|  |  |  | | | | | TERMSC | | | | | | | | | | | | | | | | | | | Microprinting of Liver Micro-organ for Drug Metabolism Study. Methods Mol Biol. 2011;671:219-38 Authors: Chang RC, Emami K, Jeevarajan A, Wu H, Sun W In their normal in vivo matrix milieu, tissues assume complex well-organized 3D architectures. Therefore, a primary aim in the tissue engineering design process is to fabricate an optimal analog of the in vivo scenario, in which the precise configuration and composition of cells and bioactive matrix components can establish the well-defined biomimetic microenvironments that promote cell-cell and cell-matrix interactions. With the advent and refinements in microfabricated systems which can present physical and chemical cues to cells in a controllable and reproducible fashion unrealizable with conventional tissue culture, high-fidelity, high-throughput in vitro models are achieved. The convergence of solid freeform fabrication (SFF) technologies, namely microprinting, along with microfabrication techniques, a 3D microprinted micro-organ, can serve as an in vitro platform for cell culture, drug screening, or to elicit further biological insights. This chapter firstly details the principles, methods, and applications that undergird the fabrication process development and adaptation of microfluidic devices for the creation of a drug screening model. This model involves the combinatorial setup of an automated syringe-based, layered direct cell writing microprinting process with soft lithographic micropatterning techniques to fabricate a microscale in vitro device housing a chamber of microprinted 3D micro-organ that biomimics the cell's natural microenvironment for enhanced performance and functionality. In order to assess the structural formability and biological feasibility of such a micro-organ, 3D cell-encapsulated hydrogel-based tissue constructs are microprinted reproducibly in defined design patterns and biologically characterized for both viability and cell-specific function. Another key facet of the in vivo microenvironment that is recapitulated with the in vitro system is the necessary dynamic perfusion of the 3D microscale liver analog with cells probed for their collective drug metabolic function and suitability as a drug metabolism model. PMID: 20967633 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Generation of protein and cell microarrays on functionalized surfaces. Methods Mol Biol. 2011;671:207-17 Authors: Choi YS, Lee CS The technique of selective immobilization of biomolecules in defined positions or areas using a simple procedure is essential for various applications such as biosensors, biochips, biomedical microdevices, and tissue engineering. For the generation of biomolecule microarrays, it is necessary to develop a functional surface retaining protein functionality and cell viability, and an efficient patterning tool having flexibility of size and shape.In this chapter, we have presented the simple tools of protein and cell microarray based on functional-ized surface such as a spotting method with improvement of protein functionality, a functionalized silicon-based surface using photolithography, and an orthogonally polyelectrolyte-coated surface based on soft-lithography. PMID: 20967632 [PubMed - in process] | | | | | | | | | | | | | | | | | | | | | | | | | Quality of Innervation in Sensate Medial Plantar Flaps for Heel Reconstruction. Plast Reconstr Surg. 2010 Oct 14; Authors: Wan DC, Gabbay J, Levi B, Boyd JB, Granzow JW BACKGROUND:: Reconstruction of the heel represents a difficult challenge for surgeons, given the demand for thick, durable skin capable of withstanding both pressure and shear. We describe the use of a sensate medial plantar flap for heel reconstruction in three patients and document the long-term retention of sensation compared to the contralateral uninjured heel and corresponding donor site. METHODS:: A medial plantar flap was harvested to include the branch of the medial plantar nerve to the instep to preserve innervation. Sharp pain, light and deep pressure, vibration, cold temperature, as well as static and dynamic two-point discrimination were examined between six months and a year after surgery. RESULTS:: Sharp pain, vibration, and deep pressure sensation were present equally in the medial plantar flap, contralateral heel, and contralateral instep. Cold perception, light pressure, static two-point, and dynamic two-point discrimination were significantly less in the normal contralateral heel when compared to the heel reconstructed by the innervated flap. There were no significant differences in sensation between the medial plantar flap and the contralateral instep. CONCLUSIONS:: The medial plantar flap is capable of providing durable, sensate coverage of plantar hind foot defects with minimal donor site morbidity. Furthermore, that sensation remains identical to the instep donor site and superior to the normal heel pad. PMID: 20966816 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Fibrin/Hyaluronic Acid Composite Hydrogels as Appropriate Scaffolds for In Vivo Artificial Cartilage Implantation. ASAIO J. 2010 Oct 20; Authors: Rampichová M, Filová E, Varga F, Lytvynets A, Prosecká E, Koláčná L, Motlík J, Nečas A, Vajner L, Uhlík J, Amler E Hydrogels prepared from a mixture of fibrin and high-molecular weight (MW) hyaluronic acid (HA) were found to be suitable scaffolds for chondrocyte seeding and pig knee cartilage regeneration. Collagen in the hydrogels is not necessary for the formation of biomechanically stable tissue. Regenerated cartilage showed very good biomechanical and histological properties only 6 months after implantation. Notably, the quality of the healing process was dependent on the initial chondrocyte concentration of the scaffolds. These experiments were performed according to good laboratory practice (GLP). PMID: 20966745 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Molecularly imprinted nanoparticles with recognition properties towards a laminin H-Tyr-Ile-Gly-Ser-Arg-OH sequence for tissue engineering applications. Biomed Mater. 2010 Oct 22;5(6):065007 Authors: Rosellini E, Barbani N, Giusti P, Ciardelli G, Cristallini C Nanotechnology is an emerging field that promises to revolutionize medicine and is increasingly used in tissue engineering applications. Our research group proposed for the first time molecular imprinting as a new nanotechnology for the creation of advanced synthetic support structures for cell adhesion and proliferation. The aim of this work was the synthesis and characterization of molecularly imprinted polymers with recognition properties towards a laminin peptide sequence and their application as functionalization structures in the development of bioactive materials. Nanoparticles with an average diameter of 200 nm were synthesized by precipitation polymerization of methacrylic acid in the presence of the template molecule and trimethylpropane trimethacrylate as the cross-linking agent. The imprinted nanoparticles showed good performance in terms of recognition capacity and selectivity. The cytotoxicity tests showed normal vitality of C2C12 myoblasts cultured in the medium that was put in contact with the imprinted polymers. After the deposition on the polymeric film surface, the imprinted particles maintained their specific recognition and rebinding behaviour, showing an even higher quantitative binding than free nanoparticles. Preliminary in vitro cell culture tests demonstrated the ability of functionalized materials to promote cell adhesion, proliferation and differentiation, suggesting that molecular imprinting can be used as an innovative functionalization technique. PMID: 20966532 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Gadonanotubes as magnetic nanolabels for stem cell detection. Biomaterials. 2010 Oct 19; Authors: Tran LA, Krishnamurthy R, Muthupillai R, da Graça Cabreira-Hansen M, Willerson JT, Perin EC, Wilson LJ Stem cell-based therapies have emerged as a promising approach in regenerative medicine. In the development of such therapies, the demand for imaging technologies that permit the noninvasive monitoring of transplanted stem cells in vivo is growing. Here, we report the performance of gadolinium-containing carbon nanocapsules, or gadonanotubes (GNTs), as a new T(1)-weighted magnetic resonance imaging (MRI) intracellular labeling agent for pig bone marrow-derived mesenchymal stem cells (MSCs). Without the use of a transfection agent, micromolar concentrations of GNTs can deliver up to 10(9) Gd(3)(+) ions per cell without compromising cell viability, differentiation potential, proliferation pattern, and phenotype. Imaging 10 × 10(6) GNT-labeled MSCs demonstrates a nearly two-fold reduction in T(1) relaxation time when compared to unlabeled MSCs at 1.5 T in a clinical MRI scanner, which easily permits the discrimination of GNT-labeled MSCs in a T(1)-weighted MR image. It is anticipated that GNTs will allow in vivo tracking of GNT-labeled MSCs, as well as other mammalian cell types, by T(1)-weighted imaging with greater efficacy than other current technologies now allow. PMID: 20965562 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Development of Nano- and Micro-Scale Chondroitin Sulfate Particles for Controlled Growth Factor Delivery. Acta Biomater. 2010 Oct 18; Authors: Lim JJ, Hammoudi TM, Bratt-Leal AM, Hamilton SK, Kepple KL, Bloodworth NC, McDevitt TC, Temenoff JS Size scale plays an important role in the release properties and cellular presentation of drug delivery vehicles. Because negatively charged chondroitin sulfate (CS) is capable of electrostatically sequestering positively charged growth factors, CS-derived nanoscale micelles and microscale spheroids were synthesized as potential growth factor carriers to enhance differentiation of stem cells. Particles were characterized for morphology, size distribution, surface charge, cytocompatibility, as well as release of transforming growth factor-β1 (TGF-β1) and tumor necrosis factor-α (TNF-α). CS micelles were spherical and negatively charged with a bimodal distribution of 324.1 ± 8.5 nm and 73.2 ± 4.4 nm diameters, and CS microspheres possessed a rounded morphology and a diameter of 4.3 ± 0.93 μm. Positively charged TGF-β1 demonstrated minimal release after loading in CS microspheres, while negatively charged TNF-α exhibited substantial release over the first 15 hours, suggesting TGF-β1 electrostatically complexed with CS. The micelles and microparticles were found to be cytocompatible at moderate concentrations with marrow stromal cell (MSC) monolayers and within embryonic stem cell (ESC) embryoid bodies. These synthesis techniques, which allow the formation of CS-based carriers over a variety of nano- and microscale sizes, offer versatility for tailored release of positively charged growth factors and controlled CS presentation for a variety of stem cell-based applications in tissue engineering and regenerative medicine. PMID: 20965281 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | SDF-1α as a therapeutic stem cell homing factor in myocardial infarction. Pharmacol Ther. 2010 Oct 18; Authors: Ghadge S, Mühlstedt S, Ozcelik C, Bader M Myocardial infarction is associated with persistent muscle damage, scar formation and depressed cardiac performance. Recent studies have demonstrated the clinical significance of stem cell-based therapies after myocardial infarction with the aim to improve cardiac remodeling and function by inducing the reconstitution of functional myocardium and formation of new blood vessels. Stem cell homing signals play an important role in stem cell mobilization from the bone marrow to the ischemic cardiac environment and are therefore crucial for myocardial repair. To date, the most prominent stem cell homing factor is the chemokine SDF-1α/CXCL12. This protein was shown to be significantly upregulated in many experimental models of myocardial infarction and in patients suffering from ischemic cardiac diseases, suggesting the involvement in the pathophysiology of these disorders. A number of studies focused on manipulating SDF-1α and its receptor CXCR4 as central regulators of the stem cell mobilization process. Targeted expression of SDF-1α after myocardial infarction was shown to result in increased engraftment of bone marrow-derived stem cells into infarcted myocardium. This was accompanied by beneficial effects on cardiomyocyte survival, neovascularization and cardiac function. Thus, the SDF-1/CXCR4 axis seems to be a promising novel therapeutic approach to improve post-infarction therapy by attracting circulating stem cells to remain, survive and possibly differentiate in the infarct area. This review will summarize clinical trials of stem cell therapy in patients with myocardial infarction. We further discuss the basic findings about SDF-1α in stem cell recruitment and its therapeutic implications in experimental myocardial infarction. PMID: 20965212 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | At the stem of youth and health. Pharmacol Ther. 2010 Oct 18; Authors: Beltrami AP, Cesselli D, Beltrami CA Cellular senescence is a specialized form of growth arrest, confined to mitotic cells, induced by various stressful stimuli and characterized by a permanent growth arrest, resistance to apoptosis, an altered pattern of gene expression and the expression of some markers that are characteristic, although not exclusive, to the senescent state. Senescent cells profoundly modify neighboring and remote cells through the production of an altered secretome, eventually leading to inflammation, fibrosis and possibly growth of neoplastic cells. Mammalian aging has been defined as a reduction in the capacity to adequately maintain tissue homeostasis or to repair tissues after injury. Tissue homeostasis and regenerative capacity are nowadays considered to be related to the stem cell pool present in every tissue. For this reason, pathological and patho-physiological conditions characterized by altered tissue homeostasis and impaired regenerative capacity can be viewed as a consequence of the reduction in stem cell number and/or function. Last, cellular senescence is a double-edged sword, since it may inhibit the growth of transformed cells, preventing the occurrence of cancer, while it may facilitate growth of preneoplastic lesions in a paracrine fashion; therefore, interventions targeting this cell response to stress may have a profound impact on many age related pathologies, ranging from cardiovascular disease to oncology. Aim of this review is to discuss both molecular mechanisms associated with stem cell senescence and interventions that may attenuate or reverse this process. PMID: 20965211 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Community effect triggers terminal differentiation of myogenic cells derived from muscle satellite cells by quenching Smad signaling. Exp Cell Res. 2010 Oct 18; Authors: Yanagisawa M, Mukai A, Shiomi K, Song SY, Hashimoto N A high concentration of bone morphogenetic proteins (BMPs) stimulates myogenic progenitor cells to undergo heterotopic osteogenic differentiation. However, the physiological role of the Smad signaling pathway during terminal muscle differentiation has not been resolved. We report here that Smad1/5/8 was phosphorylated and activated in undifferentiated growing mouse myogenic progenitor Ric10 cells without exposure to any exogenous BMPs. The amount of phosphorylated Smad1/5/8 was severely reduced during precocious myogenic differentiation under the high cell density culture condition even in growth medium supplemented with a high concentration of serum. Inhibition of the Smad signaling pathway by dorsomorphin, an inhibitor of Smad activation, or noggin, a specific antagonist of BMP, induced precocious terminal differentiation of myogenic progenitor cells in a cell density-dependent fashion even in growth medium. In addition, Smad1/5/8 was transiently activated in proliferating myogenic progenitor cells during muscle regeneration in rats. The present results indicate that the Smad signaling pathway is involved in a critical switch between growth and differentiation of myogenic progenitor cells both in vitro and in vivo. Furthermore, precocious cell density-dependent myogenic differentiation suggests that a community effect triggers the terminal muscle differentiation of myogenic cells by quenching the Smad signaling. PMID: 20965167 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Cell therapy in amyotrophic lateral sclerosis: science and controversy. Neurologia. 2010 Oct;25(8):467-469 Authors: Galán L, Guerrero-Sola A, Gómez-Pinedo U, Matias-Guiu J Stem cell therapy is seen as a possible alternative for the treatment of different degenerative diseases, among which includes amyotrophic lateral sclerosis (ALS). Despite there being basic research works with this therapy in ALS, the mechanism of action of the implanted cells are still unclear. It is also unclear which type of cells to use (bone marrow, fat, dental pulp, etc.), or the most ideal administration route. Furthermore, clinical trials with mesenchymal stem cells are not very conclusive, therefore it has not been convincingly established as an alternative therapy in ALS or any other neurodegenerative disease. Despite the scientific evidence, several clinical trials have been conducted in the last few years that offer stem cell treatments for neurodegenerative diseases, giving rise to what is known as "cellular tourism". This phenomenon has set off alarms and reactions in the scientific community. The application of these therapies must be performed following the good clinical practice guidelines in research, evidence based methodology and international ethical and scientific recommendations. PMID: 20964996 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Fluorescence nanoparticles "quantum dots" as drug delivery system and their toxicity: a review. J Drug Target. 2010 Oct 22; Authors: Ghaderi S, Ramesh B, Seifalian AM Fluorescence nanocrystals or quantum dots (QDs) are engineered nanoparticles (NP) that have shown great promise with potential for many biological and biomedical applications, especially in drug delivery/activation and cellular imaging. The use of nanotechnology in medicine directed to drug delivery is set to expand in the coming years. However, it is unclear whether QDs, which are defined as NPs rather than small molecules, can specifically and effectively deliver drugs to molecular targets at subcellular levels. When QDs are linked to suitable ligands that are site specific, it has been shown to be brighter and photostable when compared with organic dyes. Interestingly, pharmaceutical sciences are exploiting NPs to minimize toxicity and undesirable side effects of drugs. The unforeseen hazardous properties of the carrier NPs themselves have given rise to some concern in a clinical setting. The kind of hazards encountered with this new nanotechnology materials are complex compared with conventional limitations created by traditional delivery systems. The development of cadmium-derived QDs shows great potential for treatment and diagnosis of cancer and site-directed delivery by virtue of their size-tunable fluorescence and with highly customizable surface for directing their bioactivity and targeting. However, data regarding the pharmacokinetic and toxicology studies require further investigation and development, and it poses great difficulties to ascertain the risks associated with this new technology. Additionally, nanotechnology also displays yet another inherent risk for toxic cadmium, which will enter as a new form of hazard in the biomedical field. This review will look at cadmium-derived QDs and discuss their future and their possible toxicities in a disease situation. PMID: 20964619 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Delivery of Plasmid DNA Encoding Bone Morphogenetic Protein-2 with a Biodegradable Branched Polycationic Polymer in a Critical-Size Rat Cranial Defect. Tissue Eng Part A. 2010 Oct 22; Authors: Chew SA, Kretlow JD, Spicer PP, Edwards AW, Baggett LS, Tabata Y, Kasper FK, Mikos AG This study investigated the delivery of plasmid DNA (pDNA) encoding bone morphogenetic protein-2 in the form of polyplexes with a biodegradable branched triacrylate/amine polycationic polymer (TAPP), which were complexed with gelatin microparticles (GMPs) loaded within a porous tissue engineering scaffold. More specifically, the study investigated the interplay between TAPP degradation, gelatin degradation, pDNA release and bone formation in a critical-size rat cranial defect model. The pDNA release kinetics in vitro were not affected by the crosslinking density of the GMPs but depended, rather, on the degradation rates of the TAPPs. Besides the initial release of polyplexes not bound to the GMPs and the minimal release of polyplexes through diffusion or dissociation from the GMPs, the pDNA was likely released as naked pDNA or as part of an incomplete polyplex, following the degradation of fragments of the polycationic polymer. After 30 days, significantly higher amounts of pDNA were released (93-98%) from composite scaffolds containing naked pDNA or pDNA complexed with P-AEPZ (a faster degrading TAPP) compared to those containing pDNA complexed with P-DED (a slower degrading TAPP) (74-82%). Composite scaffolds containing GMPs complexed with TAPP/pDNA polyplexes did not result in enhanced bone formation, as analyzed by microcomputed tomography and histology, in a critical-size rat cranial defect at 12 weeks post-implantation compared to those loaded with naked pDNA. The results demonstrate that polycationic polymers with a slow degradation rate can prolong the release of pDNA from the composite scaffolds and suggest that a gene delivery system comprising biodegradable polycationic polymers should be designed to release the pDNA in an intact polyplex form. PMID: 20964581 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | In vitro Biomimetic Construction of Hydroxyapatite -Porcine Acellular Dermal Matrix Composite Scaffold for MC3T3-E1 Preosteoblast Culture. Tissue Eng Part A. 2010 Oct 22; Authors: Zhao H, Wang G, Hu S, Cui J, Ren N, Liu H, Cao C, Wang J, Wang Z The application of porous hydroxyapatite-collagen (HAp-Collagen) as a bone tissue engineering scaffold is hindered by two main problems, its high cost and low initial strength. As a native 3-dimenssional collagen framework, purified porcine acellular dermal matrix (PADM) has been successfully used as a skin tissue engineering scaffold. Here we report its application as a matrix for the preparation of HAp to produce a bone tissue scaffold through a biomimetic chemical process. The hydroxyapatite-porcine acellular dermal matrix (HAp-PADM) scaffold has two-level pore structure, with large channels (approximately 100 m in diameter) inherited from the purified PADM microstructure and small pores (less than 100 nm in diameter) formed by self-assembled HAp on the channel surfaces. The obtained HAp-PADM scaffold (S15D) has a compressive elastic modulus as high as 600 kPa. The presence of HAp in sample S15D reduces the degradation rate of PADM in collagenase solution at 37 oC. After 7 day culture of MC3T3-E1 pre-osteroblasts, MTT data shows no statistically significant difference on pure PADM framework and HAp-PADM scaffold (p > 0.05). Because of its high strength and nontoxicity, its simple preparation method, and designable and tailorable properties, the HAp-PADM scaffold is expected to have great potential applications in medical treatment of bone defects. PMID: 20964580 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Human Adipose-Derived Side Population Stem Cells Cultured on Demineralized Bone Matrix for Bone Tissue Engineering. Tissue Eng Part A. 2010 Oct 22; Authors: Supronowicz P, Gill E, Trujillo A, Thula T, Zhukauskas R, Ramos T, Cobb R Background: Tissue engineering of new bone relies on the combination and application of osteoconductive, osteoinductive, and osteogenic elements. Natural scaffolds, such as demineralized bone matrix (DBM), contain collagenous networks with growth factors such as BMP-2. Stem cells from readily available sources, including discarded adipose tissue, have the propensity to differentiate into bone. The present study examines a multi-component technique consisting of a novel side population of adipose stem cells cultured on DBM for tissue engineering applications. Methods: Adipose-derived side population stem cells were cultured on DBM for up to 14 days. Cell proliferation, alkaline phosphatase activity, extracellular matrix protein production, and calcium-containing mineral deposit formation were assayed. Ectopic bone formation in a rat model was also evaluated. Results: Side population stem cells attached to and proliferated on DBM while generating markers of new bone formation. When these cell/substrate composites were implanted into an ectopic model, newly formed bone was 30% greater than that of DBM alone. Conclusions: Novel populations of adipose-derived stem cells cultured on DBM compose a system that develops new bone matrix in vitro and in vivo. This strategy provides a novel approach using naturally occurring materials for bone repair in tissue engineering applications. PMID: 20964579 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Correlation between compositional and mechanical properties of human mesenchymal stem cell-collagen microspheres during chondrogenic differentiation. Tissue Eng Part A. 2010 Oct 22; Authors: Li C, Chik T, Ngan AH, Chan S, Shum DK, Chan BP Mesenchymal stem cell (MSC) based engineering is promising for cartilage repair. However, the compositional mechanical relationship of the engineered structures has not been extensively studied, given the importance of such relationship in native cartilage tissues. In this study, a novel human MSC-collagen microsphere system was used to study the compositional mechanical relationship during in vitro chondrogenic differentiation using histological and biochemical methods and a microplate compression assay. The mechanical property was found positively correlating with newly deposited cartilage-relevant matrices, glycosaminoglycan and type II collagen, and with the collagen crosslinker density, in agreement with the presence of thick collagen bundles upon structural characterization. On the other hand, the mechanical property negatively correlates with type I collagen and total collagen, suggesting that the initial collagen matrix scaffold of the microsphere system was being remodeled by the differentiating hMSCs. This study also demonstrated the application of a simple, sensitive and non-destructive tool for monitoring the progression of chondrogenic differentiation of MSCs in tissue engineered constructs and therefore contributes to future development of novel cartilage repair strategies. PMID: 20964578 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Enhanced Reprogramming and Cardiac Differentiation of Human Keratinocytes Derived from Plucked Hair Follicles, Using a Single Excisable Lentivirus. Cell Reprogram. 2010 Oct 21; Authors: Novak A, Shtrichman R, Germanguz I, Segev H, Zeevi-Levin N, Fishman B, Mandel YE, Barad L, Domev H, Kotton D, Mostoslavsky G, Binah O, Itskovitz-Eldor J Abstract Induced pluripotent stem cells (iPSCs) represent an ideal cell source for future cell therapy and regenerative medicine. However, most iPSC lines described to date have been isolated from skin fibroblasts or other cell types that require harvesting by surgical intervention. Because it is desirable to avoid such intervention, an alternative cell source that can be readily and noninvasively isolated from patients and efficiently reprogrammed, is required. Here we describe a detailed and reproducible method to derive iPSCs from plucked human hair follicle keratinocytes (HFKTs). HFKTs were isolated from single plucked hair, then expanded and reprogrammed by a single polycistronic excisable lentiviral vector. The reprogrammed HFKTs were found to be very sensitive to human embryonic stem cell (hESC) growth conditions, generating a built-in selection with easily obtainable and very stable iPSCs. All emerging colonies were true iPSCs, with characteristics typical of human embryonic stem cells, differentiated into derivatives of all three germ layers in vitro and in vivo. Spontenaeouly differentiating functional cardiomyocytes (CMs) were successfully derived and characterized from these HFKT-iPSCs. The contracting CMs exhibited well-coordinated intracellular Ca(2+) transients and contractions that were readily responsive to β-adrenergic stimulation with isoproterenol. The introduction of Cre-recombinase to HFKT-iPSC clones was able to successfully excise the integrated vector and generate transgene-free HFKT-iPSC clone that could be better differentiated into contracting CMs, thereby revealing the desired cells for modeling human diseases. Thus, HFKTs are easily obtainable, and highly reprogrammed human cell source for all iPSC applications. PMID: 20964482 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Generation of Healthy Cloned Mice Using Enucleated Cryopreserved Oocytes. Cell Reprogram. 2010 Oct 21; Authors: Hirata S, Fukasawa H, Wakayama S, Wakayama T, Hoshi K Abstract The successful generation of cloned animals and the establishment of embryonic stem (ES) cell lines from somatic cells suggest that these techniques may be used in human regenerative medicine. However, the fact that oocytes must be donated by women undergoing infertility treatment remains a fundamental ethical objection, as they might be concerned about the potential exploitation of their genome. Here, we investigated the reprogramming potential of enucleated and cryopreserved oocytes for the development of full-term cloned mice. BDF1 strain mouse oocytes were cryopreserved at metaphase II, before and after enucleation. After thawing, cumulus cell nuclei were microinjected to generate clones. Although the rate of development of cloned embryos to the blastocyst stage using the treated oocytes was lower than that obtained using fresh oocytes, three live pups were delivered after embryo transfer into pseudopregnant females (0.4% of the oocytes used). Thus, although cryopreservation reduces the potential of oocytes, these cells retain the ability to support the full-term development of cloned embryos. In addition, the removal of DNA from human oocytes may alleviate the ethical and psychological problems for women who are undergoing infertility treatment and are considering oocyte donation for research or therapeutic purposes. PMID: 20964481 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Thermoresponsive Copolymer Nanofilms for Controlling Cell Adhesion, Growth, and Detachment. Langmuir. 2010 Oct 21; Authors: Yang L, Pan F, Zhao X, Yaseen M, Padia F, Coffey P, Freund A, Yang L, Liu T, Ma X, Lu JR This study reports the development and use of a novel thermoresponsive polymeric nanofilm for controlling cell adhesion and growth at 37 °C, and then cell detachment for cell recovery by subsequent temperature drop to the ambient temperature, without enzymatic cleavage or mechanical scraping. A copolymer, poly(N-isopropylacrylamide-co-hydroxypropyl methacrylate-co-3-(trimethoxysilyl)propyl methacrylate) (abbreviated PNIPAAm copolymer), was synthesized by free radical polymerization. The thermoresponses of the copolymer in aqueous solution were demonstrated by dynamic light scattering (DLS) through detecting the sensitive changes of copolymer aggregation against temperature. The DLS measurements revealed the lower critical solution temperature (LCST) at approximately 30 °C. The PNIPAAm film stability and robustness was provided through silyl cross-linking within the film and with the hydroxyl groups on the substrate surface. Film thickness, stability, and reversibility with respect to temperature switches were examined by spectroscopic ellipsometry (SE), atomic force microscopy (AFM), and contact angle measurements. The results confirmed the high extent of thermosensitivity and structural restoration based on the alterations of film thickness and surface wettability. The effective control of adhesion, growth, and detachment of HeLa and HEK293 cells demonstrated the physical controllability and cellular compatibility of the copolymer nanofilms. These PNIPAAm copolymer nanofilms could open up a convenient interfacial mediation for cell film production and cell expansion by nonenzymatic and nonmechanical cell recovery. PMID: 20964301 [PubMed - as supplied by publisher] | | | | | | | | | | | | | | | | | | | | | | | | | Expression profile of mRNAs encoding core circadian regulatory proteins in human subcutaneous adipose tissue: correlation with age and body mass index. Int J Obes (Lond). 2009 Sep;33(9):971-7 Authors: Wu X, Xie H, Yu G, Hebert T, Goh BC, Smith SR, Gimble JM OBJECTIVE: Circadian mechanisms underlie the physiology of mammals as an adaptation to the earth's rotation on its axis. Highly conserved core circadian regulatory proteins (CCRPs) maintain an oscillatory expression profile in the central and peripheral tissues. The CCRP include both a positive and negative arm, as well as downstream transcriptional regulators. Recent studies in murine models have determined that the mRNAs encoding the CCRP are present in multiple adipose tissue depots and exhibit a robust oscillatory expression profile. This study set out to examine the expression of CCRP mRNAs in human subcutaneous adipose tissues. DESIGN: Retrospective analysis of total RNA isolated from subcutaneous adipose tissue. SUBJECTS: A total of 150 healthy female and male lean (body mass index (BMI) <25), overweight (BMI between 25 and 29.99) or obese (BMI >30) subjects of varied ethnic backgrounds undergoing elective liposuction or surgical procedures. RESULTS: The expression of the CCRP mRNAs displayed a significant correlation between each other and mRNAs representative of adipogenic biomarkers. Hierarchical cluster analyses of mRNAs isolated from the cohort of female Caucasian subjects (n=116) identified three major clusters based on expression of downstream CCRP mRNAs. The mRNAs encoding D site of albumin promoter-binding protein (DBP), E4 promoter-binding protein 4 (E4BP4), PPARgamma coactivator-1beta (PGC-1beta) and Rev-erbalpha were negatively correlated with BMI in a lean cluster (n=66), positively correlated with BMI in a younger overweight/obese cluster (n=19), and not significantly correlated with BMI in an older, overweight/obese cluster (n=31). CONCLUSIONS: These data confirm and extend findings that link the CCRP and circadian mechanisms to the risk of obesity. PMID: 19597517 [PubMed - indexed for MEDLINE] | | | | | | | | | |  | |  | |  |
No comments:
Post a Comment