is synonymous with the estimated penetration of the beam (as controlled by the employment of an attenuation shield). PMID: 20849986 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Biomimetic composite coating on rapid prototyped scaffolds for bone tissue engineering. Acta Biomater. 2010 Sep 14; Authors: Arafat MT, Lam CX, Ekaputra AK, Wong SY, Li X, Gibson I The objective of this present study is to improve functional performance of rapid prototyped scaffolds for bone tissue engineering through biomimetic composite coating. Rapid prototyped poly(ε-caprolactone)/tri-calcium phosphate (PCL/TCP) scaffolds were fabricated using screw extrusion system (SES). The fabricated PCL/TCP scaffolds were coated with carbonated hydroxyapatite (CHA)-gelatin composite via biomimetic co-precipitation. The structure of the prepared CHA-gelatin composite coating was studied by scanning electron microscopy (SEM), X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy. Compressive mechanical testing revealed that coating process did not have any detrimental effect on the mechanical property of the scaffolds. The cell-scaffold interaction was studied by culturing porcine bone marrow stromal cells (BMSCs) on the scaffolds, and assessing proliferation, bone related gene and protein expression capabilities of BMSCs. Confocal laser microscopy and SEM images of the cell-scaffold constructs showed uniform distribution of cell-sheet and accumulation of extracellular matrix at the interior of CHA-gelatin composite coated PCL/TCP scaffolds. The proliferation rate of BMSCs on CHA-gelatin composite coated PCL/TCP scaffolds was about 2.3 times and 1.7 times higher than that on the PCL/TCP scaffolds and CHA coated PCL/TCP scaffolds, respectively, by day 10. Furthermore, reverse transcription polymerase chain reaction and western blot analysis revealed that CHA-gelatin composite coated PCL/TCP scaffolds stimulate osteogenic differentiation of BMSCs the most, compared to PCL/TCP scaffolds and CHA coated PCL/TCP scaffolds. These results demonstrate that CHA-gelatin composite coated rapid prototyped PCL/TCP scaffolds are promising for bone tissue engineering. PMID: 20849985 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Aligned natural-synthetic polyblend nanofibers for peripheral nerve regeneration. Acta Biomater. 2010 Sep 14; Authors: Wang CY, Zhang KH, Fan CY, Mo XM, Ruan HJ, Li FF Peripheral nerve regeneration remains a significant clinical challenge to researchers. Progress in the design of tissue engineering scaffolds provides an alternative approach for neural regeneration. In this study, aligned silk fibroin (SF) blended poly(L-lactic acid-co-ε-caprolactone) [P(LLA-CL)] nanofibrous scaffolds were fabricated via electrospinning methods, and then reeled into aligned nerve guidance conduit (NGC) for promoting nerve regeneration. The aligned SF/P(LLA-CL) NGC was used as a bridge implanting across a 10 mm defect of sciatic nerve in rats, and the outcome of regenerated nerve at 4 and 8 weeks was evaluated by a combination of electrophysiological assessment, histological and immunohistological analysis, as well as electron microscopy. The electrophysiological examination showed that the functional recovery of regenerated nerve in the SF/P(LLA-CL) NGC group was superior to that in the P(LLA-CL) NGC group. The morphological analysis also indicated that the regenerated nerve in SF/P(LLA-CL) NGC was more mature. All the results demonstrated that the aligned SF/P(LLA-CL) NGC could greatly promote peripheral nerve regeneration in comparison with the aligned P(LLA-CL) NGC, thus raising a potential application in nerve regeneration. PMID: 20849984 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Comparative study of the chondrogenic potential of human bone marrow stromal cells, neonatal chondrocytes and adult chondrocytes. Biochem Biophys Res Commun. 2010 Sep 14; Authors: Saha S, Kirkham J, Wood D, Curran S, Yang X Cartilage tissue engineering is still a major clinical challenge with optimization of a suitable source of cells for cartilage repair/regeneration not yet fully addressed. The aims of this study were to compare and contrast the differences in chondrogenic behaviour between human bone marrow stromal cells (HBMSCs), human neonatal and adult chondrocytes to further our understanding of chondroinduction relative to cell maturity and to identify factors that promote chondrogenesis and maintain functional homeostasis. Cells were cultured in monolayer in either chondrogenic or basal medium, recapitulating procedures used in existing clinical procedures for cell based therapies. Cell doubling time, morphology and alkaline phosphatase specific activity (ALPSA) were determined at different time points. Expression of chondrogenic markers (SOX9, ACAN and COL2A1) was compared via real time polymerase chain reaction. Amongst the three cell types studied, HBMSCs had the highest ALPSA in basal culture and lowest ALPSA in chondrogenic media. Neonatal chondrocytes were the most proliferative and adult chondrocytes had the lowest ALPSA in basal media. Gene expression analysis revealed a difference in the temporal expression of chondrogenic markers which were up regulated in chondrogenic medium compared to levels in basal medium. Of the three cell types studied, adult chondrocytes offer a more promising cell source for cartilage tissue engineering. This comparative study revealed differences between the microenvironment of all three cell types and provides useful information to inform cell-based therapies for cartilage regeneration. PMID: 20849819 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Isolation of cell-free bacterial inclusion bodies. Microb Cell Fact. 2010 Sep 17;9(1):71 Authors: Rodriguez-Carmona E, Cano-Garrido O, Seras-Franzoso J, Villaverde A, Garcia-Fruitos E ABSTRACT: BACKGROUND: Bacterial inclusion bodies are submicron protein clusters usually found in recombinant bacteria that have been traditionally considered as undesirable products from protein production processes. However, being fully biocompatible, they have been recently characterized as nanoparticulate inert materials useful as scaffolds for tissue engineering, with potentially wider applicability in biomedicine and material sciences. Current protocols for inclusion body isolation from Escherichia coli usually offer between 95 to 99 % of protein recovery, what in practical terms, might imply extensive bacterial cell contamination, not compatible with the use of inclusion bodies in biological interfaces. RESULTS: Using an appropriate combination of chemical and mechanical cell disruption methods we have established a convenient procedure for the recovery of bacterial inclusion bodies with undetectable levels of viable cell contamination, below 10-1 cfu/ml, keeping the particulate organization of these aggregates regarding size and protein folding features. CONCLUSIONS: The application of the developed protocol allows obtaining bacterial free inclusion bodies suitable for use in mammalian cell cultures and other biological interfaces. PMID: 20849629 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Gelatin-Based Laser Direct-Write Technique for the Precise Spatial Patterning of Cells. Tissue Eng Part C Methods. 2010 Sep 20; Authors: Schiele NR, Chrisey DB, Corr DT Laser direct-writing provides a method to pattern living cells in vitro, to study various cell-cell interactions, and build cellular constructs. However, the materials typically used may limit its long-term application. By utilizing gelatin coatings on the print ribbon and growth surface, we developed a new approach for laser cell printing that overcomes the limitations of Matrigel™. Gelatin is free of growth factors and extraneous matrix components that may interfere with cellular processes under investigation. Gelatin-based laser direct-write was able to successfully pattern human dermal fibroblasts with high post-transfer viability (91% ± 3%) and no observed double strand DNA damage. As seen with atomic force microscopy, gelatin offers a unique benefit in that it is present temporarily to allow cell transfer, but melts and is removed with incubation to reveal the desired application-specific growth surface. This provides unobstructed cellular growth after printing. Monitoring cell location following transfer, we show that melting and removal of gelatin does not affect cellular placement; cells maintained registry within 5.6 μm ± 2.5 μm to the initial pattern. This study demonstrates the effectiveness of gelatin in laser direct-writing to create spatially-precise cell patterns with the potential for applications in tissue engineering, stem cell, and cancer research. PMID: 20849381 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Polyhydroxybutyrate and its Copolymer with Polyhydroxyvalerate as Biomaterials: Influence on Progression of Stem Cell Cycle. Biomacromolecules. 2010 Sep 17; Authors: Ahmed T, Marçal H, Lawless M, Wanandy NS, Chiu A, Foster LJ Poly(3-hydroxybutyrate) (PHB) and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) are biopolyesters reported to provide favorable microenvironments for cell culture and possess potential for tissue engineering applications. Both biopolymers have been investigated for applications in a variety of medical scenarios, including nerve and bone repair. This study investigated the influence these biomaterials exerted on cell cycle progression of olfactory ensheathing cells (OECs) and mesenchymal stem cells (MSCs) commonly used in the engineering of nerve and bone tissues. Cell cycle regulation is important for cell survival; analysis revealed that the biomaterials induced significant cell cycle progression in both MSCs and OECs. Significantly higher percentages of cells were cycled at synthesis (S) phase of the cycle on PHBV films compared to PHB, with MSCs more susceptible than OECs. Furthermore, detection of early stages of apoptotic activation showed significant differences in the two cell populations exhibiting necrosis and apoptosis when cultivated on the biomaterials. OECs compromised on PHB (5.6%) and PHBV (2.5%) compared to MSCs with 12.6% on PHB and 17% on PHBV. Significant differences in crystallinity and surface rugosity were determined between films of the two biomaterials, 88% and 1.12 μm for PHB and 76% and 0.72 μm for PHBV. While changes in surface properties may have influenced cell adhesion, the work presented here suggests that application of these biomaterials in tissue engineering are specific to cell type and requires a detailed investigation at the cell-material interface. PMID: 20849100 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Bioprinting vessel-like constructs using hyaluronan hydrogels crosslinked with tetrahedral polyethylene glycol tetracrylates. Biomaterials. 2010 Aug;31(24):6173-81 Authors: Skardal A, Zhang J, Prestwich GD Bioprinting enables deposition of cells and biomaterials into spatial orientations and complexities that mirror physiologically relevant geometries. To facilitate the development of bioartificial vessel-like grafts, two four-armed polyethylene glycol (PEG) derivatives with different PEG chain lengths, TetraPEG8 and TetraPEG13, were synthesized from tetrahedral pentaerythritol derivatives. The TetraPEGs are unique multi-armed PEGs with a compact and symmetrical core. The TetraPEGs were converted to tetra-acrylate derivatives (TetraPAcs) which were used in turn to co-crosslink thiolated hyaluronic acid and gelatin derivatives into extrudable hydrogels for printing tissue constructs. First, the hydrogels produced by TetraPAc crosslinking showed significantly higher shear storage moduli when compared to PEG diacrylate (PEGDA)-crosslinked synthetic extracellular matrices (sECMs) of similar composition. These stiffer hydrogels have rheological properties more suited to bioprinting high-density cell suspensions. Second, TetraPAc-crosslinked sECMs were equivalent or superior to PEGDA-crosslinked gels in supporting cell growth and proliferation. Third, the TetraPac sECMs were employed in a proof-of-concept experiment by encapsulation of NIH 3T3 cells in sausage-like hydrogel macrofilaments. These macrofilaments were then printed into tubular tissue constructs by layer-by-layer deposition using the Fab@Home printing system. LIVE/DEAD viability/cytotoxicity-stained cross-sectional images showed the bioprinted cell structures to be viable in culture for up to 4 weeks with little evidence of cell death. Thus, biofabrication of cell suspensions in TetraPAc sECMs demonstrates the feasibility of building bioartificial blood vessel-like constructs for research and potentially clinical uses. PMID: 20546891 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Isolation, differentiation and characterization of vascular cells derived from human embryonic stem cells. Nat Protoc. 2010;5(6):1115-26 Authors: Levenberg S, Ferreira LS, Chen-Konak L, Kraehenbuehl TP, Langer R Herein, we describe a protocol for the isolation of human embryonic stem cells (hESCs)-derived vascular cells at various stages of development. The cells are isolated from 10 to 15-d-old human embryoid bodies (EBs) cultured in suspension. After dissociation, cells are labeled with anti-CD34 or anti-CD31 (PECAM1) antibody and separated from the cell mixture by magnetic-activated cell separation (MACS) or fluorescent-activated cell sorting (FACS). Isolated vascular cells are then cultured in media conditions that support specific differentiation and expansion pathways. The resulting vascular cell populations contain >80% endothelial-like or smooth muscle-like cells. Assuming typical initial cell adhesion and proliferation rates, the entire procedure can be completed within 1.5 months. Vascular cells isolated and differentiated under the described conditions may constitute a potential cell source for therapeutic application toward repair of ischemic tissues, preparation of tissue-engineered vascular grafts and design of cellular kits for drug screening applications. PMID: 20539287 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | Effect of mechanical factors on the function of engineered human blood microvessels in microfluidic collagen gels. Biomaterials. 2010 Aug;31(24):6182-9 Authors: Price GM, Wong KH, Truslow JG, Leung AD, Acharya C, Tien J This work examines how mechanical signals affect the barrier function and stability of engineered human microvessels in microfluidic type I collagen gels. Constructs that were exposed to chronic low flow displayed high permeabilities to bovine serum albumin and 10 kDa dextran, numerous focal leaks, low size selectivity, and short lifespan of less than one week. Higher flows promoted barrier function and increased longevity; at the highest flows, the barrier function rivaled that observed in vivo, and all vessels survived to day 14. By studying the physiology of microvessels of different geometries, we established that shear stress and transmural pressure were the dominant mechanical signals that regulated barrier function and vascular stability, respectively. In microvessels that were exposed to high flow, elevation of intracellular cyclic AMP further increased the selectivity of the barrier and strongly suppressed cell proliferation. Computational models that incorporated stress dependence successfully predicted vascular phenotype. Our results indicate that the mechanical microenvironment plays a major role in the functionality and stability of engineered human microvessels in microfluidic collagen gels. PMID: 20537705 [PubMed - indexed for MEDLINE] | | | | | | | | | | | | | | | | | | | | | | | | | A strategy for fabrication of a three-dimensional tissue construct containing uniformly distributed embryoid body-derived cells as a cardiac patch. Biomaterials. 2010 Aug;31(24):6218-27 Authors: Huang CC, Liao CK, Yang MJ, Chen CH, Hwang SM, Hung YW, Chang Y, Sung HW Growing three-dimensional (3D) scaffolds that contain more than a few layers of seeded cells in vitro is crucial for the creation of thick and viable cardiac tissues in vivo. Embryonic stem cells (ESCs) have been used as an alternative cell source for cardiac repair; however, dissociated ESCs show poor viability in the scaffold and do not form the embryoid body (EB)-like structures. In this study, a strategy intended for cultivating EB-derived cells (EBDCs) uniformly in a porous 3D tissue scaffold was developed. This strategy employed techniques of formation of spherically symmetric EBs in a thermo-responsive hydrogel system, production of cell sheets of EBDCs in a similar hydrogel system coated with collagen and fabrication of sliced porous tissue scaffolds. The prepared EBs were collected and plated evenly in the cell-sheet culture system. After 8 days in culture, a continuous sheet of EBDCs with cell beating was obtained; our qPCR and flow cytometric analyses showed that the collagen-coated on the cell-sheet culture system can significantly enhance the population of cardiac-lineage cells. The produced EBDC sheets were then sandwiched into the sliced porous tissue scaffold. After reculture, the seeded EBDCs were redistributed uniformly throughout the scaffold, with a significant increase in mechanical strength. Cardiac-specific myosin heavy chain and alpha-actinin were expressed for some cells grown in the scaffold, while connexin 43 was clearly expressed at the cell borders. Additional studies such as employing purification techniques to enrich the population of cardiomyocytes are needed to further improve the developed tissue constructs as a bioengineered cardiac patch. PMID: 20537702 [PubMed - indexed for MEDLINE] | | | | | | | | | |  | |  | |  |
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