| Isolation and characterization of umbilical cord blood hematopoietic stem cells.
August 26, 2009 at 10:13 am
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| | Isolation and characterization of umbilical cord blood hematopoietic stem cells. J Med Assoc Thai. 2009 Jun;92 Suppl 3:S88-94 Authors: Chularojmontri L, Wattanapitayakul SK Umbilical cord blood (UCB) has recently represented another rich source for hematopoietic stem cells (HSCs). Recent clinical studies have shown that UCB stem cells can potentially be used in place of HSCs from bone marrow as well as in basic research in regenerative medicine. This article will describe the methods for isolation and characterization of HSCs from UCB. UCB were obtained from umbilical vessels at the time of delivery. The HSCs were isolated from UCB using a density-gradient centrifugation method, CD34-immunomagnetic separation, and finally fluorescent-activated cell sorting (FACS). Functional assays were evaluated for the ability of multipotent progenitors to differentiate to lineage-specific committed cells and heterogeneous hierarchy of pluripotent cells. Approximately 1% of CD34+ cells were isolated and sorted from mononucleated cells. Functional assays revealed that the CD34+ subpopulation gave rise to several hematopoietic cell lineages including CFU-E, BFU-E, CFU-G, CFU-M, CFU-GM, and CFU-GEMM. These cells also maintained their stemness as evaluated by primitive long-term culture initiating cells (LTC-IC). The basic methods in HSC isolation and characterization employing gradient isolation, CD34-immunomagnetic separation, FACS, and functional assays are important in the area of stem cell investigation and applications. PMID: 19705548 [PubMed - in process] |
| Collagen/silk fibroin bi-template-induced biomimetic bone-like substitutes.
August 26, 2009 at 10:13 am
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| | Collagen/silk fibroin bi-template-induced biomimetic bone-like substitutes. J Biomed Mater Res A. 2009 Aug 24; Authors: Wang J, Zhou W, Hu W, Zhou L, Wang S, Zhang S A novel bi-template-induced co-assembly method was used to fabricate biomimetic bone substitute materials, collagen-fibroin/hydroxyapatite (COL-SF/HA) composite by using a combination of Type I collagen (COL) and silk fibroin (SF) molecular templates. As a control, collagen/hydroxyapatite (COL/HA) and silk fibroin/hydroxyapatite (SF/HA) composites were also synthesized via single-template assembly technology. The structure and morphology of the resulting assembly composites were investigated by XRD, FTIR, TEM, and TGA. Their sizes and size distributions were measured by dynamic light scattering (DLS). The results indicated that the mineral phases in COL-SF/HA, COL-HA, and SF-HA composites were needle-like nano-hydroxyapatite crystals. In comparison to those in COL-HA and SF-HA, the mineral phase in COL-SF/HA displayed smaller size and more narrow distribution. Of all the above biomimetic composites, the hydroxyapatite was well assembled with molecular template(s), and the organic content of the composite was about 12-20%, which was quite similar to the natural bone in composition. Circular dichroism (CD) and SDS-PAGE were used to examine the secondary structure and subunit composition of template proteins. The results revealed that the spatial structure of co-assembly template proteins played a pivotal role in controlling and regulating hydroxyapatite crystal nucleation and growth. On the basis of the experimental results aforementioned, a possible co-assembly mechanism for the hydroxyapatite growing on fibrous bi-template proteins was suggested. (c) 2009 Wiley Periodicals, Inc. J Biomed Mater Res, 2009. PMID: 19705470 [PubMed - as supplied by publisher] |
| Hydro-spinning: A novel technology for making alginate/chitosan fibrous scaffold.
August 26, 2009 at 10:13 am
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| | Hydro-spinning: A novel technology for making alginate/chitosan fibrous scaffold. J Biomed Mater Res A. 2009 Aug 24; Authors: Wang JZ, Huang XB, Xiao J, Yu WT, Wang W, Xie WY, Zhang Y, Ma XJ Alginate/chitosan polyelectrolyte complex (PEC) hybrid fibers are promising materials for scaffold-making in tissue engineering. In this study, a new method termed "hydro-spinning" was developed to make alginate/chitosan hybrid fibers. In hydro-spinning, a chitosan solution was pumped into a flowing sodium alginate solution and sheared into streamlines. These elongated streamlines subsequently transformed into alginate/chitosan PEC ribbon-like fibers before breaking up into pieces. Average diameter and chitosan content of the fibers correlated positively with the chitosan concentration used in spinning. These hybrid fibers showed a high water-absorbability of around 50-fold to 60-fold of water to their dry weight and could retain their integrity after saturation in minimum essential medium (MEM) medium for 30 days. In vitro culture experiments demonstrated that these fibers were able to support the three-dimensional growth of MCF-7, suggesting the potential applications of these fibers in biomedical and bioengineering fields such as tissue engineering. (c) 2009 Wiley Periodicals, Inc. J Biomed Mater Res, 2009. PMID: 19705468 [PubMed - as supplied by publisher] |
| Fabrication and characterization of nano composite scaffold of poly(L: -lactic acid)/hydroxyapatite.
August 26, 2009 at 10:13 am
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| | Fabrication and characterization of nano composite scaffold of poly(L: -lactic acid)/hydroxyapatite. J Mater Sci Mater Med. 2009 Aug 25; Authors: Wang X, Song G, Lou T To mimic the nano-fibrous structure of the natural extracellular matrix, a nano composite scaffold of poly(L: -lactic acid)/hydroxyapatite(PLLA/HAP) was fabricated by a thermally induced phase separation method. The characterization of the composite scaffold showed that the scaffold had a nano-fibrous PLLA network (fiber size 100-750 nm), an interconnective microporous structure (1-10 mum) and high porosity (>90%). HAP was homogeneously distributed in the scaffold, as a result, the compressive modulus of PLLA/HAP (80:20, w/w) increased to 3.15-fold compared with that of a pure PLLA scaffold. Incorporating HAP into PLLA network also buffered the pH decline in vitro degradation and enhanced the protein adsorption of the composite scaffold significantly. The new nano composite scaffold is potentially a very promising scaffold for tissue engineering. PMID: 19705258 [PubMed - as supplied by publisher] |
| Endless Possibilities: Stem Cells and the Vision for Toxicology Testing in the 21st Century.
August 26, 2009 at 10:13 am
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| | Endless Possibilities: Stem Cells and the Vision for Toxicology Testing in the 21st Century. Toxicol Sci. 2009 Aug 24; Authors: Chapin RE, Stedman DB The National Research Council's toxicity testing vision lays out a bold future for our field. It depends heavily on computational algorithms based on the latest knowledge of cellular biochemistry and protein interaction pathways, exposing human cells to novel compounds in vitro, and being able to understand the changes seen. At the same time, significant strides are being made in our understanding of the control, production, and "behaviour" of stem cells. While stem cells offer seemingly limitless possibilities for regenerative medicine, they have already delivered new assays to predict embryofetal developmental toxicity in vitro. In addition to providing a model of cells undergoing differentiation and proliferation, stem cells will play a major role by giving rise to many of the differentiated cell types on which this new vision depends. These will not be pure populations of single cell types, but mixtures of cells much more representative of tissues in vitro. Moving from cells alone in a culture dish towards the more physiologic condition of multiple cell types being able to interact to maintain homeostasis in the face of a disequilibrating force (like a toxic exposure) will lead us towards more useful and correct predictions of in vivo toxicities. Despite the seemingly insurmountable hurdles, persistence and creativity are on our side. We expect that a long series of successive iterations of predictive models will eventually yield a working process that approximates the NRC's vision and delivers on the promise of faster evaluation of chemicals with reduced animal use. PMID: 19703945 [PubMed - as supplied by publisher] |
| Functional characterization of cardiomyocytes derived from murine induced pluripotent stem cells in vitro.
August 26, 2009 at 10:13 am
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| | Functional characterization of cardiomyocytes derived from murine induced pluripotent stem cells in vitro. FASEB J. 2009 Aug 24; Authors: Kuzmenkin A, Liang H, Xu G, Pfannkuche K, Eichhorn H, Fatima A, Luo H, Saric T, Wernig M, Jaenisch R, Hescheler J Several types of terminally differentiated somatic cells can be reprogrammed into a pluripotent state by ectopic expression of Klf4, Oct3/4, Sox2, and c-Myc. Such induced pluripotent stem (iPS) cells have great potential to serve as an autologous source of cells for tissue repair. In the process of developing iPS-cell-based therapies, the major goal is to determine whether differentiated cells derived from iPS cells, such as cardiomyocytes (CMs), have the same functional properties as their physiological in vivo counterparts. Therefore, we differentiated murine iPS cells to CMs in vitro and characterized them by RT-PCR, immunocytochemistry, and electrophysiology. As key markers of cardiac lineages, transcripts for Nkx2.5, alphaMHC, Mlc2v, and cTnT could be identified. Immunocytochemical stainings revealed the presence of organized sarcomeric actinin but the absence of mature atrial natriuretic factor. We examined characteristics and developmental changes of action potentials, as well as functional hormonal regulation and sensitivity to channel blockers. In addition, we determined expression patterns and functionality of cardiac-specific voltage-gated Na(+), Ca(2+), and K(+) channels at early and late differentiation stages and compared them with CMs derived from murine embryonic stem cells (ESCs) as well as with fetal CMs. We conclude that iPS cells give rise to functional CMs in vitro, with established hormonal regulation pathways and functionally expressed cardiac ion channels; CMs generated from iPS cells have a ventricular phenotype; and cardiac development of iPS cells is delayed compared with maturation of native fetal CMs and of ESC-derived CMs. This difference may reflect the incomplete reprogramming of iPS cells and should be critically considered in further studies to clarify the suitability of the iPS model for regenerative medicine of heart disorders.-Kuzmenkin, A., Liang, H., Xu, G., Pfannkuche, K., Eichhorn, H., Fatima, A., Luo, H., Sarić, T., Wernig, M., Jaenisch, R., Hescheler, J. Functional characterization of cardiomyocytes derived from murine induced pluripotent stem cells in vitro. PMID: 19703934 [PubMed - as supplied by publisher] |
| Fabrication and characterization of a recombinant fibronectin/cadherin bio-inspired ceramic surface and its influence on adhesion and ossification in vitro.
August 26, 2009 at 10:13 am
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| | Fabrication and characterization of a recombinant fibronectin/cadherin bio-inspired ceramic surface and its influence on adhesion and ossification in vitro. Acta Biomater. 2009 Aug 21; Authors: Zhang Y, Xiang Q, Dong S, Li C, Zhou Y This study investigates the effects of a bio-inspired ceramic surface modified with a novel recombinant protein on surface parameters and cell behavior. The surface of a biphasic calcium phosphate (BCP) ceramic was functionalized with a recombinant protein spanning the fragments of fibronectin module III7-10 and extracellular domains 1-2 of cadherin 11 (rFN/CDH) using a dimethyl 3, 3'-dithiobispropionimidate (DTBP)-crosslinking method. The surface was characterized by SEM, XPS and protein adsorption and surface density measurements. The material exhibited desirable properties for cell adhesion and proliferation. The effects of the surface on the adhesion and proliferation of human mesenchymal stem cells (hMSC) were investigated using a cell adhesion centrifugal assay and MTT methods. The data demonstrated that the adhesive capacity and proliferation rate were significantly improved as compared with FN and CDH positive controls. Moreover, the rFN/CDH bio-inspired ceramic surface also induced osteoblastic differentiation, as evidenced by the higher alkaline phosphatase activity and osteocalcin mRNA expression level of hMSC cultured in osteogenic media for 7-10 days. Furthermore, a functional blocking assay with a site-specific antibody against phosphotyrosine 397 (pY397) of focal adhesion kinase revealed that pY397 is involved in adhesion and ossification. These results suggest that the rFN/CDH bio-inspired BCP surface possesses enhanced functionality in adhesion, proliferation and ossification and may be a promising scaffold for tissue engineering. PMID: 19703596 [PubMed - as supplied by publisher] |
| Preparation and characterization of polysaccharidic microbeads by microfluidic technique: application to the encapsulation of Sertoli cells.
August 26, 2009 at 10:13 am
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| | Preparation and characterization of polysaccharidic microbeads by microfluidic technique: application to the encapsulation of Sertoli cells. Acta Biomater. 2009 Aug 21; Authors: Capretto L, Mazzitelli S, Luca G, Nastruzzi C Polysaccharides (e.g. alginate or agarose) represent a class of polymers commonly employed for the preparation of microparticles for cell entrapment and tissue engineering applications. The present work describes the production and characterization of microbeads, by a microfluidic approach, constituted of alginate and alginate/agarose blends, for the encapsulation of eukaryotic cells. The general production strategy is based on the formation of water in oil multiphase flow by a "Y" junction squeezing mechanism. The presented data demonstrate that the gelation step represents the crucial point for the production of morphologically excellent microbeads. In this respect, microfluidic methods appear to be an effective procedure for the production of microbeads intended for cell encapsulation as proved by the high viability and maintenance of functional capability demonstrated by the encapsulated Sertoli cells. PMID: 19703594 [PubMed - as supplied by publisher] |
| Anterior-posterior patterning of neural differentiated embryonic stem cells by canonical Wnts, Fgfs, Bmp4 and their respective antagonists.
August 26, 2009 at 10:13 am
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| | Anterior-posterior patterning of neural differentiated embryonic stem cells by canonical Wnts, Fgfs, Bmp4 and their respective antagonists. Dev Growth Differ. 2009 Aug 23; Authors: Hendrickx M, Van XH, Leyns L Embryonic stem (ES) cells are pluripotent and can differentiate into every cell type of the body. Next to their potential in regenerative medicine, they are excellent tools to study embryonic development. In this work the processes of neural induction and neural patterning along the antero-posterior (A/P) body axis are studied and evidence suggests a two step mechanism for these events. First, neural induction occurs by default in the primitive ectoderm, forming anterior neural tissue and thereafter, a series of factors can posteriorize this anterior neurectoderm. In a gain-of-function/loss-of-function approach using mouse ES cells, we show that Fgf2 has the strongest caudalizing potential of all Fgfs tested. Furthermore, Bmp4 and Wnt3a, but not Wnt1, can caudalize the neurectodermal cells. The effect of the antagonists of these factors was also examined and though Dkk1 and Noggin clearly have an effect that opposes that of Wnt3a and Bmp4 respectively, they fail to anteriorize the neurectoderm. The patterning effect of SU5402, an Fgf receptor inhibitor, was rather limited. These data confirm that in the mouse, two steps are involved in neural patterning and we show that while Fgf4, Fgf8 and Wnt1 have no strong patterning effect, Fgf2, Wnt3a and Bmp4 are strong posteriorizing factors. PMID: 19703209 [PubMed - as supplied by publisher] |
| Regulatory roles of beta-catenin and AP-1 on osteoprotegerin production in interleukin-1alpha-stimulated periodontal ligament cells.
August 26, 2009 at 10:13 am
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| | Regulatory roles of beta-catenin and AP-1 on osteoprotegerin production in interleukin-1alpha-stimulated periodontal ligament cells. Oral Microbiol Immunol. 2009 Oct;24(5):384-9 Authors: Suda T, Nagasawa T, Wara-Aswapati N, Kobayashi H, Iwasaki K, Yashiro R, Hormdee D, Nitta H, Ishikawa I, Izumi Y BACKGROUND: Periodontitis is a chronic inflammatory disease characterized by the enhanced expression of inflammatory mediators leading to alveolar bone resorption. Osteoprotegerin (OPG) plays a suppressive role in cytokine-induced osteoclastogenesis. In osteoblasts, OPG expression is upregulated by beta-catenin but downregulated by the transcription factor activator protein-1 (AP-1; c-fos/c-jun). The purpose of this study was to examine the roles of beta-catenin and AP-1 in interleukin-1alpha (IL-1alpha) -induced OPG production in human gingival fibroblasts (hGFs) and periodontal ligament (PDL) cells. METHODS: Expression of c-fos and c-jun messenger RNA was measured by reverse transcription-polymerase chain reaction and OPG production was analysed by enzyme-linked immunosorbent assay. The nuclear AP-1 activity was quantified using an AP-1 microplate assay. The effect of the Wnt canonical pathway on OPG production was evaluated using small interfering (si) RNA for beta-catenin and the effect of AP-1 on OPG production was evaluated using the AP-1 inhibitor curcumin. RESULTS: Levels of c-fos messenger RNA and nuclear AP-1 activity were higher in PDL cells than in hGFs. When stimulated with IL-1alpha, PDL cells had significantly higher c-fos expression and lower OPG production compared with hGFs. The siRNA for beta-catenin suppressed the IL-1alpha-induced OPG production in both PDL cells and hGFs, whereas the AP-1 inhibitor curcumin augmented the IL-1alpha-induced OPG production in PDL cells, but not in hGFs. CONCLUSION: The present study suggests that beta-catenin enhances IL-1alpha-induced OPG production in both PDL cells and hGFs, whereas AP-1 suppresses IL-1alpha-induced OPG production in PDL cells. Higher expression of c-fos in PDL cells than in hGFs may implicate a role of PDL cells in alveolar bone resorption in periodontitis. PMID: 19702951 [PubMed - in process] |
| PREPARATION OF CARDIAC EXTRACELLULAR MATRIX FROM AN INTACT PORCINE HEART.
August 26, 2009 at 10:13 am
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| | PREPARATION OF CARDIAC EXTRACELLULAR MATRIX FROM AN INTACT PORCINE HEART. Tissue Eng Part C Methods. 2009 Aug 25; Authors: Wainwright JM, Czajka CA, Patel UB, Freytes DO, Tobita K, Gilbert TW, Badylak SF Whole organ engineering would benefit from a 3-dimensional scaffold produced from intact organ specific extracellular matrix (ECM). The microenvironment and architecture provided by such a scaffold would likely support site appropriate cell differentiation and spatial organization. The methods to produce such scaffolds from intact organs require customized decellularization protocols. In the present study, intact adult porcine hearts were successfully decellularized in less than 10 hours by using pulsatile retrograde aortic perfusion. Serial perfusion of an enzymatic, non-ionic detergent, ionic detergent, and acid solution with hypotonic and hypertonic rinses were used to systematically remove cellular content. The resultant cardiac ECM (C-ECM) retained collagen, elastin, and glycosaminoglycans, and mechanical integrity. C-ECM supported the formation of organized chicken cardiomyocyte sarcomere structure in vitro. The intact decellularized porcine heart provides a tissue engineering template that may be beneficial for future pre-clinical studies and eventual clinical applications. PMID: 19702513 [PubMed - as supplied by publisher] |
| GELLAN GUM INJECTABLE HYDROGELS FOR CARTILAGE TISSUE ENGINEERING APPLICATIONS: IN VITRO STUDIES AND PRELIMINARY IN VIVO EVALUATION.
August 26, 2009 at 10:13 am
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| | GELLAN GUM INJECTABLE HYDROGELS FOR CARTILAGE TISSUE ENGINEERING APPLICATIONS: IN VITRO STUDIES AND PRELIMINARY IN VIVO EVALUATION. Tissue Eng Part A. 2009 Aug 25; Authors: Oliveira JT, Santos TC, Martins L, Picciochi R, Marques A, Castro AG, Neves NM, Mano J, Reis RL Gellan gum is a polysaccharide that we have previously proposed for applications in the cartilage tissue engineering field. In this work, gellan gum hydrogels were tested for their ability to be used as injectable systems able to deliver and maintain chondrocytes by in situ gelation, and support cell viability and production of extracellular matrix. Rheological measurements determined that the sol-gel transition occurred near the body temperature at 39şC, upon temperature decrease, in approximately 20 seconds. Gellan gum discs showing a storage compression modulus of around 80 kPa at a frequency of 1 Hz by dynamic mechanical analysis (DMA). Human articular chondrocytes were encapsulated in the gels, cultured in vitro for total periods of 56 days, and analysed for cells viability and extracellular matrix (ECM) production. Calcein AM staining showed that cell kept viable after 14 days and the histological analysis and real-time quantitative PCR revealed that hyaline-like cartilage ECM was synthesised. Finally, the in vivo performance of the gellan gum hydrogels, in terms of induced inflammatory reaction and integration into the host tissue, was evaluated by subcutaneous implantation in Balb/c mice for 21 days. Histological analysis showed a residual fibrotic capsule at the end of the experiments. DMA revealed the gels were stable throughout the experiments while evidencing a tendency for decreasing mechanical properties, which was consistent with weight measurements. Altogether, the results demonstrate the adequacy of gellan gum hydrogels for non-invasive injectable applications towards the formation of a functional cartilage tissue engineered construct. PMID: 19702512 [PubMed - as supplied by publisher] |
| Mesenchymal progenitor Cells Derived from Synovium and Infrapatellar Fat Pad as a Source for Superficial Zone Cartilage Tissue Engineering: Analysis of Superficial Zone Protein (SZP)/Lubricin Expression.
August 26, 2009 at 10:13 am
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| | Mesenchymal progenitor Cells Derived from Synovium and Infrapatellar Fat Pad as a Source for Superficial Zone Cartilage Tissue Engineering: Analysis of Superficial Zone Protein (SZP)/Lubricin Expression. Tissue Eng Part A. 2009 Aug 25; Authors: Lee SY, Nakagawa T, Reddi AH Superficial zone protein (SZP) is a boundary lubricant of articular cartilage in joints. As SZP at the surface of articular cartilage plays an important role in the normal function of synovial joints, the localization of SZP-secreting cells at the surface of tissue-engineered cartilage is prerequisite. The aim of this study is the identification of suitable progenitor cell sources for tissue engineering of superficial zone cartilage. We investigated whether mesenchymal progenitor cells (MPCs) from synovium and infrapatellar fat pad (IFP) have the potential for secretion of SZP following chondrogenic differentiation in an aggregate pellet culture system. SZP was immunolocalized in pellets from synovium-MPCs and IFP-MPCs. The ELISA analysis of SZP demonstrated that chondrogenically differentiated synovium-MPC and IFP-MPC pellets secreted SZP into media. Real-time PCR analysis showed significant up-regulation of SZP mRNA in synovium-MPC and IFP-MPC pellets after chondrogenic differentiation. The synovium-MPCs demonstrated the higher colony-forming, proliferative, and chondrogenic potential, and exhibited greater SZP-secretion following chondrogenic induction compared to IFP-MPCs. In conclusion, both synovium and IFP are promising cell sources for tissue engineering of superficial zone cartilage. PMID: 19702511 [PubMed - as supplied by publisher] |
| Formation of human capillaries in vitro: The engineering of pre-vascularized matrices.
August 26, 2009 at 10:13 am
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| | Formation of human capillaries in vitro: The engineering of pre-vascularized matrices. Tissue Eng Part A. 2009 Aug 25; Authors: Montaño I, Schiestl C, Schneider J, Pontiggia L, Luginbühl JF, Boettcher S, Biedermann T, Braziulis E, Meuli M, Reichmann E Initial take, development, and function of transplanted engineered tissue substitutes are crucially dependent on rapid and adequate blood perfusion. Therefore, the development of rapidly and efficiently vascularized tissue grafts is vital for tissue engineering and regenerative medicine. Here we report on the construction of a network of highly organotypic capillaries in engineered tissue substitutes. We employed a 3D culture system consisting of human microvascular endothelial cells (HuMECs). These were reproducibly expanded at high purity and subsequently seeded into biodegradable, fibrin-based hydrogels. The process of capillary formation in vitro followed the principles of both angiogenesis and postnatal vasculogenesis and a distinct sequence of other developmental steps that closely resemble embryonic neovascularization. Capillary lumen formation in vitro was initiated by the deposition of a basement membrane and intensive pinocytosis, followed by the generation of intracellular vacuoles, successive fusion of these vacuoles, and finally the formation of a long, continuous lumen. After transplantation the vascular structures were stabilized by mural cells of the recipient animal. Our findings suggest that the in vitro engineering of pre-vascularized (interstitial) matrices, is within reach. PMID: 19702510 [PubMed - as supplied by publisher] |
| Regeneration of myocardium--dawn of a new era!
August 26, 2009 at 10:13 am
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| | Regeneration of myocardium--dawn of a new era! J Assoc Physicians India. 2009 Apr;57:312-24, 329-31 Authors: Shah VK, Shali KK The notion of repairing or regenerating lost myocardium via cell based therapies is highly appealing. The identification of adult bone marrow stem cells and their supportive pre-clinical data fueled the interest in utilizing these cells for physiological relevant cardiomyogenesis. Enthusiasm for cardiac regeneration via cell therapy has further increased by many encouraging reports in both animal and human studies. Further intensive research in basic science paralleled with clinical trials may make cardiovascular regenerative medicine a reality in fighting against congestive heart failure, the leading cause of morbidity and mortality associated with loss of functional cardiomyocytes. Here we review the preclinical phase to clinical phase of stem cell therapy for myocardium and provide a brief overview on unresolved issue and mechanistic insight of the repair. PMID: 19702038 [PubMed - in process] |
| An in vitro skin irritation test (SIT) using the EpiDerm reconstructed human epidermal (RHE) model.
August 26, 2009 at 10:13 am
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| | An in vitro skin irritation test (SIT) using the EpiDerm reconstructed human epidermal (RHE) model. J Vis Exp. 2009;(29): Authors: Kandárová H, Hayden P, Klausner M, Kubilus J, Sheasgreen J The EpiDerm Skin Irritation test (EpiDerm SIT) was developed and validated for in vitro skin irritation testing of chemicals, including cosmetic and pharmaceutical ingredients. The EpiDerm SIT utilizes the 3D in vitro reconstructed human epidermal (RHE) model EpiDerm. The procedure described in this protocol allows for discrimination between irritants of GHS category 2 and non-irritants. The test is performed over the course of a 4 day time period, consisting of pre-incubation, 60 minute exposure, 42 hour post-incubation and MTT viability assay. After tissue receipt and overnight pre-incubation (Day 0), tissues are topically exposed to the test chemicals (Day 1), which can be liquid, semisolid, solid or waxy. Three tissues are used for each test chemical, as well as for the positive control (5% aq. SDS solution), and a negative control (DPBS). Chemical exposure lasts for 60 minutes, 35 min of which the tissues are kept in an incubator at 37 degrees C. The test substances are then removed from the tissue surface by an extensive washing procedure. The tissue inserts are blotted and transferred to fresh medium. After a 24 hr incubation period (Day 2), the medium is exchanged. The medium can be saved for further analysis of cytokines or other endpoints of interest. After the medium exchange, tissues are incubated for an additional 18 hours. At the end of the entire 42 h post-incubation (day 3), the tissues are transferred into yellow MTT solution and incubated for 3 hours. The resultant purple-blue formazan salt, formed mainly by mitochondrial metabolism, is extracted for 2 hours using isopropanol. The optical density of the extracted formazan is determined using a spectrophotometer. A chemical is classified as an irritant if the tissue viability relative to the negative control treated tissues is reduced below 50%. This procedure can be used as full replacement of the in vivo rabbit skin irritation test for hazard identification and labeling of chemicals in line with EU regulations. PMID: 19597414 [PubMed - indexed for MEDLINE] |
| Carbon nanotubes: biomaterial applications.
August 26, 2009 at 10:13 am
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| | Carbon nanotubes: biomaterial applications. Chem Soc Rev. 2009 Jul;38(7):1897-903 Authors: Saito N, Usui Y, Aoki K, Narita N, Shimizu M, Hara K, Ogiwara N, Nakamura K, Ishigaki N, Kato H, Taruta S, Endo M There is increasing interest in the unique biological and medical properties of carbon nanotubes (CNTs), and it is expected that biomaterials incorporating CNTs will be developed for clinical use. There has been a great deal of progress in improving the various properties of CNTs for use in biomaterials and for promotion of tissue regeneration as scaffold materials. The effects of CNTs on cells and tissues are extremely important for their use in biomaterials. This tutorial review clarifies the current state of knowledge in the interdisciplinary field of CNT-based nanobiotechnology to determine whether CNTs may be useful in biomaterials. Future perspectives in this rapidly developing field will also be discussed. PMID: 19551170 [PubMed - indexed for MEDLINE] |
| Status of novel drug delivery technology for phytotherapeutics.
August 26, 2009 at 10:13 am
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| | Status of novel drug delivery technology for phytotherapeutics. Expert Opin Drug Deliv. 2009 Jun;6(6):625-37 Authors: Musthaba SM, Baboota S, Ahmed S, Ahuja A, Ali J Herbal medicines have been widely used all over the world since ancient times and have been recognized by physicians and patients for their better therapeutic value as they have fewer adverse effects as compared with modern medicines. However, phytotherapeutics needs a scientific approach to deliver the components in a sustained manner to increase patient compliance and avoid repeated administration. This can be achieved by designing novel drug delivery systems for herbal constituents. Novel drug delivery systems not only reduce the repeated administration to overcome non-compliance, but also help to increase the therapeutic value by reducing toxicity and increasing the bioavailability, and so on. Recently, pharmaceutical scientists have shifted their focus to designing a drug delivery system for herbal medicines using a scientific approach. For a long time herbal medicines were not considered for development as novel formulations owing to lack of scientific justification and processing difficulties, such as standardization, extraction and identification of individual drug components in complex polyherbal systems. However, modern phytopharmaceutical research solves the scientific needs for herbal medicines as in modern medicine, which gives way for developing novel formulations such as nanoparticles, microemulsions, matrix systems, solid dispersions, liposomes, solid lipid nanoparticles, and so on. This article summarizes various drug delivery technologies for herbal actives, which are gaining more attention for better therapeutic response. PMID: 19505192 [PubMed - indexed for MEDLINE] |
| Coculture of synovium-derived stem cells and nucleus pulposus cells in serum-free defined medium with supplementation of transforming growth factor-beta1: a potential application of tissue-specific stem cells in disc regeneration.
August 26, 2009 at 10:13 am
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| | Coculture of synovium-derived stem cells and nucleus pulposus cells in serum-free defined medium with supplementation of transforming growth factor-beta1: a potential application of tissue-specific stem cells in disc regeneration. Spine (Phila Pa 1976). 2009 May 20;34(12):1272-80 Authors: Chen S, Emery SE, Pei M STUDY DESIGN: A coculture of synovium-derived stem cells (SDSCs) and nucleus pulposus cells (NPCs) in a serum-free pellet system was treated with varying doses of transforming growth factor beta (TGF-beta). Cultures of either SDSCs or NPCs alone served as controls. OBJECTIVE: The aim was to assess the feasibility of using SDSCs to supplement and replenish NPC population for disc regeneration. SUMMARY OF BACKGROUND DATA: SDSCs have been proven to be a tissue-specific type of mesenchymal stem cell capable of chondrogenesis. NPCs are chondrocyte-like cells with a high ratio of aggrecan. However, the capacity of SDSCs to complement the NPC population is not known. METHODS: SDSCs were negatively isolated from porcine knee joint synovial tissue and NPCs were isolated from porcine lumbar spines (L1-L5). SDSCs and NPCs were cocultured (50:50) in a serum-free pellet system with the supplementation of varying doses (0, 3, 10, and 30 ng/mL) of TGF-beta1 for 14 days. SDSCs or NPCs cultured alone served as controls. Chondrogenic differentiation markers were evaluated by histology, immunohistochemistry, biochemistry, and TaqMan PCR. RESULTS: The coculture of SDSCs and NPCs in a pellet system displayed comparable differentiation properties (high levels of collagen II, aggrecan and Sox 9, a low level of collagen I, and no collagen X detectable) to NPCs alone when treated with high doses of TGF-beta1. Moreover, the coculture and NPCs alone shared a similar higher ratio of aggrecan to collagen II. Hypoxia-inducible factor 1alpha (HIF-1alpha) was also observed to be up-regulated in coculture pellets at day 7 and had decreased at day 14 with the time of pellet tissue maturation. CONCLUSION: SDSCs may act as a potential mesenchymal stem cell candidate for NP regeneration. Further studies are needed to evaluate the in vivo effect of SDSCs on disc regeneration. PMID: 19455002 [PubMed - indexed for MEDLINE] |
| The curiously misunderstood role of evidence in designing new technology.
August 26, 2009 at 10:13 am
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| | The curiously misunderstood role of evidence in designing new technology. Rejuvenation Res. 2009 Apr;12(2):75-6 Authors: de Grey AD PMID: 19415977 [PubMed - indexed for MEDLINE] |
| Expression of Delta-like 1 in the splenic non-hematopoietic cells is essential for marginal zone B cell development.
August 26, 2009 at 10:13 am
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| | Expression of Delta-like 1 in the splenic non-hematopoietic cells is essential for marginal zone B cell development. Immunol Lett. 2008 Nov 16;121(1):33-7 Authors: Sheng Y, Yahata T, Negishi N, Nakano Y, Habu S, Hozumi K, Ando K The Notch ligand Delta-like 1 (Dll1) is critical for the generation of marginal zone (MZ) B cells in the spleen. However, the precise mechanism underlying the differentiation of MZB cells is unclear. To determine whether hematopoietic cells or non-hematopoietic cells provides the Dll1-mediated signals to primitive hematopoietic cells, we transplanted lineage(-)c-kit(+)Sca-1(+) (KSL) bone marrow cells derived from wild-type (Dll1(+/+)) GFP-transgenic mice into lethally irradiated Dll1 conditional knockout (cKO) mice. After transplantation, we examined the kinetics of hematopoietic reconstitution and found that although the frequency of stem/progenitor subsets and of more mature lymphoid, myeloid, and erythroid lineages were normal, the donor-derived hematopoietic cells failed to differentiate into MZB cells. We further demonstrated that while the splenic stromal cells of wild-type mice expressed Dll1 molecule, the splenic stromal cells of recipient Dll1 cKO mice deleted the expression of Dll1. These results suggesting that the expression of Dll1 in splenic non-hematopoietic stromal cells, but not hematopoietic cells, is essential for the development of MZB cells. PMID: 18786568 [PubMed - indexed for MEDLINE] | | | 
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