| Green Process to Prepare Silk Fibroin/Gelatin Biomaterial Scaffolds.
November 20, 2009 at 6:24 am
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| | Green Process to Prepare Silk Fibroin/Gelatin Biomaterial Scaffolds. Macromol Biosci. 2009 Nov 18; Authors: Lu Q, Zhang X, Hu X, Kaplan DL A new all-aqueous and green process is described to form three-dimensional porous silk fibroin matrices with control of structural and morphological features. Silk-based scaffolds are prepared using lyophilization. Gelatin is added to the aqueous silk fibroin solution to change the silk fibroin conformation and silk fibroin-water interactions through adjusting the hydrophilic interactions in silk fibroin-gelatin-water systems to restrain the formation of separate sheet like structures in the material, resulting in a more homogenous structure. Water annealing is used to generate insolubility in the silk fibroin-gelatin scaffold system, thereby avoiding the use of organic solvents such as methanol to lock in the beta-sheet structure. The adjusting of the concentration of gelatin, as well as the concentration of silk fibroin, leads to control of morphological and functional properties of the scaffolds. The scaffolds were homogeneous in terms of interconnected pores, with pore sizes ranging from 100 to 600 microm, depending on the concentration of silk fibroin used in the process. At the same time, the morphology of the scaffolds changed from lamellar sheets to porous structures based on the increase in gelatin content. Compared with salt-leaching aqueous-derived scaffolds and hexafluoroisopropanol (HFIP)-derived scaffolds, these freeze-dried scaffolds had a lower content of beta-sheet, resulting in more hydrophilic features. Most of gelatin was entrapped in the silk fibroin-gelatin scaffolds, without the burst release in PBS solution. During in vitro cell culture, these silk fibroin-gelatin scaffolds had improved cell-compatibility than salt-leaching silk fibroin scaffolds. This new process provides useful silk fibroin-based scaffold systems for use in tissue engineering. Furthermore, the whole process is green, including all-aqueous, room temperature and pressure, and without the use of toxic chemicals or solvents, offering new ways to load bioactive drugs or growth factors into the process. PMID: 19924684 [PubMed - as supplied by publisher] |
| The amniotic membrane as a source of stem cells.
November 20, 2009 at 6:24 am
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| | The amniotic membrane as a source of stem cells. Histol Histopathol. 2010 Jan;25(1):91-98 Authors: Insausti CL, Blanquer M, Bleda P, Iniesta P, Majado MJ, Castellanos G, Moraleda JM Cellular therapy has emerged as a new potential tool for curing a wide range of degenerative diseases and tissue necrosis. Embryonic stem cells possess potential for differentiation into a wide range of cell lineages, but the ethical issues associated with establishment of this human cell line have to be resolved prior to any use. The bone marrow (BM) is the usual source of adult stem cells for hematopoietic stem cell transplants and cellular therapy, but the BM harvest is a surgical procedure that requires general anesthesia or sedation, and there seems to be a reduction of the proliferative potential and differentiation capacity of the marrow mesenchymal stem cells in older donors. For these reasons there is an increasing interest in other sources of stem cells from adult and fetal tissues. The amniotic membrane (AM) or amnion is a tissue of particular interest because its cells possess characteristics of stem cells with multipotent differentiation ability, and because of low immunogenicity and easy procurement from the placenta, which is a discarded tissue after parturition, thus avoiding the current controversies associated with the use of human embryonic stem cells. Therefore, amniotic membrane has been proposed as a good candidate to be used in cellular therapy and regenerative medicine. PMID: 19924645 [PubMed - as supplied by publisher] |
| Osteogenic activity of MG63 cells on bone-like hydroxyapatite/collagen nanocomposite sponges.
November 20, 2009 at 6:24 am
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| | Osteogenic activity of MG63 cells on bone-like hydroxyapatite/collagen nanocomposite sponges. J Mater Sci Mater Med. 2009 Nov 19; Authors: Yoshida T, Kikuchi M, Koyama Y, Takakuda K The hydroxyapatite/collagen (HAp/Col) sponge with 95% (v/v) porosity was prepared by freeze-drying of a HAp/Col fiber suspension. MG63 cells were seeded onto the HAp/Col sponge and cultured under a pressure/perfusion condition with osteogenic supplements. A collagen (Col) sponge was used as a control. The cells with sponge were examined by a histology, total DNA content and gene expression. The cells showed good attachment and proliferation everywhere in the HAp/Col sponge, while the cells mainly proliferated at the peripheral part of the Col sponge. Thus, total DNA content in the HAp/Col sponges reached 1.8 times greater than that in the Col sponges at Day 21. Further, the cells and extracellular matrix only in the HAp/Col sponge were calcified, although the cells in both sponge evenly expressed osteogenic gene. These results suggest that the HAp/Col sponge could be useful as a scaffold for bone tissue engineering. PMID: 19924517 [PubMed - as supplied by publisher] |
| Systems-level dynamic analyses of fate change in murine embryonic stem cells.
November 20, 2009 at 6:24 am
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| | Systems-level dynamic analyses of fate change in murine embryonic stem cells. Nature. 2009 Nov 19;462(7271):358-362 Authors: Lu R, Markowetz F, Unwin RD, Leek JT, Airoldi EM, Macarthur BD, Lachmann A, Rozov R, Ma'ayan A, Boyer LA, Troyanskaya OG, Whetton AD, Lemischka IR Molecular regulation of embryonic stem cell (ESC) fate involves a coordinated interaction between epigenetic, transcriptional and translational mechanisms. It is unclear how these different molecular regulatory mechanisms interact to regulate changes in stem cell fate. Here we present a dynamic systems-level study of cell fate change in murine ESCs following a well-defined perturbation. Global changes in histone acetylation, chromatin-bound RNA polymerase II, messenger RNA (mRNA), and nuclear protein levels were measured over 5 days after downregulation of Nanog, a key pluripotency regulator. Our data demonstrate how a single genetic perturbation leads to progressive widespread changes in several molecular regulatory layers, and provide a dynamic view of information flow in the epigenome, transcriptome and proteome. We observe that a large proportion of changes in nuclear protein levels are not accompanied by concordant changes in the expression of corresponding mRNAs, indicating important roles for translational and post-translational regulation of ESC fate. Gene-ontology analysis across different molecular layers indicates that although chromatin reconfiguration is important for altering cell fate, it is preceded by transcription-factor-mediated regulatory events. The temporal order of gene expression alterations shows the order of the regulatory network reconfiguration and offers further insight into the gene regulatory network. Our studies extend the conventional systems biology approach to include many molecular species, regulatory layers and temporal series, and underscore the complexity of the multilayer regulatory mechanisms responsible for changes in protein expression that determine stem cell fate. PMID: 19924215 [PubMed - as supplied by publisher] |
| Genetic ablation of caveolin-1 increases neural stem cell proliferation in the subventricular zone (SVZ) of the adult mouse brain.
November 20, 2009 at 6:24 am
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| | Genetic ablation of caveolin-1 increases neural stem cell proliferation in the subventricular zone (SVZ) of the adult mouse brain. Cell Cycle. 2009 Dec 28;8(23) Authors: Jasmin JF, Yang M, Iacovitti L, Lisanti MP Adult neural stem cells are self-renewing multipotent cells that have the potential to replace dysfunctional and/or dying neuronal cells at the site of brain injury or degeneration. Caveolins are well-known tumor-suppressor genes that were recently found to be involved in the regulation of stem cell proliferation. For instance, ablation of the caveolin-1 (Cav-1) gene in mice markedly increases the proliferation of intestinal and mammary stem cells. However, the roles of caveolins in the proliferation of adult neural stem cells still remain unknown. In this study, dual-label immunofluorescence analysis of the proliferation marker, Ki67, and the stem cell markers, nestin and Sox2, was performed on brains of 8 week-old wild-type (WT) and Cav-1 knockout (KO) mice. Our results demonstrate an increased number of Ki67-positive nuclei in the subventricular zone (SVZ) of Cav-1 KO brains. Importantly, our dual-label immunofluorescence analyses demonstrate increased co-localization of Ki67 with both nestin and Sox2 in the SVZ of Cav-1 KO brains. Remarkably similar results were also obtained with Cav-2 and Cav-3 KO mouse brains as well, with increased proliferation of adult neural stem cells. Thus, the SVZ of caveolin KO mouse brains displays an increased proliferation of adult neural stem cells. Caveolin proteins might represent new crucial regulators of adult neural stem cell proliferation. PMID: 19923909 [PubMed - as supplied by publisher] |
| The reverse Warburg effect: Aerobic glycolysis in cancer associated fibroblasts and the tumor stroma.
November 20, 2009 at 6:24 am
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| | The reverse Warburg effect: Aerobic glycolysis in cancer associated fibroblasts and the tumor stroma. Cell Cycle. 2009 Dec 5;8(23) Authors: Pavlides S, Whitaker-Menezes D, Castello-Cros R, Flomenberg N, Witkiewicz AK, Frank PG, Casimiro MC, Wang C, Fortina P, Addya S, Pestell RG, Martinez-Outschoorn UE, Sotgia F, Lisanti MP Here, we propose a new model for understanding the Warburg effect in tumor metabolism. Our hypothesis is that epithelial cancer cells induce the Warburg effect (aerobic glycolysis) in neighboring stromal fibroblasts. These cancer-associated fibroblasts, then undergo myo-fibroblastic differentiation, and secrete lactate and pyruvate (energy metabolites resulting from aerobic glycolysis). Epithelial cancer cells could then take up these energy-rich metabolites and use them in the mitochondrial TCA cycle, thereby promoting efficient energy production (ATP generation via oxidative phosphorylation), resulting in a higher proliferative capacity. In this alternative model of tumorigenesis, the epithelial cancer cells instruct the normal stroma to transform into a wound-healing stroma, providing the necessary energy-rich micro-environment for facilitating tumor growth and angiogenesis. In essence, the fibroblastic tumor stroma would directly feed the epithelial cancer cells, in a type of host-parasite relationship. We have termed this new idea the "Reverse Warburg Effect." In this scenario, the epithelial tumor cells "corrupt" the normal stroma, turning it into a factory for the production of energy-rich metabolites. This alternative model is still consistent with Warburg's original observation that tumors show a metabolic shift towards aerobic glycolysis. In support of this idea, unbiased proteomic analysis and transcriptional profiling of a new model of cancer-associated fibroblasts (caveolin-1 (Cav-1) deficient stromal cells), shows the upregulation of both (1) myo-fibroblast markers and (2) glycolytic enzymes, under normoxic conditions. We validated the expression of these proteins in the fibroblastic stroma of human breast cancer tissues that lack stromal Cav-1. Importantly, a loss of stromal Cav-1 in human breast cancers is associated with tumor recurrence, metastasis, and poor clinical outcome. Thus, an absence of stromal Cav-1 may be a biomarker for the "Reverse Warburg Effect," explaining its powerful predictive value. PMID: 19923890 [PubMed - as supplied by publisher] |
| Molecular Signaling Networks That Choreograph Epimorphic Fin Regeneration in Zebrafish - A Mini-Review.
November 20, 2009 at 6:24 am
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| | Molecular Signaling Networks That Choreograph Epimorphic Fin Regeneration in Zebrafish - A Mini-Review. Gerontology. 2009 Nov 18; Authors: Tal TL, Franzosa JA, Tanguay RL This short review provides a current synopsis of caudal fin regeneration in zebrafish with an emphasis on the molecular signaling networks that dictate epimorphic regeneration. At the outset, the fundamentals of caudal fin architecture and the stages of epimorphic regeneration are described. This is followed by a detailed look at the main networks implicated in fin regeneration, namely the Wnt, fibroblast growth factor, activin-betaA, retinoic acid and hedgehog signaling pathways. Throughout this mini-review, these molecular networks are examined through the lens of wound healing, blastema formation or regenerative outgrowth, three of the main stages of epimorphic regeneration. Next, the emerging role of noncoding RNAs as regulators of regeneration and mechanisms of regenerative termination are discussed. Finally, the implications for future research and the broader field of regenerative medicine are examined. PMID: 19923791 [PubMed - as supplied by publisher] |
| On the Determination of Darcy Permeability Coefficients for a Microporous Tissue Scaffold.
November 20, 2009 at 6:24 am
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| | On the Determination of Darcy Permeability Coefficients for a Microporous Tissue Scaffold. Tissue Eng Part C Methods. 2009 Nov 18; Authors: Wang Y, Tomlins PE, Coombes AG, Rides M Structural characterization of porous tissue scaffolds is challenging due to their complexity. Most investigators report the porosity of the material together with an estimate of the mean pore size and the pore size distribution. The usefulness of these measures is limited, especially in predicting the time-dependent permeation characteristics of a biodegradable, cell-seeded scaffold. A potential solution to this problem is to measure the permeability of the matrix and determine the Darcy permeability coefficient. Darcy permeability coefficients of 3.1 x 10(-14) and 6.3 x 10(-14) m(2) were measured for air and water, respectively, in microporous polycaprolactone scaffolds by monitoring fluid flow in response to a range of pressure differentials. Permeability coefficients for phosphate-buffered saline (5.3 x 10(-14) m(2)), glucose (5.7 x 10(-14) m(2)), and bovine serum albumin (1.8 x 10(-14) m(2)) were obtained by monitoring the change in concentration of molecular probes. This approach revealed the efficiency of transport of glucose molecules through the porous material and the existence of protein-scaffold interactions that resulted in protein retention and a reduction in fluid permeation rate. Darcy permeability measurements can provide valuable insights concerning the transport properties of nutrients, metabolites, and polypeptide growth factors in porous tissue engineering scaffolds and a method of quality assurance in scaffold processing. PMID: 19922263 [PubMed - as supplied by publisher] |
| Optimization of Platelet Isolation and Extraction of Autogenous TGF-ss in Cartilage Tissue Engineering.
November 20, 2009 at 6:24 am
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| | Optimization of Platelet Isolation and Extraction of Autogenous TGF-ss in Cartilage Tissue Engineering. Artif Cells Blood Substit Immobil Biotechnol. 2009 Dec;37(6):265-272 Authors: Staudenmaier R, Froelich K, Birner M, Kindermann J, Hoang NT, Pueschel RC, Mandlik V Abstract Platelets are enriched with Transforming Growth Factor-ss (TGF-ss). However, information is limited concerning TGF-ss's effects at the molecular level. Nevertheless, it has been demonstrated that TGF-ss activates cell proliferation and its postitive influence on cartilage formation has been proven within the field of Tissue Engineering (TE). As Platelet Rich Plasma (PRP) contains TGF-ss, it was the purpose of this study to optimize PRP-isolation for further TGF-ss extraction. Red blood cell count (RBC) was separated from whole blood by centrifugation. From the supernatant PRP and platelet poor plasma (PPP) layer, the latter supernatant was re-centrifuged to extract PRP. Various experimental series were run to investigate influences concerning anticoagulating alternatives, different amounts of buffer, various centrifugal forces, or substituting centrifugation for sedimentation. TGF-ss levels were determined using ELISA. The technique of platelet-/ TGF-ss-extraction described here proves to be more effective than other methods, is easily repeatable and nottime-consuming, which predisposes it for TE requirements. PMID: 19922166 [PubMed - as supplied by publisher] |
| In situ tissue engineering of canine skull with guided bone regeneration.
November 20, 2009 at 6:24 am
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| | In situ tissue engineering of canine skull with guided bone regeneration. Acta Otolaryngol. 2009;129(12):1509-1518 Authors: Umeda H, Kanemaru SI, Yamashita M, Ohno T, Suehiro A, Tamura Y, Hirano S, Nakamura T, Omori K, Ito J Conclusion: Calcium alginate (CA) membrane prevents excessive fibrous tissue intrusion and/or dislocation of a bone scaffold. However, CA membrane did not always accelerate cranial bone regeneration. Objective: We previously reported skull regeneration using a bone substitute material (BSM), which consisted of collagen-coated beta-tricalcium phosphate and autologous bone fragments, and bone marrow-derived stromal cells (BSCs). However, excessive fibrous tissue intrusion or dislocation of the BSM occasionally interrupted bone regeneration. To avoid such problems, we examined CA membrane, which is useful for guided bone regeneration (GBR), to investigate whether this material maintains the bone regenerative space. Materials and methods: Bone defects (2x2 cm) were created in the skulls of 12 adult beagle dogs using the same clinical procedure. Four experimental models were tested with or without BSM plus BSCs or CA membrane. In group I, the original free bone flap was replaced at the defect. In group II, after replacing the bone flap, the defect was covered with CA membrane. In group III, BSM plus BSCs were used as a gap filler. In group IV, BSM plus BSCs and CA membrane were applied. Histological examinations were performed 3 and 6 months after the operation. Results: In groups I and II, bone regeneration was not observed but fibrous tissue intrusion was prevented in group II. Bone neogenesis was more observed in group III than in group IV at 3 months (p<0.05). At 6 months, the regenerated areas were larger than those observed at 3 months, but the differences between groups III and IV were not statistically significant. PMID: 19922106 [PubMed - as supplied by publisher] | | | 
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