WorldWideScience

Sample records for cell type-specific long-term

  1. Induction of long-term potentiation and long-term depression is cell-type specific in the spinal cord.

    Science.gov (United States)

    Kim, Hee Young; Jun, Jaebeom; Wang, Jigong; Bittar, Alice; Chung, Kyungsoon; Chung, Jin Mo

    2015-04-01

    The underlying mechanism of chronic pain is believed to be changes in excitability in spinal dorsal horn (DH) neurons that respond abnormally to peripheral input. Increased excitability in pain transmission neurons, and depression of inhibitory neurons, are widely recognized in the spinal cord of animal models of chronic pain. The possible occurrence of 2 parallel but opposing forms of synaptic plasticity, long-term potentiation (LTP) and long-term depression (LTD) was tested in 2 types of identified DH neurons using whole-cell patch-clamp recordings in mouse spinal cord slices. The test stimulus was applied to the sensory fibers to evoke excitatory postsynaptic currents in identified spinothalamic tract neurons (STTn) and GABAergic neurons (GABAn). Afferent conditioning stimulation (ACS) applied to primary afferent fibers with various stimulation parameters induced LTP in STTn but LTD in GABAn, regardless of stimulation parameters. These opposite responses were further confirmed by simultaneous dual patch-clamp recordings of STTn and GABAn from a single spinal cord slice. Both the LTP in STTn and the LTD in GABAn were blocked by an NMDA receptor antagonist, AP5, or an intracellular Ca chelator, BAPTA. Both the pattern and magnitude of intracellular Ca after ACS were almost identical between STTn and GABAn based on live-cell calcium imaging. The results suggest that the intense sensory input induces an NMDA receptor-dependent intracellular Ca increase in both STTn and GABAn, but produces opposing synaptic plasticity. This study shows that there is cell type-specific synaptic plasticity in the spinal DH. PMID:25785524

  2. Induction of long-term potentiation and long-term depression is cell-type specific in the spinal cord

    OpenAIRE

    Kim, Hee Young; Jun, Jaebeom; Wang, Jigong; Bittar, Alice; Chung, Kyungsoon; Chung, Jin Mo

    2015-01-01

    Abstract The underlying mechanism of chronic pain is believed to be changes in excitability in spinal dorsal horn (DH) neurons that respond abnormally to peripheral input. Increased excitability in pain transmission neurons, and depression of inhibitory neurons, are widely recognized in the spinal cord of animal models of chronic pain. The possible occurrence of 2 parallel but opposing forms of synaptic plasticity, long-term potentiation (LTP) and long-term depression (LTD) was tested in 2 ty...

  3. Layer- and cell-type-specific subthreshold and suprathreshold effects of long-term monocular deprivation in rat visual cortex.

    Science.gov (United States)

    Medini, Paolo

    2011-11-23

    Connectivity and dendritic properties are determinants of plasticity that are layer and cell-type specific in the neocortex. However, the impact of experience-dependent plasticity at the level of synaptic inputs and spike outputs remains unclear along vertical cortical microcircuits. Here I compared subthreshold and suprathreshold sensitivity to prolonged monocular deprivation (MD) in rat binocular visual cortex in layer 4 and layer 2/3 pyramids (4Ps and 2/3Ps) and in thick-tufted and nontufted layer 5 pyramids (5TPs and 5NPs), which innervate different extracortical targets. In normal rats, 5TPs and 2/3Ps are the most binocular in terms of synaptic inputs, and 5NPs are the least. Spike responses of all 5TPs were highly binocular, whereas those of 2/3Ps were dominated by either the contralateral or ipsilateral eye. MD dramatically shifted the ocular preference of 2/3Ps and 4Ps, mostly by depressing deprived-eye inputs. Plasticity was profoundly different in layer 5. The subthreshold ocular preference shift was sevenfold smaller in 5TPs because of smaller depression of deprived inputs combined with a generalized loss of responsiveness, and was undetectable in 5NPs. Despite their modest ocular dominance change, spike responses of 5TPs consistently lost their typically high binocularity during MD. The comparison of MD effects on 2/3Ps and 5TPs, the main affected output cells of vertical microcircuits, indicated that subthreshold plasticity is not uniquely determined by the initial degree of input binocularity. The data raise the question of whether 5TPs are driven solely by 2/3Ps during MD. The different suprathreshold plasticity of the two cell populations could underlie distinct functional deficits in amblyopia.

  4. Long-term depression: a cell biological view

    OpenAIRE

    Sheng, Morgan; Ertürk, Ali

    2014-01-01

    Recent studies of the molecular mechanisms of long-term depression (LTD) suggest a crucial role for the signalling pathways of apoptosis (programmed cell death) in the weakening and elimination of synapses and dendritic spines. With this backdrop, we suggest that LTD can be considered as the electrophysiological aspect of a larger cell biological programme of synapse involution, which uses localized apoptotic mechanisms to sculpt synapses and circuits without causing cell death.

  5. Long-term survival in small-cell lung cancer

    DEFF Research Database (Denmark)

    Lassen, U; Osterlind, K; Hansen, M;

    1995-01-01

    PURPOSE: To describe in patients with small-cell lung cancer (SCLC) the characteristics of those who survive for > or = 5 years, to identify long-term prognostic factors, to analyze survival data of 5-year survivors, and to study 10-year survival in patients entered before 1981. PATIENTS AND METH......PURPOSE: To describe in patients with small-cell lung cancer (SCLC) the characteristics of those who survive for > or = 5 years, to identify long-term prognostic factors, to analyze survival data of 5-year survivors, and to study 10-year survival in patients entered before 1981. PATIENTS......, liver and bone marrow metastases, and elevated lactate dehydrogenase (LDH) and alkaline phosphatase levels were all negative prognostic factors. The 5-year survival rate was 3.5% (limited-stage disease, 4.8%; extensive-stage disease, 2.3%), and the 10-year survival rate was 1.8% (limited-stage disease......, especially tobacco-related cancers and other tobacco-related diseases....

  6. Type-specific cell line models for type-specific ovarian cancer research.

    Directory of Open Access Journals (Sweden)

    Michael S Anglesio

    Full Text Available BACKGROUND: OVARIAN CARCINOMAS CONSIST OF AT LEAST FIVE DISTINCT DISEASES: high-grade serous, low-grade serous, clear cell, endometrioid, and mucinous. Biomarker and molecular characterization may represent a more biologically relevant basis for grouping and treating this family of tumors, rather than site of origin. Molecular characteristics have become the new standard for clinical pathology, however development of tailored type-specific therapies is hampered by a failure of basic research to recognize that model systems used to study these diseases must also be stratified. Unrelated model systems do offer value for study of biochemical processes but specific cellular context needs to be applied to assess relevant therapeutic strategies. METHODS: We have focused on the identification of clear cell carcinoma cell line models. A panel of 32 "ovarian cancer" cell lines has been classified into histotypes using a combination of mutation profiles, IHC mutation-surrogates, and a validated immunohistochemical model. All cell lines were identity verified using STR analysis. RESULTS: Many described ovarian clear cell lines have characteristic mutations (including ARID1A and PIK3CA and an overall molecular/immuno-profile typical of primary tumors. Mutations in TP53 were present in the majority of high-grade serous cell lines. Advanced genomic analysis of bona-fide clear cell carcinoma cell lines also support copy number changes in typical biomarkers such at MET and HNF1B and a lack of any recurrent expressed re-arrangements. CONCLUSIONS: As with primary ovarian tumors, mutation status of cancer genes like ARID1A and TP53 and a general immuno-profile serve well for establishing histotype of ovarian cancer cell We describe specific biomarkers and molecular features to re-classify generic "ovarian carcinoma" cell lines into type specific categories. Our data supports the use of prototype clear cell lines, such as TOV21G and JHOC-5, and questions the use of

  7. Monitoring Astrocytic Proteome Dynamics by Cell Type-Specific Protein Labeling.

    Directory of Open Access Journals (Sweden)

    Anke Müller

    Full Text Available The ability of the nervous system to undergo long-term plasticity is based on changes in cellular and synaptic proteomes. While many studies have explored dynamic alterations in neuronal proteomes during plasticity, there has been less attention paid to the astrocytic counterpart. Indeed, progress in identifying cell type-specific proteomes is limited owing to technical difficulties. Here, we present a cell type-specific metabolic tagging technique for a mammalian coculture model based on the bioorthogonal amino acid azidonorleucine and the mutated Mus musculus methionyl-tRNA synthetaseL274G enabling azidonorleucine introduction into de novo synthesized proteins. Azidonorleucine incorporation resulted in cell type-specific protein labeling and retained neuronal or astrocytic cell viability. Furthermore, we were able to label astrocytic de novo synthesized proteins and identified both Connexin-43 and 60S ribosomal protein L10a upregulated upon treatment with Brain-derived neurotrophic factor in astrocytes of a neuron-glia coculture. Taken together, we demonstrate the successful dissociation of astrocytic from neuronal proteomes by cell type-specific metabolic labeling offering new possibilities for the analyses of cell type-specific proteome dynamics.

  8. Long-term three-dimensional perfusion culture of human adult bone marrow mononuclear cells in bioreactors.

    Science.gov (United States)

    Schmelzer, Eva; Finoli, Anthony; Nettleship, Ian; Gerlach, Jörg C

    2015-04-01

    The construction and long-term maintenance of three-dimensional in vitro bone marrow models is of great interest but still quite challenging. Here we describe the use of a multi-compartment hollow-fiber membrane based three-dimensional perfusion bioreactor for long-term culture of whole human bone marrow mononuclear cells. We also investigated bioreactors with incorporated open-porous foamed hydroxyapatite scaffolds, mimicking the in vivo bone matrix. Cells in bioreactors with and without scaffolds were cultured to 6 weeks and compared to Petri dish controls. Cells were analyzed for gene expression, surface markers by flow cytometry, metabolic activity, hematopoietic potential, viability, and attachment by immunocytochemistry. Cells in bioreactors were metabolic active during long-term culture. The percentages of hematopoietic stem cell and mature endothelial cell fractions were maintained in bioreactors. The expression of most of the analyzed genes stabilized and increased after long-term culture of 6 weeks. Compared to Petri dish culture controls, bioreactor perfusion culture improved in both the short and long-term, the colony formation unit capacity of hematopoietic progenitors. Cells attached to the ample surface area provided by hydroxyapatite scaffolds. The implementation of a hydroxyapatite scaffold did not influence colony formation capacity, percentages of cell type specific fractions, gene expression, cell viability or metabolic turnover when compared to control cells cultured in bioreactors without scaffolds. In conclusion, three-dimensional perfusion bioreactor culture enables long-term maintenance of primary human bone marrow cells, with hydroxyapatite scaffolds providing an in vivo-like scaffold for three-dimensional culture.

  9. Miniature Bioreactor System for Long-Term Cell Culture

    Science.gov (United States)

    Gonda, Steve R.; Kleis, Stanley J.; Geffert, Sandara K.

    2010-01-01

    A prototype miniature bioreactor system is designed to serve as a laboratory benchtop cell-culturing system that minimizes the need for relatively expensive equipment and reagents and can be operated under computer control, thereby reducing the time and effort required of human investigators and reducing uncertainty in results. The system includes a bioreactor, a fluid-handling subsystem, a chamber wherein the bioreactor is maintained in a controlled atmosphere at a controlled temperature, and associated control subsystems. The system can be used to culture both anchorage-dependent and suspension cells, which can be either prokaryotic or eukaryotic. Cells can be cultured for extended periods of time in this system, and samples of cells can be extracted and analyzed at specified intervals. By integrating this system with one or more microanalytical instrument(s), one can construct a complete automated analytical system that can be tailored to perform one or more of a large variety of assays.

  10. Immune reactivity of cells from long-term rat renal allograft survivors

    Energy Technology Data Exchange (ETDEWEB)

    Weiss, A.; Stuart, F.P.; Fitch, F.W.

    1978-11-01

    Lewis rats receiving an LBN kidney allograft demonstrate no signs of rejection if they are pretreated with donor spleen cells and antiserum reactive with the donor alloantigen. We examined the cellular reactivity of long-term kidney allograft survivors. Normal proliferative and cytolytic responses were obtained with spleen cells from long-term survivors, in marked contrast to the diminished responses of cells from neonatally tolerant rats or the heightened cytolytic response of cells from rats that had rejected a renal allograft. Serum from long-term renal allograft survivors as well as serum obtained from rats at the time of transplantation did not suppress proliferative or cytolytic responses of normal cells. The results of this study suggest that long-term renal allograft survivors possess the precursors of those cells which are responsible for proliferative and cytolytic responses in mixed leukocyte cultures, but that they have not been sensitized to their renal allograft.

  11. A Sealed Preparation for Long-Term Observations of Cultured Cells

    OpenAIRE

    sprotocols

    2014-01-01

    Authors: Greenfield Sluder, Joshua J. Nordberg, Frederick J. Miller and Edward H. Hinchcliffe This protocol was adapted from “A Sealed Preparation for Long-Term Observations of Cultured Cells,” Chapter 18, in *Live Cell Imaging* (eds. Goldman and Spector). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2005. ### INTRODUCTION The continuous long-term observation of cultured cells on the microscope has always been a technically demanding undertaking. This protocol...

  12. Hippocampal Place Cell Firing Patterns can Induce Long-Term Synaptic Plasticity In Vitro

    OpenAIRE

    Isaac, John T. R.; Buchanan, Katherine A.; Muller, Robert U.; Mellor, Jack R.

    2009-01-01

    In the hippocampus, synaptic strength between pyramidal cells is modifiable by NMDA receptor (NMDAR)-dependent long-term potentiation (LTP) and long-term depression (LTD), both of which require coincident pre- and postsynaptic activity. In vivo, many pyramidal cells exhibit location-specific activity patterns and are known as “place cells”. The combination of these factors suggests that synaptic plasticity will be induced at synapses connecting place cells with overlapping firing fields, sinc...

  13. Long-Term Performance of Uranium Tailings Disposal Cells - 13340

    Energy Technology Data Exchange (ETDEWEB)

    Bostick, Kent; Daniel, Anamary; Pill, Ken [Professional Project Services, Inc., 1100 Bethel Valley Road, Oak Ridge, TN, 37922 (United States); Tachiev, Georgio; Noosai, Nantaporn; Villamizar, Viviana [Florida International University, 10555 W. Flagler St., EC 2100, Miami FL, 33174 (United States)

    2013-07-01

    Recently, there has been interest in the performance and evolution of Uranium Mill Tailings Remedial Action (UMTRA) Project disposal cell covers because some sites are not compliant with groundwater standards. Field observations of UMTRA disposal cells indicate that rock covers tend to become vegetated and that saturated conductivities in the upper portion of radon barriers may increase due to freeze/thaw cycles and biointrusion. This paper describes the results of modeling that addresses whether these potential changes and transient drainage of moisture in the tailings affect overall performance of the disposal cells. A numerical unsaturated/saturated 3-dimensional flow model was used to simulate whether increases in saturated conductivities in radon barriers with rock covers affect the overall performance of the disposal cells using field data from the Shiprock, NM, UMTRA site. A unique modeling approach allowed simulation with daily climatic conditions to determine changes in moisture and moisture flux from the disposal cell. Modeling results indicated that increases in the saturated conductivity at the top of radon barrier do not influence flux from the tailings with time because the tailings behave similar hydraulically to the radon barrier. The presence of a thin layer of low conductivity material anywhere in the cover or tailings restricts flux in the worst case to the saturated conductivity of that material. Where materials are unsaturated at depth within the radon barrier of tailings slimes, conductivities are typically less than 10{sup -8} centimeters per second. If the low conductivity layer is deep within the disposal cell, its saturated properties are less likely to change with time. The significance of this modeling is that operation and maintenance of the disposal cells can be minimized if they are allowed to progress to a natural condition with some vegetation and soil genesis. Because the covers and underlying tailings have a very low saturated

  14. GABABR-Dependent Long-Term Depression at Hippocampal Synapses between CB1-Positive Interneurons and CA1 Pyramidal Cells.

    Science.gov (United States)

    Jappy, Dave; Valiullina, Fliza; Draguhn, Andreas; Rozov, Andrei

    2016-01-01

    Activity induced long lasting modifications of synaptic efficacy have been extensively studied in excitatory synapses, however, long term plasticity is also a property of inhibitory synapses. Inhibitory neurons in the hippocampal CA1 region can be subdivided according to the compartment they target on the pyramidal cell. Some interneurons preferentially innervate the perisomatic area and axon hillock of the pyramidal cells while others preferentially target dendritic branches and spines. Another characteristic feature allowing functional classification of interneurons is cell type specific expression of different neurochemical markers and receptors. In the hippocampal CA1 region, nearly 90% of the interneurons expressing cannabinoid type 1 receptors (CB1R) also express cholecystokinin (CCK). Therefore, the functional presence of CB1 receptors can be used for identification of the inhibitory input from CCK positive (CCK+) interneurons to CA1 pyramidal cells. The goal of this study was to explore the nature of long term plasticity at the synapses between interneurons expressing CB1Rs (putative CCK+) and pyramidal neurons in the CA1 region of the hippocampus in vitro. We found that theta burst stimulation triggered robust long-term depression (LTD) at this synapse. The locus of LTD induction was postsynaptic and required activation of GABAB receptors. We also showed that LTD at this synaptic connection involves GABABR-dependent suppression of adenylyl cyclase and consequent reduction of PKA activity. In this respect, CB1+ to pyramidal cell synapses differ from the majority of the other hippocampal inhibitory connections where theta burst stimulation results in long-term potentiation. PMID:26858602

  15. Method and Apparatus for a Miniature Bioreactor System for Long-Term Cell Culture

    Science.gov (United States)

    Kleis, Stanley J. (Inventor); Geffert, Sandra K. (Inventor); Gonda, Steve R. (Inventor)

    2015-01-01

    A bioreactor and method that permits continuous and simultaneous short, moderate, or long term cell culturing of one or more cell types or tissue in a laminar flow configuration is disclosed, where the bioreactor supports at least two laminar flow zones, which are isolated by laminar flow without the need for physical barriers between the zones. The bioreactors of this invention are ideally suited for studying short, moderate and long term studies of cell cultures and the response of cell cultures to one or more stressors such as pharmaceuticals, hypoxia, pathogens, or any other stressor. The bioreactors of this invention are also ideally suited for short, moderate or long term cell culturing with periodic cell harvesting and/or medium processing for secreted cellular components.

  16. Ni/YSZ microstructure optimization for long-term stability of solid oxide electrolysis cells

    DEFF Research Database (Denmark)

    Hauch, Anne; Brodersen, Karen; Karas, Filip;

    2014-01-01

    In the last decade there has been a renewed and increased interest in electrolysis using solid oxide cells (SOC). So far the vast majority of results reported on long-term durability of solid oxide electrolysis cells (SOEC) have been obtained using SOC produced and optimized for fuel cell operation......; i.e. solid oxide fuel cells (SOFC). However, previous long-term tests have shown that the stability behavior of the Ni/yttria-stabilized-zirconia (Ni/YSZ) fuel electrode may fall out quite differently depending on whether the cell is operated in fuel cell or electrolysis mode at otherwise similar...... test conditions. Initial work has shown significant microstructural changes of the Ni/YSZ electrode close to the electrolyte interface after long-term steam electrolysis test at -1 A/cm2 at 800 C. The results indicate that it will be advantageous to optimize the electrode structure with the aim...

  17. Tracking genetically engineered lymphocytes long-term reveals the dynamics of T cell immunological memory.

    Science.gov (United States)

    Oliveira, Giacomo; Ruggiero, Eliana; Stanghellini, Maria Teresa Lupo; Cieri, Nicoletta; D'Agostino, Mattia; D'Agostino, Mattio; Fronza, Raffaele; Lulay, Christina; Dionisio, Francesca; Mastaglio, Sara; Greco, Raffaella; Peccatori, Jacopo; Aiuti, Alessandro; Ambrosi, Alessandro; Biasco, Luca; Bondanza, Attilio; Lambiase, Antonio; Traversari, Catia; Vago, Luca; von Kalle, Christof; Schmidt, Manfred; Bordignon, Claudio; Ciceri, Fabio; Bonini, Chiara

    2015-12-01

    Long-lasting immune protection from pathogens and cancer requires the generation of memory T cells able to survive long-term. To unravel the immunological requirements for long-term persistence of human memory T cells, we characterized and traced, over several years, T lymphocytes genetically modified to express the thymidine kinase (TK) suicide gene that were infused in 10 patients after haploidentical hematopoietic stem cell transplantation (HSCT). At 2 to 14 years after infusion and in the presence of a broad and resting immune system, we could still detect effectors/effector memory (TEM/EFF), central memory (TCM), and stem memory (TSCM) TK(+) cells, circulating at low but stable levels in all patients. Longitudinal analysis of cytomegalovirus (CMV)- and Flu-specific TK(+) cells indicated that antigen recognition was dominant in driving in vivo expansion and persistence at detectable levels. The amount of infused TSCM cells positively correlated with early expansion and with the absolute counts of long-term persisting gene-marked cells. By combining T cell sorting with sequencing of integration (IS), TCRα and TCRβ clonal markers, we showed that T cells retrieved long-term were enriched in clones originally shared in different memory T cell subsets, whereas dominant long-term clonotypes appeared to preferentially originate from infused TSCM and TCM clones. Together, these results indicate that long-term persistence of gene-modified memory T cells after haploidentical HSCT is influenced by antigen exposure and by the original phenotype of infused cells. Cancer adoptive immunotherapy might thus benefit from cellular products enriched in lymphocytes with an early-differentiated phenotype. PMID:26659572

  18. GATA-3 REGULATES THE SELF-RENEWAL OF LONG-TERM HEMATOPOIETIC STEM CELLS

    Science.gov (United States)

    Frelin, Catherine; Herrington, Robert; Janmohamed, Salima; Barbara, Mary; Tran, Gary; Paige, Christopher J.; Benveniste, Patricia; Zuñiga-Pflücker, Juan-Carlos; Souabni, Abdallah; Busslinger, Meinrad; Iscove, Norman N

    2016-01-01

    Gata3 is expressed and required for differentiation and function throughout the T lymphocyte lineage. Despite evidence it may also be expressed in multipotent hematopoietic stem cells (HSC), any role in these cells has remained unclear. Here we show GATA3 was cytoplasmic in quiescent long-term stem cells from steady state bone marrow, but relocated to the nucleus when HSC cycle. Relocation depended on p38-MAPK signaling and was associated with diminished capacity for long-term reconstitution upon transfer to irradiated mice. Deletion of Gata3 enhanced repopulating capacity and augmented self-renewal of long term HSC in cell-autonomous fashion, without affecting cell cycle. These observations position Gata3 as a regulator of the balance between self-renewal and differentiation in HSC acting downstream of the p38 signaling pathway. PMID:23974957

  19. Stevioside counteracts the alpha-cell hypersecretion caused by long-term palmitate exposure

    DEFF Research Database (Denmark)

    Hong, J; Chen, L; Jeppesen, P B;

    2006-01-01

    Long-term exposure to fatty acids impairs beta-cell function in type 2 diabetes, but little is known about the chronic effects of fatty acids on alpha-cells. We therefore studied the prolonged impact of palmitate on alpha-cell function and on the expression of genes related to fuel metabolism. We...

  20. Long-term maintenance of in vitro cultured honeybee (Apis mellifera embryonic cells

    Directory of Open Access Journals (Sweden)

    Aamodt Randi M

    2006-03-01

    Full Text Available Abstract Background In vitro cultivation of cells allows novel investigation of in vivo- mechanisms and is a helpful tool in developmental biology, biochemistry and functional genomics. Numerous cell lines of insect species, e.g., silkworm and mosquito, have been reported. However, this is not the case for successful long-term cultivation of cells in honeybees. Results Methods for cultivation of honeybee embryonic cells are discussed here. Pre-gastrula stage embryos were used to initiate cultures, and cells were reared on 96-wells microplates with Grace insect medium, supplemented with Fetal Bovine Serum. Cells proliferated in clusters, and maintained viable and mitotic for more than three months. Conclusion We report here, for the first time, long-term cultivation of honeybee cells. Results represent a highly useful in vitro-system for studying a model organism of increasing importance in areas such as aging, sociality and neurobiology.

  1. Human breast microvascular endothelial cells retain phenotypic traits in long-term finite life span culture

    DEFF Research Database (Denmark)

    Sigurdsson, Valgardur; Fridriksdottir, Agla J R; Kjartansson, Jens;

    2007-01-01

    Attempts to study endothelial-epithelial interactions in the human breast have been hampered by lack of protocols for long-term cultivation of breast endothelial cells (BRENCs). The aim of this study was to establish long-term cultures of BRENCs and to compare their phenotypic traits with the tis......Attempts to study endothelial-epithelial interactions in the human breast have been hampered by lack of protocols for long-term cultivation of breast endothelial cells (BRENCs). The aim of this study was to establish long-term cultures of BRENCs and to compare their phenotypic traits...... with the tissue of origin. Microvasculature was localized in situ by immunohistochemistry in breast samples. From this tissue, collagen-rich stroma and adipose tissue were dissected mechanically and further disaggregated to release microvessel organoids. BRENCs were cultured from these organoids in endothelial......-galactosidase staining. We demonstrate here that breast microvasculature may serve as a large-scale source for expansion of BRENCs with molecular and functional traits preserved. These cells will form the basis for studies on the role of endothelial cells in breast morphogenesis....

  2. Long-term outcome of Hurler syndrome patients after hematopoietic cell transplantation : an international multicenter study

    NARCIS (Netherlands)

    Aldenhoven, Mieke; Wynn, Robert F.; Orchard, Paul J.; O'Meara, Anne; Veys, Paul; Fischer, Alain; Valayannopoulos, Vassili; Neven, Benedicte; Rovelli, Attilio; Prasad, Vinod K.; Tolar, Jakub; Allewelt, Heather; Jones, Simon A.; Parini, Rossella; Renard, Marleen; Bordon, Victoria; Wulffraat, Nico M.; de Koning, Tom J.; Shapiro, Elsa G.; Kurtzberg, Joanne; Boelens, Jaap Jan

    2015-01-01

    Mucopolysaccharidosis type I-Hurler syndrome (MPS-IH) is a lysosomal storage disease characterized by multisystem morbidity and death in early childhood. Although hematopoietic cell transplantation (HCT) has been performed in these patients for more than 30 years, large studies on the long-term outc

  3. Long-term outcome of Hurler syndrome patients after hematopoietic cell transplantation : An international multicenter study

    NARCIS (Netherlands)

    Aldenhoven, Mieke; Wynn, Robert F.; Orchard, Paul J.; O'Meara, Anne; Veys, Paul; Fischer, Alain; Valayannopoulos, Vassili; Neven, Benedicte; Rovelli, Attilio; Prasad, Vinod K.; Tolar, Jakub; Allewelt, Heather; Jones, Simon A.; Parini, Rossella; Renard, Marleen; Bordon, Victoria; Wulffraat, Nico M.; de Koning, Tom J.; Shapiro, Elsa G.; Kurtzberg, Joanne; Boelens, Jaap Jan

    2015-01-01

    Mucopolysaccharidosis type I-Hurler syndrome (MPS-IH) is a lysosomal storage disease characterized by multisystem morbidity and death in early childhood. Although hematopoietic cell transplantation (HCT) has been performed in these patients for more than 30 years, large studies on the long-term outc

  4. Intranasal mesenchymal stem cell treatment for neonatal brain damage : long-term cognitive and sensorimotor improvement

    NARCIS (Netherlands)

    Donega, Vanessa; van Velthoven, Cindy T J; Nijboer, Cora H; van Bel, Frank; Kas, Martien J H; Kavelaars, Annemieke; Heijnen, Cobi J

    2013-01-01

    Mesenchymal stem cell (MSC) administration via the intranasal route could become an effective therapy to treat neonatal hypoxic-ischemic (HI) brain damage. We analyzed long-term effects of intranasal MSC treatment on lesion size, sensorimotor and cognitive behavior, and determined the therapeutic wi

  5. Long-term surveillance plan for the Burro Canyon disposal cell Slick Rock, Colorado

    International Nuclear Information System (INIS)

    This long-term surveillance plan (LTSP) describes the U.S. Department of Energy (DOE) long-term care program for the Uranium Mill Tailings Remedial Action (UMTRA) Project Burro Canyon disposal cell in San Miguel County, Colorado. The U.S. Nuclear Regulatory Commission (NRC) developed regulations for the issuance of a general license for the custody and long-term care of UMTRA Project disposal sites in 10 CFR Part 40. The purpose of this general license is to ensure that the UMTRA Project disposal sites are cared for in a manner that protects the public health and safety and the environment. Before each disposal site is licensed, the NRC requires the DOE to submit a site-specific LTSP. The DOE prepared this LTSP to meet this requirement for the Burro Canyon disposal cell. The general license becomes effective when the NRC concurs with the DOE's determination that remedial action is complete at the Burro Canyon disposal cell and the NRC formally accepts this LTSP. Attachment 1 contains the concurrence letters from NRC. This LTSP describes the long-term surveillance program the DOE has implemented to ensure that the Burro Canyon disposal cell performs as designed. The program is based on site inspections to identify threats to disposal cell integrity. Ground water monitoring will not be required at the Burro Canyon disposal cell because the ground water protection strategy is supplemental standards based on low yield from the uppermost aquifer. The LTSP is based on the UMTRA Project's long-term surveillance program guidance and meets the requirements of 10 CFR 40.27(b) and 40 CFR 192.03

  6. Phenotypic and Functional Characterization of Long-Term Cryopreserved Human Adipose-derived Stem Cells

    OpenAIRE

    Yong, Kar Wey; Pingguan-Murphy, Belinda; Xu, Feng; Abas, Wan Abu Bakar Wan; Choi, Jane Ru; Omar, Siti Zawiah; Azmi, Mat Adenan Noor; Chua, Kien Hui; Safwani, Wan Kamarul Zaman Wan

    2015-01-01

    Cryopreservation represents an effective technique to maintain the functional properties of human adipose-derived stem cells (ASCs) and allows pooling of cells via long-term storage for clinical applications, e.g., cell-based therapies. It is crucial to reduce freezing injury during the cryopreservation process by loading the ASCs with the optimum concentration of suitable cryoprotective agents (CPAs). In this study, human ASCs were preserved for 3 months in different combinations of CPAs, in...

  7. Freedom of expression: cell-type-specific gene profiling.

    Science.gov (United States)

    Otsuki, Leo; Cheetham, Seth W; Brand, Andrea H

    2014-01-01

    Cell fate and behavior are results of differential gene regulation, making techniques to profile gene expression in specific cell types highly desirable. Many methods now enable investigation at the DNA, RNA and protein level. This review introduces the most recent and popular techniques, and discusses key issues influencing the choice between these such as ease, cost and applicability of information gained. Interdisciplinary collaborations will no doubt contribute further advances, including not just in single cell type but single-cell expression profiling.

  8. Cell-type specific four-component hydrogel.

    Science.gov (United States)

    Aberle, Timo; Franke, Katrin; Rist, Elke; Benz, Karin; Schlosshauer, Burkhard

    2014-01-01

    In the field of regenerative medicine we aim to develop implant matrices for specific tissue needs. By combining two per se, cell-permissive gel systems with enzymatic crosslinkers (gelatin/transglutaminase and fibrinogen/thrombin) to generate a blend (technical term: quattroGel), an unexpected cell-selectivity evolved. QuattroGels were porous and formed cavities in the cell diameter range, possessed gelation kinetics in the minute range, viscoelastic properties and a mechanical strength appropriate for general cell adhesion, and restricted diffusion. Cell proliferation of endothelial cells, chondrocytes and fibroblasts was essentially unaffected. In contrast, on quattroGels neither endothelial cells formed vascular tubes nor did primary neurons extend neurites in significant amounts. Only chondrocytes differentiated properly as judged by collagen isoform expression. The biophysical quattroGel characteristics appeared to leave distinct cell processes such as mitosis unaffected and favored differentiation of sessile cells, but hampered differentiation of migratory cells. This cell-type selectivity is of interest e.g. during articular cartilage or invertebral disc repair, where pathological innervation and angiogenesis represent adverse events in tissue engineering. PMID:24475174

  9. Cell-type specific four-component hydrogel.

    Directory of Open Access Journals (Sweden)

    Timo Aberle

    Full Text Available In the field of regenerative medicine we aim to develop implant matrices for specific tissue needs. By combining two per se, cell-permissive gel systems with enzymatic crosslinkers (gelatin/transglutaminase and fibrinogen/thrombin to generate a blend (technical term: quattroGel, an unexpected cell-selectivity evolved. QuattroGels were porous and formed cavities in the cell diameter range, possessed gelation kinetics in the minute range, viscoelastic properties and a mechanical strength appropriate for general cell adhesion, and restricted diffusion. Cell proliferation of endothelial cells, chondrocytes and fibroblasts was essentially unaffected. In contrast, on quattroGels neither endothelial cells formed vascular tubes nor did primary neurons extend neurites in significant amounts. Only chondrocytes differentiated properly as judged by collagen isoform expression. The biophysical quattroGel characteristics appeared to leave distinct cell processes such as mitosis unaffected and favored differentiation of sessile cells, but hampered differentiation of migratory cells. This cell-type selectivity is of interest e.g. during articular cartilage or invertebral disc repair, where pathological innervation and angiogenesis represent adverse events in tissue engineering.

  10. Cell-Type Specific Four-Component Hydrogel

    OpenAIRE

    Timo Aberle; Katrin Franke; Elke Rist; Karin Benz; Burkhard Schlosshauer

    2014-01-01

    In the field of regenerative medicine we aim to develop implant matrices for specific tissue needs. By combining two per se, cell-permissive gel systems with enzymatic crosslinkers (gelatin/transglutaminase and fibrinogen/thrombin) to generate a blend (technical term: quattroGel), an unexpected cell-selectivity evolved. QuattroGels were porous and formed cavities in the cell diameter range, possessed gelation kinetics in the minute range, viscoelastic properties and a mechanical strength appr...

  11. Long-term outcome of Hurler syndrome patients after hematopoietic cell transplantation: an international multicenter study.

    Science.gov (United States)

    Aldenhoven, Mieke; Wynn, Robert F; Orchard, Paul J; O'Meara, Anne; Veys, Paul; Fischer, Alain; Valayannopoulos, Vassili; Neven, Benedicte; Rovelli, Attilio; Prasad, Vinod K; Tolar, Jakub; Allewelt, Heather; Jones, Simon A; Parini, Rossella; Renard, Marleen; Bordon, Victoria; Wulffraat, Nico M; de Koning, Tom J; Shapiro, Elsa G; Kurtzberg, Joanne; Boelens, Jaap Jan

    2015-03-26

    Mucopolysaccharidosis type I-Hurler syndrome (MPS-IH) is a lysosomal storage disease characterized by multisystem morbidity and death in early childhood. Although hematopoietic cell transplantation (HCT) has been performed in these patients for more than 30 years, large studies on the long-term outcome of patients with MPS-IH after HCT are lacking. The goal of this international study was to identify predictors of the long-term outcome of patients with MPS-IH after successful HCT. Two hundred seventeen patients with MPS-IH successfully engrafted with a median follow-up age of 9.2 years were included in this retrospective analysis. Primary endpoints were neurodevelopmental outcomes and growth. Secondary endpoints included neurologic, orthopedic, cardiac, respiratory, ophthalmologic, audiologic, and endocrinologic outcomes. Considerable residual disease burden was observed in the majority of the transplanted patients with MPS-IH, with high variability between patients. Preservation of cognitive function at HCT and a younger age at transplantation were major predictors for superior cognitive development posttransplant. A normal α-l-iduronidase enzyme level obtained post-HCT was another highly significant predictor for superior long-term outcome in most organ systems. The long-term prognosis of patients with MPS-IH receiving HCT can be improved by reducing the age at HCT through earlier diagnosis, as well as using exclusively noncarrier donors and achieving complete donor chimerism. PMID:25624320

  12. A random-effects model for long-term degradation analysis of solid oxide fuel cells

    International Nuclear Information System (INIS)

    Solid oxide fuel cells (SOFCs) are electrochemical devices converting the chemical energy into electricity with high efficiency and low pollutant emissions. Tough very promising, this technology is still in a developing phase, and degradation at the cell/stack level with operating time is still an issue of major concern. Methods to directly observe degradation modes and to measure their evolution over time are difficult to implement, and indirect performance indicators are adopted, typically related to voltage measurements in long-term tests. In order to describe long-term degradation tests, three components of the voltage measurements should be modelled: the smooth decay of voltage over time for each single unit; the variability of voltage decay among units; and the high-frequency small fluctuations of voltage due to experimental noise and lack of fit. In this paper, we propose an empirical random-effects regression model of polynomial type enabling to evaluate separately these three types of variability. Point and interval estimates are also derived for some performance measures, such as the mean voltage, the prediction of cell voltage, the reliability function and the cell-to-cell variability in SOFC stacks. Finally, the proposed methodology is applied to two real case-studies of long-term degradation tests of SOFC stacks. - Highlights: • We propose an empirical random-effects model for SOFC cells voltage in long runs. • Some SOFC performance and manufacturing quality measures are derived. • An application to two real case-studies of long-term degradation tests is provided. • The reliability function of SOFCs and its lower confidence limit are computed

  13. Cell search algorithms for the 3G long-term evolution

    Institute of Scientific and Technical Information of China (English)

    SU Huan; ZHANG Jian-hua

    2007-01-01

    This article presents downlink initial synchronization and cell identification algorithms for long term evolution (LTE) of third-generation (3G) mobile communication systems, which are based on synchronization channel (SCH) and cell specific pilot symbols, respectively. The key features of the proposed scheme are: it can improve performance of the frequency synchronization through oversampling of the SCH, it can support a large number of target cells by modulating a cell-specific pilot sequence over two symbols within a subframe, and it can guarantee cell identification performance by maximally ratio combining the frequency domain differential cross-correlation. Simulations show that the proposed scheme has a potential use in 3G LTE.

  14. Investigating Striatal Function through Cell-Type-Specific Manipulations

    OpenAIRE

    Kreitzer, Anatol C.; Berke, Joshua D.

    2011-01-01

    The striatum integrates convergent input from the cortex, thalamus, and midbrain, and has a powerful influence over motivated behavior via outputs to downstream basal ganglia nuclei. Although the anatomy and physiology of distinct classes of striatal neurons has been intensively studied, the specific functions of these cell subpopulations have been more difficult to address. Recently, application of new methodologies for perturbing activity and signaling in different cell types in vivo has be...

  15. Cell type-specific neuroprotective activity of untranslocated prion protein.

    Directory of Open Access Journals (Sweden)

    Elena Restelli

    Full Text Available BACKGROUND: A key pathogenic role in prion diseases was proposed for a cytosolic form of the prion protein (PrP. However, it is not clear how cytosolic PrP localization influences neuronal viability, with either cytotoxic or anti-apoptotic effects reported in different studies. The cellular mechanism by which PrP is delivered to the cytosol of neurons is also debated, and either retrograde transport from the endoplasmic reticulum or inefficient translocation during biosynthesis has been proposed. We investigated cytosolic PrP biogenesis and effect on cell viability in primary neuronal cultures from different mouse brain regions. PRINCIPAL FINDINGS: Mild proteasome inhibition induced accumulation of an untranslocated form of cytosolic PrP in cortical and hippocampal cells, but not in cerebellar granules. A cyclopeptolide that interferes with the correct insertion of the PrP signal sequence into the translocon increased the amount of untranslocated PrP in cortical and hippocampal cells, and induced its synthesis in cerebellar neurons. Untranslocated PrP boosted the resistance of cortical and hippocampal neurons to apoptotic insults but had no effect on cerebellar cells. SIGNIFICANCE: These results indicate cell type-dependent differences in the efficiency of PrP translocation, and argue that cytosolic PrP targeting might serve a physiological neuroprotective function.

  16. Long-term perturbation of hemopoiesis after moderate damage to stem cells

    International Nuclear Information System (INIS)

    Hematological indices of BDF1 mice characterizing regeneration in a complex fashion (peripheral cell counts, bone marrow CFU-S, GM-CFU, CFU-F, and the turnover state of bone marrow CFU-S) were followed periodically for 11-14 weeks after 0.5 Gy acute gamma irradiation in order to obtain detailed information about the nonsteady state of murine hemopoiesis after moderate damage to stem cells, demonstrated in previous experiments. Virtually complete hematological regeneration was followed by long-term increase in the proliferation rate of CFU-S, oscillation in some parameters (granulocyte count, CFU-S level) or an overshooting, sometimes multiwavelike regeneration in others (reticulocyte count, thrombocyte count, and GM-CFU and CFU-F levels). These data suggest a long-term perturbation of hemopoiesis and its environment after moderate damage to the bone marrow

  17. Long-term quantitative phase-contrast imaging of living cells by digital holographic microscopy

    Science.gov (United States)

    Liu, S.; Pan, F.; Wang, Z.; Wang, F.; Rong, L.; Shang, P.; Xiao, W.

    2011-04-01

    The dynamic analysis of biological living samples is one of the particular interests in life sciences. An improved digital holographic microscope for long-term quantitative phase-contrast imaging of living cells is presented in this paper. The optical configuration is optimized in the form of a free-space-fiber hybrid system which promotes the flexibility of imaging in complex or semi-enclosed experimental environment. Aberrations compensation is implemented taking into account the additional phase aberration induced by liquid culture medium in long-term observation. The proposed approach is applied to investigate living samples of MC3T3-E1 and MLO-Y4 cells. The experimental results demonstrate its availability in the analysis of cellular changes.

  18. Long-term proliferation in culture and germline transmission of mouse male germline stem cells.

    Science.gov (United States)

    Kanatsu-Shinohara, Mito; Ogonuki, Narumi; Inoue, Kimiko; Miki, Hiromi; Ogura, Atsuo; Toyokuni, Shinya; Shinohara, Takashi

    2003-08-01

    Spermatogenesis is a complex process that originates in a small population of spermatogonial stem cells. Here we report the in vitro culture of spermatogonial stem cells that proliferate for long periods of time. In the presence of glial cell line-derived neurotrophic factor, epidermal growth factor, basic fibroblast growth factor, and leukemia inhibitory factor, gonocytes isolated from neonatal mouse testis proliferated over a 5-month period (>10(14)-fold) and restored fertility to congenitally infertile recipient mice following transplantation into seminiferous tubules. Long-term spermatogonial stem cell culture will be useful for studying spermatogenesis mechanism and has important implications for developing new technology in transgenesis or medicine.

  19. Long-term live cell microscopy studies of lipid droplet fusion dynamics in adipocytes[S

    OpenAIRE

    Jüngst, Christian; Klein, Matthias; Zumbusch, Andreas

    2013-01-01

    During the adipogenic differentiation process of mesenchymal stem cells, lipid droplets (LDs) grow slowly by transferring lipids between each other. Recent findings hint at the possibility that a fusion pore is involved. In this study, we analyze lipid transfer data obtained in long-term label-free microscopy studies in the framework of a Hagen-Poiseuille model. The data obtained show a LD fusion process in which the lipid transfer directionality depends on the size difference between LDs, wh...

  20. Cell Type-Specific Modulation of Respiratory Chain Supercomplex Organization.

    Science.gov (United States)

    Sun, Dayan; Li, Bin; Qiu, Ruyi; Fang, Hezhi; Lyu, Jianxin

    2016-01-01

    Respiratory chain complexes are organized into large supercomplexes among which supercomplex In + IIIn + IVn is the only one that can directly transfer electrons from NADH to oxygen. Recently, it was reported that the formation of supercomplex In + IIIn + IVn in mice largely depends on their genetic background. However, in this study, we showed that the composition of supercomplex In + IIIn + IVn is well conserved in various mouse and human cell lines. Strikingly, we found that a minimal supercomplex In + IIIn, termed "lowest supercomplex" (LSC) in this study because of its migration at the lowest position close to complex V dimers in blue native polyacrylamide gel electrophoresis, was associated with complex IV to form a supercomplex In + IIIn + IVn in some, but not all of the human and mouse cells. In addition, we observed that the 3697G>A mutation in mitochondrial-encoded NADH dehydrogenase 1 (ND1) in one patient with Leigh's disease specifically affected the assembly of supercomplex In + IIIn + IVn containing LSC, leading to decreased cellular respiration and ATP generation. In conclusion, we showed the existence of LSC In + IIIn + IVn and impairment of this supercomplex causes disease. PMID:27338358

  1. Immune responsiveness and incidence of reticulum cell sarcoma in long-term syngeneic radiation chimeras

    International Nuclear Information System (INIS)

    Long-term syngeneic radiation chimeras displayed a very low incidence of reticulum cell sarcoma as compared with control mice. Immune reactivity of these animals was studied in vivo by anti-dinitrophenyl antibody titer and affinity and in vitro by mitotic responsiveness to phytohemagglutinin, concanavalin A and lipopolysaccharide. Antibody titer and affinity as well as the response to T lectins were found to be increased in chimeras. These results were attributed to increased function of mature T2 cells, which could explain the reduced incidence of reticulum cell sarcoma in chimeras

  2. CD34 expression on long-term repopulating hematopoietic stem cells changes during developmental stages.

    Science.gov (United States)

    Matsuoka, S; Ebihara, Y; Xu, M; Ishii, T; Sugiyama, D; Yoshino, H; Ueda, T; Manabe, A; Tanaka, R; Ikeda, Y; Nakahata, T; Tsuji, K

    2001-01-15

    The CD34 antigen serves as an important marker for primitive hematopoietic cells in therapeutic transplantation of hematopoietic stem cells (HSC) and gene therapy, but it has remained an open question as to whether or not most HSC express CD34. Using a competitive long-term reconstitution assay, the results of this study confirm developmental changes in CD34 expression on murine HSC. In fetuses and neonates, CD34 was expressed on Lin(-)c-Kit(+) long-term repopulating HSC of bone marrow (BM), liver, and spleen. However, CD34 expression on HSC decreased with aging, and in mice older than 10 weeks, HSC were most enriched in the Lin(-)c-Kit(+)CD34(-) marrow cell fraction. A second transplantation was performed from primary recipients who were transplanted with neonatal Lin(-)c-Kit(+) CD34(high) HSC marrow. Although donor-type HSC resided in CD34-expressing cell fraction in BM cells of the first recipients 4 weeks after the first transplantation, the stem cell activity had shifted to Lin(-)c-Kit(+)CD34(-) cells after 16 weeks, indicating that adult Lin(-)c-Kit(+)CD34(-) HSC are the progeny of neonatal CD34-expresssing HSC. Assays for colony-forming cells showed that hematopoietic progenitor cells, unlike HSC, continue to express CD34 throughout murine development. The present findings are important because the clinical application of HSC can be extended, in particular as related to CD34-enriched HSC and umbilical cord blood HSC.

  3. Characterization of vascular smooth muscle cell phenotype in long-term culture.

    Science.gov (United States)

    Absher, M; Woodcock-Mitchell, J; Mitchell, J; Baldor, L; Low, R; Warshaw, D

    1989-02-01

    Studies of bovine carotid artery smooth muscle cells, during long-term in vitro subcultivation (up to 100 population doublings), have revealed phenotypic heterogeneity among cells, as characterized by differences in proliferative behavior, cell morphology, and contractile-cytoskeletal protein profiles. In vivo, smooth muscle cells were spindle-shaped and expressed desmin and alpha-smooth muscle actin (50% of total actin) as their predominant cytoskeletal and contractile proteins. Within 24 h of culture, vimentin rather than desmin was the predominant intermediate filament protein, with little change in alpha-actin content. Upon initial subcultivation, all cells were flattened and fibroblastic in appearance with a concomitant fivefold reduction in alpha-actin content, whereas the beta and gamma nonmuscle actins predominated. In three out of four cell lines studied, fluctuations in proliferative activity were observed during the life span of the culture. These spontaneous fluctuations in proliferation were accompanied by coordinated changes in morphology and contractile-cytoskeletal protein profiles. During periods of enhanced proliferation a significant proportion of cells reverted to their original spindle-shaped morphology with a simultaneous increase in alpha-actin content (20 to 30% of total actin). These results suggest that in long-term culture smooth muscle cells undergo spontaneous modulations in cell phenotype and may serve as a useful model for studying the regulation of intracellular protein expression.

  4. Long term maintenance of myeloid leukemic stem cells cultured with unrelated human mesenchymal stromal cells

    Directory of Open Access Journals (Sweden)

    Sawa Ito

    2015-01-01

    Full Text Available Mesenchymal stromal cells (MSCs support the growth and differentiation of normal hematopoietic stem cells (HSCs. Here we studied the ability of MSCs to support the growth and survival of leukemic stem cells (LSCs in vitro. Primary leukemic blasts isolated from the peripheral blood of 8 patients with acute myeloid leukemia (AML were co-cultured with equal numbers of irradiated MSCs derived from unrelated donor bone marrow, with or without cytokines for up to 6 weeks. Four samples showed CD34+CD38− predominance, and four were predominantly CD34+CD38+. CD34+ CD38− predominant leukemia cells maintained the CD34+ CD38− phenotype and were viable for 6 weeks when co-cultured with MSCs compared to co-cultures with cytokines or medium only, which showed rapid differentiation and loss of the LSC phenotype. In contrast, CD34+ CD38+ predominant leukemic cells maintained the CD34+CD38+ phenotype when co-cultured with MSCs alone, but no culture conditions supported survival beyond 4 weeks. Cell cycle analysis showed that MSCs maintained a higher proportion of CD34+ blasts in G0 than leukemic cells cultured with cytokines. AML blasts maintained in culture with MSCs for up to 6 weeks engrafted NSG mice with the same efficiency as their non-cultured counterparts, and the original karyotype persisted after co-culture. Chemosensitivity and transwell assays suggest that MSCs provide pro-survival benefits to leukemic blasts through cell–cell contact. We conclude that MSCs support long-term maintenance of LSCs in vitro. This simple and inexpensive approach will facilitate basic investigation of LSCs and enable screening of novel therapeutic agents targeting LSCs.

  5. Autonomous Inter Cell Interference Avoidance under Fractional Load for Downlink Long Term Evolution

    DEFF Research Database (Denmark)

    Kumar, S.; Monghal, Guillaume Damien; Nin, Jaume;

    2009-01-01

    avoidance schemes under fractional load (FL) conditions in the downlink for 3rd generation partnership project (3GPP) long term evolution (LTE) are proposed. The proposed schemes do not require any inter-cell signaling for ICIC; rather the decision about the allocation of the spectrum in order to avoid......The main source of interference in OFDMA system in downlink is inter-cell interference, which can severely limit the throughput of users near the cell edge. The inter-cell interference coordination (ICIC) is one method to improve the performance. In this paper autonomous inter-cell interference...... the inter- cell interference is taken based on the information available within the cell itself. We show that the schemes for spectral resource selection is important for FL scenario to avoid high BLER. The proposed schemes further improve the SINR condition therefore higher cell throughput and coverage...

  6. Fetal hepatic progenitors support long-term expansion of hematopoietic stem cells.

    Science.gov (United States)

    Chou, Song; Flygare, Johan; Lodish, Harvey F

    2013-05-01

    We have developed a coculture system that establishes DLK(+) fetal hepatic progenitors as the authentic supportive cells for expansion of hematopoietic stem (HSCs) and progenitor cells. In 1-week cultures supplemented with serum and supportive cytokines, both cocultured DLK(+) fetal hepatic progenitors and their conditioned medium supported rapid expansion of hematopoietic progenitors and a small increase in HSC numbers. In 2- and 3-week cultures DLK(+) cells, but not their conditioned medium, continuously and significantly (>20-fold) expanded both hematopoietic stem and progenitor cells. Physical contact between HSCs and DLK(+) cells was crucial to maintaining this long-term expansion. Similar HSC expansion (approximately sevenfold) was achieved in cocultures using a serum-free, low cytokine- containing medium. In contrast, DLK(-) cells are incapable of expanding hematopoietic cells, demonstrating that hepatic progenitors are the principle supportive cells for HSC expansion in the fetal liver.

  7. B-cell Lymphoma in retrieved femoral heads: a long term follow up

    Directory of Open Access Journals (Sweden)

    van Kemenade Folkert J

    2009-05-01

    Full Text Available Abstract Background A relatively high incidence of pathological conditions in retrieved femoral heads, including a group of patients having low grade B-cell lymphoma, has been described before. At short term follow up none of these patients with low-grade B-cell lymphoma showed evidence of systemic disease. However, the long term follow up of these patients is not known. Methods From November 1994 up to and including December 2005 we screened all femoral heads removed at the time of primary total hip replacement histopathologically and included them in the bone banking protocol according to the guidelines of the American Associations of Tissue Banks (AATB and the European Association of Musculo-Skeletal Transplantation (EAMST. We determined the percentage of B-cell lymphoma in all femoral heads and in the group that fulfilled all criteria of the bone banking protocol and report on the long-term follow-up. Results Of 852 femoral heads fourteen (1.6% were highly suspicious for low-grade B-cell lymphoma. Of these 852 femoral heads, 504 were eligible for bone transplantation according to the guidelines of the AATB and the EAMST. Six femoral heads of this group of 504 were highly suspicious for low-grade B-cell lymphoma (1.2%. At long term follow up two (0.2% of all patients developed systemic malignant disease and one of them needed medical treatment for her condition. Conclusion In routine histopathological screening we found variable numbers of low-grade B-cell lymphoma throughout the years, even in a group of femoral heads that were eligible for bone transplantation. Allogenic transmission of malignancy has not yet been reported on, but surviving viruses are proven to be transmissible. Therefore, we recommend the routine histopathological evaluation of all femoral heads removed at primary total hip arthroplasty as a tool for quality control, whether the femoral head is used for bone banking or not.

  8. Impact of process variations and long term degradation on 6T-SRAM cells

    Directory of Open Access Journals (Sweden)

    Th. Fischer

    2007-06-01

    Full Text Available In modern deep-submicron CMOS technologies voltage scaling can not keep up with the scaling of the dimensions of transistors. Therefore the electrical fields inside the transistors are not constant anymore, while scaling down the device area. The rising electrical fields bring up reliability problems, such as hot carrier injection. Also other long term degradation mechanisms like Negative Bias Temperature Instability (NBTI come into the focus of circuit design.

    Along with process device parameter variations (threshold voltage, mobility, variations due to the degradation of devices form a big challenge for designers to build circuits that both yield high under the influence of process variations and remain functional with respect to long term device drift.

    In this work we present the influence of long term degradation and process variations on the performance of SRAM core-cells and parametric yield of SRAM arrays. For different use cases we show the performance degradation depending on temperature and supply voltage.

  9. Long-Term Culture of Genome-Stable Bipotent Stem Cells from Adult Human Liver

    OpenAIRE

    Huch, Meritxell; Gehart, Helmuth; van Boxtel, Ruben; Hamer, Karien; Blokzijl, Francis; Verstegen, Monique M.A.; Ellis, Ewa; van Wenum, Martien; Fuchs, Sabine A.; de Ligt, Joep; van de Wetering, Marc; SASAKI, Nobuo; Boers, Susanne J.; Kemperman, Hans; de Jonge, Jeroen

    2015-01-01

    Summary Despite the enormous replication potential of the human liver, there are currently no culture systems available that sustain hepatocyte replication and/or function in vitro. We have shown previously that single mouse Lgr5+ liver stem cells can be expanded as epithelial organoids in vitro and can be differentiated into functional hepatocytes in vitro and in vivo. We now describe conditions allowing long-term expansion of adult bile duct-derived bipotent progenitor cells from human live...

  10. Long-Term Depression at Parallel Fiber to Golgi Cell Synapses

    OpenAIRE

    Robberechts, Quinten; Wijnants, Mike; Giugliano, Michele; De Schutter, Erik

    2010-01-01

    Golgi cells (GoCs) are the primary inhibitory interneurons of the granular layer of the cerebellum. Their inhibition of granule cells is central to operate the relay of excitatory inputs to the cerebellar cortex. Parallel fibers (PFs) establish synapses to the GoCs in the molecular layer; these synapses contain AMPA, N-methyl-d-aspartate (NMDA), and mostly group II metabotropic glutamate receptors. Long-term changes in the efficacy of synaptic transmission at the PF-GoC synapse have not been ...

  11. Measuring stem cell frequency in epidermis: A quantitative in vivo functional assay for long-term repopulating cells

    Science.gov (United States)

    Schneider, T. E.; Barland, C.; Alex, A. M.; Mancianti, M. L.; Lu, Y.; Cleaver, J. E.; Lawrence, H. J.; Ghadially, R.

    2003-09-01

    Epidermal stem cells play a central role in tissue homeostasis, wound repair, tumor initiation, and gene therapy. A major impediment to the purification and molecular characterization of epidermal stem cells is the lack of a quantitative assay for cells capable of long-term repopulation in vivo, such as exists for hematopoietic cells. The tremendous strides made in the characterization and purification of hematopoietic stem cells have been critically dependent on the availability of competitive transplantation assays, because these assays permit the accurate quantitation of long-term repopulating cells in vivo. We have developed an analogous functional assay for epidermal stem cells, and have measured the frequency of functional epidermal stem cells in interfollicular epidermis. These studies indicate that cells capable of long-term reconstitution of a squamous epithelium reside in the interfollicular epidermis. We find that the frequency of these long-term repopulating cells is 1 in 35,000 total epidermal cells, or in the order of 1 in 104 basal epidermal cells, similar to that of hematopoietic stem cells in the bone marrow, and much lower than previously estimated in epidermis. Furthermore, these studies establish a novel functional assay that can be used to validate immunophenotypic markers and enrichment strategies for epidermal stem cells, and to quantify epidermal stem cells in various keratinocyte populations. Thus further studies using this type of assay for epidermis should aid in the progress of cutaneous stem cell-targeted gene therapy, and in more basic studies of epidermal stem cell regulation and differentiation.

  12. Long-term nonprogression and broad HIV-1-specific proliferative T-cell responses

    Directory of Open Access Journals (Sweden)

    Nesrina eImami

    2013-03-01

    Full Text Available Complex mechanisms underlying the maintenance of fully functional, proliferative, HIV-1-specific T-cell responses involve processes from early T-cell development through to the final stages of T-cell differentiation and antigen recognition. Virus-specific proliferative CD4 and CD8 T-cell responses, important for the control of infection, are observed in some HIV-1+ patients during early stages of disease, and are maintained in long-term nonprogressing subjects. In the vast majority of HIV-1+ patients, full immune functionality is lost when proliferative HIV-1-specific T-cell responses undergo a variable progressive decline throughout the course of chronic infection. This appears irreparable despite administration of potent combination antiretroviral therapy, which to date is non-curative, necessitating life-long administration and the development of effective, novel, therapeutic interventions. While a sterilising cure, involving clearance of virus from the host, remains a primary aim, a functional cure may be a more feasible goal with considerable impact on worldwide HIV-1 infection. Such an approach would enable long-term co-existence of host and virus in the absence of toxic and costly drugs. Effective immune homeostasis coupled with a balanced response appropriately targeting conserved viral antigens, in a manner that avoids hyperactivation and exhaustion, may prove to be the strongest correlate of durable viral control. This review describes novel concepts underlying full immune functionality in the context of HIV-1 infection, which may be utilised in future strategies designed to improve upon existing therapy. The aim will be to induce long-term nonprogressor or elite controller status in every infected host, through immune-mediated control of viraemia and reduction of viral reservoirs, leading to lower HIV-1 transmission rates.

  13. Long-term culture of human odontoma-derived cells with a Rho kinase inhibitor.

    Science.gov (United States)

    Uzawa, Katsuhiro; Kasamatsu, Atsushi; Saito, Tomoaki; Takahara, Toshikazu; Minakawa, Yasuyuki; Koike, Kazuyuki; Yamatoji, Masanobu; Nakashima, Dai; Higo, Morihiro; Sakamoto, Yosuke; Shiiba, Masashi; Tanzawa, Hideki

    2016-09-10

    Because of cellular senescence/apoptosis, no effective culture systems are available to maintain replication of cells from odontogenic tumors especially for odontoma, and, thus, the ability to isolate human odontoma-derived cells (hODCs) for functional studies is needed. The current study was undertaken to develop an approach to isolate hODCs and fully characterize the cells in vitro. The hODCs were cultured successfully with a Rho-associated protein kinase inhibitor (Y-27632) for an extended period with stabilized lengths of the telomeres to sustain a similar phenotype/property as the primary tumoral cells. While the hODCs showed stable long-term expansion with expression of major dental epithelial markers including dentin sialophosphoprotein (DSPP) even in the three-dimensional microenvironment, they lack the specific markers for the characteristics of stem cells. Moreover, cells from dental pulp showed significant up-regulation of DSPP when co-cultured with the hODCs, while control fibroblasts with the hODCs did not. Taken together, we propose that the hODCs can be isolated and expanded over the long term with Y-27632 to investigate not only the development of the hODCs but also other types of benign human tumors.

  14. Cell proliferation as a long-term prognostic factor in diffuse large-cell lymphomas.

    Science.gov (United States)

    Silvestrini, R; Costa, A; Boracchi, P; Giardini, R; Rilke, F

    1993-05-01

    The relevance of cell proliferation rate--defined as the 3H-thymidine labeling index (3H-dT LI)--in predicting response to treatment (complete remission, CR), freedom from progression (FFP) and overall survival (OS) was evaluated in 86 patients with diffuse large-cell lymphoma (DLCL). The biologic variable was not associated with most of the established clinical factors, such as gender and age of the patient, performance status, B symptoms, tumor bulk, or extranodal disease, but was directly related to stage. 3H-dT LI significantly predicted short- and long-term clinical outcome. In fact, more patients with slowly proliferating DLCL reached CR and had longer median FFP and OS than patients with rapidly proliferating DLCL. Multiple-regression analysis to evaluate the relative contribution of the different biologic and clinical variables in predicting CR, FFP and OS showed that 3H-dT LI and Ann Arbor stage were the only 2 stable factors, which retained their prognostic significance even in the presence of other conventional factors, and that 3H-dT LI was the most powerful as an indicator of risk of death in DLCL patients.

  15. Long-Term Culture of Genome-Stable Bipotent Stem Cells from Adult Human Liver

    Science.gov (United States)

    Huch, Meritxell; Gehart, Helmuth; van Boxtel, Ruben; Hamer, Karien; Blokzijl, Francis; Verstegen, Monique M.A.; Ellis, Ewa; van Wenum, Martien; Fuchs, Sabine A.; de Ligt, Joep; van de Wetering, Marc; Sasaki, Nobuo; Boers, Susanne J.; Kemperman, Hans; de Jonge, Jeroen; Ijzermans, Jan N.M.; Nieuwenhuis, Edward E.S.; Hoekstra, Ruurdtje; Strom, Stephen; Vries, Robert R.G.; van der Laan, Luc J.W.; Cuppen, Edwin; Clevers, Hans

    2015-01-01

    Summary Despite the enormous replication potential of the human liver, there are currently no culture systems available that sustain hepatocyte replication and/or function in vitro. We have shown previously that single mouse Lgr5+ liver stem cells can be expanded as epithelial organoids in vitro and can be differentiated into functional hepatocytes in vitro and in vivo. We now describe conditions allowing long-term expansion of adult bile duct-derived bipotent progenitor cells from human liver. The expanded cells are highly stable at the chromosome and structural level, while single base changes occur at very low rates. The cells can readily be converted into functional hepatocytes in vitro and upon transplantation in vivo. Organoids from α1-antitrypsin deficiency and Alagille syndrome patients mirror the in vivo pathology. Clonal long-term expansion of primary adult liver stem cells opens up experimental avenues for disease modeling, toxicology studies, regenerative medicine, and gene therapy. PMID:25533785

  16. Long-term multilineage engraftment of autologous genome-edited hematopoietic stem cells in nonhuman primates.

    Science.gov (United States)

    Peterson, Christopher W; Wang, Jianbin; Norman, Krystin K; Norgaard, Zachary K; Humbert, Olivier; Tse, Collette K; Yan, Jenny J; Trimble, Richard G; Shivak, David A; Rebar, Edward J; Gregory, Philip D; Holmes, Michael C; Kiem, Hans-Peter

    2016-05-19

    Genome editing in hematopoietic stem and progenitor cells (HSPCs) is a promising novel technology for the treatment of many human diseases. Here, we evaluated whether the disruption of the C-C chemokine receptor 5 (CCR5) locus in pigtailed macaque HSPCs by zinc finger nucleases (ZFNs) was feasible. We show that macaque-specific CCR5 ZFNs efficiently induce CCR5 disruption at levels of up to 64% ex vivo, 40% in vivo early posttransplant, and 3% to 5% in long-term repopulating cells over 6 months following HSPC transplant. These genome-edited HSPCs support multilineage engraftment and generate progeny capable of trafficking to secondary tissues including the gut. Using deep sequencing technology, we show that these ZFNs are highly specific for the CCR5 locus in primary cells. Further, we have adapted our clonal tracking methodology to follow individual CCR5 mutant cells over time in vivo, reinforcing that CCR5 gene-edited HSPCs are capable of long-term engraftment. Together, these data demonstrate that genome-edited HSPCs engraft, and contribute to multilineage repopulation after autologous transplantation in a clinically relevant large animal model, an important step toward the development of stem cell-based genome-editing therapies for HIV and potentially other diseases as well. PMID:26980728

  17. Efficient long-term cryopreservation of pluripotent stem cells at −80 °C

    Science.gov (United States)

    Yuan, Ye; Yang, Ying; Tian, Yuchen; Park, Jinkyu; Dai, Aihua; Roberts, R. Michael; Liu, Yang; Han, Xu

    2016-01-01

    Current long term cryopreservation of cell stocks routinely requires the use of liquid nitrogen (LN2), because commonly used cryopreservation media containing cell membrane permeating cryoprotectants are thermally unstable when frozen at higher storage temperatures, e.g. −80 °C. This instability leads to ice recrystallization, causing progressive loss of cell viability over time under the storage conditions provided by most laboratory deep freezers. The dependency on LN2 for cell storage significantly increases operational expense and raises issues related to impaired working efficiency and safety. Here we demonstrate that addition of Ficoll 70 to cryoprotectant solutions significantly improves system thermal stability at the working temperature (~−80 °C) of laboratory deep freezers. Moreover, a medium comprised of Ficoll 70 and dimethyl sulfoxide (DMSO) in presence or absence of fetal bovine serum (FBS) can provide reliable cryopreservation of various kinds of human and porcine pluripotent stem cells at −80 °C for periods that extend up to at least one year, with the post-thaw viability, plating efficiency, and full retention of pluripotent phenotype comparable to that achieved with LN2 storage. These results illustrate the practicability of a promising long-term cryopreservation method that completely eliminates the need for LN2. PMID:27694817

  18. Cell-type specific light-mediated transcript regulation in the multicellular alga Volvox carteri

    OpenAIRE

    Kianianmomeni, Arash

    2014-01-01

    Background The multicellular green alga Volvox carteri makes use of none less than 13 photoreceptors, which are mostly expressed in a cell-type specific manner. This gives reason to believe that trasncriptome pattern of each cell type could change differentially in response to environmental light. Here, the cell-type specific changes of various transcripts from different pathways in response to blue, red and far-red light were analyzed. Results In response to different light qualities, distin...

  19. Phenotypic and functional characterization of human mammary stem/progenitor cells in long term culture.

    Directory of Open Access Journals (Sweden)

    Devaveena Dey

    Full Text Available BACKGROUND: Cancer stem cells exhibit close resemblance to normal stem cells in phenotype as well as function. Hence, studying normal stem cell behavior is important in understanding cancer pathogenesis. It has recently been shown that human breast stem cells can be enriched in suspension cultures as mammospheres. However, little is known about the behavior of these cells in long-term cultures. Since extensive self-renewal potential is the hallmark of stem cells, we undertook a detailed phenotypic and functional characterization of human mammospheres over long-term passages. METHODOLOGY: Single cell suspensions derived from human breast 'organoids' were seeded in ultra low attachment plates in serum free media. Resulting primary mammospheres after a week (termed T1 mammospheres were subjected to passaging every 7th day leading to the generation of T2, T3, and T4 mammospheres. PRINCIPAL FINDINGS: We show that primary mammospheres contain a distinct side-population (SP that displays a CD24(low/CD44(low phenotype, but fails to generate mammospheres. Instead, the mammosphere-initiating potential rests within the CD44(high/CD24(low cells, in keeping with the phenotype of breast cancer-initiating cells. In serial sphere formation assays we find that even though primary (T1 mammospheres show telomerase activity and fourth passage T4 spheres contain label-retaining cells, they fail to initiate new mammospheres beyond T5. With increasing passages, mammospheres showed an increase in smaller sized spheres, reduction in proliferation potential and sphere forming efficiency, and increased differentiation towards the myoepithelial lineage. Significantly, staining for senescence-associated beta-galactosidase activity revealed a dramatic increase in the number of senescent cells with passage, which might in part explain the inability to continuously generate mammospheres in culture. CONCLUSIONS: Thus, the self-renewal potential of human breast stem cells is

  20. Three-dimensional photopatterning of hydrogels using stereolithography for long-term cell encapsulation.

    Science.gov (United States)

    Chan, Vincent; Zorlutuna, Pinar; Jeong, Jae Hyun; Kong, Hyunjoon; Bashir, Rashid

    2010-08-21

    Cell-encapsulated hydrogels with complex three-dimensional (3D) structures were fabricated from photopolymerizable poly(ethylene glycol) diacrylate (PEGDA) using modified 'top-down' and 'bottoms-up' versions of a commercially available stereolithography apparatus (SLA). Swelling and mechanical properties were measured for PEGDA hydrogels with molecular weights (M(w)) ranging from 700 to 10 000 Daltons (Da). Long-term viability of encapsulated NIH/3T3 cells was quantitatively evaluated using an MTS assay and shown to improve over 14 days by increasing the M(w) of the hydrogels. Addition of adhesive RGDS peptide sequences resulted in increased cell viability, proliferation, and spreading compared to pristine PEG hydrogels of the same M(w). Spatial 3D layer-by-layer cell patterning was successfully demonstrated, and the feasibility of depositing multiple cell types and material compositions into distinct layers was established.

  1. Short and long-term conservation of blood and bone marrow cells for clinical use

    International Nuclear Information System (INIS)

    The development of methods to conserve bone marrow, blood and blood components is necessary to ensure adequate supplies of these at times of radiation accidents or during radiation therapy with accompanying destruction of the haemopoietic tissues and essential blood elements. In addition, with adequate supplies of stored blood, treatment of astronauts affected by radiation, particularly when interplanetary flights become feasible, will be possible and therapy for patients with blood dyscrasias can be greatly expanded. The creation and expansion of blood banks for long-term storage of haemotherapeutic products is necessarily dependent on improved methods of conservation of blood and bone marrow cells

  2. A zero-flow microfluidics for long-term cell culture and detection

    Science.gov (United States)

    Sang, Shengbo; Tang, Xiaoliang; Feng, Qiliang; Jian, Aoqun; Zhang, Wendong

    2015-04-01

    A zero-flow microfluidic design is proposed in this paper, which can be used for long-term cell culture and detection, especially for a lab-on-chip integrated with a biosensor. It consists of two parts: a main microchannel; and a circle microchamber. The Finite Element Method (FEM) was employed to predict the fluid transport properties for a minimum fluid flow disturbance. Some commonly used microfluidic structures were also analysed systematically to prove the designed structure. Then the designed microfluidics was fabricated. Based on the simulations and experiments, this design provides a continuous flow environment, with a relatively stable and low shear stress atmosphere, similar to a zero-flow environment. Furthermore, the nutrients maintaining cells' normal growth can be taken into the chamber through the diffusion effect. It also proves that the microfluidics can realize long-term cell culture and detection. The application of the structure in the field of biological microelectromechenical systems (BioMEMS) will provide a research foundation for microfluidic technology.

  3. Tritium contamination of hematopoietic stem cells alters long-term hematopoietic reconstitution

    International Nuclear Information System (INIS)

    Purpose: In vivo effects of tritium contamination are poorly documented. Here, we study the effects of tritiated Thymidine ([3H] Thymidine) or tritiated water (HTO) contamination on the biological properties of hematopoietic stem cells (HSC). Materials and methods: Mouse HSC were contaminated with concentrations of [3H] Thymidine ranging from 0.37-37.03 kBq/ml or of HTO ranging from 5-50 kBq/ml. The biological properties of contaminated HSC were studied in vitro after HTO contamination and in vitro and in vivo after [3H] Thymidine contamination. Results: Proliferation, viability and double-strand breaks were dependent on [3H] Thymidine or HTO concentrations used for contamination but in vitro myeloid differentiation of HSC was not affected by [3H] Thymidine contamination. [3H] Thymidine contaminated HSC showed a compromised long-term capacity of hematopoietic reconstitution and competition experiments showed an up to two-fold decreased capacity of contaminated HSC to reconstitute hematopoiesis. These defects were not due to impaired homing in bone marrow but to an initial decreased proliferation rate of HSC. Conclusion: These results indicate that contaminations of HSC with doses of tritium that do not result in cell death, induce short-term effects on proliferation and cell cycle and long-term effects on hematopoietic reconstitution capacity of contaminated HSC. (authors)

  4. Cytomegalovirus shapes long-term immune reconstitution after allogeneic stem cell transplantation

    Science.gov (United States)

    Itzykson, Raphael; Robin, Marie; Moins-Teisserenc, Helene; Delord, Marc; Busson, Marc; Xhaard, Aliénor; de Fontebrune, Flore Sicre; de Latour, Régis Peffault; Toubert, Antoine; Socié, Gérard

    2015-01-01

    Immune reconstitution after allogeneic stem cell transplantation is a dynamic and complex process depending on the recipient and donor characteristics, on the modalities of transplantation, and on the occurrence of graft-versus-host disease. Multivariate methods widely used for gene expression profiling can simultaneously analyze the patterns of a great number of biological variables on a heterogeneous set of patients. Here we use these methods on flow cytometry assessment of up to 25 lymphocyte populations to analyze the global pattern of long-term immune reconstitution after transplantation. Immune patterns were most distinct from healthy controls at six months, and had not yet fully recovered as long as two years after transplant. The two principal determinants of variability were linked to the balance of B and CD8+ T cells and of natural killer and B cells, respectively. Recipient’s cytomegalovirus serostatus, cytomegalovirus replication, and chronic graft-versus-host disease were the main factors shaping the immune pattern one year after transplant. We identified a complex signature of under- and over-representation of immune populations dictated by recipient’s cytomegalovirus seropositivity. Finally, we identified dimensions of variance in immune patterns as significant predictors of long-term non-relapse mortality, independently of chronic graft-versus-host disease. PMID:25261095

  5. Ultra-structural morphology of long-term cultivated white adipose tissue-derived stem cells.

    Science.gov (United States)

    Varga, Ivan; Miko, Michal; Oravcová, Lenka; Bačkayová, Tatiana; Koller, Ján; Danišovič, Ľuboš

    2015-12-01

    White adipose tissue was long perceived as a passive lipid storage depot but it is now considered as an active and important endocrine organ. It also harbours not only adipocytes and vascular cells but also a wide array of immunologically active cells, including macrophages and lymphocytes, which may induce obesity-related inflammation. Recently, adipose tissue has been reported as a source of adult mesenchymal stem cells with wide use in regenerative medicine and tissue engineering. Their relatively non-complicated procurement and collection (often performed as liposuction during aesthetic surgery) and grand plasticity support this idea even more. We focused our research on exploring the issues of isolation and long-term cultivation of mesenchymal stem cells obtained from adipose tissue. Ultra-structural morphology of the cells cultivated in vitro has been studied and analysed in several cultivation time periods and following serial passages--up to 30 passages. In the first passages they had ultra-structural characteristics of cells with high proteosynthetic activity. Within the cytoplasm, big number of small lipid droplets and between them, sparsely placed, small and inconspicuous, electron-dense, lamellar bodies, which resembled myelin figures were observed. The cells from the later passages contained high number of lamellar electron-dense structures, which filled out almost the entire cytoplasm. In between, mitochondria were often found. These bodies were sometimes small and resembled myelin figures, but several of them reached huge dimensions (more than 1 µm) and their lamellar structure was not distinguishable. We did not have an answer to the question about their function, but they probably represented the evidence of active metabolism of lipids present in the cytoplasm of these cells or represented residual bodies, which arise after the breakdown of cellular organelles, notably mitochondria during long-term cultivation.

  6. General approach for in vivo recovery of cell type-specific effector gene sets.

    Science.gov (United States)

    Barsi, Julius C; Tu, Qiang; Davidson, Eric H

    2014-05-01

    Differentially expressed, cell type-specific effector gene sets hold the key to multiple important problems in biology, from theoretical aspects of developmental gene regulatory networks (GRNs) to various practical applications. Although individual cell types of interest have been recovered by various methods and analyzed, systematic recovery of multiple cell type-specific gene sets from whole developing organisms has remained problematic. Here we describe a general methodology using the sea urchin embryo, a material of choice because of the large-scale GRNs already solved for this model system. This method utilizes the regulatory states expressed by given cells of the embryo to define cell type and includes a fluorescence activated cell sorting (FACS) procedure that results in no perturbation of transcript representation. We have extensively validated the method by spatial and qualitative analyses of the transcriptome expressed in isolated embryonic skeletogenic cells and as a consequence, generated a prototypical cell type-specific transcriptome database.

  7. A simple and efficient method for the long-term preservation of plant cell suspension cultures

    Directory of Open Access Journals (Sweden)

    Boisson Anne-Marie

    2012-01-01

    Full Text Available Abstract Background The repeated weekly subculture of plant cell suspension is labour intensive and increases the risk of variation from parental cells lines. Most of the procedures to preserve cultures are based on controlled freezing/thawing and storage in liquid nitrogen. However, cells viability after unfreezing is uncertain. The long-term storage and regeneration of plant cell cultures remains a priority. Results Sycamore (Acer pseudoplatanus and Arabidopsis cell were preserved over six months as suspensions cultures in a phosphate-free nutrient medium at 5°C. The cell recovery monitored via gas exchange measurements and metabolic profiling using in vitro and in vivo 13C- and 31P-NMR took a couple of hours, and cell growth restarted without appreciable delay. No measurable cell death was observed. Conclusion We provide a simple method to preserve physiologically homogenous plant cell cultures without subculture over several months. The protocol based on the blockage of cell growth and low culture temperature is robust for heterotrophic and semi-autotrophic cells and should be adjustable to cell lines other than those utilised in this study. It requires no specialized equipment and is suitable for routine laboratory use.

  8. A zero-flow microfluidics for long-term cell culture and detection

    Directory of Open Access Journals (Sweden)

    Shengbo Sang

    2015-04-01

    Full Text Available A zero-flow microfluidic design is proposed in this paper, which can be used for long-term cell culture and detection, especially for a lab-on-chip integrated with a biosensor. It consists of two parts: a main microchannel; and a circle microchamber. The Finite Element Method (FEM was employed to predict the fluid transport properties for a minimum fluid flow disturbance. Some commonly used microfluidic structures were also analysed systematically to prove the designed structure. Then the designed microfluidics was fabricated. Based on the simulations and experiments, this design provides a continuous flow environment, with a relatively stable and low shear stress atmosphere, similar to a zero-flow environment. Furthermore, the nutrients maintaining cells’ normal growth can be taken into the chamber through the diffusion effect. It also proves that the microfluidics can realize long-term cell culture and detection. The application of the structure in the field of biological microelectromechenical systems (BioMEMS will provide a research foundation for microfluidic technology.

  9. Long-term Survival, Organ Function, and Malignancy after Hematopoietic Stem Cell Transplantation for Fanconi Anemia.

    Science.gov (United States)

    Bonfim, Carmem; Ribeiro, Lisandro; Nichele, Samantha; Bitencourt, Marco; Loth, Gisele; Koliski, Adriana; Funke, Vaneuza A M; Pilonetto, Daniela V; Pereira, Noemi F; Flowers, Mary E D; Velleuer, Eunike; Dietrich, Ralf; Fasth, Anders; Torres-Pereira, Cassius C; Pedruzzi, Paola; Eapen, Mary; Pasquini, Ricardo

    2016-07-01

    We report on long-term survival in 157 patients with Fanconi anemia (FA) who survived 2 years or longer after their first transplantation with a median follow-up of 9 years. Marrow failure (80%) was the most common indication for transplantation. There were 20 deaths beyond 2 years after transplantation, with 12 of the deaths occurring beyond 5 years after transplantation. Donor chimerism was available for 149 patients: 112 (76%) reported > 95% chimerism, 27 (18%) reported 90% to 95% chimerism, and 8 (5%) reported 20% to 89% donor chimerism. Two patients have donor chimerism. The 10- and 15-year probabilities of survival were 90% and 79%, respectively. Results of multivariate analysis showed higher mortality risks for transplantations before 2003 (hazard ratio [HR], 7.87; P = .001), chronic graft-versus-host disease (GVHD) (HR, 3.80; P = .004) and squamous cell carcinoma after transplantation (HR, 38.17; P < .0001). The predominant cause of late mortality was squamous cell carcinoma, with an incidence of 8% and 14% at 10 and 15 years after transplantation, respectively, and was more likely to occur in those with chronic GVHD. Other causes of late mortality included chronic GVHD, infection, graft failure, other cancers, and hemorrhage. Although most patients are disease free and functional long term, our data support aggressive surveillance for long periods to identify those at risk for late mortality. PMID:26976241

  10. Long-term Survival, Organ Function, and Malignancy after Hematopoietic Stem Cell Transplantation for Fanconi Anemia.

    Science.gov (United States)

    Bonfim, Carmem; Ribeiro, Lisandro; Nichele, Samantha; Bitencourt, Marco; Loth, Gisele; Koliski, Adriana; Funke, Vaneuza A M; Pilonetto, Daniela V; Pereira, Noemi F; Flowers, Mary E D; Velleuer, Eunike; Dietrich, Ralf; Fasth, Anders; Torres-Pereira, Cassius C; Pedruzzi, Paola; Eapen, Mary; Pasquini, Ricardo

    2016-07-01

    We report on long-term survival in 157 patients with Fanconi anemia (FA) who survived 2 years or longer after their first transplantation with a median follow-up of 9 years. Marrow failure (80%) was the most common indication for transplantation. There were 20 deaths beyond 2 years after transplantation, with 12 of the deaths occurring beyond 5 years after transplantation. Donor chimerism was available for 149 patients: 112 (76%) reported > 95% chimerism, 27 (18%) reported 90% to 95% chimerism, and 8 (5%) reported 20% to 89% donor chimerism. Two patients have < 20% donor chimerism. The 10- and 15-year probabilities of survival were 90% and 79%, respectively. Results of multivariate analysis showed higher mortality risks for transplantations before 2003 (hazard ratio [HR], 7.87; P = .001), chronic graft-versus-host disease (GVHD) (HR, 3.80; P = .004) and squamous cell carcinoma after transplantation (HR, 38.17; P < .0001). The predominant cause of late mortality was squamous cell carcinoma, with an incidence of 8% and 14% at 10 and 15 years after transplantation, respectively, and was more likely to occur in those with chronic GVHD. Other causes of late mortality included chronic GVHD, infection, graft failure, other cancers, and hemorrhage. Although most patients are disease free and functional long term, our data support aggressive surveillance for long periods to identify those at risk for late mortality.

  11. Late Mortality and Causes of Death among Long-Term Survivors after Allogeneic Stem Cell Transplantation.

    Science.gov (United States)

    Atsuta, Yoshiko; Hirakawa, Akihiro; Nakasone, Hideki; Kurosawa, Saiko; Oshima, Kumi; Sakai, Rika; Ohashi, Kazuteru; Takahashi, Satoshi; Mori, Takehiko; Ozawa, Yukiyasu; Fukuda, Takahiro; Kanamori, Heiwa; Morishima, Yasuo; Kato, Koji; Yabe, Hiromasa; Sakamaki, Hisashi; Taniguchi, Shuichi; Yamashita, Takuya

    2016-09-01

    We sought to assess the late mortality risks and causes of death among long-term survivors of allogeneic hematopoietic stem cell transplantation (HCT). The cases of 11,047 relapse-free survivors of a first HCT at least 2 years after HCT were analyzed. Standardized mortality ratios (SMR) were calculated and specific causes of death were compared with those of the Japanese population. Among relapse-free survivors at 2 years, overall survival percentages at 10 and 15 years were 87% and 83%, respectively. The overall risk of mortality was significantly higher compared with that of the general population. The risk of mortality was significantly higher from infection (SMR = 57.0), new hematologic malignancies (SMR = 2.2), other new malignancies (SMR = 3.0), respiratory causes (SMR = 109.3), gastrointestinal causes (SMR = 3.8), liver dysfunction (SMR = 6.1), genitourinary dysfunction (SMR = 17.6), and external or accidental causes (SMR = 2.3). The overall annual mortality rate showed a steep decrease from 2 to 5 years after HCT; however, the decrease rate slowed after 10 years but was still higher than that of the general population at 20 years after HCT. SMRs in the earlier period of 2 to 4 years after HCT and 5 years or longer after HCT were 16.1 and 7.4, respectively. Long-term survivors after allogeneic HCT are at higher risk of mortality from various causes other than the underlying disease that led to HCT. Screening and preventive measures should be given a central role in reducing the morbidity and mortality of HCT recipients on long-term follow-up. PMID:27246369

  12. Cell search scheme for long-term evolution of TD-SCDMA system

    Institute of Scientific and Technical Information of China (English)

    SU Huan; ZHANG Jian-hua; LI Ke

    2008-01-01

    Cell search is an important aspect for 3G long-term evolution (LTE). This article deals with cell search in the time-division-synchronou code-division multiple access (TD-SCDMA) LTE system. On the basis of the synchronization channel (SCH) and cell specific reference symbols (CSRSs), the proposed cell search procedure includes five stages: frame detection and coarse timing, coarse carrier frequency offset (CFO) estimation, fine timing, fine CFO estimation, and cell identification. The key features of the proposed method are as follows: first, the neighboring three cells' CSRSs are frequency division multiplexed (FD) to mitigate inter-cell interference. Second, the frequency domain differential cross-correlation, computed from CSRSs are maximally ratio combined for cell identification. Finally, the large set Kasami sequences are quadrature phase shift key (QPSK) modulated to be cell specific sequences (CSSs), to support a large number of target cells. Simulations show that the FD method is better than the code division multiplexed (CD) method.

  13. Long-term, large scale cryopreservation of insect cells at -80 °C.

    Science.gov (United States)

    Vyletova, Lucie; Rennalls, La'Verne P; Wood, Kirstin J L; Good, Valerie M

    2016-03-01

    Standard tissue culture methods advise freezing cells in small aliquots (≤1 × 10(7) cells in 1 mL), and storing in liquid nitrogen. This is inconvenient for laboratories culturing large quantities of insect cells for recombinant baculovirus expression, owing to the length of time taken to produce large scale cultures from small aliquots of cells. Liquid nitrogen storage requires use of specialized cryovials, personal protective equipment and oxygen monitoring systems. This paper describes the long-term, large scale cryopreservation of 8 × 10(8) insect cells at -80 °C, using standard 50 mL conical tubes to contain a 40 mL cell suspension. Sf9, Sf21 and High 5 cells were recovered with a viability > 90 % after storage for one year under these conditions, which compared favorably with the viability of cells stored in liquid nitrogen for the same length of time. Addition of green fluorescent protein encoding baculovirus demonstrated that cells were "expression ready" immediately post thaw. Our method enables large scale cultures to be recovered rapidly from stocks cryopreserved at -80 °C, thus avoiding the inconvenience, hazards and expense associated with liquid nitrogen.

  14. Pancreatic small cells: Analysis of quiescence, long-term maintenance and insulin expression in vitro

    International Nuclear Information System (INIS)

    We have previously identified a novel population of small cells in human and canine pancreas characterized by immature morphology, quiescence, and a glucose-responsive insulin secretion. Based on their immature phenotype and predominant presence in small islets, we have hypothesized that small cells serve as islet progenitors. This hypothesis remains untested, however, due to persistent quiescence and scarcity of small cells in vitro. We have recently developed a culture medium that allowed for modest small cell proliferation. In this study we characterized the expression of genes potentially involved in small cell growth regulation by Q-RT-PCR. Our results suggest that quiescence of small cells correlates with up-regulation of Cdk inhibitors p27Kip1, p16INK4a and p21CIP1, PTEN, Hep27 and Foxo1a and with down-regulation of c-Myc and the receptors for EGF, FGF2 and HGF. The exit from quiescence correlates with activation of EGFR expression and down-regulation of p27Kip1 and p16INK4a. We also report here that small cells can be maintained in long-term non-adherent cultures preserving insulin and glucagon production for up to 208 days. Therefore, expansion of small cells in vitro may have a significant potential for the treatment of diabetes. This study is an important step in understanding the mechanisms involved in small cell growth regulation, which is required to fully evaluate their functional potential

  15. Human cytomegalovirus gene expression in long-term infected glioma stem cells.

    Directory of Open Access Journals (Sweden)

    Estefania Fiallos

    Full Text Available The most common adult primary brain tumor, glioblastoma (GBM, is characterized by fifteen months median patient survival and has no clear etiology. We and others have identified the presence of human cytomegalovirus (HCMV gene products endogenously expressed in GBM tissue and primary cells, with a subset of viral genes being consistently expressed in most samples. Among these viral genes, several have important oncomodulatory properties, regulating tumor stemness, proliferation, immune evasion, invasion and angiogenesis. These findings lead us to hypothesize that a specific HCMV gene signature may be associated with GBM pathogenesis. To investigate this hypothesis, we used glioma cell lines and primary glioma stem-like cells (GSC infected with clinical and laboratory HCMV strains and measured relative viral gene expression levels along several time points up to 15 weeks post-infection. While HCMV gene expression was detected in several infected glioma lines through week 5 post-infection, only HCMV-infected GSC expressed viral gene products 15 weeks post-infection. Efficiency of infection across time was higher in GSC compared to cell lines. Importantly, HCMV-infected GSC outlived their uninfected counterparts, and this extended survival was paralleled by increased tumorsphere frequency and upregulation of stemness regulators, such as SOX2, p-STAT3, and BMX (a novel HCMV target identified in this study. Interleukin 6 (IL-6 treatment significantly upregulated HCMV gene expression in long-term infected glioma cultures, suggesting that pro-inflammatory signaling in the tumor milieu may further augment HCMV gene expression and subsequent tumor progression driven by viral-induced cellular signaling. Together, our data support a critical role for long-term, low-level HCMV infection in promoting survival, stemness, and proliferation of GSC that could significantly contribute to GBM pathogenesis.

  16. Long-term live cell imaging and automated 4D analysis of drosophila neuroblast lineages.

    Directory of Open Access Journals (Sweden)

    Catarina C F Homem

    Full Text Available The developing Drosophila brain is a well-studied model system for neurogenesis and stem cell biology. In the Drosophila central brain, around 200 neural stem cells called neuroblasts undergo repeated rounds of asymmetric cell division. These divisions typically generate a larger self-renewing neuroblast and a smaller ganglion mother cell that undergoes one terminal division to create two differentiating neurons. Although single mitotic divisions of neuroblasts can easily be imaged in real time, the lack of long term imaging procedures has limited the use of neuroblast live imaging for lineage analysis. Here we describe a method that allows live imaging of cultured Drosophila neuroblasts over multiple cell cycles for up to 24 hours. We describe a 4D image analysis protocol that can be used to extract cell cycle times and growth rates from the resulting movies in an automated manner. We use it to perform lineage analysis in type II neuroblasts where clonal analysis has indicated the presence of a transit-amplifying population that potentiates the number of neurons. Indeed, our experiments verify type II lineages and provide quantitative parameters for all cell types in those lineages. As defects in type II neuroblast lineages can result in brain tumor formation, our lineage analysis method will allow more detailed and quantitative analysis of tumorigenesis and asymmetric cell division in the Drosophila brain.

  17. Reversibility of β-Cell-Specific Transcript Factors Expression by Long-Term Caloric Restriction in db/db Mouse

    Directory of Open Access Journals (Sweden)

    Chunjun Sheng

    2016-01-01

    Full Text Available Type 2 diabetes (T2D is characterized by β-cell dedifferentiation, but underlying mechanisms remain unclear. The purpose of the current study was to explore the mechanisms of β-cell dedifferentiation with and without long-term control of calorie intake. We used a diabetes mouse model (db/db to analyze the changes in the expression levels of β-cell-specific transcription factors (TFs and functional factors with long-term caloric restriction (CR. Our results showed that chronic euglycemia was maintained in the db/db mice with long-term CR intervention, and β-cell dedifferentiation was significantly reduced. The expression of Glut2, Pdx1, and Nkx6.1 was reversed, while MafA expression was significantly increased with long-term CR. GLP-1 pathway was reactivated with long-term CR. Our work showed that the course of β-cell dedifferentiation can intervene by long-term control of calorie intake. Key β-cell-specific TFs and functional factors play important roles in maintaining β-cell differentiation. Targeting these factors could optimize T2D therapies.

  18. Stromal cells in long-term murine bone marrow culture: FACS studies and origin of stromal cells in radiation chimeras

    International Nuclear Information System (INIS)

    Adherent layers from hematopoietically active long-term bone marrow cultures (LTBMC), incubated with fluorescent beads, were analyzed for autofluorescence and phagocytic ability, using a fluorescence-activated cell sorter (FACS). Four groups of cells were separated from the adherent layers, including a group of large polygonal fibroblastoid stromal cells. Long-term chimeras were made by lethal irradiation of CBA/Ca (CBA) and C57Bl6/J (B6) mice and repopulation with phosphoglycerate kinase (PGK-1) alloenzyme-congenic bone marrow cells. Hematopoietically active LTBMC were established from such chimeras, and donor and host contributions of FACS-sorted adherent-layer cells were measured. While macrophages and other hematopoietic cells were of donor origin, the fibroblastoid stromal cells were mainly or entirely host derived

  19. Long-Term Degradation Testing of High-Temperature Electrolytic Cells

    Energy Technology Data Exchange (ETDEWEB)

    C.M. Stoots; J.E. O' Brien; J.S. Herring; G.K. Housley; D.G. Milobar; M.S. Sohal

    2009-08-01

    The Idaho National Laboratory (INL) has been researching the application of solid-oxide electrolysis cell for large-scale hydrogen production from steam over a temperature range of 800 to 900ºC. The INL has been testing various solid oxide cell designs to characterize their electrolytic performance operating in the electrolysis mode for hydrogen production. Some results presented in this report were obtained from cells, with an active area of 16 cm2 per cell. The electrolysis cells are electrode-supported, with ~10 µm thick yttria-stabilized zirconia (YSZ) electrolytes, ~1400 µm thick nickel-YSZ steam-hydrogen electrodes, and manganite (LSM) air-oxygen electrodes. The experiments were performed over a range of steam inlet mole fractions (0.1 to 0.6), gas flow rates, and current densities (0 to 0.6 A/cm2). Steam consumption rates associated with electrolysis were measured directly using inlet and outlet dewpoint instrumentation. On a molar basis, the steam consumption rate is equal to the hydrogen production rate. Cell performance was evaluated by performing DC potential sweeps at 800, 850, and 900°C. The voltage-current characteristics are presented, along with values of area-specific resistance as a function of current density. Long-term cell performance is also assessed to evaluate cell degradation. Details of the custom single-cell test apparatus developed for these experiments are also presented. NASA, in conjunction with the University of Toledo, has developed a new cell concept with the goals of reduced weight and high power density. This report presents results of the INL's testing of this new solid oxide cell design as an electrolyzer. Gas composition, operating voltage, and other parameters were varied during testing. Results to date show the NASA cell to be a promising design for both high power-to-weight fuel cell and electrolyzer applications.

  20. Differential effects of HOXB4 on nonhuman primate short- and long-term repopulating cells.

    Directory of Open Access Journals (Sweden)

    Xiao-Bing Zhang

    2006-05-01

    Full Text Available BACKGROUND: Hematopoietic stem cells (HSCs or repopulating cells are able to self-renew and differentiate into cells of all hematopoietic lineages, and they can be enriched using the CD34 cell surface marker. Because of this unique property, HSCs have been used for HSC transplantation and gene therapy applications. However, the inability to expand HSCs has been a significant limitation for clinical applications. Here we examine, in a clinically relevant nonhuman primate model, the ability of HOXB4 to expand HSCs to potentially overcome this limitation. METHODS AND FINDINGS: Using a competitive repopulation assay, we directly compared in six animals engraftment of HOXB4GFP (HOXB4 green fluorescent protein and control (yellow fluorescent protein [YFP]-transduced and expanded CD34+ cells. In three animals, cells were infused after a 3-d transduction culture, while in three other animals cells were infused after an additional 6-9 d of ex vivo expansion. We demonstrate that HOXB4 overexpression resulted in superior engraftment in all animals. The most dramatic effect of HOXB4 was observed early after transplantation, resulting in an up to 56-fold higher engraftment compared to the control cells. At 6 mo after transplantation, the proportion of marker gene-expressing cells in peripheral blood was still up to 5-fold higher for HOXB4GFP compared to YFP-transduced cells. CONCLUSIONS: These data demonstrate that HOXB4 overexpression in CD34+ cells has a dramatic effect on expansion and engraftment of short-term repopulating cells and a significant, but less pronounced, effect on long-term repopulating cells. These data should have important implications for the expansion and transplantation of HSCs, in particular for cord blood transplantations where often only suboptimal numbers of HSCs are available.

  1. Long-term stabilization of organic solar cells using hydroperoxide decomposers as additives

    Science.gov (United States)

    Turkovic, Vida; Engmann, Sebastian; Tsierkezos, Nikos; Hoppe, Harald; Madsen, Morten; Rubahn, Horst-Günter; Ritter, Uwe; Gobsch, Gerhard

    2016-03-01

    Stability of organic solar cells (OPV) remains a big problem on the way to their commercialization. Different approaches are being investigated: development of intrinsically more photochemically stable materials, optimization of encapsulation, and implementation of getter and UV blocking layers. In this study, we investigate stabilization of OPV devices using hydroperoxide decomposers as stabilizing additives. A set of five commercially available additives of organophosphorus, organosulfur, Ni chelate, and blocked thiol type are compared, ternary blended into the active layer, under exposure to aging under ISOS-3 degradation conditions. Improvements in long-term performance of OPV devices were observed upon stabilization with Advapak NEO-1120, lifetime was prolonged by a factor of 1.7, and accumulated power generation increased by a factor of 1.4. The stabilizing mechanisms are discussed using spectroscopic and microscopic measurements.

  2. Long-term effects of the transient COD concentration on the performance of microbial fuel cells.

    Science.gov (United States)

    Mateo, S; Gonzalez Del Campo, A; Lobato, J; Rodrigo, M; Cañizares, P; Fernandez-Morales, F J

    2016-07-01

    In this work, the long-term effects of transient chemical oxygen demands (COD) concentrations over the performance of a microbial fuel cell were studied. From the obtained results, it was observed that the repetitive change in the COD loading rate during 12 h conditioned the behavior of the system during periods of up to 7 days. The main modifications were the enhancement of the COD consumption rate and the exerted current. These enhancements yielded increasing Coulombic efficiencies (CEs) when working with COD concentrations of 300 mg/L, but constant CEs when working with COD concentrations from 900 to 1800 mg/L. This effect could be explained by the higher affinity for the substrate of Geobacter than that of the nonelectrogenic organisms such as Clostridia. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:883-890, 2016. PMID:27072066

  3. Long-term cognitive effects of human stem cell transplantation in the irradiated brain

    Science.gov (United States)

    Acharya, Munjal M.; Martirosian, Vahan; Christie, Lori-Ann; Limoli, Charles L.

    2016-01-01

    Purpose Radiotherapy remains a primary treatment modality for the majority of central nervous system tumors, but frequently leads to debilitating cognitive dysfunction. Given the absence of satisfactory solutions to this serious problem, we have used human stem cell therapies to ameliorate radiation-induced cognitive impairment. Here, past studies have been extended to determine whether engrafted cells provide even longer-term benefits to cognition. Materials and methods Athymic nude rats were cranially irradiated (10 Gy) and subjected to intrahippocampal transplantation surgery 2 days later. Human embryonic stem cells (hESC) or human neural stem cells (hNSC) were transplanted, and animals were subjected to cognitive testing on a novel place recognition task 8 months later. Results Grafting of hNSC was found to provide long lasting cognitive benefits over an 8-month post-irradiation interval. At this protracted time, hNSC grafting improved behavioral performance on a novel place recognition task compared to irradiated animals not receiving stem cells. Engrafted hESC previously shown to be beneficial following a similar task, 1 and 4 months after irradiation, were not found to provide cognitive benefits at 8 months. Conclusions Our findings suggest that hNSC transplantation promotes the long-term recovery of the irradiated brain, where intrahippocampal stem cell grafting helps to preserve cognitive function. PMID:24882389

  4. Propagation of human germ stem cells in long-term culture

    Directory of Open Access Journals (Sweden)

    Abbas Khodadadi

    2013-01-01

    Full Text Available Background: Spermatogonial stem cells (SSCs, a subset of undifferentiated type A spermatogonia, are the foundation of complex process of spermatogenesis and could be propagated in vitro culture conditions for long time for germ cell transplantation and fertility preservation. Objective: The aim of this study was in vitro propagation of human spermatogonial stem cells (SSCs and improvement of presence of human Germ Stem Cells (hGSCs were assessed by specific markers POU domain, class 5, transcription factor 1 (POU5F1, also known as Octamer-binding transcription factor 4 (Oct-4 and PLZF (Promyelocytic leukaemia zinc finger protein. Materials and Methods: Human testicular cells were isolated by enzymatic digestion (Collagenase IV and Trypsin. Germ cells were cultured in Stem-Pro 34 media supplemented by growth factors such as glial cell line-derived neurotrophic factor, basic fibroblast growth factor, epidermal growth factor and leukemia inhibitory factor to support self-renewal divisions. Germline stem cell clusters were passaged and expanded every week. Immunofluorecent study was accomplished by Anti-Oct4 antibody through the culture. The spermatogonial stem cells genes expression, PLZF, was studied in testis tissue and germ stem cells entire the culture. Results: hGSCs clusters from a brain dead patient developed in testicular cell culture and then cultured and propagated up to 6 weeks. During the culture Oct4 were a specific marker for identification of hGSCs in testis tissue. Expression of PLZF was applied on RNA level in germ stem cells. Conclusion: hGSCs indicated by SSCs specific marker can be cultured and propagated for long-term in vitro conditions.

  5. Quantitative analysis of cell-type specific gene expression in the green alga Volvox carteri

    Directory of Open Access Journals (Sweden)

    Hallmann Armin

    2006-12-01

    Full Text Available Abstract Background The multicellular alga Volvox carteri possesses only two cell types: mortal, motile somatic cells and potentially immortal, immotile reproductive cells. It is therefore an attractive model system for studying how cell-autonomous cytodifferentiation is programmed within a genome. Moreover, there are ongoing genome projects both in Volvox carteri and in the closely related unicellular alga Chlamydomonas reinhardtii. However, gene sequencing is only the beginning. To identify cell-type specific expression and to determine relative expression rates, we evaluate the potential of real-time RT-PCR for quantifying gene transcript levels. Results Here we analyze a diversified pool of 39 target genes by real-time RT-PCR for each cell type. This gene pool contains previously known genes with unknown localization of cellular expression, 28 novel genes which are described in this study for the first time, and a few known, cell-type specific genes as a control. The respective gene products are, for instance, part of photosynthesis, cellular regulation, stress response, or transport processes. We provide expression data for all these genes. Conclusion The results show that quantitative real-time RT-PCR is a favorable approach to analyze cell-type specific gene expression in Volvox, which can be extended to a much larger number of genes or to developmental or metabolic mutants. Our expression data also provide a basis for a detailed analysis of individual, previously unknown, cell-type specifically expressed genes.

  6. Mobile Cell Selection In 4G Long Term Evolution-Advanced (LTE-A Networks

    Directory of Open Access Journals (Sweden)

    Murtadha Ali Nsaif Shukur

    2016-08-01

    Full Text Available With the high demands for broadband mobile wireless communications and the emergence of new wireless multimedia applications constitute the motivation to the development of broadband wireless access technologies in recent years. The Long Term Evolution/System Architecture Evolution (LTE/SAE system has been specified by the Third Generation Partnership Project (3GPP on the way towards fourth-generation (4G mobile to ensure 3GPP keeping the dominance of the cellular communication technologies. Through the design and optimization of new radio access techniques and a further evolution of the LTE-A systems, Cell selection is the process of determining the cell(s that provide service to each mobile station. By study the potential benefits of global cell selection versus the current local mobile SNR-based decision protocol. In particular, and present the new possibility available in OFDMA & SC-FDMA based systems, such as IEEE 802.16m and LTEAdvanced, of satisfying the minimal demand of a mobile station simultaneously by more than one base station. After formalized the problems as an optimization problem; it's present how the mobile unit establishes this connection with the strongest cell station in vicinity. To do this, the mobile unit has to overcome the challenges of estimating the channel to communicate with the cell site and frequency synchronization. Also, multiple mobile units communicate to the same receiver and from various distances. Hence, it is up to the mobile to synchronize itself appropriately to the base stations. LTE-A uses two signals, the Primary Synchronization Signal and the Secondary Synchronization Signal sequentially to determine which of the available cell sites, a mobile would lock in to it. While inter-cell interference (ICI one of problems for the downlink and uplink of multi-cell systems (in general and OFDMA& SC-FDMA networks (in particular.

  7. Feline Neural Progenitor Cells I: Long-Term Expansion under Defined Culture Conditions

    Directory of Open Access Journals (Sweden)

    Jing Yang

    2012-01-01

    Full Text Available Neural progenitor cells (NPCs of feline origin (cNPCs have demonstrated utility in transplantation experiments, yet are difficult to grow in culture beyond the 1 month time frame. Here we use an enriched, serum-free base medium (Ultraculture and report the successful long-term propagation of these cells. Primary cultures were derived from fetal brain tissue and passaged in DMEM/F12-based or Ultraculture-based proliferation media, both in the presence of EGF + bFGF. Cells in standard DMEM/F12-based medium ceased to proliferate by 1-month, whereas the cells in the Ultraculture-based medium continued to grow for at least 5 months (end of study with no evidence of senescence. The Ultraculture-based cultures expressed lower levels of progenitor and lineage-associated markers under proliferation conditions but retained multipotency as evidenced by the ability to differentiate into neurons and glia following growth factor removal in the presence of FBS. Importantly, later passage cNPCs did not develop chromosomal aberrations.

  8. A novel whole-cell mechanism for long-term memory enhancement.

    Directory of Open Access Journals (Sweden)

    Iris Reuveni

    Full Text Available Olfactory-discrimination learning was shown to induce a profound long-lasting enhancement in the strength of excitatory and inhibitory synapses of pyramidal neurons in the piriform cortex. Notably, such enhancement was mostly pronounced in a sub-group of neurons, entailing about a quarter of the cell population. Here we first show that the prominent enhancement in the subset of cells is due to a process in which all excitatory synapses doubled their strength and that this increase was mediated by a single process in which the AMPA channel conductance was doubled. Moreover, using a neuronal-network model, we show how such a multiplicative whole-cell synaptic strengthening in a sub-group of cells that form a memory pattern, sub-serves a profound selective enhancement of this memory. Network modeling further predicts that synaptic inhibition should be modified by complex learning in a manner that much resembles synaptic excitation. Indeed, in a subset of neurons all GABAA-receptors mediated inhibitory synapses also doubled their strength after learning. Like synaptic excitation, Synaptic inhibition is also enhanced by two-fold increase of the single channel conductance. These findings suggest that crucial learning induces a multiplicative increase in strength of all excitatory and inhibitory synapses in a subset of cells, and that such an increase can serve as a long-term whole-cell mechanism to profoundly enhance an existing Hebbian-type memory. This mechanism does not act as synaptic plasticity mechanism that underlies memory formation but rather enhances the response of already existing memory. This mechanism is cell-specific rather than synapse-specific; it modifies the channel conductance rather than the number of channels and thus has the potential to be readily induced and un-induced by whole-cell transduction mechanisms.

  9. Cell tropism predicts long-term nucleotide substitution rates of mammalian RNA viruses.

    Directory of Open Access Journals (Sweden)

    Allison L Hicks

    2014-01-01

    Full Text Available The high rates of RNA virus evolution are generally attributed to replication with error-prone RNA-dependent RNA polymerases. However, these long-term nucleotide substitution rates span three orders of magnitude and do not correlate well with mutation rates or selection pressures. This substitution rate variation may be explained by differences in virus ecology or intrinsic genomic properties. We generated nucleotide substitution rate estimates for mammalian RNA viruses and compiled comparable published rates, yielding a dataset of 118 substitution rates of structural genes from 51 different species, as well as 40 rates of non-structural genes from 28 species. Through ANCOVA analyses, we evaluated the relationships between these rates and four ecological factors: target cell, transmission route, host range, infection duration; and three genomic properties: genome length, genome sense, genome segmentation. Of these seven factors, we found target cells to be the only significant predictors of viral substitution rates, with tropisms for epithelial cells or neurons (P<0.0001 as the most significant predictors. Further, one-tailed t-tests showed that viruses primarily infecting epithelial cells evolve significantly faster than neurotropic viruses (P<0.0001 and P<0.001 for the structural genes and non-structural genes, respectively. These results provide strong evidence that the fastest evolving mammalian RNA viruses infect cells with the highest turnover rates: the highly proliferative epithelial cells. Estimated viral generation times suggest that epithelial-infecting viruses replicate more quickly than viruses with different cell tropisms. Our results indicate that cell tropism is a key factor in viral evolvability.

  10. Induced Neural Stem Cells Achieve Long-Term Survival and Functional Integration in the Adult Mouse Brain

    OpenAIRE

    Kathrin Hemmer; Mingyue Zhang; Thea van Wüllen; Marna Sakalem; Natalia Tapia; Aidos Baumuratov; Christian Kaltschmidt; Barbara Kaltschmidt; Hans R. Schöler; Weiqi Zhang; Jens C. Schwamborn

    2014-01-01

    Summary Differentiated cells can be converted directly into multipotent neural stem cells (i.e., induced neural stem cells [iNSCs]). iNSCs offer an attractive alternative to induced pluripotent stem cell (iPSC) technology with regard to regenerative therapies. Here, we show an in vivo long-term analysis of transplanted iNSCs in the adult mouse brain. iNSCs showed sound in vivo long-term survival rates without graft overgrowths. The cells displayed a neural multilineage potential with a clear ...

  11. Visualizing tropoelastin in a long-term human elastic fibre cell culture model.

    Science.gov (United States)

    Halm, M; Schenke-Layland, K; Jaspers, S; Wenck, H; Fischer, F

    2016-02-04

    Elastin is an essential protein found in a variety of tissues where resilience and flexibility are needed, such as the skin and the heart. When aiming to engineer suitable implants, elastic fibres are needed to allow adequate tissue renewal. However, the visualization of human elastogenesis remains in the dark. To date, the visualization of human tropoelastin (TE) production in a human cell context and its fibre assembly under live cell conditions has not been achieved. Here, we present a long-term cell culture model of human dermal fibroblasts expressing fluorescence-labelled human TE. We employed a lentiviral system to stably overexpress Citrine-labelled TE to build a fluorescent fibre network. Using immunofluorescence, we confirmed the functionality of the Citrine-tagged TE. Furthermore, we visualized the fibre assembly over the course of several days using confocal microscopy. Applying super resolution microscopy, we were able to investigate the inner structure of the elastin-fibrillin-1 fibre network. Future investigations will allow the tracking of TE produced under various conditions. In tissue engineering applications the fluorescent fibre network can be visualized under various conditions or it serves as a tool for investigating fibre degradation processes in disease-in-a-dish-models.

  12. Visualizing tropoelastin in a long-term human elastic fibre cell culture model

    Science.gov (United States)

    Halm, M.; Schenke-Layland, K.; Jaspers, S.; Wenck, H.; Fischer, F.

    2016-01-01

    Elastin is an essential protein found in a variety of tissues where resilience and flexibility are needed, such as the skin and the heart. When aiming to engineer suitable implants, elastic fibres are needed to allow adequate tissue renewal. However, the visualization of human elastogenesis remains in the dark. To date, the visualization of human tropoelastin (TE) production in a human cell context and its fibre assembly under live cell conditions has not been achieved. Here, we present a long-term cell culture model of human dermal fibroblasts expressing fluorescence-labelled human TE. We employed a lentiviral system to stably overexpress Citrine-labelled TE to build a fluorescent fibre network. Using immunofluorescence, we confirmed the functionality of the Citrine-tagged TE. Furthermore, we visualized the fibre assembly over the course of several days using confocal microscopy. Applying super resolution microscopy, we were able to investigate the inner structure of the elastin–fibrillin-1 fibre network. Future investigations will allow the tracking of TE produced under various conditions. In tissue engineering applications the fluorescent fibre network can be visualized under various conditions or it serves as a tool for investigating fibre degradation processes in disease-in-a-dish-models. PMID:26842906

  13. Quantum dot-sized organic fluorescent dots for long-term cell tracing

    Science.gov (United States)

    Li, Kai; Tang, Ben Zhong; Liu, Bin

    2014-03-01

    Fluorescence techniques have been extensively employed to develop non-invasive methodologies for tracking and understanding complex biological processes both in vitro and in vivo, which is of high importance in modern life science research. Among a variety of fluorescent probes, inorganic semiconductor quantum dots (QDs) have shown advantages in terms of better photostability, larger Stokes shift and more feasible surface functionalization. However, their intrinsic toxic heavy metal components and unstable fluorescence at low pH greatly impede the applications of QDs in in vivo studies. In this work, we developed novel fluorescent probes that can outperform currently available QD based probes in practice. Using conjugated oligomer with aggregation-induced emission characteristics as the fluorescent domain and biocompatible lipid-PEG derivatives as the encapsulation matrix, the obtained organic dots have shown higher brightness, better stability in biological medium and comparable size and photostability as compared to their counterparts of inorganic QDs. More importantly, unlike QD-based probes, the organic fluorescent dots do not blink, and also do not contain heavy metal ions that could be potentially toxic when applied for living biosubstrates. Upon surface functionalization with a cell-penetrating peptide, the organic dots greatly outperform inorganic quantum dots in both in vitro and in vivo long-term cell tracing studies, which will be beneficial to answer crucial questions in stem cell/immune cell therapies. Considering the customized fluorescent properties and surface functionalities of the organic dots, a series of biocompatible organic dots will be developed to serve as a promising platform for multifarious bioimaging tasks in future.

  14. KCNK5 is Functionally Down-Regulated Upon Long-Term Hypotonicity in Ehrlich Ascites Tumor Cells

    DEFF Research Database (Denmark)

    Kirkegaard, S. S.; Wulff, Tune; Gammeltoft, S.;

    2013-01-01

    Background/Aims: Regulatory volume decrease (RVD) in response to acute cell swelling is well described and KCNK5 (also known as TASK-2 or K2P5.1) has been shown to be the volume sensitive K+ channel in Ehrlich cells. Very little is, on the other hand, known about the effects of long-term hypotoni...... physiological impairment of KCNK5 in Ehrlich cells after long-term hypotonic stimulation is predominantly due to down-regulation of the KCNK5 protein synthesis.© 2013 S. Karger AG, Basel...

  15. Long-term arsenic monitoring with an Enterobacter cloacae microbial fuel cell.

    Science.gov (United States)

    Rasmussen, Michelle; Minteer, Shelley D

    2015-12-01

    A microbial fuel cell was constructed with biofilms of Enterobacter cloacae grown on the anode. Bioelectrocatalysis was observed when the biofilm was grown in media containing sucrose as the carbon source and methylene blue as the mediator. The presence of arsenic caused a decrease in bioelectrocatalytic current. Biofilm growth in the presence of arsenic resulted in lower power outputs whereas addition of arsenic showed no immediate result in power output due to the short term arsenic resistance of the bacteria and slow transport of arsenic across cellular membranes to metabolic enzymes. Calibration curves plotted from the maximum current and maximum power of power curves after growth show that this system is able to quantify both arsenate and arsenate with low detection limits (46 μM for arsenate and 4.4 μM for arsenite). This system could be implemented as a method for long-term monitoring of arsenic concentration in environments where arsenic contamination could occur and alter the metabolism of the organisms resulting in a decrease in power output of the self-powered sensor.

  16. Neurotoxicity in long-term survivors of small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Lee, J.S.; Umsawasdi, T.; Lee, Y.Y.; Barkley, H.T. Jr.; Murphy, W.K.; Welch, S.; Valdivieso, M.

    1986-03-01

    Chronic central nervous system neurotoxicity was studied in 38 long-term survivors (greater than or equal to 3 years) of small cell lung cancer who were treated at the University of Texas M. D. Anderson Hospital and Tumor Institute at Houston between 1971 and 1980. All but one patient received combination chemotherapy with or without chest irradiation. Twenty-four patients received whole brain irradiation (Group I), 22 for elective and two for therapeutic purposes, while 14 did not (Group II). Abnormalities in computed tomographic (CT) scans of the brain were more frequently observed in Group I than in Group II (70% vs. 0%, p less than 0.01). Clinical central nervous system neurotoxicity developed in three patients in Group I, while none developed in patients in Group II (p less than 0.05). Patients who received methotrexate and procarbazine after whole brain irradiation were at a higher risk for clinical central nervous system neurotoxicity (p less than 0.05), and for development of periventricular white matter changes in CT brain scans (p less than 0.05) than were patients in Group II. Impaired methylation of the myelin sheath is proposed as a possible underlying pathogenic mechanism.

  17. CDy6, a photostable probe for long-term real-time visualization of mitosis and proliferating cells.

    Science.gov (United States)

    Jeong, Yun-Mi; Duanting, Zhai; Hennig, Holger; Samanta, Animesh; Agrawalla, Bikram Keshari; Bray, Mark-Anthony; Carpenter, Anne E; Chang, Young-Tae

    2015-02-19

    Long-term real-time visualization of lysosomal dynamics has been challenging at the onset of mitosis due to the lack of fluorescent probes enabling convenient imaging of dividing cells. We developed a long-term real-time photostable mitotic or proliferating marker, CDy6, a BODIPY-derived compound of designation yellow 6, which labels lysosome. In long-term real-time, CDy6 displayed a sharp increase in intensity and change in localization in mitosis, improved photostability, and decreased toxicity compared with other widely used lysosomal and DNA markers, and the ability to label cells in mouse xenograft models. Therefore, CDy6 may open new possibilities to target and trace lysosomal contents during mitosis and to monitor cell proliferation, which can further our knowledge of the basic underlying biological mechanisms in the management of cancer.

  18. Short- and long-term effects of T-cell modulating agents in experimental autoimmunity

    International Nuclear Information System (INIS)

    Due to the easy and reliable induction of a disease condition with many of the features present in human autoimmunity, mercury-induced autoimmunity (mHgAI) in rodents is a favourable autoimmune model. Genetically susceptible (H-2s) mice develop in response to mercury (Hg) a systemic autoimmune condition with antinucleolar antibodies (ANoA) targeting the protein fibrillarin, transient polyclonal B-cell activation, hyperimmunoglobulinemia, and systemic immune-complex (IC) deposits. In order to study the short- and long-term effects of treatment with immunomodulating agents on the disease parameters in HgAI, groups of B10.S (H-2s) mice were given 6 mg HgCl2/l drinking water for 22 weeks. Three weeks initial treatment with cyclosporin A (CyA), a high dose of tacrolimus (HD tacrolimus), or anti-CD4 monoclonal antibody (a-CD4) inhibited induction of ANoA and IC deposit by Hg. This effect persisted for the subsequent 19 weeks when the mice were only treated with Hg. Initial treatment with anti-IL-4 monoclonal antibody (a-IL-4) for 3 weeks inhibited induction of IgE and IC deposits by Hg, but not ANoA. However, subsequent treatment with Hg without a-IL-4 for 19 weeks induced IC deposits. The T-cell modulating agents aggravated some of the HgAI disease parameters: a-CD4 stimulated the polyclonal B-cell activation, a-IL-4 increased the IgG antichromatin antibody response, and a low dose of tacrolimus (LD tacrolimus) enhanced the ANoA, the polyclonal B-cell activation, and the IC deposits. We conclude that a short initial treatment with a-CD4 or CyA efficiently protects against induction of systemic autoimmunity for an extended period of time. However, some of the T-cell modulating agents, especially a low dose of tacrolimus, aggravate autoimmune manifestations not only during ongoing treatment, but also after treatment with these agents has ceased

  19. Toxicological Effects of Caco-2 Cells Following Short-Term and Long-Term Exposure to Ag Nanoparticles

    Science.gov (United States)

    Chen, Ni; Song, Zheng-Mei; Tang, Huan; Xi, Wen-Song; Cao, Aoneng; Liu, Yuanfang; Wang, Haifang

    2016-01-01

    Extensive utilization increases the exposure of humans to Ag nanoparticles (NPs) via the oral pathway. To comprehensively address the action of Ag NPs to the gastrointestinal systems in real situations, i.e., the long-term low-dose exposure, we evaluated and compared the toxicity of three Ag NPs (20–30 nm with different surface coatings) to the human intestine cell Caco-2 after 1-day and 21-day exposures, using various biological assays. In both the short- and long-term exposures, the variety of surface coating predominated the toxicity of Ag NPs in a descending order of citrate-coated Ag NP (Ag-CIT), bare Ag NP (Ag-B), and poly (N-vinyl-2-pyrrolidone)-coated Ag NP (Ag-PVP). The short-term exposure induced cell growth inhibition and death. The cell viability loss appeared after cells were exposed to 0.7 μg/mL Ag-CIT, 0.9 μg/mL Ag-B or >1.0 μg/mL Ag-PVP for 24 h. The short-term and higher-dose exposure also induced reactive oxygen species (ROS) generation, mitochondrial damage, cell membrane leakage, apoptosis, and inflammation (IL-8 level). The long-term exposure only inhibited the cell proliferation. After 21-day exposure to 0.4 μg/mL Ag-CIT, the cell viability dropped to less than 50%, while cells exposed to 0.5 μg/mL Ag-PVP remained normal as the control. Generally, 0.3 μg/mL is the non-toxic dose for the long-term exposure of Caco-2 cells to Ag NPs in this study. However, cells presented inflammation after exposure to Ag NPs with the non-toxic dose in the long-term exposure. PMID:27338357

  20. Toxicological Effects of Caco-2 Cells Following Short-Term and Long-Term Exposure to Ag Nanoparticles

    Directory of Open Access Journals (Sweden)

    Ni Chen

    2016-06-01

    Full Text Available Extensive utilization increases the exposure of humans to Ag nanoparticles (NPs via the oral pathway. To comprehensively address the action of Ag NPs to the gastrointestinal systems in real situations, i.e., the long-term low-dose exposure, we evaluated and compared the toxicity of three Ag NPs (20–30 nm with different surface coatings to the human intestine cell Caco-2 after 1-day and 21-day exposures, using various biological assays. In both the short- and long-term exposures, the variety of surface coating predominated the toxicity of Ag NPs in a descending order of citrate-coated Ag NP (Ag-CIT, bare Ag NP (Ag-B, and poly (N-vinyl-2-pyrrolidone-coated Ag NP (Ag-PVP. The short-term exposure induced cell growth inhibition and death. The cell viability loss appeared after cells were exposed to 0.7 μg/mL Ag-CIT, 0.9 μg/mL Ag-B or >1.0 μg/mL Ag-PVP for 24 h. The short-term and higher-dose exposure also induced reactive oxygen species (ROS generation, mitochondrial damage, cell membrane leakage, apoptosis, and inflammation (IL-8 level. The long-term exposure only inhibited the cell proliferation. After 21-day exposure to 0.4 μg/mL Ag-CIT, the cell viability dropped to less than 50%, while cells exposed to 0.5 μg/mL Ag-PVP remained normal as the control. Generally, 0.3 μg/mL is the non-toxic dose for the long-term exposure of Caco-2 cells to Ag NPs in this study. However, cells presented inflammation after exposure to Ag NPs with the non-toxic dose in the long-term exposure.

  1. Cell type-specific responses of peripheral blood mononuclear cells to silver nanoparticles.

    Science.gov (United States)

    Greulich, C; Diendorf, J; Gessmann, J; Simon, T; Habijan, T; Eggeler, G; Schildhauer, T A; Epple, M; Köller, M

    2011-09-01

    Silver nanoparticles (Ag-NP) are increasingly used in biomedical applications because of their remarkable antimicrobial activity. In biomedicine, Ag-NP are coated onto or embedded in wound dressings, surgical instruments and bone substitute biomaterials, such as silver-containing calcium phosphate cements. Free Ag-NP and silver ions are released from these coatings or after the degradation of a biomaterial, and may come into close contact with blood cells. Despite the widespread use of Ag-NP as an antimicrobial agent, there is a serious lack of information on the biological effects of Ag-NP on human blood cells. In this study, the uptake of Ag-NP by peripheral monocytes and lymphocytes (T-cells) was analyzed, and the influence of nanosilver on cell biological functions (proliferation, the expression of adhesion molecules, cytokine release and the generation of reactive oxygen species) was studied. After cell culture in the presence of monodispersed Ag-NP (5-30μgml(-1) silver concentration), agglomerates of nanoparticles were detected within monocytes (CD14+) but not in T-cells (CD3+) by light microscopy, flow cytometry and combined focused ion beam/scanning electron microscopy. The uptake rate of nanoparticles was concentration dependent, and the silver agglomerates were typically found in the cytoplasm. Furthermore, a concentration-dependent activation (e.g. an increased expression of adhesion molecule CD54) of monocytes at Ag-NP concentrations of 10-15μgml(-1) was observed, and cytotoxicity of Ag-NP-treated monocytes was observed at Ag-NP levels of 25μgml(-1) and higher. However, no modulation of T-cell proliferation was observed in the presence of Ag-NP. Taken together, our results provide the first evidence for a cell-type-specific uptake of Ag-NP by peripheral blood mononuclear cells (PBMC) and the resultant cellular responses after exposure.

  2. Long-Term Outcomes of Sacrococcygeal Germ Cell Tumors in Infancy and Childhood

    Directory of Open Access Journals (Sweden)

    Rangsan Niramis

    2015-01-01

    Full Text Available Purpose. The aim of this study was to evaluate long-term outcomes of sacrococcygeal germ cell tumors (SC-GCTs over a 15-year period. Materials and Methods. A retrospective review was conducted of all pediatric patients treated for SC-GCTs at our hospital from 1998 to 2012. Results. Fifty-seven patients were treated for SC-GCTs with the most common in Altman’s classification type I. Age at surgery ranged from one day to 5.6 years. Tumor resection and coccygectomy were primarily performed in about 84% of the cases. Pathology revealed mature, immature, malignant sacrococcygeal teratomas (SCTs, and endodermal sinus tumors (ESTs in 41 (72%, 4 (77%, 6 (10.5%, and 6 (10.5%, respectively. Recurrence of discase occurred in 3 of 41 patients with mature teratomas (7.3%; 2 recurrences with mature teratomas and one recurrence with EST. Five of 6 malignant SCTs and 3 of 6 ESTs responded well to the treatment. Alpha-fetoprotein (AFP level was elevated in both malignant teratomas and ESTs. No immediate patient death was noted in any of the 57 cases, but 4 patients with malignant tumors and distant metastasis succumbed at home within 2 years of the initial treatment. Conclusion. Benign SCTs have a significant recurrence rate of approximately 7%. Close follow-up with serial AFP level monitoring should be done for 5 years after initial tumor resection and coccygectomy. The survival rate for malignant SC-GCTs with distant metastasis was unfavorable in the present study.

  3. Phenotypic and Functional Properties of Porcine Dedifferentiated Fat Cells during the Long-Term Culture In Vitro

    OpenAIRE

    Xuewu Peng; Tongxing Song; Xiaoming Hu; Yuanfei Zhou; Hongkui Wei; Jian Peng; Siwen Jiang

    2015-01-01

    It has been proved that terminally differentiated mature adipocytes possess abilities to dedifferentiate into fibroblast-like progeny cells with self-renewal and multiple differentiation, termed dedifferentiated fat (DFAT) cells. However, the biological properties of DFAT cells during long-term culture in vitro have not been elucidated. Here, we obtained fibroblast-like morphology of porcine DFAT cells by ceiling culture. During the dedifferentiation process, round mature adipocytes with sing...

  4. Long term observations in combined modality therapy for limited stage small cell lung cancer

    International Nuclear Information System (INIS)

    Purpose/Objective: With the discovery that patients with small cell lung cancer (SCLC) exhibit a high level of sensitivity to both chemotherapy and radiotherapy, the treatment of SCLC became a model for the success of combined modality treatment. In this retrospective review, we analyze the outcomes and patterns of failure when patients are treated with chemotherapy and thoracic irradiation. The relative values of sequential and concurrent chemotherapy, in conjunction with chest irradiation, are assessed. The potential benefit of prophylactic cranial irradiation is explored. The impact of prognostic factors for long term survival of SCLC patients are examined to identify pretreatment patient characteristics and treatment parameters which might predict for a favorable outcome. Materials and Methods: We identified 190 patients treated at M.D. Anderson Cancer Center from January 1985 to December 1992 with curative intent for limited stage SCLC. Prognostic factors were determined using univariate and multivariate analysis. The significant covariates for each outcome endpoint were evaluated. Probabilities of local failure, overall survival, relapse-free survival, and distant metastasis-free survival were calculated from the time of treatment using actuarial life table analysis. Results: The median age was 61, with 51% males. There were 119 patients treated sequentially, and 71 concurrently. The Karnofsky Performance Status was >= 90 in 48% of patients in the concurrent cohort, vs. 35% of the sequential group. Prophylactic cranial irradiation (PCI) was delivered in 117 cases (62%). There were 51 long term survivors, defined as survival >=36 months. The median follow-up in surviving patients was 75 months. At the time of the analysis, 166 patients (87%) had expired. The crude 2 and 3 year survival rate for the entire group was 38.4% and 26.8%, respectively. The actuarial 2-year survival was 39.9%, and at 3 years the actuarial survival was 27.8%. The median actuarial

  5. Tumorigenic heterogeneity in cancer stem cells evolved from long-term cultures of telomerase-immortalized human mesenchymal stem cells

    DEFF Research Database (Denmark)

    Burns, Jorge S; Abdallah, Basem M; Guldberg, Per;

    2005-01-01

    Long-term cultures of telomerase-transduced adult human mesenchymal stem cells (hMSC) may evolve spontaneous genetic changes leading to tumorigenicity in immunodeficient mice (e.g., hMSC-TERT20). We wished to clarify whether this unusual phenotype reflected a rare but dominant subpopulation...... or if the stem cell origin allowed most cells to behave as cancer stem cells. Cultures of the hMSC-TERT20 strain at population doubling 440 were highly clonogenic (94%). From 110 single-cell clones expanded by 20 population doublings, 6 underwent detailed comparison. Like the parental population, each clone had...... tumorigenicity correlated with good viability plus capillary morphogenesis on serum starvation and high cyclin D1 expression. Thus, hMSC-TERT20 clones represent cancer stem cells with hierarchical tumorigenicity, providing new models to explore the stem cell hypothesis for cancer....

  6. Long-term depression of climbing fiber-evoked calcium transients in Purkinje cell dendrites.

    NARCIS (Netherlands)

    J.T. Weber (John); C.I. de Zeeuw (Chris); C.R.W. Hansel (Christian); D.J. Linden (David)

    2003-01-01

    textabstractIn recent years much has been learned about the molecular requirements for inducing long-term synaptic depression (LTD) in various brain regions. However, very little is known about the consequences of LTD induction for subsequent signaling events in postsynaptic neurons. We have address

  7. Accumulation of apoptosis-insensitive human bone marrow-mesenchymal stromal cells after long-term expansion.

    Science.gov (United States)

    Jeong, Sin-Gu; Cho, Goang-Won

    2016-07-01

    Cells undergo replicative senescence during in vitro expansion, which is induced by the accumulation of cellular damage caused by excessive reactive oxygen species. In this study, we investigated whether long-term-cultured human bone marrow mesenchymal stromal cells (MSCs) are insensitive to apoptotic stimulation. To examine this, we established replicative senescent cells from long-term cultures of human bone marrow MSCs. Senescent cells were identified based on declining population doublings, increased expression of senescence markers p16 and p53 and increased senescence-associated β-gal activity. In cell viability assays, replicative senescent MSCs in late passages (i.e. 15-19 passages) resisted damage induced by oxidative stress more than those in early passages did (i.e. 7-10 passages). This resistance occurred via caspase-9 and caspase-3 rather than via caspase-8. The senescent cells are gradually accumulated during long-term expansion. The oxidative stress-sensitive proteins ataxia-telangiectasia mutated and p53 were phosphorylated, and the expression of apoptosis molecules Bax increased, and Bcl-2 decreased in early passage MSCs; however, the expression of the apoptotic molecules did less change in response to apoptotic stimulation in late-passage MSCs, suggesting that the intrinsic apoptotic signalling pathway was not induced by oxidative stress in long-term-cultured MSCs. Based on these results, we propose that some replicative senescent cells may avoid apoptosis signalling via impairment of signalling molecules and accumulation during long-term expansion. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27212655

  8. Melatonin Treatment Improves Mesenchymal Stem Cells Therapy by Preserving Stemness during Long-term In Vitro Expansion

    Science.gov (United States)

    Shuai, Yi; Liao, Li; Su, Xiaoxia; Yu, Yang; Shao, Bingyi; Jing, Huan; Zhang, Xinjing; Deng, Zhihong; Jin, Yan

    2016-01-01

    Mesenchymal stem cells (MSCs) are promising candidates for tissue regeneration and disease treatment. However, long-term in vitro passaging leads to stemness loss of MSCs, resulting in failure of MSCs therapy. Here, we report a melatonin-based strategy to improve cell therapy of in vitro cultured MSCs. Among four small molecules with anti-aging and stem cell-protection properties (rapamycin, resveratrol, quercetin and melatonin), colony forming, proliferation, and osteogenic differentiation assay showed that melatonin was the most efficient to preserve self-renewal and differentiation properties of rat bone marrow MSCs (BMMSCs) after long-term passaging. Functional assays confirmed melatonin treatment did not affect the colony forming, proliferation and osteogenic differentiation of BMMSCs cultured for 1 or 4 passages, but largely prevented the decline of self-renew and differentiation capacity of BMMSCs cultured for 15 passages in vitro. Furthermore, heterotopic osteogenesis assay, critical size calvarial defects repair assay, osteoporosis treatment and experimental colitis therapy assay strongly certified that melatonin preserved the therapeutic effect of long-term passaged BMMSCs on bone regeneration and immunotherapy in vivo. Mechanistically, melatonin functioned by activating antioxidant defense system, inhibiting the pathway of cell senescence, and preserving the expression of gene governing the stemness. Taken together, our findings showed that melatonin treatment efficiently prevented the dysfunction and therapeutic failure of BMMSCs after long-term passaging, providing a practical strategy to improve the application of BMMSCs in tissue engineering and cytotherapy. PMID:27570559

  9. Melatonin Treatment Improves Mesenchymal Stem Cells Therapy by Preserving Stemness during Long-term In Vitro Expansion.

    Science.gov (United States)

    Shuai, Yi; Liao, Li; Su, Xiaoxia; Yu, Yang; Shao, Bingyi; Jing, Huan; Zhang, Xinjing; Deng, Zhihong; Jin, Yan

    2016-01-01

    Mesenchymal stem cells (MSCs) are promising candidates for tissue regeneration and disease treatment. However, long-term in vitro passaging leads to stemness loss of MSCs, resulting in failure of MSCs therapy. Here, we report a melatonin-based strategy to improve cell therapy of in vitro cultured MSCs. Among four small molecules with anti-aging and stem cell-protection properties (rapamycin, resveratrol, quercetin and melatonin), colony forming, proliferation, and osteogenic differentiation assay showed that melatonin was the most efficient to preserve self-renewal and differentiation properties of rat bone marrow MSCs (BMMSCs) after long-term passaging. Functional assays confirmed melatonin treatment did not affect the colony forming, proliferation and osteogenic differentiation of BMMSCs cultured for 1 or 4 passages, but largely prevented the decline of self-renew and differentiation capacity of BMMSCs cultured for 15 passages in vitro. Furthermore, heterotopic osteogenesis assay, critical size calvarial defects repair assay, osteoporosis treatment and experimental colitis therapy assay strongly certified that melatonin preserved the therapeutic effect of long-term passaged BMMSCs on bone regeneration and immunotherapy in vivo. Mechanistically, melatonin functioned by activating antioxidant defense system, inhibiting the pathway of cell senescence, and preserving the expression of gene governing the stemness. Taken together, our findings showed that melatonin treatment efficiently prevented the dysfunction and therapeutic failure of BMMSCs after long-term passaging, providing a practical strategy to improve the application of BMMSCs in tissue engineering and cytotherapy. PMID:27570559

  10. Cell-type-specific, Aptamer-functionalized Agents for Targeted Disease Therapy.

    Science.gov (United States)

    Zhou, Jiehua; Rossi, John J

    2014-06-17

    One hundred years ago, Dr. Paul Ehrlich popularized the "magic bullet" concept for cancer therapy in which an ideal therapeutic agent would only kill the specific tumor cells it targeted. Since then, "targeted therapy" that specifically targets the molecular defects responsible for a patient's condition has become a long-standing goal for treating human disease. However, safe and efficient drug delivery during the treatment of cancer and infectious disease remains a major challenge for clinical translation and the development of new therapies. The advent of SELEX technology has inspired many groundbreaking studies that successfully adapted cell-specific aptamers for targeted delivery of active drug substances in both in vitro and in vivo models. By covalently linking or physically functionalizing the cell-specific aptamers with therapeutic agents, such as siRNA, microRNA, chemotherapeutics or toxins, or delivery vehicles, such as organic or inorganic nanocarriers, the targeted cells and tissues can be specifically recognized and the therapeutic compounds internalized, thereby improving the local concentration of the drug and its therapeutic efficacy. Currently, many cell-type-specific aptamers have been developed that can target distinct diseases or tissues in a cell-type-specific manner. In this review, we discuss recent advances in the use of cell-specific aptamers for targeted disease therapy, as well as conjugation strategies and challenges.

  11. Fast stack activation procedure and effective long-term storage for high-performance polymer electrolyte membrane fuel cell

    Science.gov (United States)

    Yang, Seung Yong; Seo, Dong-Jun; Kim, Myeong-Ri; Seo, Min Ho; Hwang, Sun-Mi; Jung, Yong-Min; Kim, Beom-Jun; Yoon, Young-Gi; Han, Byungchan; Kim, Tae-Young

    2016-10-01

    Time-saving stack activation and effective long-term storage are one of most important issues that must be resolved for the commercialization of polymer electrolyte membrane fuel cell (PEMFC). Herein, we developed the cost-effective stack activation method to finish the whole activation within 30 min and the long-term storage method by using humidified N2 without any significant decrease in cell's performance for 30 days. Specifically, the pre-activation step with the direct injection of DI water into the stack and storage at 65 or 80 °C for 2 h increases the distinctive phase separation between the hydrophobic and hydrophilic regions in Nafion membrane, which significantly reduces the total activation time within 30 min. Additionally, the long-term storage with humidified N2 has no effect on the Pt oxidation and drying of Nafion membrane for 30 days due to its exergonic reaction in the cell. As a result, the high water content in Nafion membrane and the decrease of Pt oxidation are the critical factors that have a strong influence on the activation and long-term storage for high-performance PEMFC.

  12. Impact of process variations and long term degradation on 6T-SRAM cells

    OpenAIRE

    Fischer, Th.; Olbrich, A.; G. Georgakos; Lemaitre, B.; D. Schmitt-Landsiedel

    2007-01-01

    In modern deep-submicron CMOS technologies voltage scaling can not keep up with the scaling of the dimensions of transistors. Therefore the electrical fields inside the transistors are not constant anymore, while scaling down the device area. The rising electrical fields bring up reliability problems, such as hot carrier injection. Also other long term degradation mechanisms like Negative Bias Temperature Instability (NBTI) come into the focus of circuit design.

    Along wit...

  13. General and cell-type specific mechanisms target TRPP2/PKD-2 to cilia.

    Science.gov (United States)

    Bae, Young-Kyung; Qin, Hongmin; Knobel, Karla M; Hu, Jinghua; Rosenbaum, Joel L; Barr, Maureen M

    2006-10-01

    Ciliary localization of the transient receptor potential polycystin 2 channel (TRPP2/PKD-2) is evolutionarily conserved, but how TRPP2 is targeted to cilia is not known. In this study, we characterize the motility and localization of PKD-2, a TRPP2 homolog, in C. elegans sensory neurons. We demonstrate that GFP-tagged PKD-2 moves bidirectionally in the dendritic compartment. Furthermore, we show a requirement for different molecules in regulating the ciliary localization of PKD-2. PKD-2 is directed to moving dendritic particles by the UNC-101/adaptor protein 1 (AP-1) complex. When expressed in non-native neurons, PKD-2 remains in cell bodies and is not observed in dendrites or cilia, indicating that cell-type specific factors are required for directing PKD-2 to the dendrite. PKD-2 stabilization in cilia and cell bodies requires LOV-1, a functional partner and a TRPP1 homolog. In lov-1 mutants, PKD-2 is greatly reduced in cilia and forms abnormal aggregates in neuronal cell bodies. Intraflagellar transport (IFT) is not essential for PKD-2 dendritic motility or access to the cilium, but may regulate PKD-2 ciliary abundance. We propose that both general and cell-type-specific factors govern TRPP2/PKD-2 subcellular distribution by forming at least two steps involving somatodendritic and ciliary sorting decisions. PMID:16943275

  14. Long Term Aggresome Accumulation Leads to DNA Damage, p53-dependent Cell Cycle Arrest, and Steric Interference in Mitosis.

    Science.gov (United States)

    Lu, Meng; Boschetti, Chiara; Tunnacliffe, Alan

    2015-11-13

    Juxtanuclear aggresomes form in cells when levels of aggregation-prone proteins exceed the capacity of the proteasome to degrade them. It is widely believed that aggresomes have a protective function, sequestering potentially damaging aggregates until these can be removed by autophagy. However, most in-cell studies have been carried out over a few days at most, and there is little information on the long term effects of aggresomes. To examine these long term effects, we created inducible, single-copy cell lines that expressed aggregation-prone polyglutamine proteins over several months. We present evidence that, as perinuclear aggresomes accumulate, they are associated with abnormal nuclear morphology and DNA double-strand breaks, resulting in cell cycle arrest via the phosphorylated p53 (Ser-15)-dependent pathway. Further analysis reveals that aggresomes can have a detrimental effect on mitosis by steric interference with chromosome alignment, centrosome positioning, and spindle formation. The incidence of apoptosis also increased in aggresome-containing cells. These severe defects developed gradually after juxtanuclear aggresome formation and were not associated with small cytoplasmic aggregates alone. Thus, our findings demonstrate that, in dividing cells, aggresomes are detrimental over the long term, rather than protective. This suggests a novel mechanism for polyglutamine-associated developmental and cell biological abnormalities, particularly those with early onset and non-neuronal pathologies.

  15. Induced neural stem cells achieve long-term survival and functional integration in the adult mouse brain.

    Science.gov (United States)

    Hemmer, Kathrin; Zhang, Mingyue; van Wüllen, Thea; Sakalem, Marna; Tapia, Natalia; Baumuratov, Aidos; Kaltschmidt, Christian; Kaltschmidt, Barbara; Schöler, Hans R; Zhang, Weiqi; Schwamborn, Jens C

    2014-09-01

    Differentiated cells can be converted directly into multipotent neural stem cells (i.e., induced neural stem cells [iNSCs]). iNSCs offer an attractive alternative to induced pluripotent stem cell (iPSC) technology with regard to regenerative therapies. Here, we show an in vivo long-term analysis of transplanted iNSCs in the adult mouse brain. iNSCs showed sound in vivo long-term survival rates without graft overgrowths. The cells displayed a neural multilineage potential with a clear bias toward astrocytes and a permanent downregulation of progenitor and cell-cycle markers, indicating that iNSCs are not predisposed to tumor formation. Furthermore, the formation of synaptic connections as well as neuronal and glial electrophysiological properties demonstrated that differentiated iNSCs migrated, functionally integrated, and interacted with the existing neuronal circuitry. We conclude that iNSC long-term transplantation is a safe procedure; moreover, it might represent an interesting tool for future personalized regenerative applications. PMID:25241741

  16. Induced Neural Stem Cells Achieve Long-Term Survival and Functional Integration in the Adult Mouse Brain

    Directory of Open Access Journals (Sweden)

    Kathrin Hemmer

    2014-09-01

    Full Text Available Differentiated cells can be converted directly into multipotent neural stem cells (i.e., induced neural stem cells [iNSCs]. iNSCs offer an attractive alternative to induced pluripotent stem cell (iPSC technology with regard to regenerative therapies. Here, we show an in vivo long-term analysis of transplanted iNSCs in the adult mouse brain. iNSCs showed sound in vivo long-term survival rates without graft overgrowths. The cells displayed a neural multilineage potential with a clear bias toward astrocytes and a permanent downregulation of progenitor and cell-cycle markers, indicating that iNSCs are not predisposed to tumor formation. Furthermore, the formation of synaptic connections as well as neuronal and glial electrophysiological properties demonstrated that differentiated iNSCs migrated, functionally integrated, and interacted with the existing neuronal circuitry. We conclude that iNSC long-term transplantation is a safe procedure; moreover, it might represent an interesting tool for future personalized regenerative applications.

  17. Reprogramming suppresses premature senescence phenotypes of Werner syndrome cells and maintains chromosomal stability over long-term culture.

    Directory of Open Access Journals (Sweden)

    Akira Shimamoto

    Full Text Available Werner syndrome (WS is a premature aging disorder characterized by chromosomal instability and cancer predisposition. Mutations in WRN are responsible for the disease and cause telomere dysfunction, resulting in accelerated aging. Recent studies have revealed that cells from WS patients can be successfully reprogrammed into induced pluripotent stem cells (iPSCs. In the present study, we describe the effects of long-term culture on WS iPSCs, which acquired and maintained infinite proliferative potential for self-renewal over 2 years. After long-term cultures, WS iPSCs exhibited stable undifferentiated states and differentiation capacity, and premature upregulation of senescence-associated genes in WS cells was completely suppressed in WS iPSCs despite WRN deficiency. WS iPSCs also showed recapitulation of the phenotypes during differentiation. Furthermore, karyotype analysis indicated that WS iPSCs were stable, and half of the descendant clones had chromosomal profiles that were similar to those of parental cells. These unexpected properties might be achieved by induced expression of endogenous telomerase gene during reprogramming, which trigger telomerase reactivation leading to suppression of both replicative senescence and telomere dysfunction in WS cells. These findings demonstrated that reprogramming suppressed premature senescence phenotypes in WS cells and WS iPSCs could lead to chromosomal stability over the long term. WS iPSCs will provide opportunities to identify affected lineages in WS and to develop a new strategy for the treatment of WS.

  18. Ligation-free ribosome profiling of cell type-specific translation in the brain.

    Science.gov (United States)

    Hornstein, Nicholas; Torres, Daniela; Das Sharma, Sohani; Tang, Guomei; Canoll, Peter; Sims, Peter A

    2016-01-01

    Ribosome profiling has emerged as a powerful tool for genome-wide measurements of translation, but library construction requires multiple ligation steps and remains cumbersome relative to more conventional deep-sequencing experiments. We report a new, ligation-free approach to ribosome profiling that does not require ligation. Library construction for ligation-free ribosome profiling can be completed in one day with as little as 1 ng of purified RNA footprints. We apply ligation-free ribosome profiling to mouse brain tissue to identify new patterns of cell type-specific translation and test its ability to identify translational targets of mTOR signaling in the brain.

  19. In Vivo Transplantation of Enteric Neural Crest Cells into Mouse Gut; Engraftment, Functional Integration and Long-Term Safety.

    Directory of Open Access Journals (Sweden)

    Julie E Cooper

    Full Text Available Enteric neuropathies are severe gastrointestinal disorders with unsatisfactory outcomes. We aimed to investigate the potential of enteric neural stem cell therapy approaches for such disorders by transplanting mouse enteric neural crest cells (ENCCs into ganglionic and aganglionic mouse gut in vivo and analysing functional integration and long-term safety.Neurospheres generated from yellow fluorescent protein (YFP expressing ENCCs selected from postnatal Wnt1-cre;R26R-YFP/YFP murine gut were transplanted into ganglionic hindgut of wild-type littermates or aganglionic hindgut of Ednrbtm1Ywa mice (lacking functional endothelin receptor type-B. Intestines were then assessed for ENCC integration and differentiation using immunohistochemistry, cell function using calcium imaging, and long-term safety using PCR to detect off-target YFP expression.YFP+ ENCCs engrafted, proliferated and differentiated into enteric neurons and glia within recipient ganglionic gut. Transplanted cells and their projections spread along the endogenous myenteric plexus to form branching networks. Electrical point stimulation of endogenous nerve fibres resulted in calcium transients (F/F0 = 1.16 ± 0.01;43 cells, n = 6 in YFP+ transplanted ENCCs (abolished with TTX. Long-term follow-up (24 months showed transplanted ENCCs did not give rise to tumours or spread to other organs (PCR negative in extraintestinal sites. In aganglionic gut ENCCs similarly spread and differentiated to form neuronal and glial networks with projections closely associated with endogenous neural networks of the transition zone.Transplanted ENCCs successfully engrafted into recipient ganglionic and aganglionic gut showing appropriate spread, localisation and, importantly, functional integration without any long-term safety issues. This study provides key support for the development and use of enteric neural stem cell therapies.

  20. Isolation and Assessment of Single Long-Term Reconstituting Hematopoietic Stem Cells from Adult Mouse Bone Marrow.

    Science.gov (United States)

    Kent, David G; Dykstra, Brad J; Eaves, Connie J

    2016-01-01

    Hematopoietic stem cells with long-term repopulating activity can now be routinely obtained at purities of 40% to 50% from suspensions of adult mouse bone marrow. Here we describe robust protocols for both their isolation as CD45(+) EPCR(+) CD150(+) CD48(-) (ESLAM) cells using multiparameter cell sorting and for tracking their clonal growth and differentiation activity in irradiated mice transplanted with single ESLAM cells. The simplicity of these procedures makes them attractive for characterizing the molecular and biological properties of individual hematopoietic stem cells with unprecedented power and precision. © 2016 by John Wiley & Sons, Inc. PMID:27532815

  1. The roadmap for estimation of cell-type-specific neuronal activity from non-invasive measurements.

    Science.gov (United States)

    Uhlirova, Hana; Kılıç, Kıvılcım; Tian, Peifang; Sakadžić, Sava; Gagnon, Louis; Thunemann, Martin; Desjardins, Michèle; Saisan, Payam A; Nizar, Krystal; Yaseen, Mohammad A; Hagler, Donald J; Vandenberghe, Matthieu; Djurovic, Srdjan; Andreassen, Ole A; Silva, Gabriel A; Masliah, Eliezer; Kleinfeld, David; Vinogradov, Sergei; Buxton, Richard B; Einevoll, Gaute T; Boas, David A; Dale, Anders M; Devor, Anna

    2016-10-01

    The computational properties of the human brain arise from an intricate interplay between billions of neurons connected in complex networks. However, our ability to study these networks in healthy human brain is limited by the necessity to use non-invasive technologies. This is in contrast to animal models where a rich, detailed view of cellular-level brain function with cell-type-specific molecular identity has become available due to recent advances in microscopic optical imaging and genetics. Thus, a central challenge facing neuroscience today is leveraging these mechanistic insights from animal studies to accurately draw physiological inferences from non-invasive signals in humans. On the essential path towards this goal is the development of a detailed 'bottom-up' forward model bridging neuronal activity at the level of cell-type-specific populations to non-invasive imaging signals. The general idea is that specific neuronal cell types have identifiable signatures in the way they drive changes in cerebral blood flow, cerebral metabolic rate of O2 (measurable with quantitative functional Magnetic Resonance Imaging), and electrical currents/potentials (measurable with magneto/electroencephalography). This forward model would then provide the 'ground truth' for the development of new tools for tackling the inverse problem-estimation of neuronal activity from multimodal non-invasive imaging data.This article is part of the themed issue 'Interpreting BOLD: a dialogue between cognitive and cellular neuroscience'. PMID:27574309

  2. Mouse pancreas tissue slice culture facilitates long-term studies of exocrine and endocrine cell physiology in situ.

    Directory of Open Access Journals (Sweden)

    Anja Marciniak

    Full Text Available Studies on pancreatic cell physiology rely on the investigation of exocrine and endocrine cells in vitro. Particularly, in the case of the exocrine tissue these studies have suffered from a reduced functional viability of acinar cells in culture. As a result not only investigations on dispersed acinar cells and isolated acini were limited in their potential, but also prolonged studies on pancreatic exocrine and endocrine cells in an intact pancreatic tissue environment were unfeasible. To overcome these limitations, we aimed to establish a pancreas tissue slice culture platform to allow long-term studies on exocrine and endocrine cells in the intact pancreatic environment. Mouse pancreas tissue slice morphology was assessed to determine optimal long-term culture settings for intact pancreatic tissue. Utilizing optimized culture conditions, cell specificity and function of exocrine acinar cells and endocrine beta cells were characterized over a culture period of 7 days. We found pancreas tissue slices cultured under optimized conditions to have intact tissue specific morphology for the entire culture period. Amylase positive intact acini were present at all time points of culture and acinar cells displayed a typical strong cell polarity. Amylase release from pancreas tissue slices decreased during culture, but maintained the characteristic bell-shaped dose-response curve to increasing caerulein concentrations and a ca. 4-fold maximal over basal release. Additionally, endocrine beta cell viability and function was well preserved until the end of the observation period. Our results show that the tissue slice culture platform provides unprecedented maintenance of pancreatic tissue specific morphology and function over a culture period for at least 4 days and in part even up to 1 week. This analytical advancement now allows mid -to long-term studies on the cell biology of pancreatic disorder pathogenesis and therapy in an intact surrounding in situ.

  3. Phenotypic and Functional Properties of Porcine Dedifferentiated Fat Cells during the Long-Term Culture In Vitro

    Directory of Open Access Journals (Sweden)

    Xuewu Peng

    2015-01-01

    Full Text Available It has been proved that terminally differentiated mature adipocytes possess abilities to dedifferentiate into fibroblast-like progeny cells with self-renewal and multiple differentiation, termed dedifferentiated fat (DFAT cells. However, the biological properties of DFAT cells during long-term culture in vitro have not been elucidated. Here, we obtained fibroblast-like morphology of porcine DFAT cells by ceiling culture. During the dedifferentiation process, round mature adipocytes with single large lipid droplets changed into spindle-shaped cells accompanied by the adipogenic markers PPARγ, aP2, LPL, and Adiponectin significant downregulation. Flow cytometric analysis showed that porcine DFAT cells displayed similar cell-surface antigen profile to mesenchymal stem cells (MSCs. Furthermore, different passages of porcine DFAT cells during long-term culture in vitro retained high levels of cell viabilities (>97%, efficient proliferative capacity including population doubling time ranged from 20 h to 22 h and population doubling reached 47.40±1.64 by 58 days of culture. In addition, porcine DFAT cells maintained the multiple differentiation capabilities into adipocytes, osteoblasts, and skeletal myocytes and displayed normal chromosomal karyotypes for prolonged passaging. Therefore, porcine DFAT cells may be a novel model of stem cells for studying the functions of gene in the different biological events.

  4. Cell type-specific bipolar cell input to ganglion cells in the mouse retina.

    Science.gov (United States)

    Neumann, S; Hüser, L; Ondreka, K; Auler, N; Haverkamp, S

    2016-03-01

    Many distinct ganglion cell types, which are the output elements of the retina, were found to encode for specific features of a visual scene such as contrast, color information or movement. The detailed composition of retinal circuits leading to this tuning of retinal ganglion cells, however, is apart from some prominent examples, largely unknown. Here we aimed to investigate if ganglion cell types in the mouse retina receive selective input from specific bipolar cell types or if they sample their synaptic input non-selectively from all bipolar cell types stratifying within their dendritic tree. To address this question we took an anatomical approach and immunolabeled retinae of two transgenic mouse lines (GFP-O and JAM-B) with markers for ribbon synapses and type 2 bipolar cells. We morphologically identified all green fluorescent protein (GFP)-expressing ganglion cell types, which co-stratified with type 2 bipolar cells and assessed the total number of bipolar input synapses and the proportion of synapses deriving from type 2 bipolar cells. Only JAM-B ganglion cells received synaptic input preferentially from bipolar cell types other than type 2 bipolar cells whereas the other analyzed ganglion cell types sampled their bipolar input most likely from all bipolar cell terminals within their dendritic arbor.

  5. The Drosophila cell adhesion molecule Klingon is required for long-term memory formation and is regulated by Notch

    OpenAIRE

    Matsuno, Motomi; Horiuchi, Junjiro; Tully, Tim; Saitoe, Minoru

    2008-01-01

    The ruslan (rus) mutant was previously identified in a behavioral screen for mutants defective in long-lasting memory, which consists of two consolidated memory types, anesthesia-resistant memory, and protein synthesis-dependent long-term memory (LTM). We demonstrate here that rus is a new allele of klingon (klg), which encodes a homophilic cell adhesion molecule. Klg is acutely required for LTM but not anesthesia-resistant memory formation, and Klg expression increases upon LTM induction. LT...

  6. Long-term Disease Free and Successful Pregnancy in a Woman with Gonadal Dysgenesis and Malignant Germ Cell Tumor

    OpenAIRE

    Azamsadat Mousavi; Mitra Gilani; Shirin Goodarzi; Ensieh Tehraninejad; Hayedeh Haeri

    2012-01-01

    Objective: To report a case of long-term disease free and successful pregnancy after fertility sparing staging surgery with adjuvant chemotherapy in a 46,Xy gonadal dysgenetic with malignant germ cell tumor.Materials and methods: A case report from a university hospital about a 19-year-old female with 46,XY karyotype ( Swyer syndrome). The patient underwent bilateral gonadectomy and staging with uterus preservation. Six course adjuvant chemotherapy with VBP (Vinblastin, Bleomycin, Cisplatin) ...

  7. The female gametophyte: an emerging model for cell type-specific systems biology in plant development

    Directory of Open Access Journals (Sweden)

    Marc William Schmid

    2015-11-01

    Full Text Available Systems biology, a holistic approach describing a system emerging from the interactions of its molecular components, critically depends on accurate qualitative determination and quantitative measurements of these components. Development and improvement of large-scale profiling methods (omics now facilitates comprehensive measurements of many relevant molecules. For multicellular organisms, such as animals, fungi, algae, and plants, the complexity of the system is augmented by the presence of specialized cell types and organs, and a complex interplay within and between them. Cell type-specific analyses are therefore crucial for the understanding of developmental processes and environmental responses. This review first gives an overview of current methods used for large-scale profiling of specific cell types exemplified by recent advances in plant biology. The focus then lies on suitable model systems to study plant development and cell type specification. We introduce the female gametophyte of flowering plants as an ideal model to study fundamental developmental processes. Moreover, the female reproductive lineage is of importance for the emergence of evolutionary novelties such as an unequal parental contribution to the tissue nurturing the embryo or the clonal production of seeds by asexual reproduction (apomixis. Understanding these processes is not only interesting from a developmental or evolutionary perspective, but bears great potential for further crop improvement and the simplification of breeding efforts. We finally highlight novel methods, which are already available or which will likely soon facilitate large-scale profiling of the specific cell types of the female gametophyte in both model and non-model species. We conclude that it may take only few years until an evolutionary systems biology approach toward female gametogenesis may decipher some of its biologically most interesting and economically most valuable processes.

  8. Analysis of cell-type-specific gene expression during mouse spermatogenesis

    DEFF Research Database (Denmark)

    Almstrup, Kristian; Nielsen, John E; Hansen, Martin Asser;

    2004-01-01

    In rodents, changes in gene expression during spermatogenesis can be monitored by sampling testis from each day during postnatal development. However, changes in gene expression at the tissue level can reflect changes in the concentration of an mRNA in a specific cell type, changes in volume....... Combining results from these techniques allows determination of the cell-type-specific gene-expression patterns of many genes during spermatogenesis. Differential display was used to determine expression profiles with high sensitivity and independent of prior knowledge of the sequence, whereas DNA arrays...... quickly assess the expression profiles of all the genes. This identified three groups of gene-expression profiles. The major group corresponds to genes that are upregulated in spermatocytes during either the mid- or late- pachytene phase of spermatogenesis (stages VII-XI). This pachytene cluster...

  9. Long-Term In Vitro Expansion of Salivary Gland Stem Cells Driven by Wnt Signals

    Science.gov (United States)

    Maimets, Martti; Rocchi, Cecilia; Bron, Reinier; Pringle, Sarah; Kuipers, Jeroen; Giepmans, Ben N.G.; Vries, Robert G.J.; Clevers, Hans; de Haan, Gerald; van Os, Ronald; Coppes, Robert P.

    2015-01-01

    Summary Adult stem cells are the ultimate source for replenishment of salivary gland (SG) tissue. Self-renewal ability of stem cells is dependent on extrinsic niche signals that have not been unraveled for the SG. The ductal compartment in SG has been identified as the location harboring stem cells. Here, we report that rare SG ductal EpCAM+ cells express nuclear β-catenin, indicating active Wnt signaling. In cell culture experiments, EpCAMhigh cells respond potently to Wnt signals stimulating self-renewal and long-term expansion of SG organoids, containing all differentiated SG cell types. Conversely, Wnt inhibition ablated long-term organoid cultures. Finally, transplantation of cells pre-treated with Wnt agonists into submandibular glands of irradiated mice successfully and robustly restored saliva secretion and increased the number of functional acini in vivo. Collectively, these results identify Wnt signaling as a key driver of adult SG stem cells, allowing extensive in vitro expansion and enabling restoration of SG function upon transplantation. PMID:26724906

  10. Long-Term In Vitro Expansion of Salivary Gland Stem Cells Driven by Wnt Signals

    Directory of Open Access Journals (Sweden)

    Martti Maimets

    2016-01-01

    Full Text Available Adult stem cells are the ultimate source for replenishment of salivary gland (SG tissue. Self-renewal ability of stem cells is dependent on extrinsic niche signals that have not been unraveled for the SG. The ductal compartment in SG has been identified as the location harboring stem cells. Here, we report that rare SG ductal EpCAM+ cells express nuclear β-catenin, indicating active Wnt signaling. In cell culture experiments, EpCAMhigh cells respond potently to Wnt signals stimulating self-renewal and long-term expansion of SG organoids, containing all differentiated SG cell types. Conversely, Wnt inhibition ablated long-term organoid cultures. Finally, transplantation of cells pre-treated with Wnt agonists into submandibular glands of irradiated mice successfully and robustly restored saliva secretion and increased the number of functional acini in vivo. Collectively, these results identify Wnt signaling as a key driver of adult SG stem cells, allowing extensive in vitro expansion and enabling restoration of SG function upon transplantation.

  11. WI-38 cell long-term quiescence model system: a valuable tool to study molecular events that regulate growth.

    Science.gov (United States)

    Soprano, K J

    1994-04-01

    A number of cell culture model systems have been used to study the regulation of cell cycle progression at the molecular level. In this paper we describe the WI-38 cell long-term quiescence model system. By modulating the length of time that WI-38 cells are density arrested, it is possible to proportionately alter the length of the prereplicative or G-1 phase which the cell traverses after growth factor stimulation in preparation for entry into DNA synthesis. Through studies aimed at understanding the cause and molecular nature of the prolongation of the prereplicative phase, we have determined that gene expression plays an important role in establishing growth factor "competence" and that once the cell becomes "competent" there is a defined order to the molecular events that follow during the remainder of G-1. More specifically, we have determined that the prolongation represents a delay in the ability of long term quiescent cells to become fully "competent" to respond to growth factors which regulate progression through G-1 into S. This prolongation appears to occur as a result of changes during long term quiescence in the ability of immediate early G-1 specific genes (such as c-myc) to activate the expression of early G-1 specific genes (such as ornithine decarboxylase). While ODC is the first and thus far only growth associated gene identified as a target of c-myc (and the Myc/Max protein complex), it is likely that further studies in this model system will reveal other early G-1 growth regulatory genes. We anticipate that future follow-up studies in this model system will provide additional valuable information about the function of growth-regulatory genes in controlling growth factor responsiveness and cell cycle progression.

  12. Multiple origins of spontaneously arising micronuclei in HeLa cells: Direct evidence from long-term live cell imaging

    International Nuclear Information System (INIS)

    Although micronuclei (MNi) are extensively used to evaluate genotoxic effects and chromosome instability, the most basic issue regarding their origins has not been completely addressed due to limitations of traditional methods. Recently, long-term live cell imaging was developed to monitor the dynamics of single cell in a real-time and high-throughput manner. In the present study, this state-of-the-art technique was employed to examine spontaneous micronucleus (MN) formation in untreated HeLa cells. We demonstrate that spontaneous MNi are derived from incorrectly aligned chromosomes in metaphase (displaced chromosomes, DCs), lagging chromosomes (LCs) and broken chromosome bridges (CBs) in later mitotic stages, but not nuclear buds in S phase. However, most of bipolar mitoses with DCs (91.29%), LCs (73.11%) and broken CBs (88.93%) did not give rise to MNi. Our data also show directly, for the first time, that MNi could originate spontaneously from (1) MNi already presented in the mother cells; (2) nuclear fragments that appeared during mitosis with CB; and (3) chromosomes being extruded into a minicell which fused with one of the daughter cells later. Quantitatively, most of MNi originated from LCs (63.66%), DCs (10.97%) and broken CBs (9.25%). Taken together, these direct evidences show that there are multiple origins for spontaneously arising MNi in HeLa cells and each mechanism contributes to overall MN formation to different extents

  13. Multiple origins of spontaneously arising micronuclei in HeLa cells: Direct evidence from long-term live cell imaging

    Energy Technology Data Exchange (ETDEWEB)

    Rao Xiaotang; Zhang Yingyin; Yi Qiyi; Hou Heli; Xu Bo; Chu Liang; Huang Yun; Zhang Wenrui [Laboratory of Molecular and Cell Genetics, Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027 (China); Fenech, Michael [CSIRO Human Nutrition, PO Box 10041, Adelaide BC, Adelaide, SA 5000 (Australia); Shi Qinghua [Laboratory of Molecular and Cell Genetics, Hefei National Laboratory for Physical Sciences at Microscale and School of Life Sciences, University of Science and Technology of China, Hefei, Anhui 230027 (China)], E-mail: qshi@ustc.edu.cn

    2008-11-10

    Although micronuclei (MNi) are extensively used to evaluate genotoxic effects and chromosome instability, the most basic issue regarding their origins has not been completely addressed due to limitations of traditional methods. Recently, long-term live cell imaging was developed to monitor the dynamics of single cell in a real-time and high-throughput manner. In the present study, this state-of-the-art technique was employed to examine spontaneous micronucleus (MN) formation in untreated HeLa cells. We demonstrate that spontaneous MNi are derived from incorrectly aligned chromosomes in metaphase (displaced chromosomes, DCs), lagging chromosomes (LCs) and broken chromosome bridges (CBs) in later mitotic stages, but not nuclear buds in S phase. However, most of bipolar mitoses with DCs (91.29%), LCs (73.11%) and broken CBs (88.93%) did not give rise to MNi. Our data also show directly, for the first time, that MNi could originate spontaneously from (1) MNi already presented in the mother cells; (2) nuclear fragments that appeared during mitosis with CB; and (3) chromosomes being extruded into a minicell which fused with one of the daughter cells later. Quantitatively, most of MNi originated from LCs (63.66%), DCs (10.97%) and broken CBs (9.25%). Taken together, these direct evidences show that there are multiple origins for spontaneously arising MNi in HeLa cells and each mechanism contributes to overall MN formation to different extents.

  14. Radioresistant cell strain of human fibrosarcoma cells obtained after long-term exposure to X-rays

    International Nuclear Information System (INIS)

    A radioresistant cell strain from human fibrosarcoma HT1080 has been obtained after prolonged exposure to x-rays for 7 months (2 Gy per day, 5 days per week). This new strain, HT1080R, differs from HT1080 in a significantly increased ability of clonogenical survival, with coefficient α decreasing from 0.161 to 0.123 Gy-1 and coefficient β decreasing from 0.0950 to 0.0565 Gy-2. Furthermore, the radioresistance of HT1080R proved to be stable in long-term passaged cultures as well as in frozen samples. Differences between the two cell lines are also observed in the G-banded karyotype; the new cell line shows monosomy of chromosome 17 and loss of 5p+ and 11q+ present in the parental cells. These data suggest that the radioresistance may have been caused by radiation-induced cell mutation and that the resistant cells may have been selected by repeated irradiations. In order to characterize this new strain, the ability of the cells to rejoin DNA double-strand breaks, the cell cycle distribution and the amount of apoptosis after irradiation have been estimated; however, no differences are observed between these two cell strains. Although the mechanism of the elevated radioresistance remains unknown, this pair of cell strains can provide a new model system for further investigations with regard to the mechanisms of cellular radioresistance. The results also show that any type of irradiation similar to the schedules used in radiotherapy can lead to the formation and selection of more radioresistant cell clones in vitro, a phenomenon with possible implications for radiotherapy. (orig.)

  15. Cell-type specific mechanisms of D-serine uptake and release in the brain

    Directory of Open Access Journals (Sweden)

    Magalie eMartineau

    2014-05-01

    Full Text Available Accumulating evidence during the last decade established that D-serine is a key signaling molecule utilized by neurons and astroglia in the mammalian central nervous system. D-serine is increasingly appreciated as the main physiological endogenous coagonist for synaptic NMDA receptors at central excitatory synapses; it is mandatory for long-term changes in synaptic strength, memory, learning, and social interactions. Alterations in the extracellular levels of D-serine leading to disrupted cell-cell signaling are a trademark of many chronic or acute neurological (i.e. Alzheimer disease, epilepsy, stroke and psychiatric (i.e. schizophrenia disorders, and are associated with addictive behavior (i.e. cocaine addiction. Indeed, fine tuning of the extracellular levels of D-serine, achieved by various molecular machineries and signaling pathways, is necessary for maintenance of accurate NMDA receptor functions. Here, we review the experimental data supporting the notion that astroglia and neurons use different pathways to regulate levels of extracellular D-serine.

  16. Long-term cultures of testicular biopsies from boys with cryptorchidism: effect of FSH and LH on the number of germ cells

    DEFF Research Database (Denmark)

    Larsen, Hans-Peter Ejler; Thorup, Jørgen; Skovgaard, Lene Theil;

    2002-01-01

    A long-term culture system of testicular biopsies from boys with undescended testes was established to evaluate the effect of gonadotrophins on germ cell survival and growth.......A long-term culture system of testicular biopsies from boys with undescended testes was established to evaluate the effect of gonadotrophins on germ cell survival and growth....

  17. Labeling and imaging of human mesenchymal stem cells with quantum dot bioconjugates during proliferation and osteogenic differentiation in long term.

    Science.gov (United States)

    Shah, B; Clark, P; Stroscio, M; Mao, J

    2006-01-01

    Quantum dots (QDs) are semiconductor nanocrystals that serve as promising alternatives to organic dyes for cell labeling. Because of their unique spectral, physical and chemical properties, QDs are useful for concurrently monitoring several intercellular and intracellular interactions in live normal cells and cancer cells over periods ranging from less than a second to over several days (several divisions of cells). Here, peptide CGGGRGD is immobilized on CdSe-ZnS QDs coated with carboxyl groups by cross linking with amine groups. These conjugates are directed by the peptide to bind with selected integrins on the membrane of human Mesenchymal stem cells. Upon overnight incubation with optimal concentration, QDs effectively labeled all the cells. Here, we report long-term labeling of human bone-marrow-derived mesenchymal stem cells (hMSCs) with RGD-conjugated QDs during self replication and differentiation into osteogenic cell lineages.

  18. Upregulation of CD11A on Hematopoietic Stem Cells Denotes the Loss of Long-Term Reconstitution Potential

    Directory of Open Access Journals (Sweden)

    John W. Fathman

    2014-11-01

    Full Text Available Small numbers of hematopoietic stem cells (HSCs generate large numbers of mature effector cells through the successive amplification of transiently proliferating progenitor cells. HSCs and their downstream progenitors have been extensively characterized based on their cell-surface phenotype and functional activities during transplantation assays. These cells dynamically lose and acquire specific sets of surface markers during differentiation, leading to the identification of markers that allow for more refined separation of HSCs from early hematopoietic progenitors. Here, we describe a marker, CD11A, which allows for the enhanced purification of mouse HSCs. We show through in vivo transplantations that upregulation of CD11A on HSCs denotes the loss of their long-term reconstitution potential. Surprisingly, nearly half of phenotypic HSCs (defined as Lin−KIT+SCA-1+CD150+CD34− are CD11A+ and lack long-term self-renewal potential. We propose that CD11A+Lin−KIT+SCA-1+CD150+CD34− cells are multipotent progenitors and CD11A−Lin−KIT+SCA-1+CD150+CD34− cells are true HSCs.

  19. Protein conservation and variation suggest mechanisms of cell type-specific modulation of signaling pathways.

    Science.gov (United States)

    Schaefer, Martin H; Yang, Jae-Seong; Serrano, Luis; Kiel, Christina

    2014-06-01

    Many proteins and signaling pathways are present in most cell types and tissues and yet perform specialized functions. To elucidate mechanisms by which these ubiquitous pathways are modulated, we overlaid information about cross-cell line protein abundance and variability, and evolutionary conservation onto functional pathway components and topological layers in the pathway hierarchy. We found that the input (receptors) and the output (transcription factors) layers evolve more rapidly than proteins in the intermediary transmission layer. In contrast, protein expression variability decreases from the input to the output layer. We observed that the differences in protein variability between the input and transmission layer can be attributed to both the network position and the tendency of variable proteins to physically interact with constitutively expressed proteins. Differences in protein expression variability and conservation are also accompanied by the tendency of conserved and constitutively expressed proteins to acquire somatic mutations, while germline mutations tend to occur in cell type-specific proteins. Thus, conserved core proteins in the transmission layer could perform a fundamental role in most cell types and are therefore less tolerant to germline mutations. In summary, we propose that the core signal transmission machinery is largely modulated by a variable input layer through physical protein interactions. We hypothesize that the bow-tie organization of cellular signaling on the level of protein abundance variability contributes to the specificity of the signal response in different cell types. PMID:24922536

  20. Protein conservation and variation suggest mechanisms of cell type-specific modulation of signaling pathways.

    Directory of Open Access Journals (Sweden)

    Martin H Schaefer

    2014-06-01

    Full Text Available Many proteins and signaling pathways are present in most cell types and tissues and yet perform specialized functions. To elucidate mechanisms by which these ubiquitous pathways are modulated, we overlaid information about cross-cell line protein abundance and variability, and evolutionary conservation onto functional pathway components and topological layers in the pathway hierarchy. We found that the input (receptors and the output (transcription factors layers evolve more rapidly than proteins in the intermediary transmission layer. In contrast, protein expression variability decreases from the input to the output layer. We observed that the differences in protein variability between the input and transmission layer can be attributed to both the network position and the tendency of variable proteins to physically interact with constitutively expressed proteins. Differences in protein expression variability and conservation are also accompanied by the tendency of conserved and constitutively expressed proteins to acquire somatic mutations, while germline mutations tend to occur in cell type-specific proteins. Thus, conserved core proteins in the transmission layer could perform a fundamental role in most cell types and are therefore less tolerant to germline mutations. In summary, we propose that the core signal transmission machinery is largely modulated by a variable input layer through physical protein interactions. We hypothesize that the bow-tie organization of cellular signaling on the level of protein abundance variability contributes to the specificity of the signal response in different cell types.

  1. Species- and cell type-specific interactions between CD47 and human SIRPalpha.

    Science.gov (United States)

    Subramanian, Shyamsundar; Parthasarathy, Ranganath; Sen, Shamik; Boder, Eric T; Discher, Dennis E

    2006-03-15

    CD47 on red blood cells (RBCs) reportedly signals "self" by binding SIRPalpha on phagocytes, at least in mice. Such interactions across and within species, from mouse to human, are not yet clear and neither is the relation to cell adhesion. Using human SIRPalpha1 as a probe, antibody-inhibitable binding to CD47 was found only with human and pig RBCs (not mouse, rat, or cow). In addition, CD47-mediated adhesion of human and pig RBCs to SIRPalpha1 surfaces resists sustained forces in centrifugation (as confirmed by atomic force microscopy) but only at SIRPalpha-coating densities far above those measurable on human neutrophils, monocytes, and THP-1 macrophages. While interactions strengthen with deglycosylation of SIRPalpha1, low copy numbers explain the absence of RBC adhesion to phagocytes under physiologic conditions and imply that the interaction being studied is not responsible for red cell clearance in humans. Evidence of clustering nonetheless suggests mechanisms of avidity enhancement. Finally, using the same CD47 antibodies and soluble SIRPalpha1, bone marrow-derived mesenchymal stem cells were assayed and found to display CD47 but not bind SIRPalpha1 significantly. The results thus demonstrate that SIRPalpha-CD47 interactions, which reportedly define self, exhibit cell type specificity and limited cross-species reactivity.

  2. Pore-filled electrolyte membranes for facile fabrication of long-term stable dye-sensitized solar cells

    International Nuclear Information System (INIS)

    Graphical abstract: Display Omitted -- Highlights: •Pore-filled film electrolytes (PFEMs) were investigated for facile DSSC fabrication. •Optimal mixed solvent was suggested to enhance the long-term stability of DSSCs. •The PFEMs promised both the excellent thermal stability and energy efficiency. •Thephotovoltaic efficiency was well correlated with porous structure of substrates. -- ABSTRACT: Pore-filled electrolyte membranes (PFEMs) have been prepared by employing an optimized porous substrate and stable electrolyte composition for a facile manufacturing process of dye-sensitized solar cells (DSSCs). The PFEMs could be easily loaded into a photovoltaic device without adding a traditional electrolyte injection through a hole. In order to meet the requirements of both high energy conversion efficiency and proper long-term stability, three different solvents with high boiling point, i.e. valeronitrile, dimethyl sulfoxide, and dimethylacetamide, were appropriately mixed as a volumetric ratio of 7:2:1, respectively. As a result, similar conductivity and viscosity as well as better chemical stability were obtained compared to those of conventional 3-methoxypropionitrile-based electrolyte. In addition, linear relations were observed between the photovoltaic efficiency and porous film properties (i.e. porosity and tortuosity). The DSSC employing the PFEM doped with the mixed solvent based electrolyte exhibited the photon-to-current conversion efficiency of 6.30% at one sun condition. Moreover, the long-term stability test fixed at an elevated temperature of 85 °C exhibited outstanding durability of DSSC for 500 h

  3. Cell- and stimulus type-specific intracellular free Ca2+ signals in Arabidopsis.

    Science.gov (United States)

    Martí, María C; Stancombe, Matthew A; Webb, Alex A R

    2013-10-01

    Appropriate stimulus-response coupling requires that each signal induces a characteristic response, distinct from that induced by other signals, and that there is the potential for individual signals to initiate different downstream responses dependent on cell type. How such specificity is encoded in plant signaling is not known. One possibility is that information is encoded in signal transduction pathways to ensure stimulus- and cell type-specific responses. The calcium ion acts as a second messenger in response to mechanical stimulation, hydrogen peroxide, NaCl, and cold in plants and also in circadian timing. We use GAL4 transactivation of aequorin in enhancer trap lines of Arabidopsis (Arabidopsis thaliana) to test the hypothesis that stimulus- and cell-specific information can be encoded in the pattern of dynamic alterations in the concentration of intracellular free Ca(2+) ([Ca(2+)]i). We demonstrate that mechanically induced increases in [Ca(2+)]i are largely restricted to the epidermal pavement cells of leaves, that NaCl induces oscillatory [Ca(2+)]i signals in spongy mesophyll and vascular bundle cells, but not other cell types, and detect circadian rhythms of [Ca(2+)]i only in the spongy mesophyll. We demonstrate stimulus-specific [Ca(2+)]i dynamics in response to touch, cold, and hydrogen peroxide, which in the case of the latter two signals are common to all cell types tested. GAL4 transactivation of aequorin in specific leaf cell types has allowed us to bypass the technical limitations associated with fluorescent Ca(2+) reporter dyes in chlorophyll-containing tissues to identify the cell- and stimulus-specific complexity of [Ca(2+)]i dynamics in leaves of Arabidopsis and to determine from which tissues stress- and circadian-regulated [Ca(2+)]i signals arise.

  4. Epigenetic regulation of normal human mammary cell type-specific miRNAs

    Energy Technology Data Exchange (ETDEWEB)

    Vrba, Lukas [Univ. of Arizona, Tucson, AZ (United States). Arizona Cancer Center; Inst. of Plant Molecular Biology, Ceske Budejovice (Czech Republic). Biology Centre ASCR; Garbe, James C. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Center; Stampfer, Martha R. [Univ. of Arizona, Tucson, AZ (United States). Arizona Cancer Center; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Center; Futscher, Bernard W. [Univ. of Arizona, Tucson, AZ (United States). Arizona Cancer Center and Dept. of Pharmacology & Toxicology

    2011-08-26

    Epigenetic mechanisms are important regulators of cell type–specific genes, including miRNAs. In order to identify cell type-specific miRNAs regulated by epigenetic mechanisms, we undertook a global analysis of miRNA expression and epigenetic states in three isogenic pairs of human mammary epithelial cells (HMEC) and human mammary fibroblasts (HMF), which represent two differentiated cell types typically present within a given organ, each with a distinct phenotype and a distinct epigenotype. While miRNA expression and epigenetic states showed strong interindividual concordance within a given cell type, almost 10% of the expressed miRNA showed a cell type–specific pattern of expression that was linked to the epigenetic state of their promoter. The tissue-specific miRNA genes were epigenetically repressed in nonexpressing cells by DNA methylation (38%) and H3K27me3 (58%), with only a small set of miRNAs (21%) showing a dual epigenetic repression where both DNA methylation and H3K27me3 were present at their promoters, such as MIR10A and MIR10B. Individual miRNA clusters of closely related miRNA gene families can each display cell type–specific repression by the same or complementary epigenetic mechanisms, such as the MIR200 family, and MIR205, where fibroblasts repress MIR200C/141 by DNA methylation, MIR200A/200B/429 by H3K27me3, and MIR205 by both DNA methylation and H3K27me3. Since deregulation of many of the epigenetically regulated miRNAs that we identified have been linked to disease processes such as cancer, it is predicted that compromise of the epigenetic control mechanisms is important for this process. Overall, these results highlight the importance of epigenetic regulation in the control of normal cell type–specific miRNA expression.

  5. Imaging long-term fate of intramyocardially implanted mesenchymal stem cells in a porcine myocardial infarction model.

    Directory of Open Access Journals (Sweden)

    Emerson C Perin

    Full Text Available The long-term fate of stem cells after intramyocardial delivery is unknown. We used noninvasive, repetitive PET/CT imaging with [(18F]FEAU to monitor the long-term (up to 5 months spatial-temporal dynamics of MSCs retrovirally transduced with the sr39HSV1-tk gene (sr39HSV1-tk-MSC and implanted intramyocardially in pigs with induced acute myocardial infarction. Repetitive [(18F]FEAU PET/CT revealed a biphasic pattern of sr39HSV1-tk-MSC dynamics; cell proliferation peaked at 33-35 days after injection, in periinfarct regions and the major cardiac lymphatic vessels and lymph nodes. The sr39HSV1-tk-MSC-associated [(18F]FEAU signals gradually decreased thereafter. Cardiac lymphography studies using PG-Gd-NIRF813 contrast for MRI and near-infrared fluorescence imaging showed rapid clearance of the contrast from the site of intramyocardial injection through the subepicardial lymphatic network into the lymphatic vessels and periaortic lymph nodes. Immunohistochemical analysis of cardiac tissue obtained at 35 and 150 days demonstrated several types of sr39HSV1-tk expressing cells, including fibro-myoblasts, lymphovascular cells, and microvascular and arterial endothelium. In summary, this study demonstrated the feasibility and sensitivity of [(18F]FEAU PET/CT imaging for long-term, in-vivo monitoring (up to 5 months of the fate of intramyocardially injected sr39HSV1-tk-MSC cells. Intramyocardially transplanted MSCs appear to integrate into the lymphatic endothelium and may help improve myocardial lymphatic system function after MI.

  6. Cell-intrinsic in vivo requirement for the E47-p21 pathway in long-term hematopoietic stem cells.

    Science.gov (United States)

    Santos, Patricia M; Ding, Ying; Borghesi, Lisa

    2014-01-01

    Major regulators of long-term hematopoietic stem cell (LT-HSC) self-renewal and proliferation have been identified, but knowledge of their in vivo interaction in a linear pathway is lacking. In this study, we show a direct genetic link between the transcription factor E47 and the major cell cycle regulator p21 in controlling LT-HSC integrity in vivo under repopulation stress. Numerous studies have shown that E47 activates p21 transcription in hematopoietic subsets in vitro, and we now reveal the in vivo relevance of the E47-p21 pathway by reducing the gene dose of each factor individually (E47(het) or p21(het)) versus in tandem (E47(het)p21(het)). E47(het)p21(het) LT-HSCs and downstream short-term hematopoietic stem cells exhibit hyperproliferation and preferential susceptibility to mitotoxin compared to wild-type or single haploinsufficient controls. In serial adoptive transfers that rigorously challenge self-renewal, E47(het)p21(het) LT-HSCs dramatically and progressively decline, indicating the importance of cell-intrinsic E47-p21 in preserving LT-HSCs under stress. Transient numeric recovery of downstream short-term hematopoietic stem cells enabled the production of functionally competent myeloid but not lymphoid cells, as common lymphoid progenitors were decreased, and peripheral lymphocytes were virtually ablated. Thus, we demonstrate a developmental compartment-specific and lineage-specific requirement for the E47-p21 pathway in maintaining LT-HSCs, B cells, and T cells under hematopoietic repopulation stress in vivo.

  7. Human Neural Stem Cell Transplantation Provides Long-Term Restoration of Neuronal Plasticity in the Irradiated Hippocampus

    Science.gov (United States)

    Acharya, Munjal M.; Rosi, Susanna; Jopson, Timothy; Limoli, Charles L.

    2016-01-01

    For the majority of CNS malignancies, radiotherapy provides the best option for forestalling tumor growth, but is frequently associated with debilitating and progressive cognitive dysfunction. Despite the recognition of this serious side effect, satisfactory long-term solutions are not currently available and have prompted our efforts to explore the potential therapeutic efficacy of cranial stem cell transplants. We have demonstrated that intrahippocampal transplantation of human neural stem cells (hNSCs) can provide long-lasting cognitive benefits using an athymic rat model subjected to cranial irradiation. To explore the possible mechanisms underlying the capability of engrafted cells to ameliorate radiation-induced cognitive dysfunction we analyzed the expression patterns of the behaviorally induced activity-regulated cytoskeleton-associated protein (Arc) in the hippocampus at 1 and 8 months postgrafting. While immunohistochemical analyses revealed a small fraction (4.5%) of surviving hNSCs in the irradiated brain that did not express neuronal or astroglial makers, hNSC transplantation impacted the irradiated microenvironment of the host brain by promoting the expression of Arc at both time points. Arc is known to play key roles in the neuronal mechanisms underlying long-term synaptic plasticity and memory and provides a reliable marker for detecting neurons that are actively engaged in spatial and contextual information processing associated with memory consolidation. Cranial irradiation significantly reduced the number of pyramidal (CA1) and granule neurons (DG) expressing behaviorally induced Arc at 1 and 8 months postirradiation. Transplantation of hNSCs restored the expression of plasticity-related Arc in the host brain to control levels. These findings suggest that hNSC transplantation promotes the long-term recovery of host hippocampal neurons and indicates that one mechanism promoting the preservation of cognition after irradiation involves trophic

  8. Long-term frozen storage of stem cells: challenges and solutions

    OpenAIRE

    Harris, David

    2016-01-01

    David T Harris1-5 1University of Arizona Health Sciences Biorepository, 2Department of Immunology, 3Division of Translational Medicine, 4GMP Laboratory, University of Arizona, Tucson, 5Celebration Stem Cell Centre, Gilbert, AZ, USA Abstract: Stem cells are found in all multicellular organisms and are defined as cells that can differentiate into specialized mature cells as well as divide to produce more stem cells. Stem cells are commonly harvested for clinical and research applications from b...

  9. Cell-type specific roles for PTEN in establishing a functional retinal architecture.

    Directory of Open Access Journals (Sweden)

    Robert Cantrup

    Full Text Available BACKGROUND: The retina has a unique three-dimensional architecture, the precise organization of which allows for complete sampling of the visual field. Along the radial or apicobasal axis, retinal neurons and their dendritic and axonal arbors are segregated into layers, while perpendicular to this axis, in the tangential plane, four of the six neuronal types form patterned cellular arrays, or mosaics. Currently, the molecular cues that control retinal cell positioning are not well-understood, especially those that operate in the tangential plane. Here we investigated the role of the PTEN phosphatase in establishing a functional retinal architecture. METHODOLOGY/PRINCIPAL FINDINGS: In the developing retina, PTEN was localized preferentially to ganglion, amacrine and horizontal cells, whose somata are distributed in mosaic patterns in the tangential plane. Generation of a retina-specific Pten knock-out resulted in retinal ganglion, amacrine and horizontal cell hypertrophy, and expansion of the inner plexiform layer. The spacing of Pten mutant mosaic populations was also aberrant, as were the arborization and fasciculation patterns of their processes, displaying cell type-specific defects in the radial and tangential dimensions. Irregular oscillatory potentials were also observed in Pten mutant electroretinograms, indicative of asynchronous amacrine cell firing. Furthermore, while Pten mutant RGC axons targeted appropriate brain regions, optokinetic spatial acuity was reduced in Pten mutant animals. Finally, while some features of the Pten mutant retina appeared similar to those reported in Dscam-mutant mice, PTEN expression and activity were normal in the absence of Dscam. CONCLUSIONS/SIGNIFICANCE: We conclude that Pten regulates somal positioning and neurite arborization patterns of a subset of retinal cells that form mosaics, likely functioning independently of Dscam, at least during the embryonic period. Our findings thus reveal an unexpected

  10. Mosaic Analysis with Double Markers Reveals Cell-Type-Specific Paternal Growth Dominance

    Directory of Open Access Journals (Sweden)

    Simon Hippenmeyer

    2013-03-01

    Full Text Available Genomic imprinting leads to preferred expression of either the maternal or paternal alleles of a subset of genes. Imprinting is essential for mammalian development, and its deregulation causes many diseases. However, the functional relevance of imprinting at the cellular level is poorly understood for most imprinted genes. We used mosaic analysis with double markers (MADM in mice to create uniparental disomies (UPDs and to visualize imprinting effects with single-cell resolution. Although chromosome 12 UPD did not produce detectable phenotypes, chromosome 7 UPD caused highly significant paternal growth dominance in the liver and lung, but not in the brain or heart. A single gene on chromosome 7, encoding the secreted insulin-like growth factor 2 (IGF2, accounts for most of the paternal dominance effect. Mosaic analyses implied additional imprinted loci on chromosome 7 acting cell autonomously to transmit the IGF2 signal. Our study reveals chromosome- and cell-type specificity of genomic imprinting effects.

  11. Long-Term Self-Renewal of Postnatal Muscle-derived Stem Cells

    OpenAIRE

    Deasy, B. M.; Gharaibeh, B. M.; Pollett, J. B.; Jones, M.M.; Lucas, M. A.; Kanda, Y.; Huard, J

    2005-01-01

    The ability to undergo self-renewal is a defining characteristic of stem cells. Self-replenishing activity sustains tissue homeostasis and regeneration. In addition, stem cell therapy strategies require a heightened understanding of the basis of the self-renewal process to enable researchers and clinicians to obtain sufficient numbers of undifferentiated stem cells for cell and gene therapy. Here, we used postnatal muscle-derived stem cells to test the basic biological assumption of unlimited...

  12. Cell-type specificity of ChIP-predicted transcription factor binding sites

    Directory of Open Access Journals (Sweden)

    Håndstad Tony

    2012-08-01

    Full Text Available Abstract Background Context-dependent transcription factor (TF binding is one reason for differences in gene expression patterns between different cellular states. Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq identifies genome-wide TF binding sites for one particular context—the cells used in the experiment. But can such ChIP-seq data predict TF binding in other cellular contexts and is it possible to distinguish context-dependent from ubiquitous TF binding? Results We compared ChIP-seq data on TF binding for multiple TFs in two different cell types and found that on average only a third of ChIP-seq peak regions are common to both cell types. Expectedly, common peaks occur more frequently in certain genomic contexts, such as CpG-rich promoters, whereas chromatin differences characterize cell-type specific TF binding. We also find, however, that genotype differences between the cell types can explain differences in binding. Moreover, ChIP-seq signal intensity and peak clustering are the strongest predictors of common peaks. Compared with strong peaks located in regions containing peaks for multiple transcription factors, weak and isolated peaks are less common between the cell types and are less associated with data that indicate regulatory activity. Conclusions Together, the results suggest that experimental noise is prevalent among weak peaks, whereas strong and clustered peaks represent high-confidence binding events that often occur in other cellular contexts. Nevertheless, 30-40% of the strongest and most clustered peaks show context-dependent regulation. We show that by combining signal intensity with additional data—ranging from context independent information such as binding site conservation and position weight matrix scores to context dependent chromatin structure—we can predict whether a ChIP-seq peak is likely to be present in other cellular contexts.

  13. Long-Term Stability and Safety of Transgenic Cultured Epidermal Stem Cells in Gene Therapy of Junctional Epidermolysis Bullosa

    Directory of Open Access Journals (Sweden)

    Laura De Rosa

    2014-01-01

    Full Text Available We report a long-term follow-up (6.5 years of a phase I/II clinical trial envisaging the use of autologous genetically modified cultured epidermal stem cells for gene therapy of junctional epidermolysis bullosa, a devastating genetic skin disease. The critical goals of the trial were to evaluate the safety and long-term persistence of genetically modified epidermis. A normal epidermal-dermal junction was restored and the regenerated transgenic epidermis was found to be fully functional and virtually indistinguishable from a normal control. The epidermis was sustained by a discrete number of long-lasting, self-renewing transgenic epidermal stem cells that maintained the memory of the donor site, whereas the vast majority of transduced transit-amplifying progenitors were lost within the first few months after grafting. These data pave the way for the safe use of epidermal stem cells in combined cell and gene therapy for genetic skin diseases.

  14. Evaluation of Bronchiolar and Alveolar Cell Injuries Induced by Short- and Long-term Exposure to Sidestream Smoke

    OpenAIRE

    Kwon, Kun-Young; Jung, Hye-Ra; Hwang, Ilseon; Choi, Won-Il

    2012-01-01

    Background We investigated effects of short- and long-term exposure to sidestream smoke on the bronchiolar and alveolar cells in Sprague-Dawley rats. Methods Rats were divided into five experimental groups: groups 1, 2, and 3 (1-month exposure to 3, 5, and 7 cigarettes a day, respectively), groups 4 and 5 (3- and 6 month exposure to five cigarettes a day, respectively). We examined the morphologic changes, the expressions of tumor necrosis factor α (TNF-α), tumor growth factor β1 (TGF-β1), in...

  15. Long-Term Extracorporeal Membrane Oxygenation as Bridging Strategies to Lung Transplantation in Rapidly Devastating Isolated Langerhans Cell Histiocytosis.

    Science.gov (United States)

    Sacco, Oliviero; Moscatelli, Andrea; Conte, Massimo; Grasso, Chiara; Magnano, Gian Michele; Sementa, Angela Rita; Martelli, Alberto; Rossi, Giovanni A

    2016-05-01

    Isolated pulmonary involvement in pediatric Langerhans cell histiocytosis (LCH) is extremely rare. While the multisystem-LCH course varies from spontaneous remission to rapid deterioration with lethal outcome, single system involvement is generally associated with favorable prognosis. A child with isolated pulmonary LCH had an extremely rapid progression leading to respiratory failure, despite treatment with prednisone and vinblastine. Since lung hyperinflation and cystic degeneration contraindicated conventional mechanical ventilation, extracorporeal membrane oxygenation (ECMO) was chosen for 50 days as a bridge to lung transplantation. The mechanisms involved in disease progression and the usefulness of long-term ECMO are discussed. PMID:26840616

  16. [Long-term subculture and biological characterization of the murine bone marrow endothelial cell line].

    Science.gov (United States)

    Huang, Chang; Zhu, Wen-Biao; Zhu, Hai-Ling; Wang, Bao-He; Wang, Qi-Ru

    2007-12-01

    The murine bone marrow endothelial cell line (mBMEC) has been maintained by means of subculture and cryopreservation for over 10 years since it was established in our laboratory. This study was aimed to newly identify biological characteristics of this cell line for further study. The cultured mBMEC cells were observed by inverted microscopy and transmission electron microscopy (TEM). PECAM-1 (CD31) and von Willebrand factor (vWF) were detected by immunofluorescent staining. The phagocytotic activity of the cells in culture was tested by using fluorescent acetylated low-density lipoprotein (Dil-Ac-LDL). The cell growth kinetics analysis and karyotype analysis were performed. The results showed that the adherent cells were mostly elliptical, rounded and spindle-shaped, and some of them connected to each other to form cord- and network-like arrangements in mBMEC cultures at subconfluence. The adherent cells grew up to confluence as a cobblestone-like monolayer. Several ultrastructural features of the endothelial cells could be observed in TEM sections of the cultured cells. More than 94% of mBMEC cells were positive for either CD31 or vWF. The phagocytotic ingestion of Dil-Ac-LDL occurred in 98.5% of cells. In normal culture conditions, the cells grew with a mean population doubling time of 54.6 hours and the maximal mitotic index was 38 per thousand in the rapid growth period. The colony yields were 4.33% to 7.40% depending on the plating density of cells. Karyotypes of all the cells were aneuploidy with a greater percentage of hyperdiploid. It is concluded that mBMEC cells retain the fundamental properties of endothelial cells, but the growth kinetics and biological behaviors are slightly different from those in the early days after the establishment of this cell line.

  17. Long-Term Oocyte-Like Cell Development in Cultures Derived from Neonatal Marmoset Monkey Ovary

    Directory of Open Access Journals (Sweden)

    Bentolhoda Fereydouni

    2016-01-01

    Full Text Available We use the common marmoset monkey (Callithrix jacchus as a preclinical nonhuman primate model to study reproductive and stem cell biology. The neonatal marmoset monkey ovary contains numerous primitive premeiotic germ cells (oogonia expressing pluripotent stem cell markers including OCT4A (POU5F1. This is a peculiarity compared to neonatal human and rodent ovaries. Here, we aimed at culturing marmoset oogonia from neonatal ovaries. We established a culture system being stable for more than 20 passages and 5 months. Importantly, comparative transcriptome analysis of the cultured cells with neonatal ovary, embryonic stem cells, and fibroblasts revealed a lack of germ cell and pluripotency genes indicating the complete loss of oogonia upon initiation of the culture. From passage 4 onwards, however, the cultured cells produced large spherical, free-floating cells resembling oocyte-like cells (OLCs. OLCs strongly expressed several germ cell genes and may derive from the ovarian surface epithelium. In summary, our novel primate ovarian cell culture initially lacked detectable germ cells but then produced OLCs over a long period of time. This culture system may allow a deeper analysis of early phases of female primate germ cell development and—after significant refinement—possibly also the production of monkey oocytes.

  18. Transformation of human mesenchymal stem cells in radiation carcinogenesis: long-term effect of ionizing radiation

    DEFF Research Database (Denmark)

    Christensen, Rikke; Alsner, Jan; Sørensen, Flemming Brandt;

    2008-01-01

    Increasing evidence on cancer stem cells suggest that stem cells are susceptive to carcinogenesis and consequently can be the origin of many cancers. We have recently established a telomerase-transduced human mesenchymal stem cell line and subsequently irradiated this in order to achieve malignant...

  19. CD19 CAR-targeted T cells induce long-term remission and B Cell Aplasia in an immunocompetent mouse model of B cell acute lymphoblastic leukemia.

    Directory of Open Access Journals (Sweden)

    Marco L Davila

    Full Text Available Although many adults with B cell acute lymphoblastic leukemia (B-ALL are induced into remission, most will relapse, underscoring the dire need for novel therapies for this disease. We developed murine CD19-specific chimeric antigen receptors (CARs and an immunocompetent mouse model of B-ALL that recapitulates the disease at genetic, cellular, and pathologic levels. Mouse T cells transduced with an all-murine CD3ζ/CD28-based CAR that is equivalent to the one being used in our clinical trials, eradicate B-ALL in mice and mediate long-term B cell aplasias. In this model, we find that increasing conditioning chemotherapy increases tumor eradication, B cell aplasia, and CAR-modified T cell persistence. Quantification of recipient B lineage cells allowed us to estimate an in vivo effector to endogenous target ratio for B cell aplasia maintenance. In mice exhibiting a dramatic B cell reduction we identified a small population of progenitor B cells in the bone marrow that may serve as a reservoir for long-term CAR-modified T cell stimulation. Lastly, we determine that infusion of CD8+ CAR-modified T cells alone is sufficient to maintain long-term B cell eradication. The mouse model we report here should prove valuable for investigating CAR-based and other therapies for adult B-ALL.

  20. Hematopoietic microenvironment. Origin, lineage, and transplantability of the stromal cells in long-term bone marrow cultures from chimeric mice

    Energy Technology Data Exchange (ETDEWEB)

    Perkins, S.; Fleischman, R.A.

    1988-04-01

    Studies of bone marrow transplant patients have suggested that the stromal cells of the in vitro hematopoietic microenvironment are transplantable into conditioned recipients. Moreover, in patients with myeloproliferative disorders, all of the stromal cells, which include presumptive endothelial cells, appear to be derived from hematopoietic precursors. To confirm these findings, we have constructed two chimeric mouse models: (a) traditional radiation chimeras, and (b) fetal chimeras, produced by placental injection of bone marrow into genetically anemic Wx/Wv fetuses, a technique that essentially precludes engraftment of nonhematopoietic cells. Using two-color indirect immunofluorescence, the stromal cells in long-term bone marrow culture derived from these chimeras were analyzed for donor or host origin by strain-specific H-2 antigens, and for cell lineage by a variety of other specific markers. 75-95% of the stromal cells were shown to be hematopoietic cells of the monocyte-macrophage lineage, based upon donor origin, phagocytosis, and expression of specific hematopoietic surface antigens. The remaining 5-25% of the stromal cells were exclusively host in origin. Apart from occasional fat cells, these cells uniformly expressed collagen type IV, laminin, and a surface antigen associated with endothelial cells. Since these endothelial-like cells are not transplantable into radiation or fetal chimeras, they are not derived from hematopoietic stem cells. The contrast between our findings and human studies suggests either unexpected species differences in the origin of stromal lineages or limitations in the previous methodology used to detect nonhematopoietic stromal cells.

  1. Hematopoietic microenvironment. Origin, lineage, and transplantability of the stromal cells in long-term bone marrow cultures from chimeric mice

    International Nuclear Information System (INIS)

    Studies of bone marrow transplant patients have suggested that the stromal cells of the in vitro hematopoietic microenvironment are transplantable into conditioned recipients. Moreover, in patients with myeloproliferative disorders, all of the stromal cells, which include presumptive endothelial cells, appear to be derived from hematopoietic precursors. To confirm these findings, we have constructed two chimeric mouse models: (a) traditional radiation chimeras, and (b) fetal chimeras, produced by placental injection of bone marrow into genetically anemic Wx/Wv fetuses, a technique that essentially precludes engraftment of nonhematopoietic cells. Using two-color indirect immunofluorescence, the stromal cells in long-term bone marrow culture derived from these chimeras were analyzed for donor or host origin by strain-specific H-2 antigens, and for cell lineage by a variety of other specific markers. 75-95% of the stromal cells were shown to be hematopoietic cells of the monocyte-macrophage lineage, based upon donor origin, phagocytosis, and expression of specific hematopoietic surface antigens. The remaining 5-25% of the stromal cells were exclusively host in origin. Apart from occasional fat cells, these cells uniformly expressed collagen type IV, laminin, and a surface antigen associated with endothelial cells. Since these endothelial-like cells are not transplantable into radiation or fetal chimeras, they are not derived from hematopoietic stem cells. The contrast between our findings and human studies suggests either unexpected species differences in the origin of stromal lineages or limitations in the previous methodology used to detect nonhematopoietic stromal cells

  2. Partitioning Heritability of Regulatory and Cell-Type-Specific Variants across 11 Common Diseases

    Science.gov (United States)

    Gusev, Alexander; Lee, S. Hong; Trynka, Gosia; Finucane, Hilary; Vilhjálmsson, Bjarni J.; Xu, Han; Zang, Chongzhi; Ripke, Stephan; Bulik-Sullivan, Brendan; Stahl, Eli; Ripke, Stephan; Neale, Benjamin M.; Corvin, Aiden; Walters, James T.R.; Farh, Kai-How; Holmans, Peter A.; Lee, Phil; Bulik-Sullivan, Brendan; Collier, David A.; Huang, Hailiang; Pers, Tune H.; Agartz, Ingrid; Agerbo, Esben; Albus, Margot; Alexander, Madeline; Amin, Farooq; Bacanu, Silviu A.; Begemann, Martin; Belliveau, Richard A.; Bene, Judit; Bergen, Sarah E.; Bevilacqua, Elizabeth; Bigdeli, Tim B.; Black, Donald W.; Børglum, Anders D.; Bruggeman, Richard; Buccola, Nancy G.; Buckner, Randy L.; Byerley, William; Cahn, Wiepke; Cai, Guiqing; Campion, Dominique; Cantor, Rita M.; Carr, Vaughan J.; Carrera, Noa; Catts, Stanley V.; Chambert, Kimberly D.; Chan, Raymond C.K.; Chen, Ronald Y.L.; Chen, Eric Y.H.; Cheng, Wei; Cheung, Eric F.C.; Chong, Siow Ann; Cloninger, C. Robert; Cohen, David; Cohen, Nadine; Cormican, Paul; Craddock, Nick; Crowley, James J.; Curtis, David; Davidson, Michael; Davis, Kenneth L.; Degenhardt, Franziska; Del Favero, Jurgen; DeLisi, Lynn E.; Demontis, Ditte; Dikeos, Dimitris; Dinan, Timothy; Djurovic, Srdjan; Donohoe, Gary; Drapeau, Elodie; Duan, Jubao; Dudbridge, Frank; Durmishi, Naser; Eichhammer, Peter; Eriksson, Johan; Escott-Price, Valentina; Essioux, Laurent; Fanous, Ayman H.; Farrell, Martilias S.; Frank, Josef; Franke, Lude; Freedman, Robert; Freimer, Nelson B.; Friedl, Marion; Friedman, Joseph I.; Fromer, Menachem; Genovese, Giulio; Georgieva, Lyudmila; Gershon, Elliot S.; Giegling, Ina; Giusti-Rodrguez, Paola; Godard, Stephanie; Goldstein, Jacqueline I.; Golimbet, Vera; Gopal, Srihari; Gratten, Jacob; Grove, Jakob; de Haan, Lieuwe; Hammer, Christian; Hamshere, Marian L.; Hansen, Mark; Hansen, Thomas; Haroutunian, Vahram; Hartmann, Annette M.; Henskens, Frans A.; Herms, Stefan; Hirschhorn, Joel N.; Hoffmann, Per; Hofman, Andrea; Hollegaard, Mads V.; Hougaard, David M.; Ikeda, Masashi; Joa, Inge; Julià, Antonio; Kahn, René S.; Kalaydjieva, Luba; Karachanak-Yankova, Sena; Karjalainen, Juha; Kavanagh, David; Keller, Matthew C.; Kelly, Brian J.; Kennedy, James L.; Khrunin, Andrey; Kim, Yunjung; Klovins, Janis; Knowles, James A.; Konte, Bettina; Kucinskas, Vaidutis; Kucinskiene, Zita Ausrele; Kuzelova-Ptackova, Hana; Kähler, Anna K.; Laurent, Claudine; Keong, Jimmy Lee Chee; Lee, S. Hong; Legge, Sophie E.; Lerer, Bernard; Li, Miaoxin; Li, Tao; Liang, Kung-Yee; Lieberman, Jeffrey; Limborska, Svetlana; Loughland, Carmel M.; Lubinski, Jan; Lnnqvist, Jouko; Macek, Milan; Magnusson, Patrik K.E.; Maher, Brion S.; Maier, Wolfgang; Mallet, Jacques; Marsal, Sara; Mattheisen, Manuel; Mattingsdal, Morten; McCarley, Robert W.; McDonald, Colm; McIntosh, Andrew M.; Meier, Sandra; Meijer, Carin J.; Melegh, Bela; Melle, Ingrid; Mesholam-Gately, Raquelle I.; Metspalu, Andres; Michie, Patricia T.; Milani, Lili; Milanova, Vihra; Mokrab, Younes; Morris, Derek W.; Mors, Ole; Mortensen, Preben B.; Murphy, Kieran C.; Murray, Robin M.; Myin-Germeys, Inez; Mller-Myhsok, Bertram; Nelis, Mari; Nenadic, Igor; Nertney, Deborah A.; Nestadt, Gerald; Nicodemus, Kristin K.; Nikitina-Zake, Liene; Nisenbaum, Laura; Nordin, Annelie; O’Callaghan, Eadbhard; O’Dushlaine, Colm; O’Neill, F. Anthony; Oh, Sang-Yun; Olincy, Ann; Olsen, Line; Van Os, Jim; Pantelis, Christos; Papadimitriou, George N.; Papiol, Sergi; Parkhomenko, Elena; Pato, Michele T.; Paunio, Tiina; Pejovic-Milovancevic, Milica; Perkins, Diana O.; Pietilinen, Olli; Pimm, Jonathan; Pocklington, Andrew J.; Powell, John; Price, Alkes; Pulver, Ann E.; Purcell, Shaun M.; Quested, Digby; Rasmussen, Henrik B.; Reichenberg, Abraham; Reimers, Mark A.; Richards, Alexander L.; Roffman, Joshua L.; Roussos, Panos; Ruderfer, Douglas M.; Salomaa, Veikko; Sanders, Alan R.; Schall, Ulrich; Schubert, Christian R.; Schulze, Thomas G.; Schwab, Sibylle G.; Scolnick, Edward M.; Scott, Rodney J.; Seidman, Larry J.; Shi, Jianxin; Sigurdsson, Engilbert; Silagadze, Teimuraz; Silverman, Jeremy M.; Sim, Kang; Slominsky, Petr; Smoller, Jordan W.; So, Hon-Cheong; Spencer, Chris C.A.; Stahl, Eli A.; Stefansson, Hreinn; Steinberg, Stacy; Stogmann, Elisabeth; Straub, Richard E.; Strengman, Eric; Strohmaier, Jana; Stroup, T. Scott; Subramaniam, Mythily; Suvisaari, Jaana; Svrakic, Dragan M.; Szatkiewicz, Jin P.; Sderman, Erik; Thirumalai, Srinivas; Toncheva, Draga; Tooney, Paul A.; Tosato, Sarah; Veijola, Juha

    2014-01-01

    Regulatory and coding variants are known to be enriched with associations identified by genome-wide association studies (GWASs) of complex disease, but their contributions to trait heritability are currently unknown. We applied variance-component methods to imputed genotype data for 11 common diseases to partition the heritability explained by genotyped SNPs (hg2) across functional categories (while accounting for shared variance due to linkage disequilibrium). Extensive simulations showed that in contrast to current estimates from GWAS summary statistics, the variance-component approach partitions heritability accurately under a wide range of complex-disease architectures. Across the 11 diseases DNaseI hypersensitivity sites (DHSs) from 217 cell types spanned 16% of imputed SNPs (and 24% of genotyped SNPs) but explained an average of 79% (SE = 8%) of hg2 from imputed SNPs (5.1× enrichment; p = 3.7 × 10−17) and 38% (SE = 4%) of hg2 from genotyped SNPs (1.6× enrichment, p = 1.0 × 10−4). Further enrichment was observed at enhancer DHSs and cell-type-specific DHSs. In contrast, coding variants, which span 1% of the genome, explained <10% of hg2 despite having the highest enrichment. We replicated these findings but found no significant contribution from rare coding variants in independent schizophrenia cohorts genotyped on GWAS and exome chips. Our results highlight the value of analyzing components of heritability to unravel the functional architecture of common disease. PMID:25439723

  3. Tracking the extramedullary PML-RARα-positive cell reservoirs in a preclinical model: biomarker of long-term drug efficacy.

    Science.gov (United States)

    Pokorna, Katerina; Le Pogam, Carole; Chopin, Martine; Balitrand, Nicole; Reboul, Murielle; Cassinat, Bruno; Chomienne, Christine; Padua, Rose Ann; Pla, Marika

    2013-02-01

    Using an acute promyelocytic leukemia (APL) preclinical model, we show that oncogene-specific PCR (Polymerase Chain Reaction)-based assays allow to evaluate the efficacy of immunotherapy combining all-trans retinoic acid (ATRA) and a DNA-based vaccine targeting the promyelocytic leukemia-retinoic acid receptor alpha (PML-RARα) oncogene. Kaplan-Meier survival analysis according to the peripheral blood PML-RARα normalized copy number (NCN) clearly shows that ATRA + DNA-treated mice with an NCN lower than 10 (43%) formed the group with a highly significant (p < 0.0001) survival advantage. Furthermore, a PCR assay was used to assess various tissues and organs for the presence of PML-RARα-positive cells in long-term survivors (n = 15). As expected, the majority of mice (n = 10) had no measurable tissue level of PML-RARα. However, five mice showed a weak positive signal in both the brain and spleen (n = 2), in the brain only (n = 2) and in the spleen only (n = 1). Thus tracking the oncogene-positive cells in long-term survivors reveals for the first time that extramedullary PML-RARα-positive cell reservoirs such as the brain may persist and be involved in relapses.

  4. Long-term dexamethasone treatment alters the histomorphology of acinar cells in rat parotid and submandibular glands.

    Science.gov (United States)

    Bighetti, Bruna B; d Assis, Gerson F; Vieira, Danilo C; Violato, Natalia M; Cestari, Tania M; Taga, Rumio; Bosqueiro, José R; Rafacho, Alex

    2014-10-01

    Glucocorticoids (GCs) induce insulin resistance (IR), a condition known to alter oral homeostasis. This study investigated the effects of long-term dexamethasone administration on morphofunctional aspects of salivary glands. Male Wistar rats received daily injections of dexamethasone [0.1 mg/kg body weight (b.w.), intraperitoneally] for 10 days (DEX), whereas control rats received saline. Subsequently, glycaemia, insulinaemia, insulin secretion and salivary flow were analysed. The parotid and submandibular glands were collected for histomorphometric evaluation and Western blot experiments. The DEX rats were found to be normoglycaemic, hyperinsulinaemic, insulin resistant and glucose intolerant (P glands (P salivary flux rate. The hypotrophy in both glands observed in the DEX group was associated with marked reduction in the volume of the acinar cells in these glands of 50% and 26% respectively (P acinar cells was increased in the submandibular glands of the DEX rats (P glands. The levels of proteins related to insulin and survival signalling in both glands did not differ between the groups. In conclusion, the long-term administration of dexamethasone caused IR, which was associated with significant reductions in both mass and flux rate of the salivary glands. The parotid and submandibular glands exhibited reduced acinar cell volume; however, the submandibular glands displayed acinar hyperplasia, indicating a gland-specific response to GCs. Our data emphasize that GC-based therapies and insulin-resistant states have a negative impact on salivary gland homeostasis.

  5. Uptake of silica covered Quantum Dots into living cells: Long term vitality and morphology study on hyaluronic acid biomaterials.

    Science.gov (United States)

    D'Amico, Michele; Fiorica, Calogero; Palumbo, Fabio Salvatore; Militello, Valeria; Leone, Maurizio; Dubertret, Benoit; Pitarresi, Giovanna; Giammona, Gaetano

    2016-10-01

    Quantum Dots (QDs) are promising very bright and stable fluorescent probes for optical studies in the biological field but water solubility and possible metal bio-contamination need to be addressed. In this work, a simple silica-QD hybrid system is prepared and the uptake in bovine chondrocytes living cells without any functionalization of the external protective silica shield is demonstrated. Moreover, long term treated cells vitality (up to 14days) and the transfer of silica-QDs to the next cell generations are here reported. Confocal fluorescence microscopy was also used to determine the morphology of the so labelled cells and the relative silica-QDs distribution. Finally, we employ silica-QD stained chondrocytes to characterize, as proof of concept, hydrogels obtained from an amphiphilic derivative of hyaluronic acid (HA-EDA-C18) functionalized with different amounts of the RGD peptide. PMID:27287118

  6. Uptake of silica covered Quantum Dots into living cells: Long term vitality and morphology study on hyaluronic acid biomaterials.

    Science.gov (United States)

    D'Amico, Michele; Fiorica, Calogero; Palumbo, Fabio Salvatore; Militello, Valeria; Leone, Maurizio; Dubertret, Benoit; Pitarresi, Giovanna; Giammona, Gaetano

    2016-10-01

    Quantum Dots (QDs) are promising very bright and stable fluorescent probes for optical studies in the biological field but water solubility and possible metal bio-contamination need to be addressed. In this work, a simple silica-QD hybrid system is prepared and the uptake in bovine chondrocytes living cells without any functionalization of the external protective silica shield is demonstrated. Moreover, long term treated cells vitality (up to 14days) and the transfer of silica-QDs to the next cell generations are here reported. Confocal fluorescence microscopy was also used to determine the morphology of the so labelled cells and the relative silica-QDs distribution. Finally, we employ silica-QD stained chondrocytes to characterize, as proof of concept, hydrogels obtained from an amphiphilic derivative of hyaluronic acid (HA-EDA-C18) functionalized with different amounts of the RGD peptide.

  7. Long-Term Survival of Neonatal Porcine Islets Without Sertoli Cells in Rabbits

    OpenAIRE

    Rafael Vald and eacute;s-Gonz and aacute;lez; Ana L. Rodriguez-Ventura; Briceyda Gonz and aacute;lez-Ram and iacute;rez; Benjam and iacute;n Le and oacute;n-Mancilla; Pedro Valencia; Mar and iacute;a del Carmen Garc and iacute;a de Le and oacute;n; Ruy P and eacute;rez-Tamayo

    2013-01-01

    Cell-based therapy is a promising treatment for metabolic disorders such as type-1 diabetes. Transplantation protocols have investigated several anatomical sites for cell implantation; however, some of these procedures, such as intraportal infusion, can cause organ failure or thrombosis secondarily. Bio-artificial organs could be the choice, although concerns still remain. Using a subcutaneous device, we are able to preserve neonatal porcine islets without sertoli cells in healthy New Zealand...

  8. Th17 cells confer long-term adaptive immunity to oral mucosal Candida albicans infections.

    Science.gov (United States)

    Hernández-Santos, N; Huppler, A R; Peterson, A C; Khader, S A; McKenna, K C; Gaffen, S L

    2013-09-01

    Oropharyngeal candidiasis (OPC) is an opportunistic infection caused by Candida albicans. Despite its prevalence, little is known about C. albicans-specific immunity in the oral mucosa. Vaccines against Candida generate both T helper type 1 (Th1) and Th17 responses, and considerable evidence implicates interleukin (IL)-17 in immunity to OPC. However, IL-17 is also produced by innate immune cells that are remarkably similar to Th17 cells, expressing the same markers and localizing to similar mucosal sites. To date, the relative contribution(s) of Th1, Th17, and innate IL-17-producing cells in OPC have not been clearly defined. Here, we sought to determine the nature and function of adaptive T-cell responses to OPC, using a new recall infection model. Mice subjected to infection and re-challenge with Candida mounted a robust and stable antigen-specific IL-17 response in CD4+ but not CD8+ T cells. There was little evidence for Th1 or Th1/Th17 responses. The Th17 response promoted accelerated fungal clearance, and Th17 cells could confer protection in Rag1-/- mice upon adoptive transfer. Surprisingly, CD4 deficiency did not cause OPC but was instead associated with compensatory IL-17 production by Tc17 and CD3+CD4-CD8- cells. Therefore, classic CD4+Th17 cells protect from OPC but can be compensated by other IL-17-producing cells in CD4-deficient hosts. PMID:23250275

  9. Long-term adaptation to hypoxia preserves hematopoietic stem cell function.

    Science.gov (United States)

    Chen, Jichun; Kang, Ju-Gyeong; Keyvanfar, Keyvan; Young, Neal S; Hwang, Paul M

    2016-09-01

    Molecular oxygen sustains aerobic life, but it also serves as the substrate for oxidative stress, which has been associated with the pathogenesis of disease and with aging. Compared with mice housed in normoxia (21% O2), reducing ambient oxygen to 10% O2 (hypoxia) resulted in increased hematopoietic stem cell (HSC) function as measured by bone marrow (BM) cell engraftment onto lethally irradiated recipients. The number of BM c-Kit(+)Sca-1(+)Lin(-) (KSL) cells as well as the number of cells with other hematopoietic stem and progenitor cell markers were increased in hypoxia mice, whereas the BM cells' colony-forming capacity remained unchanged. KSL cells from hypoxia mice showed a decreased level of oxidative stress and increased expression of transcription factor Gata1 and cytokine receptor c-Mpl, consistent with the observations of increased erythropoiesis and enhanced HSC engraftment. These observations demonstrate the benefit of a hypoxic HSC niche and suggest that hypoxic conditions can be further optimized to preserve stem cell integrity in vivo. PMID:27118043

  10. Long-term performance of a plant microbial fuel cell with Spartina anglica

    NARCIS (Netherlands)

    Timmers, R.A.; Strik, D.P.B.T.B.; Hamelers, H.V.M.; Buisman, C.J.N.

    2010-01-01

    The plant microbial fuel cell is a sustainable and renewable way of electricity production. The plant is integrated in the anode of the microbial fuel cell which consists of a bed of graphite granules. In the anode, organic compounds deposited by plant roots are oxidized by electrochemically active

  11. Lim homeobox genes in the Ctenophore Mnemiopsis leidyi: the evolution of neural cell type specification

    Directory of Open Access Journals (Sweden)

    Simmons David K

    2012-01-01

    Full Text Available Abstract Background Nervous systems are thought to be important to the evolutionary success and diversification of metazoans, yet little is known about the origin of simple nervous systems at the base of the animal tree. Recent data suggest that ctenophores, a group of macroscopic pelagic marine invertebrates, are the most ancient group of animals that possess a definitive nervous system consisting of a distributed nerve net and an apical statocyst. This study reports on details of the evolution of the neural cell type specifying transcription factor family of LIM homeobox containing genes (Lhx, which have highly conserved functions in neural specification in bilaterian animals. Results Using next generation sequencing, the first draft of the genome of the ctenophore Mnemiopsis leidyi has been generated. The Lhx genes in all animals are represented by seven subfamilies (Lhx1/5, Lhx3/4, Lmx, Islet, Lhx2/9, Lhx6/8, and LMO of which four were found to be represented in the ctenophore lineage (Lhx1/5, Lhx3/4, Lmx, and Islet. Interestingly, the ctenophore Lhx gene complement is more similar to the sponge complement (sponges do not possess neurons than to either the cnidarian-bilaterian or placozoan Lhx complements. Using whole mount in situ hybridization, the Lhx gene expression patterns were examined and found to be expressed around the blastopore and in cells that give rise to the apical organ and putative neural sensory cells. Conclusion This research gives us a first look at neural cell type specification in the ctenophore M. leidyi. Within M. leidyi, Lhx genes are expressed in overlapping domains within proposed neural cellular and sensory cell territories. These data suggest that Lhx genes likely played a conserved role in the patterning of sensory cells in the ancestor of sponges and ctenophores, and may provide a link to the expression of Lhx orthologs in sponge larval photoreceptive cells. Lhx genes were later co-opted into patterning more

  12. Modeling long-term host cell-Giardia lamblia interactions in an in vitro co-culture system.

    Directory of Open Access Journals (Sweden)

    Bridget S Fisher

    Full Text Available Globally, there are greater than 700,000 deaths per year associated with diarrheal disease. The flagellated intestinal parasite, Giardia lamblia, is one of the most common intestinal pathogens in both humans and animals throughout the world. While attached to the gastrointestinal epithelium, Giardia induces epithelial cell apoptosis, disrupts tight junctions, and increases intestinal permeability. The underlying cellular and molecular mechanisms of giardiasis, including the role lamina propria immune cells, such as macrophages, play in parasite control or clearance are poorly understood. Thus far, one of the major obstacles in ascertaining the mechanisms of Giardia pathology is the lack of a functionally relevant model for the long-term study of the parasite in vitro. Here we report on the development of an in vitro co-culture model which maintains the basolateral-apical architecture of the small intestine and allows for long-term survival of the parasite. Using transwell inserts, Caco-2 intestinal epithelial cells and IC-21 macrophages are co-cultured in the presence of Giardia trophozoites. Using the developed model, we show that Giardia trophozoites survive over 21 days and proliferate in a combination media of Caco-2 cell and Giardia medium. Giardia induces apoptosis of epithelial cells through caspase-3 activation and macrophages do not abrogate this response. Additionally, macrophages induce Caco-2 cells to secrete the pro-inflammatory cytokines, GRO and IL-8, a response abolished by Giardia indicating parasite induced suppression of the host immune response. The co-culture model provides additional complexity and information when compared to a single-cell model. This model will be a valuable tool for answering long-standing questions on host-parasite biology that may lead to discovery of new therapeutic interventions.

  13. Long-Term Survival of Neonatal Porcine Islets Without Sertoli Cells in Rabbits

    Directory of Open Access Journals (Sweden)

    Rafael Vald and eacute;s-Gonz and aacute;lez

    2013-04-01

    Full Text Available Cell-based therapy is a promising treatment for metabolic disorders such as type-1 diabetes. Transplantation protocols have investigated several anatomical sites for cell implantation; however, some of these procedures, such as intraportal infusion, can cause organ failure or thrombosis secondarily. Bio-artificial organs could be the choice, although concerns still remain. Using a subcutaneous device, we are able to preserve neonatal porcine islets without sertoli cells in healthy New Zealand rabbits. Devices were implanted in the back of the animals underneath the skin, and after 3 months the islets were transplanted. Histology showed the presence of inflammatory cells, predominantly eosinophils; however, insulin- and glucagon-positive cell clusters were identified inside the device at different time points for at least 90 days, and porcine C-peptide was also detected during the follow-up, indicating graft functionality. We have found that our device induces the deposition of a fibrous matrix enriched in blood vessels, which forms a good place for cell grafting, and this model is probably able to induce an immunoprivileged site. Under these conditions, transplanted porcine islet cells have the capability of producing insulin and glucagon for at least three months. [Arch Clin Exp Surg 2013; 2(2.000: 101-108

  14. Identification of Cell Type-Specific Differences in Erythropoietin Receptor Signaling in Primary Erythroid and Lung Cancer Cells

    Science.gov (United States)

    Salopiata, Florian; Depner, Sofia; Wäsch, Marvin; Böhm, Martin E.; Mücke, Oliver; Plass, Christoph; Lehmann, Wolf D.; Kreutz, Clemens; Timmer, Jens; Klingmüller, Ursula

    2016-01-01

    Lung cancer, with its most prevalent form non-small-cell lung carcinoma (NSCLC), is one of the leading causes of cancer-related deaths worldwide, and is commonly treated with chemotherapeutic drugs such as cisplatin. Lung cancer patients frequently suffer from chemotherapy-induced anemia, which can be treated with erythropoietin (EPO). However, studies have indicated that EPO not only promotes erythropoiesis in hematopoietic cells, but may also enhance survival of NSCLC cells. Here, we verified that the NSCLC cell line H838 expresses functional erythropoietin receptors (EPOR) and that treatment with EPO reduces cisplatin-induced apoptosis. To pinpoint differences in EPO-induced survival signaling in erythroid progenitor cells (CFU-E, colony forming unit-erythroid) and H838 cells, we combined mathematical modeling with a method for feature selection, the L1 regularization. Utilizing an example model and simulated data, we demonstrated that this approach enables the accurate identification and quantification of cell type-specific parameters. We applied our strategy to quantitative time-resolved data of EPO-induced JAK/STAT signaling generated by quantitative immunoblotting, mass spectrometry and quantitative real-time PCR (qRT-PCR) in CFU-E and H838 cells as well as H838 cells overexpressing human EPOR (H838-HA-hEPOR). The established parsimonious mathematical model was able to simultaneously describe the data sets of CFU-E, H838 and H838-HA-hEPOR cells. Seven cell type-specific parameters were identified that included for example parameters for nuclear translocation of STAT5 and target gene induction. Cell type-specific differences in target gene induction were experimentally validated by qRT-PCR experiments. The systematic identification of pathway differences and sensitivities of EPOR signaling in CFU-E and H838 cells revealed potential targets for intervention to selectively inhibit EPO-induced signaling in the tumor cells but leave the responses in erythroid

  15. Identification of Cell Type-Specific Differences in Erythropoietin Receptor Signaling in Primary Erythroid and Lung Cancer Cells.

    Science.gov (United States)

    Merkle, Ruth; Steiert, Bernhard; Salopiata, Florian; Depner, Sofia; Raue, Andreas; Iwamoto, Nao; Schelker, Max; Hass, Helge; Wäsch, Marvin; Böhm, Martin E; Mücke, Oliver; Lipka, Daniel B; Plass, Christoph; Lehmann, Wolf D; Kreutz, Clemens; Timmer, Jens; Schilling, Marcel; Klingmüller, Ursula

    2016-08-01

    Lung cancer, with its most prevalent form non-small-cell lung carcinoma (NSCLC), is one of the leading causes of cancer-related deaths worldwide, and is commonly treated with chemotherapeutic drugs such as cisplatin. Lung cancer patients frequently suffer from chemotherapy-induced anemia, which can be treated with erythropoietin (EPO). However, studies have indicated that EPO not only promotes erythropoiesis in hematopoietic cells, but may also enhance survival of NSCLC cells. Here, we verified that the NSCLC cell line H838 expresses functional erythropoietin receptors (EPOR) and that treatment with EPO reduces cisplatin-induced apoptosis. To pinpoint differences in EPO-induced survival signaling in erythroid progenitor cells (CFU-E, colony forming unit-erythroid) and H838 cells, we combined mathematical modeling with a method for feature selection, the L1 regularization. Utilizing an example model and simulated data, we demonstrated that this approach enables the accurate identification and quantification of cell type-specific parameters. We applied our strategy to quantitative time-resolved data of EPO-induced JAK/STAT signaling generated by quantitative immunoblotting, mass spectrometry and quantitative real-time PCR (qRT-PCR) in CFU-E and H838 cells as well as H838 cells overexpressing human EPOR (H838-HA-hEPOR). The established parsimonious mathematical model was able to simultaneously describe the data sets of CFU-E, H838 and H838-HA-hEPOR cells. Seven cell type-specific parameters were identified that included for example parameters for nuclear translocation of STAT5 and target gene induction. Cell type-specific differences in target gene induction were experimentally validated by qRT-PCR experiments. The systematic identification of pathway differences and sensitivities of EPOR signaling in CFU-E and H838 cells revealed potential targets for intervention to selectively inhibit EPO-induced signaling in the tumor cells but leave the responses in erythroid

  16. Long-Term Follow-Up after Reduced-Intensity Conditioning and Stem Cell Transplantation for Childhood Nonmalignant Disorders.

    Science.gov (United States)

    Madden, Lisa M; Hayashi, Robert J; Chan, Ka Wah; Pulsipher, Michael A; Douglas, Dorothea; Hale, Gregory A; Chaudhury, Sonali; Haut, Paul; Kasow, Kimberly A; Gilman, Andrew L; Murray, Lisa M; Shenoy, Shalini

    2016-08-01

    Reduced-intensity conditioning (RIC) before hematopoietic stem cell transplantation (HCT) in children could result in fewer complications during follow-up compared with myeloablative regimens. Hence, many RIC regimens are under investigation, but long-term follow-up is essential. We describe late follow-up beyond 2 years post-HCT in 43 children with nonmalignant disorders who underwent related or unrelated donor (56%) HCT on a multicenter study using a RIC regimen (alemtuzumab, fludarabine, and melphalan) followed by bone marrow (n = 30), peripheral blood (n = 3), or umbilical cord blood (n = 10) HCT for immune dysfunction, bone marrow failure, metabolic disorders, or hemoglobinopathy. Recipients (median age, 7.5 years; range, 3 to 26) underwent HCT 2 to 8 years (median, 3.1 years) before this report. Full donor (67%) or stable mixed chimerism (33%) was noted without late graft rejection. Five patients (12%) required systemic immunosuppression therapy (IST) beyond 2 years post-HCT for graft-versus-host disease (GVHD); 2 patients died 38 and 79 months later, whereas the others improved, enabling an IST wean. Overall, 17 complications were documented in 10 patients (23%). Complications not related to GVHD included hypothyroidism (n = 2), low grade neoplasms (n = 2), and delayed puberty (n = 1). One patient with GVHD had ovarian failure; all other postpubertal females resumed normal ovarian function. Twenty-seven of 28 school-age recipients were functioning at grade level. RIC HCT recipients thus had few regimen-related toxicities during long-term follow-up. However, objective long-term follow-up is still necessary to identify complications so timely intervention may be planned. PMID:27164064

  17. Long-term survival of exogenous embryonic stem cells in adult bone marrow

    Institute of Scientific and Technical Information of China (English)

    Yueying Li; Jing Xu; Weiping Yuan; Shaorong Gao; Tao Cheng; Yanfeng Liu; Jing He; Fengchao Wang; Sheng Liu; Yu Zhang; Zhaohui Kou; Zhenyu Ju; Guoguang Zheng

    2011-01-01

    @@ Dear Editor, Embryonic stem cell (ESC) is an attractive and promising source in regenerative medicine.However, current technologies for ESC differentiation do not ensure complete or synchronized conversion of ESCs into a specific functional tissue type for therapeutic purposes.

  18. Enhancing the survival of grafted cardiac stem cells for long-term imaging

    Energy Technology Data Exchange (ETDEWEB)

    Le, Uyenchi N.; Tae, Seong Ho; Bom, Hee Seung; Min, Jung Joon [Chonnam National University Medical School, Gwangju (Korea, Republic of)

    2007-07-01

    Heat shock treatment is known to induce the protection for cells from various environmental insults. Akt (protein kinase B) - with anti-apoptotic activity - has presently been reemerged as a critical enzyme in several signal transduction pathways involved in cell proliferation and programmed cell death. We hypothesized that thermotic treatment and Akt activity in genetically modified cardiomyoblasts would improve their survival after transplantation. Embryonic rat H9c2 cardiomyoblasts were simultaneously transfected with adenovirus containing luciferase reporter gene (MOl 50) and another containing Akt gene [MOl (0 100) ]. 5x106 harvested cells were i.m. implanted into murine skeletal muscles. Bioluminescence imaging was acquired for everyday and luciferase assay was performed to validate the imaging data. For thermotic challenge, adenovirus-mediated flue expressing H9c2 cells were subjected to great heat of 42 .deg. C for 1 hr and re-cultured at 37 .deg. C for 18 hours. Expression of heat shock protein in cells was detected in vitro by Western-blotting. 5x106 normal and shocked cells were implanted into mouse thigh (n = 5) and the animals were imaged with bioluminescence imaging system. In vitro evidences showed a high level expression of Akt and HSP in transfected H9c2 cells. Animals carrying Akt expressed bioluminescence signals until day 34 of post-implantation. The Flue activity was significantly higher in the shocked H9c2 cell-implanted rats and detected over 10 days as compared with the control group. The graft cell death was reduced by 73% at day 2 (1.46+ 10-7 p/s/cm{sup 2}/sr), 51% at day 3 (1.02+10-7 p/s/cm{sup 2}/sr), and 8% at day 10 (1.62+ 10-6 p/s/cm{sup 2}/sr). We revealed here improvement of donor cell's survival induced by the anti-apoptotic means of Akt genetic therapy or heat shock. Utility of bioluminescence imaging resulted in a potential to noninvasively and repetitively monitor implanted cardiac myoblasts over time.

  19. A Novel Whole-Cell Mechanism for Long-Term Memory Enhancement

    OpenAIRE

    Iris Reuveni; Drorit Saar; Edi Barkai

    2013-01-01

    Olfactory-discrimination learning was shown to induce a profound long-lasting enhancement in the strength of excitatory and inhibitory synapses of pyramidal neurons in the piriform cortex. Notably, such enhancement was mostly pronounced in a sub-group of neurons, entailing about a quarter of the cell population. Here we first show that the prominent enhancement in the subset of cells is due to a process in which all excitatory synapses doubled their strength and that this increase was mediate...

  20. Th17 cells confer long term adaptive immunity to oral mucosal Candida albicans infections

    OpenAIRE

    Hernández-Santos, Nydiaris; Huppler, Anna R; Peterson, Alanna C.; Khader, Shabaana A.; McKenna, Kyle C.; Sarah L Gaffen

    2012-01-01

    Oropharyngeal candidiasis (OPC) is an opportunistic infection caused by Candida albicans. Despite its prevalence, little is known about C. albicans-specific immunity in the oral mucosa. Vaccines against Candida generate both Th1 and Th17 responses, and considerable evidence implicates IL-17 in immunity to OPC. However, IL-17 is also produced by innate immune cells that are remarkably similar to Th17 cells, expressing the same markers and localizing to similar mucosal sites. To date, the relat...

  1. Comparison of Different Culture Mode for Long-term Expansion of Neural Stem Cells

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    1 Introduction The mammalian central nervous system(CNS) is incredibly complex and possesses only a limited ability to recover from damage~([1]). Fortunately, the discovery of self-renewing stem cell populations within the fetal and adult CNS has opened promising lines of inquiry. Neural stem cells (NSCs) can be cultured in two modes in vitro, suspension and monolayer. Reynolds~([2]) and other groups culture NSCs as neurospheres in suspension. Alternatively Gage cultured NSCs in monolayer. There is little s...

  2. Preliminary investigations of Spirulina effect on cancer cells: interest for long-term manned space missions

    Science.gov (United States)

    Baatout, S.; Bekaert, S.; Hendrickx, L.; Derradji, H.; Mergeay, M.

    Background In view of long haul space exploration missions the development of regenerative life support systems is of crucial importance to increase the crew autonomy and decrease the cost associated to the mass embarked Therefore in the late 80 s the European Space Agency initiated the MELiSSA project Micro-Ecological Life Support System Alternative MELiSSA has been conceived as a micro-organisms and higher plant process enabling high recycling efficiency The cyanobacteria Arthrospira sp is occupying one of the MELiSSA compartments Its genome is now being sequenced and this will help to better understand or improve its food value as well as to have a look at its putative toxic potential Aim In this study we were interested in studying the threshold of intrinsic cytotoxic effects of Spirulina dry extract from Sigma containing washed and lyophilized mixed Arthrospira strains on human cancer cells and its cell type dependency Method For that purpose we used flow cytometry to estimate cell death apoptosis and necrosis in three human leukaemic cell lines HELA cervix carcinoma IM-9 multiple myeloma K562 chronic myelogenous leukaemia Cells were cultured in the presence of an aqueous extract of Spirulina concentrations ranging from 0 to 500 mu g ml for 15 to 40 hours Apoptosis and necrosis were evaluated by annexin-V-PI staining cell size and granularity Early apoptosis was monitored by analysing the maintenance of mitochondrial membrane potential DioC 6 3 and the

  3. Short- and long-term effects of silver nanoparticles on human microvascular endothelial cells

    Institute of Scientific and Technical Information of China (English)

    Sara; Castiglioni; Clelia; Caspani; Alessandra; Cazzaniga; Jeanette; AM; Maier

    2014-01-01

    AIM: To study the response to silver nanoparticles(Ag NP) of human microvascular endothelial cells, protagonists of angiogenesis. METHODS: We cultured human microvascular endothelial cells and endothelial colony-forming cells in their corresponding growth medium. Stock solutions of Ag NP were prepared in culture medium and sonicated before use. They were added at different concentrations and for different times to culture media. The toxicity of Ag NP was investigated by measuring the reduction of yellow tetrazolium salt to dark purple formazan(MTT assay) at 575 nm. After staining with trypan blue, cell proliferation was assessed by counting viable cells. The lactate dehydrogenase leakage assay was performed on culture media by following the oxidation of NADH to NAD+ and monitoring the reaction kinetically at 340 nm. Reactive oxygen species production was quantified using 2’-7’-dichlorofluorescein diacetate. The alkaline comet assay was performed after mixing the cells with low melting-point agarose. Electrophoresis was then conducted and the samples were stained with ethidium bromide and analyzed with a fluorescence microscope.RESULTS: Ag NP are cytotoxic in a dose and time dependent fashion for HMEC. At high concentrations, Ag NP determine loss of membrane integrity as demonstrated by the increased activity of lactate dehydrogenase in the culture medium. Ag NP rapidly stimulate the formation of free radicals. However, pre-incubation with Trolox, apocynin, or N-acetyl-L-cysteine, antioxidants which have different structure and act through different mechanisms, is not sufficient to prevent cytotoxicity. Ag NP also induce DNA damage dose-dependently, as shown by comet assay. When exposed to sublethal concentrations of Ag NP for long times, the cells remain viable but are growth retarded. Interestingly, removal of Ag NP partially rescues cell growth. Also genotoxicity is reversible upon removal of Ag NP from culture medium, suggesting that no permanent

  4. Long-term Fluorometholone Topical Use Induces Ganglion Cell Damage in Rats Analyzed With Optical Coherence Tomography.

    Science.gov (United States)

    Lin, Cheng-Hui; Liao, Po-Lin; Hsiao, George; Li, Ching-Hao; Huang, Shih-Hsuan; Tsai, Chi-Hao; Wu, Man-Ru; Lin, Fan-Li; Ho, Jau-Der; Cheng, Hui-Wen; Cheng, Yu-Wen

    2015-10-01

    To determine the toxic effects of long-term topical usage of fluorometholone (FLM) on ganglion cells using a direct in vivo retinopathological Brown Norway (BN) rat model. The BN rat retinal model was investigated with a minimum of 3 rats and a maximum of 4 rats per group. Rats received vehicle and 0.02% FLM suspension via topical administration 3 times a day for 28 days. The fundus images and retinal vessels were detected on days 1, 14, and 28 using Micron III retinal imaging microscope and fundus fluorescein angiography (FFA). For retinal structures, spectral-domain optical coherence tomography (SD-OCT) images were taken after FFA on days 1, 14, and 28 using an SD-OCT Imaging System. For retinal function, electrical signal transduction of photoreceptors and bipolar cells was determined by electroretinographic (ERG) recording on days 1 and 28 and IOP detection. At the end of the experiment on day 28, immunohistochemistry and TUNEL assay were performed to investigate apoptosis in ganglion cells. Total retina and nerve fiber layer (NFL) to the inner plexiform layer (IPL) were significantly thinner following 28 days of FLM treatment. Hematoxylin and eosin stain showed that there were NFL and ganglion cell layer deformations in the FLM group. With FLM treatment, TUNEL assay showed approximately a 4.68-fold increase in apoptotic cells. Moreover, FLM decreased ERG b-wave amplitude by about 56%. Using ophthalmofundoscopy devices, after 28 days of topical administration, FLM decreased NFL-IPL and total retina thickness. This suggests that long-term FLM induces adverse effects with respect to ganglion cell apoptosis. PMID:26141393

  5. TLR-2 activation induces regulatory T cells and long-term suppression of asthma manifestations in mice.

    Directory of Open Access Journals (Sweden)

    Martijn C Nawijn

    Full Text Available Asthma is a chronic inflammatory disease of the airways characterized by variable airway obstruction and airway hyperresponsiveness (AHR. The T regulatory (Treg cell subset is critically important for the regulation of immune responses. Adoptive transfer of Treg cells has been shown to be sufficient for the suppression of airway inflammation in experimental allergic asthma. Intervention strategies aimed at expanding the Treg cell population locally in the airways of sensitized individuals are therefore of high interest as a potential therapeutic treatment for allergic airway disease. Here, we aim to test whether long-term suppression of asthma manifestations can be achieved by locally expanding the Treg cell subset via intranasal administration of a TLR-2 agonist. To model therapeutic intervention aimed at expanding the endogenous Treg population in a sensitized host, we challenged OVA-sensitized mice by OVA inhalation with concomitant intranasal instillation of the TLR-2 agonist Pam3Cys, followed by an additional series of OVA challenges. Pam3Cys treatment induced an acute but transient aggravation of asthma manifestations, followed by a reduction or loss of AHR to methacholine, depending on the time between Pam3Cys treatment and OVA challenges. In addition, Pam3Cys-treatment induced significant reductions of eosinophils and increased numbers of Treg cells in the lung infiltrates. Our data show that, despite having adverse acute effects, TLR2 agonist treatment as a therapeutic intervention induces an expansion of the Treg cell population in the lungs and results in long-term protection against manifestation of allergic asthma upon subsequent allergen provocation. Our data indicate that local expansion of Tregs in allergic airway disease is an interesting therapeutic approach that warrants further investigation.

  6. Isolated tumor endothelial cells maintain specific character during long-term culture

    Energy Technology Data Exchange (ETDEWEB)

    Matsuda, Kohei [Vascular Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Oral Pathology and Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Oral and Maxillofacial Surgery, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Ohga, Noritaka [Vascular Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Hida, Yasuhiro [Surgical Oncology, Hokkaido University Graduate School of Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Muraki, Chikara [Oral Pathology and Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Oral and Maxillofacial Surgery, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Tsuchiya, Kunihiko [Vascular Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Renal and Genitourinary Surgery, Hokkaido University Graduate School of Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Kurosu, Takuro [Vascular Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Oral Pathology and Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Oral and Maxillofacial Surgery, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Akino, Tomoshige [Vascular Biology, Hokkaido University Graduate School of Dental Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Renal and Genitourinary Surgery, Hokkaido University Graduate School of Medicine, N13 W7, Kita-ku, Sapporo 060-8586 (Japan); Shih, Shou-Ching [Pathology, Beth Israel Deaconess Medical Center, 330 Brookline Ave, Boston, MA 02215 (United States); and others

    2010-04-16

    Tumor angiogenesis is necessary for solid tumor progression and metastasis. Increasing evidence indicates that tumor endothelial cells (TECs) are more relevant to the study of tumor angiogenesis than normal endothelial cells (NECs) because their morphologies and gene expression are different from NECs. However, it is challenging to isolate and culture large numbers of pure ECs from tumor tissue since the percentage of ECs is only about 1-2% and tumor cells and fibroblasts easily overgrow them. In addition, there has been concern that isolated TECs may lose their special phenotype once they are dissociated from tumor cells. In this study, we have successfully purified murine TECs from four different human tumor xenografts and NECs from murine dermal tissue. Isolated ECs expressed endothelial markers, such as CD31, VE-cadherin (CD144), and endoglin (CD105), for more than 3 months after isolation. TECs maintained tumor endothelial-specific markers, such as tumor endothelial marker 8 (TEM8) and aminopeptidase N (APN), as in tumor blood vessels in vivo. In addition, TECs were more proliferative and motile than NECs. TECs showed a higher response to VEGF and higher expression of VEGF receptors-1 and -2 than NECs did. Stem cell antigen-1 was up-regulated in all four TECs, suggesting that they have a kind of stemness. Cultured TECs maintain distinct biological differences from NECs as in vivo. In conclusion, it was suggested that TECs are relevant material for tumor angiogenesis research.

  7. Isolated tumor endothelial cells maintain specific character during long-term culture

    International Nuclear Information System (INIS)

    Tumor angiogenesis is necessary for solid tumor progression and metastasis. Increasing evidence indicates that tumor endothelial cells (TECs) are more relevant to the study of tumor angiogenesis than normal endothelial cells (NECs) because their morphologies and gene expression are different from NECs. However, it is challenging to isolate and culture large numbers of pure ECs from tumor tissue since the percentage of ECs is only about 1-2% and tumor cells and fibroblasts easily overgrow them. In addition, there has been concern that isolated TECs may lose their special phenotype once they are dissociated from tumor cells. In this study, we have successfully purified murine TECs from four different human tumor xenografts and NECs from murine dermal tissue. Isolated ECs expressed endothelial markers, such as CD31, VE-cadherin (CD144), and endoglin (CD105), for more than 3 months after isolation. TECs maintained tumor endothelial-specific markers, such as tumor endothelial marker 8 (TEM8) and aminopeptidase N (APN), as in tumor blood vessels in vivo. In addition, TECs were more proliferative and motile than NECs. TECs showed a higher response to VEGF and higher expression of VEGF receptors-1 and -2 than NECs did. Stem cell antigen-1 was up-regulated in all four TECs, suggesting that they have a kind of stemness. Cultured TECs maintain distinct biological differences from NECs as in vivo. In conclusion, it was suggested that TECs are relevant material for tumor angiogenesis research.

  8. Red blood cell engineering in stroma and serum/plasma-free conditions and long term storage.

    Science.gov (United States)

    Kim, Hyun Ok; Baek, Eun Jung

    2012-01-01

    In vitro generation of artificial red blood cells (RBCs) is very important to overcome insufficient and unsafe blood supply. Despite recent progresses in RBCs engineering from several stem cell sources, none of them could succeed in generation of functional RBCs in the absence of serum/plasma and feeder cells. Without the elimination of serum and plasma, human RBC engineering in a large scale is impossible, especially for the future bioreactor system. Using an appropriate combination of cost-effective and safe reagents, the present study demonstrated the terminal maturation of hematopoietic stem cells into enucleated RBCs, which were functional comparable to donated human RBCs. Surprisingly, the viability of erythroid cells was higher in our serum- and feeder-free culture condition than in the previous serum-added condition. This was possible by supplementation with vitamin C in media and hypothermic conditions. Also, our report firstly presents the storability of artificial RBCs, which possibility is essential for clinical application. In summary, our report demonstrates engineering of human applicable RBCs with a dramatically enhanced viability and shelf-life in both serum- and stroma-free conditions. This innovative culture technology could contribute to the realization of the large-scale pharming of human RBCs using bioreactor systems. PMID:21902543

  9. Red blood cell engineering in stroma and serum/plasma-free conditions and long term storage.

    Science.gov (United States)

    Kim, Hyun Ok; Baek, Eun Jung

    2012-01-01

    In vitro generation of artificial red blood cells (RBCs) is very important to overcome insufficient and unsafe blood supply. Despite recent progresses in RBCs engineering from several stem cell sources, none of them could succeed in generation of functional RBCs in the absence of serum/plasma and feeder cells. Without the elimination of serum and plasma, human RBC engineering in a large scale is impossible, especially for the future bioreactor system. Using an appropriate combination of cost-effective and safe reagents, the present study demonstrated the terminal maturation of hematopoietic stem cells into enucleated RBCs, which were functional comparable to donated human RBCs. Surprisingly, the viability of erythroid cells was higher in our serum- and feeder-free culture condition than in the previous serum-added condition. This was possible by supplementation with vitamin C in media and hypothermic conditions. Also, our report firstly presents the storability of artificial RBCs, which possibility is essential for clinical application. In summary, our report demonstrates engineering of human applicable RBCs with a dramatically enhanced viability and shelf-life in both serum- and stroma-free conditions. This innovative culture technology could contribute to the realization of the large-scale pharming of human RBCs using bioreactor systems.

  10. Replica-moulded polydimethylsiloxane culture vessel lids attenuate osmotic drift in long-term cell cultures

    Indian Academy of Sciences (India)

    Axel Blau; Tanja Neumann; Christiane Ziegler; Fabio Benfenati

    2009-03-01

    An imbalance in medium osmolarity is a determinant that affects cell culture longevity. Even in humidified incubators, evaporation of water leads to a gradual increase in osmolarity overtime. We present a simple replica-moulding strategy for producing self-sealing lids adaptable to standard, small-size cell-culture vessels. They are made of polydimethylsiloxane (PDMS), a flexible, transparent and biocompatible material, which is gas-permeable but largely impermeable to water. Keeping cell cultures in a humidified 5% CO2 incubator at 37°C, medium osmolarity increased by +6.86 mosmol/kg/day in standard 35 mm Petri dishes, while PDMS lids attenuated its rise by a factor of four to changes of +1.72 mosmol/kg/ day. Depending on the lid membrane thickness, pH drifts at ambient CO2 levels were attenuated by a factor of 4 to 9. Comparative evaporation studies at temperatures below 60°C yielded a 10-fold reduced water vapour flux of 1.75 g/day/dm2 through PDMS lids as compared with 18.69 g/day/dm2 with conventional Petri dishes. Using such PDMS lids, about 2/3 of the cell cultures grew longer than 30 days in vitro. Among these, the average survival time was 69 days with the longest survival being 284 days under otherwise conventional cell culture conditions.

  11. Long-term exposure to genistein improves insulin secretory function of pancreatic β-cells

    OpenAIRE

    Fu, Zhuo; Liu, Dongmin

    2009-01-01

    We recently found that genistein, a plant-derived natural compound, is a novel cAMP signaling agonist in pancreatic β-cells. In the present study, we further show that exposure of clonal insulin secreting (INS-1E) cells to genistein for 48 h enhanced glucose-stimulated insulin secretion (GSIS), whereas insulin content was not altered, suggesting that genistein-enhanced GSIS is not due to a modulation of insulin synthesis. This genistein’s effect is protein tyrosine kinase- and KATP channel-in...

  12. Long-term culture of genome-stable bipotent stem cells from adult human liver

    NARCIS (Netherlands)

    Huch, Meritxell; Gehart, Helmuth; van Boxtel, Ruben; Hamer, Karien; Blokzijl, Francis; Verstegen, Monique M A; Ellis, Ewa; van Wenum, Martien; Fuchs, Sabine A; de Ligt, Joep; van de Wetering, Marc; Sasaki, Nobuo; Boers, Susanne J; Kemperman, Hans; de Jonge, Jeroen; Ijzermans, Jan N M; Nieuwenhuis, Edward E S; Hoekstra, Ruurdtje; Strom, Stephen; Vries, Robert R G; van der Laan, Luc J W; Cuppen, Edwin; Clevers, Hans

    2015-01-01

    Despite the enormous replication potential of the human liver, there are currently no culture systems available that sustain hepatocyte replication and/or function in vitro. We have shown previously that single mouse Lgr5+ liver stem cells can be expanded as epithelial organoids in vitro and can be

  13. Long-term survival after allogeneic haematopoietic cell transplantation for AML in remission

    DEFF Research Database (Denmark)

    Sengeløv, H; Gerds, T A; Brændstrup, P;

    2013-01-01

    We report the results of non-myeloablative (NM) and myeloablative (MA) conditioning for haematopoietic cell transplantation in 207 consecutive AML patients at a single institution. A total of 122 patients were transplanted in first CR (CR1) and 67 in second CR (CR2). MA conditioning was given to 60...

  14. Small airways dysfunction in long-term survivors of pediatric stem cell transplantation

    DEFF Research Database (Denmark)

    Uhlving, Hilde Hylland; Mathiesen, Sidsel; Buchvald, Frederik;

    2015-01-01

    BACKGROUND: Chronic graft-versus-host disease (cGvHD) in the lungs is a life-threatening complication of allogeneic hematopoietic stem cell transplantation (HSCT). Pulmonary cGvHD is initiated in the peripheral airways, and diagnosis may be delayed by low sensitivity of standard pulmonary functio...

  15. Comparison of Different Culture Mode for Long-term Expansion of Neural Stem Cells

    Institute of Scientific and Technical Information of China (English)

    Ke ZHENG; Dan GE; Tian-Qing LIU; Xue-Hu MA

    2005-01-01

    @@ 1 Introduction The mammalian central nervous system(CNS) is incredibly complex and possesses only a limited ability to recover from damage[1]. Fortunately, the discovery of self-renewing stem cell populations within the fetal and adult CNS has opened promising lines of inquiry.

  16. Study of the long-term operation of a vanadium/oxygen fuel cell

    Science.gov (United States)

    Noack, Jens; Cognard, Gwenn; Oral, Meryem; Küttinger, Michael; Roznyatovskaya, Nataliya; Pinkwart, Karsten; Tübke, Jens

    2016-09-01

    A vanadium/oxygen fuel cell (VOFC) with a geometrically active area of 51 cm2 and two membranes was discontinuously operated over a period of over 676 h with 47 successive tests at room temperature with a current density of 19.6 mA/cm2 in order to investigate signs of ageing. As well as measuring cell voltages, the test setup was also used to measure anode and redox potentials as well as cell and half-cell impedances. The performance data of the VOFC fluctuated widely over the course of the test period, due to different V2+ concentrations and instabilities of the starting solutions on the one hand and complex changes in cathode conditions on the other. The desired behaviour of the anode reactions was achieved primarily through improved methods for producing the V2+ solutions, and remained stable at the end of the experiments. The kinetics of the cathode reactions were temporarily increased by purging with 2 M H2SO4, however their performance decreased over time. The VOFC had symptoms of ageing by complex and overlaid changes in the cathode's triple phase boundary layer and in the special conditions between the two electrodes and membranes.

  17. LONG-TERM SURVIVAL OF SMALL-CELL LUNG-CANCER PATIENTS AFTER CHEMOTHERAPY

    NARCIS (Netherlands)

    VANDERGAAST, A; POSTMUS, PE; BURGHOUTS, J; VANBOLHUIS, C; STAM, J; SPLINTER, TAW

    1993-01-01

    Eighty-one patients with small cell lung cancer (SCLC) with a survival Of more than 2 years start of chemotherapy were studied. Twenty-six of the 28 patients who died of relapsed SCLC had in relapsed before two years and of the 55 who had not then only two (4%) relapsed subsequently. It is stressed

  18. Promoting long-term survival of insulin-producing cell grafts that differentiate from adipose tissue-derived stem cells to cure type 1 diabetes.

    Directory of Open Access Journals (Sweden)

    Shuzi Zhang

    Full Text Available BACKGROUND: Insulin-producing cell clusters (IPCCs have recently been generated in vitro from adipose tissue-derived stem cells (ASCs to circumvent islet shortage. However, it is unknown how long they can survive upon transplantation, whether they are eventually rejected by recipients, and how their long-term survival can be induced to permanently cure type 1 diabetes. IPCC graft survival is critical for their clinical application and this issue must be systematically addressed prior to their in-depth clinical trials. METHODOLOGY/PRINCIPAL FINDINGS: Here we found that IPCC grafts that differentiated from murine ASCs in vitro, unlike their freshly isolated islet counterparts, did not survive long-term in syngeneic mice, suggesting that ASC-derived IPCCs have intrinsic survival disadvantage over freshly isolated islets. Indeed, β cells retrieved from IPCC syngrafts underwent faster apoptosis than their islet counterparts. However, blocking both Fas and TNF receptor death pathways inhibited their apoptosis and restored their long-term survival in syngeneic recipients. Furthermore, blocking CD40-CD154 costimulation and Fas/TNF signaling induced long-term IPCC allograft survival in overwhelming majority of recipients. Importantly, Fas-deficient IPCC allografts exhibited certain immune privilege and enjoyed long-term survival in diabetic NOD mice in the presence of CD28/CD40 joint blockade while their islet counterparts failed to do so. CONCLUSIONS/SIGNIFICANCE: Long-term survival of ASC-derived IPCC syngeneic grafts requires blocking Fas and TNF death pathways, whereas blocking both death pathways and CD28/CD40 costimulation is needed for long-term IPCC allograft survival in diabetic NOD mice. Our studies have important clinical implications for treating type 1 diabetes via ASC-derived IPCC transplantation.

  19. Long-Term Treatment with Erlotinib for EGFR Wild-Type Non-Small Cell Lung Cancer: A Case Report.

    Science.gov (United States)

    Polychronidou, Genovefa; Papakotoulas, Pavlos

    2013-01-01

    The epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) gefitinib and erlotinib are known to have greater efficacy in EGFR mutation-positive non-small cell lung cancer (NSCLC), although erlotinib also has activity in wild-type disease. We report the successful long-term maintenance treatment of a patient with EGFR wild-type NSCLC with gefitinib and later erlotinib. The patient (male; 44 years old; smoker) was diagnosed with EGFR wild-type NSCLC after computer tomography had revealed a mediastinal mass, and histology and mutation testing had identified the tumor as an EGFR wild-type grade 3 adenocarcinoma. The patient received multiple rounds of chemotherapy, followed by gefitinib maintenance (3 years). Later on, he received erlotinib maintenance and developed a persistent rash (grade 1/2) that lasted throughout the treatment. The patient's condition has remained stable on erlotinib for more than 5 years, with no evidence of progression. We describe the patient's disease course and treatment in the context of EGFR TKI therapy and the prognostic factors for long-term clinical outcomes of NSCLC, including the development of erlotinib-induced rash.

  20. Biocompatible fluorescent supramolecular nanofibrous hydrogel for long-term cell tracking and tumor imaging applications

    Science.gov (United States)

    Wang, Huaimin; Mao, Duo; Wang, Youzhi; Wang, Kai; Yi, Xiaoyong; Kong, Deling; Yang, Zhimou; Liu, Qian; Ding, Dan

    2015-11-01

    Biocompatible peptide-based supramolecular hydrogel has recently emerged as a new and promising system for biomedical applications. In this work, Rhodamine B is employed as a new capping group of self-assembling peptide, which not only provides the driving force for supramolecular nanofibrous hydrogel formation, but also endows the hydrogel with intrinsic fluroescence signal, allowing for various bioimaging applications. The fluorescent peptide nanofibrous hydrogel can be formed via disulfide bond reduction. After dilution of the hydrogel with aqueous solution, the fluorescent nanofiber suspension can be obtained. The resultant nanofibers are able to be internalized by the cancer cells and effectively track the HeLa cells for as long as 7 passages. Using a tumor-bearing mouse model, it is also demonstrated that the fluorescent supramolecular nanofibers can serve as an efficient probe for tumor imaging in a high-contrast manner.

  1. Microbial electrolysis cells for production of methane from CO2: long-term performance and perspectives

    OpenAIRE

    Eerten-Jansen, van, M.C.A.A.; Heijne, ter, A.; C J N Buisman; Hamelers, H.V.M.

    2012-01-01

    A methane-producing microbial electrolysis cell (MEC) is a technology to convert CO2 into methane, using electricity as an energy source and microorganisms as the catalyst. A methane-producing MEC provides the possibility to increase the fuel yield per hectare of land area, when the CO2 produced in biofuel production processes is converted to additional fuel methane. Besides increasing fuel yield per hectare of land area, this also results in more efficient use of land area, water, and nutrie...

  2. Ultrastructural study of long-term canine distemper virus infection in tissue culture cells.

    OpenAIRE

    Narang, H K

    1982-01-01

    The morphogenesis of canine distemper virus was studied in Vero cell cultures for 43 days post-inoculation. Active replication of the virus was observed by electron microscopy and assay from 12 h after inoculation on, and peak production was observed on days 5, 14, and 22. From day 28 on, constant but smaller amounts of infectious virus were detected. Two ultrastructural types of intracytoplasmic nucleoprotein filaments were observed; although they first appeared at different times, their sub...

  3. Characterization and Long-Term Testing of Solid Oxide Electrolyzer Cells

    OpenAIRE

    Schiller, Günter; Hörlein, Michael; Tietz, Frank; Friedrich, K. Andreas

    2014-01-01

    A reliable energy supply which is based on increasing shares of sustainable and renewable energy sources, such as wind power and solar energy, requires appropriate storage technologies. Hydrogen as energy carrier, produced by water electrolysis using electric current from regenerative energy sources, offers a high potential in this respect. A very efficient option to produce hydrogen in this way is high-temperature steam electrolysis based on solid oxide electrolyzer cells (SOEC). This techno...

  4. Long-term persistence of T cell memory to HBsAg after hepatitis B vaccination

    Institute of Scientific and Technical Information of China (English)

    Ru-Xiang Wang; Greet J. Boland; Jan van Hattum; Gijsbert C. de Gast

    2004-01-01

    AIM: To determine if the T cell memory to HBsAg can persist for a long time after hepatitis B (HB) vaccination.METHODS: Thirty one vaccine recipients who were healthcare workers (18 females and 13 males aged 34-58 years) from Utrecht University Hospital, Netherlands, and had previously Received a standard course of vaccination for hepatitis B were investigated and another 9 unvaccinated healthy volunteers from the same hospital were used as the control. Blood samples were taken just before the experiment to test serum anti-HBs levels and the subjects were classified into different groups according to their serum titers of anti-HBs and vaccination history. Their peripheral blood mononuclear cells (pBrvMc) were isolated from freshly heparinized venous blood and the proliferative response of Tlymphocytes to the recombinant hepatitis B surface antigen(HBsAg) was investigated.RESULTS: Positive serum anti-HBs was found in 61.3%(19/31) vaccine recipients and a significant in vitro lymphocyte proliferative response to recombinant HBsAg was observed in all the vaccinees with positive anti-HBs. Serum anti-HBs level ≤10 IU/L was found in 38.7% (12/31)subjects. In this study, we specially focused on lymphocyte proliferative response to recombinant HBsAg in those vaccine recipients with serum anti-HBsAg less than 10 IU/L.Most of them had Received a standard course of vaccination about 10 years before. T lymphocyte proliferative response was found positive in 7 of the 12 vaccine recipients. These results confirmed that HBsAg-specific memory T cells remained detectable in the circulation for a long time after vaccination, even when serum anti-HBs level had been undetectable.CONCLUSION: The T cell memory to HBsAg can persist for at least 10 years after HB vaccination. Further booster injection is not necessary in healthy responders to HB vaccine.

  5. Cytomegalovirus shapes long-term immune reconstitution after allogeneic stem cell transplantation

    OpenAIRE

    Itzykson, Raphael; Robin, Marie; Moins-Teisserenc, Helene; Delord, Marc; Busson, Marc; Xhaard, Aliénor; de Fontebrune, Flore Sicre; de Latour, Régis Peffault; Toubert, Antoine; Socié, Gérard

    2015-01-01

    Immune reconstitution after allogeneic stem cell transplantation is a dynamic and complex process depending on the recipient and donor characteristics, on the modalities of transplantation, and on the occurrence of graft-versus-host disease. Multivariate methods widely used for gene expression profiling can simultaneously analyze the patterns of a great number of biological variables on a heterogeneous set of patients. Here we use these methods on flow cytometry assessment of up to 25 lymphoc...

  6. Alcohol impairs long-term depression at the cerebellar parallel fiber-Purkinje cell synapse

    OpenAIRE

    Belmeguenai, A.; Botta, Paolo; Weber, John; Carta, Mario; De Ruiter, Martijn; De Zeeuw, Chris; Valenzuela, Fernando; Hansel, Christian

    2008-01-01

    textabstractAcute alcohol consumption causes deficits in motor coordination and gait, suggesting an involvement of cerebellar circuits, which play a role in the fine adjustment of movements and in motor learning. It has previously been shown that ethanol modulates inhibitory transmission in the cerebellum and affects synaptic transmission and plasticity at excitatory climbing fiber (CF) to Purkinje cell synapses. However, it has not been examined thus far how acute ethanol application affects...

  7. Nanodefects of membranes cause destruction of packed red blood cells during long-term storage

    Energy Technology Data Exchange (ETDEWEB)

    Kozlova, Elena, E-mail: waterlake@mail.ru [V.A. Negovsky Scientific Research Institute of General Reanimatology, Moscow (Russian Federation); I.M. Sechenov First Moscow State Medical University, Moscow (Russian Federation); Chernysh, Aleksandr [V.A. Negovsky Scientific Research Institute of General Reanimatology, Moscow (Russian Federation); I.M. Sechenov First Moscow State Medical University, Moscow (Russian Federation); Moroz, Victor; Sergunova, Victoria; Gudkova, Olga; Kuzovlev, Artem [V.A. Negovsky Scientific Research Institute of General Reanimatology, Moscow (Russian Federation)

    2015-10-01

    Packed red blood cells (PRBC) are used for blood transfusion. PRBC were stored for 30 days under 4 °C in hermetic blood bags with CPD anticoagulant-preservative solution. Hematocrit was 50–55%. The distortions of PRBC membranes nanostructure and cells morphology during storage were studied by atomic force microscopy. Basic measurements were performed at the day 2, 6, 9, 16, 23 and 30 of storage and additionally 2–3 days after it. Topological defects occurred on RBC membranes by day 9. They appeared as domains with grain-like structures (“grains”) sized up to 200 nm. These domains were appeared in almost all cells. Later these domains merged and formed large defects on cells. It was the formation of domains with the “grains” which was onset process leading eventually to destruction of PRBC. Possible mechanisms of transformation of PRBC and their membrane are related to the alterations of spectrin cytoskeleton. During this storage period potassium ions and lactat concentrations increased, pH decreased, intracellular concentration of reduced glutathione diminished in the preservative solution. Changes of PRBC morphology were detected within the entire period of PRBC storage. Discocytes predominated at the days 1 and 2. By day 30 PRBC transformed into irreversible echinocytes and spheroechinocytes. Study of defects of membranes nanostructure may form the basis of assessing the quality of the stored PRBC. This method may allow to work out the best recommendations for blood transfusion. - Highlights: • Domains with “grains” are formed on membranes surface on 9–16 days of PRBC storage. • The development of domains is the reason of irreversible changes of PRBC structure. • The origin of domains is the consequence of alterations of spectrin cytoskeleton. • Study of nanostructure may form basis of assessing the quality of the stored PRBC.

  8. Genotype instability during long-term subculture of lymphoblastoid cell lines.

    Science.gov (United States)

    Oh, Ji Hee; Kim, Young Jin; Moon, Sanghoon; Nam, Hye-Young; Jeon, Jae-Pil; Lee, Jong Ho; Lee, Jong-Young; Cho, Yoon Shin

    2013-01-01

    Epstein-Barr virus (EBV)-transformed lymphoblastoid cell lines (LCLs) promise to address the challenge posed by the limited availability of primary cells needed as a source of genomic DNA for genetic studies. However, the genetic stability of LCLs following prolonged culture has never been rigorously investigated. To evaluate genotypic errors caused by EBV integration into human chromosomes, we isolated genomic DNA from human peripheral blood mononuclear cells and LCLs collected from 20 individuals and genotyped the DNA samples using the Affymetrix 500K SNP array set. Genotype concordance measurements between two sources of DNA from the same individual indicated that genotypic discordance is negligible in early-passage LCLs (50 passages). Analysis of concordance on a chromosome-by-chromosome basis identified genomic regions with a high frequency of genotypic errors resulting from the loss of heterozygosity observed in late-passage LCLs. Our findings suggest that, although LCLs harvested during early stages of propagation are a reliable source of genomic DNA for genetic studies, investigations that involve genotyping of the entire genome should not use DNA from late-passage LCLs.

  9. LONG-TERM PERFORMANCE OF SOLID OXIDE STACKS WITH ELECTRODE-SUPPORTED CELLS OPERATING IN THE STEAM ELECTROLYSIS MODE

    Energy Technology Data Exchange (ETDEWEB)

    J. E. O' Brien; R. C. O' Brien; X. Zhang; G. Tao; B. J. Butler

    2011-11-01

    Performance characterization and durability testing have been completed on two five-cell high-temperature electrolysis stacks constructed with advanced cell and stack technologies. The solid oxide cells incorporate a negative-electrode-supported multi-layer design with nickel-zirconia cermet negative electrodes, thin-film yttria-stabilized zirconia electrolytes, and multi-layer lanthanum ferrite-based positive electrodes. The per-cell active area is 100 cm2. The stack is internally manifolded with compliant mica-glass seals. Treated metallic interconnects with integral flow channels separate the cells. Stack compression is accomplished by means of a custom spring-loaded test fixture. Initial stack performance characterization was determined through a series of DC potential sweeps in both fuel cell and electrolysis modes of operation. Results of these sweeps indicated very good initial performance, with area-specific resistance values less than 0.5 ?.cm2. Long-term durability testing was performed with A test duration of 1000 hours. Overall performance degradation was less than 10% over the 1000-hour period. Final stack performance characterization was again determined by a series of DC potential sweeps at the same flow conditions as the initial sweeps in both electrolysis and fuel cell modes of operation. A final sweep in the fuel cell mode indicated a power density of 0.356 W/cm2, with average per-cell voltage of 0.71 V at a current of 50 A.

  10. Long-term effects of autologous bone marrow stem cell treatment in acute myocardial infarction: factors that may influence outcomes.

    Directory of Open Access Journals (Sweden)

    David M Clifford

    Full Text Available AIMS: To investigate whether there are important sources of heterogeneity between the findings of different clinical trials which administer autologous stem cell treatment for acute myocardial infarction (AMI and to evaluate what factors may influence the long-term effects of this treatment. METHODS AND RESULTS: MEDLINE (1950-January 2011, EMBASE (1974-January 2011, CENTRAL (The Cochrane Library 2011, Issue 1, CINAHL (1982-January 2011, and ongoing trials registers were searched for randomised trials of bone marrow stem cells as treatment for AMI. Hand-searching was used to screen recent, relevant conference proceedings (2005-2010/11. Meta-analyses were conducted using random-effects models and heterogeneity between subgroups was assessed using chi-squared tests. Planned analyses included length of follow-up, timing of cell infusion and dose, patient selection, small trial size effect, methodological quality, loss of follow-up and date of publication. Thirty-three trials with a total of 1,765 participants were included. There was no evidence of bias due to publication or time-lag, methodological quality of included studies, participant drop-out, duration of follow-up or date of the first disclosure of results. However, in long-term follow-ups the treatment seemed more effective when administered at doses greater than 10(8 cells and to patients with more severe heart dysfunction. CONCLUSIONS: Evaluation of heterogeneity between trials has not identified significant sources of bias in this study. However, clinical differences between trials are likely to exist which should be considered when undertaking future trials.

  11. Factors predicting long-term survival in low-risk diffuse large B-cell lymphoma

    DEFF Research Database (Denmark)

    Møller, Michael B; Pedersen, Niels T; Christensen, Bjarne E

    2003-01-01

    The International Prognostic Index (IPI) is widely used for risk stratification of patients with diffuse large B-cell lymphoma (DLBCL). However, even among patients with low-risk disease, according to the IPI a substantial proportion of patients ultimately succumb to their disease. Using mature...... at 5 years and 10 years was 85% and 75%, respectively. Stage II was associated with poor response to treatment (P=0.044). In a multivariate analysis, Stage II (P=0.001) and age >50 years (P=0.043) were independently associated with poor outcome. Patients without these adverse factors had an excellent...

  12. Cesium Enhances Long-Term Stability of Lead Bromide Perovskite-Based Solar Cells

    OpenAIRE

    Kulbak, Michael; Gupta, Satyajit; Kedem, Nir; Levine, Igal; Bendikov, Tatyana; Hodes, Gary; Cahen, David

    2015-01-01

    Direct comparison between perovskite-structured hybrid organic-inorganic - methyl ammonium lead bromide (MAPbBr3) and all-inorganic cesium lead bromide (CsPbBr3), allows identifying possible fundamental differences in their structural, thermal and electronic characteristics. Both materials possess a similar direct optical band-gap, but CsPbBr3 demonstrates a higher thermal stability than MAPbBr3. In order to compare device properties we fabricated solar cells, with similarly synthesized MAPbB...

  13. Long-term kinetics of T cell production in HIV-infected subjects treated with highly active antiretroviral therapy

    Science.gov (United States)

    Fleury, S.; Rizzardi, G. P.; Chapuis, A.; Tambussi, G.; Knabenhans, C.; Simeoni, E.; Meuwly, J.-Y.; Corpataux, J.-M.; Lazzarin, A.; Miedema, F.; Pantaleo, G.

    2000-01-01

    The long-term kinetics of T cell production following highly active antiretroviral therapy (HAART) were investigated in blood and lymph node in a group of HIV-infected subjects at early stage of established infection and prospectively studied for 72 wk. Before HAART, CD4 and CD8 T cell turnover was increased. However, the total number of proliferating CD4+ T lymphocytes, i.e., CD4+Ki67+ T lymphocytes, was not significantly different in HIV-infected (n = 73) and HIV-negative (n = 15) subjects, whereas proliferating CD8+Ki67+ T lymphocytes were significantly higher in HIV-infected subjects. After HAART, the total body number of proliferating CD4+Ki67+ T lymphocytes increased over time and was associated with an increase of both naive and memory CD4+ T cells. The maximal increase (2-fold) was observed at week 36, whereas at week 72 the number of proliferating CD4+ T cells dropped to baseline levels, i.e., before HAART. The kinetics of the fraction of proliferating CD4 and CD8 T cells were significantly correlated with the changes in the total body number of these T cell subsets. These results demonstrate a direct relationship between ex vivo measures of T cell production and quantitative changes in total body T lymphocyte populations. This study provides advances in the delineation of the kinetics of T cell production in HIV infection in the presence and/or in the absence of HAART. PMID:10805798

  14. Beta-cell function in isolated human pancreatic islets in long-term tissue culture

    DEFF Research Database (Denmark)

    Nielsen, Jens Høiriis

    1981-01-01

    Human pancreatic islets were isolated by collagenase treatment of pancreatic tissue obtained from 27 individuals aged 12 to 69 years. The islets were maintained free floating in tissue culture medium RPMI 1640 supplemented with calf or human serum. In two cases the insulin production was followed...... adult human pancreatic tissue and that their beta-cell function can be maintained for up to two years. The variation in insulin production rate could not be ascribed to age or sex and may reflect both physiological and methodological factors....... up to nearly two years. The insulin production rate of the individual islet preparations varied between 0.2 and 8 ng per islet per day. No significant correlation with donor age or sex was found. The glucose concentration in the medium influenced the insulin release in a dose dependent manner. The...

  15. Long-term survival in uterine clear cell carcinoma and uterine papillary serous carcinoma.

    Science.gov (United States)

    Lindahl, Bengt; Persson, Jan; Ranstam, Jonas; Willén, Roger

    2010-09-01

    Uterine clear cell carcinoma (UCC) and uterine papillary serous carcinoma (UPSC) are rare entities that differ in clinical behavior from endometrial adenocarcinoma. Compared with endometrioid adenocarcinoma, they more often metastasize early and more commonly in the upper abdomen including the omentum. Treatment programs of UCC and UPSC at different stages vary and range from no adjuvant therapy in stage Ia to a wide variety of chemotherapies and radiotherapies in more advanced stages. This study presents the outcome of 109 patients with UCC or UPSC treated according to essentially the same treatment program from May 1993 to December 2004. Most patients were treated with a simple hysterectomy with no further adjuvant treatment. In stage Ia, 2/46 patients died of their disease and amongst all the stages, 30/109 patients died of their disease. These survival outcomes are comparable to or better than those presented previously. PMID:20944161

  16. Predicting the ageing and the long-term durability of organic polymer solar cells

    Science.gov (United States)

    Gardette, Jean-Luc; Rivaton, Agnès; Thérias, Sandrine; Chambon, Sylvain; Manceau, Matthieu; Gaume, Julien

    2010-06-01

    Organic solar cells based on conductive polymers exhibit a unique combination of properties which include low cost, flexibility and large surface processability. Organic photovoltaic could then prevail for some applications alongside silicon, such as nomad or indoor. To achieve this objective, the sustainability of the initial properties in conditions of use of the cell is required, since it could be a lock to the emergence of these devices in the market. The polymers used in solar cells are indeed known to exhibit low resistance to environmental constraints, in particular to the combined action of sunlight, oxygen and water. We present recent results on both the accelerated artificial and the natural outdoors ageing of MDMO-PPV (Poly[2-methoxy-5-(3',7'-dimethyloctyloxy)-1,4-Phenylenevinylene) and P3HT/PCBM blends poly(3-hexylthiophene) (P3HT) (methano-fullerene[6,6]-phenyl C61-butyric acid methyl ester) ([60] PCBM). The influence of various parameters such as the temperature and the presence of oxygen were studied. The modifications of the chemical structure of both the components of the blend were monitored by spectroscopic analysis (infrared, UV-visible), the morphology of the blends was analysed by AFM and XRD and the photovoltaic performances all along the exposure were recorded. Two important results have been pointed out: on one hand, the Achilles heel of the chemical structure of MDMO-PPV and P3HT under the impact of light has been evidenced. On the other hand, it has been shown that P3HT:PCBM blends are much more stable than MDMO:PCBM blends whatever the conditions of ageing are. Results show that a convenient encapsulation can ensure a promising lifetime of P3HT/PCBM blends in real conditions of use. This work also focuses on this last point and proposes to study and try to understand the behavior of the materials used in the active layer when submitted to photoaging and thermal aging in the absence of oxygen. To fulfil very good encapsulation, glass

  17. Construction of cell type-specific logic models of signaling networks using CellNOpt.

    Science.gov (United States)

    Morris, Melody K; Melas, Ioannis; Saez-Rodriguez, Julio

    2013-01-01

    Mathematical models are useful tools for understanding protein signaling networks because they provide an integrated view of pharmacological and toxicological processes at the molecular level. Here we describe an approach previously introduced based on logic modeling to generate cell-specific, mechanistic and predictive models of signal transduction. Models are derived from a network encoding prior knowledge that is trained to signaling data, and can be either binary (based on Boolean logic) or quantitative (using a recently developed formalism, constrained fuzzy logic). The approach is implemented in the freely available tool CellNetOptimizer (CellNOpt). We explain the process CellNOpt uses to train a prior knowledge network to data and illustrate its application with a toy example as well as a realistic case describing signaling networks in the HepG2 liver cancer cell line.

  18. Facial stimulation induces long-term depression at cerebellar molecular layer interneuron–Purkinje cell synapses in vivo in mice

    Directory of Open Access Journals (Sweden)

    De-Lai eQiu

    2015-06-01

    Full Text Available Cerebellar long-term synaptic plasticity has been proposed to provide a cellular mechanism for motor learning. Numerous studies have demonstrated the induction and mechanisms of synaptic plasticity at parallel fiber–Purkinje cell (PF–PC, parallel fiber–molecular layer interneurons (PF–MLI and mossy fiber–granule cell (MF–GC synapses, but no study has investigated sensory stimulation-evoked synaptic plasticity at MLI–PC synapses in the cerebellar cortex of living animals. We studied the expression and mechanism of MLI–PC GABAergic synaptic plasticity induced by a train of facial stimulation in urethane-anesthetized mice by cell-attached recordings and pharmacological methods. We found that 1 Hz, but not a 2 Hz or 4 Hz, facial stimulation induced a long-term depression (LTD of GABAergic transmission at MLI–PC synapses, which was accompanied with a decrease in the stimulation-evoked pause of spike firing in PCs, but did not induce a significant change in the properties of the sensory-evoked spike events of MLIs. The MLI–PC GABAergic LTD could be prevented by blocking cannabinoid type 1 (CB1 receptors, and could be pharmacologically induced by a CB1 receptor agonist. Additionally, 1 Hz facial stimulation delivered in the presence of a metabotropic glutamate receptor 1 (mGluR1 antagonist, JNJ16259685, still induced the MLI–PC GABAergic LTD, whereas blocking N-methyl-D-aspartate (NMDA receptors during 1 Hz facial stimulation abolished the expression of MLI–PC GABAergic LTD. These results indicate that sensory stimulation can induce an endocannabinoid (eCB-dependent LTD of GABAergic transmission at MLI–PC synapses via activation of NMDA receptors in cerebellar cortical Crus II in vivo in mice. Our results suggest that the sensory stimulation-evoked MLI–PC GABAergic synaptic plasticity may play a critical role in motor learning in animals.

  19. Analysis of the Long-term Effect of Intraoperative Radiotherapy (IORT) for Non-Small Cell Lung Carcinoma (NSCLC)

    Institute of Scientific and Technical Information of China (English)

    Guxia Zhou; Tiwen Zeng; Lianyuan Wang; Lin Ma

    2007-01-01

    OBJECTIVE To analyze the long-term effects of treatment with an operation+postoperative irradiation (A group) and an operation+intraoperative radiotherapy+postoperative irradiation (B group) in non-small cell lung cancer patients.METHODS Through a prospective randomized clinical trial,a total of 154 patients with non-small cell lung carcinoma were divided into two groups of 77 cases.Among the 154 cases,there were 134 squamous carcinomas,17 adenocarcinomas and 3 adeno-squamous carcinomas.TNM staging:there were 17 in Stage Ⅰ.76 in Stage Ⅱ and 61 in Stage Ⅲ.Adosage of 15~25 Gy IORT,energy 9~16 MeV electrons,was delivered to the fumors.The doses given were 40~60 Gy postoperation.RESULTS The local control rates in A and B groups were 49.4% and 62.3% respectively (P<0.05).The survivals at 3,5 and 7 years for group A were 40.3%,27.3%,and 5.2% and for group B 44.2%,28.6% and 6.5% (P>0.05).There were 16 deaths from radiotherapy complications,with 2 cases in group A and 14 in group B.CONCLUSION IORT+postoperative irradiation can enhance the local control rate of non-small cell lung cancer patients and reduce the recurrentrates.but it can not improve long-term survival.KEYWORDS:lung neoplasms/surgery,lung neoplasms/radiotherapy,radiotherapy intraoperative,prognosis.

  20. Stable Marking and Transgene Expression Without Progression to Monoclonality in Canine Long-Term Hematopoietic Repopulating Cells Transduced with Lentiviral Vectors

    OpenAIRE

    Enssle, Joerg; Trobridge, Grant D.; Keyser, Kirsten A.; Ironside, Christina; Beard, Brian C; Kiem, Hans-Peter

    2010-01-01

    Lentiviral gene transfer vectors have a number of potential advantages over gammaretroviral vectors including more efficient transduction of nondividing cells, a more favorable integration site profile, and the ability to accommodate large transgenes. Here, we present long-term follow-up data of animals that received lentivirus-transduced CD34-enriched cells. Six long-term surviving dogs were available for analysis. Transgene expression was analyzed from at least 12 months to more than 5 year...

  1. Long Term Study of Protective Mechanisms of Human Adipose Derived Mesenchymal Stem Cells on Cisplatin Induced Kidney injury in Sprague-Dawley Rats

    OpenAIRE

    Elhusseini, Fatma M; Saad, Mohamed-Ahdy A.A; Anber, Nahla; Elghannam, Doaa; Sobh, Mohamed-Ahmed; ALSAYED, AZIZA; El-dusoky, Sara; SHEASHAA, HUSSEIN; Abdel-Ghaffar, Hassan; Sobh, Mohamed

    2016-01-01

    Background/Aims: Long-term evaluation of cisplatin induced nephrotoxicity and the probable renal protective activities of stem cells are lacking up until now. We evaluated the early and long-term role of human adipose derived mesenchymal stem cells (ADMSCs) in prevention or amelioration of cisplatin induced acute kidney injury (AKI) in Sprague-Dawley rats. For this, we determined the kidney tissue level of oxidative stress markers in conjugation with a renal histopathological scoring system o...

  2. Impact of radiotherapy on pain relief and recalcification in plasma cell neoplasms. Long-term experience

    Energy Technology Data Exchange (ETDEWEB)

    Balducci, Mario; Chiesa, Silvia; Manfrida, Stefania [Univ. Hospital ' Gemelli' , Rome (IT). Radiotherapy Dept.] (and others)

    2011-02-15

    Purpose: To evaluate the impact of radiotherapy on pain relief and on recalcification in patients with osteolytic lesions due to plasma cell neoplasm. Patients and Methods: Pain relief was evaluated according to a 0-10 verbal numerical rating scale (NRS) and recalcification was measured using radiological imaging. Results: From 1996-2007, 52 patients were treated. Median total dose was 38 Gy (range, 16-50 Gy). Pain before radiotherapy was reported by 45 of 52 (86.5%) patients as being severe (8 {<=} NRS {<=} 10) in 5 (11%), moderate (5 {<=} NRS {<=} 7) in 27 (60%), and mild in 13 (29%). Pain relief was achieved in 41 of 45 patients (91%): complete relief was obtained in 21 (51.2%) and partial relief in 20 patients (48.8%); patients with severe pain experienced resolution and none presented an increase of pain. Drugs reduction/suspension was achieved in 7 of the 21 patients with complete response. Of 42 patients evaluable for recalcification, 21 (50%) presented a radiological response, which was identified as complete in 16 (38%). Conclusion: Our data confirm the effectiveness of radiotherapy for pain relief, including a reduction in drug intake, and on recalcification, thus, supporting its use in a multidisciplinary approach. (orig.)

  3. A two-compartment microfluidic device for long-term live cell detection based on surface plasmon resonance.

    Science.gov (United States)

    Deng, Shijie; Yu, Xinglong; Liu, Ran; Chen, Weixing; Wang, Peng

    2016-07-01

    A two-compartment microfluidic device integrated with a surface plasmon resonance (SPR) interferometric imaging system has been developed for long-term and real-time cell detection. The device uses a porous membrane sandwiched between two chambers to obtain an exact medium exchange rate and minimal fluid shear stress for cell culture. The two-compartment device was optimized by COMSOL simulations and fabricated using Poly (dimethylsiloxane) elastomer replica molding methods. To confirm the capability of the microfluidic device to maintain the cell physiological environment over long intervals, HeLa cells were cultured in the device for up to 48 h. The cell proliferation process was monitored by both SPR and microscopic time-lapse imaging. The SPR response showed four phases with different growth rates, and agreed well with the time-lapse imaging. Furthermore, real-time detection of cell behaviors under different doses of Paclitaxel and Cisplatin was performed. The SPR responses revealed dose-dependent inhibitions of cell proliferation, with distinct drug action kinetics. PMID:27570574

  4. Cell-Type Specific Inactivation of Hippocampal CA1 Disrupts Location-Dependent Object Recognition in the Mouse

    Science.gov (United States)

    Haettig, Jakob; Sun, Yanjun; Wood, Marcelo A.; Xu, Xiangmin

    2013-01-01

    The allatostatin receptor (AlstR)/ligand inactivation system enables potent regulation of neuronal circuit activity. To examine how different cell types participate in memory formation, we have used this system through Cre-directed, cell-type specific expression in mouse hippocampal CA1 in vivo and examined functional effects of inactivation of…

  5. Characterization of long-term extension of isolated cell walls from growing cucumber hypocotyls

    Science.gov (United States)

    Cosgrove, D. J.

    1989-01-01

    Walls from frozen-thawed cucumber (Cucumis sativus L.) hypocotyls extend for many hours when placed in tension under acidic conditions. This study examined whether such "creep" is a purely physical process dependent on wall viscoelasticity alone or whether enzymatic activities are needed to maintain wall extension. Chemical denaturants inhibited wall creep, some acting reversibly and others irreversibly. Brief (15 s) boiling in water irreversibly inhibited creep, as did pre-incubation with proteases. Creep exhibited a high Q10 (3.8) between 20 degrees and 30 degrees C, with slow inactivation at higher temperatures, whereas the viscous flow of pectin solutions exhibited a much lower Q10 (1.35). On the basis of its temperature sensitivity, involvement of pectic gel-sol transitions was judged to be of little importance in creep. Pre-incubation of walls in neutral pH irreversibly inactivated their ability to creep, with a half-time of about 40 min. At 1 mM, Cu2+, Hg2+ and Al3+ were strongly inhibitory whereas most other cations, including Ca2+, had little effect. Sulfhydryl-reducing agents strongly stimulated creep, apparently by stabilizing wall enzyme(s). The physical effects of these treatments on polymer interactions were examined by Instron and stress-relaxation analyses. Some treatments, such as pH and Cu2+, had significant effects on wall viscoelasticity, but others had little or no apparent effect, thus implicating an enzymatic creep mechanism. The results indicate that creep depends on relatively rugged enzymes that are firmly attached to or entangled in the wall. The sensitivity of creep to SH-reducing agents indicates that thiol reduction of wall enzymes might provide a control mechanism for endogenous cell growth.

  6. Long-term change in pulmonary function after definitive radiotherapy for non-small cell lung cancer

    DEFF Research Database (Denmark)

    Schytte, T.; Bentzen, S. M.; Brink, C.;

    2015-01-01

    Purpose/Objective: Radiotherapy (RT) for non-small cell lung cancer (NSCLC) may cause late toxicities, such as heart toxicity, changes in pulmonary function (PF) and lung fibrosis, but late toxicity data are scarce in the literature for this category of patients. The objective of this study...... was to analyze long term PF changes after definitive RT for patients with NSCLC. PF was measured by FEV1 and FVC as assessed by spirometry. Materials and Methods: This is a single institution study of patients receiving definitive RT for NSCLC between 1996 and 2010. A total of 556 consecutive treated patients...... finding was that high V60 was associated with less decline in FVC, but FVC may be a less reliable parameter for pulmonary damage. (Figure Presented) Conclusions: Patients experience a decline in FEV1 after the 12-month value following definitive RT for NSCLC. In a multivariable analysis, deterioration...

  7. Long-term cathode performance and the microbial communities that develop in microbial fuel cells fed different fermentation endproducts

    KAUST Repository

    Kiely, Patrick D.

    2011-01-01

    To better understand how cathode performance and substrates affected communities that evolved in these reactors over long periods of time, microbial fuel cells were operated for more than 1year with individual endproducts of lignocellulose fermentation (acetic acid, formic acid, lactic acid, succinic acid, or ethanol). Large variations in reactor performance were primarily due to the specific substrates, with power densities ranging from 835±21 to 62±1mW/m3. Cathodes performance degraded over time, as shown by an increase in power of up to 26% when the cathode biofilm was removed, and 118% using new cathodes. Communities that developed on the anodes included exoelectrogenic families, such as Rhodobacteraceae, Geobacteraceae, and Peptococcaceae, with the Deltaproteobacteria dominating most reactors. Pelobacter propionicus was the predominant member in reactors fed acetic acid, and it was abundant in several other MFCs. These results provide valuable insights into the effects of long-term MFC operation on reactor performance. © 2010 Elsevier Ltd.

  8. Prion protein is expressed on long-term repopulating hematopoietic stem cells and is important for their self-renewal.

    Science.gov (United States)

    Zhang, Cheng Cheng; Steele, Andrew D; Lindquist, Susan; Lodish, Harvey F

    2006-02-14

    Although the wild-type prion protein (PrP) is abundant and widely expressed in various types of tissues and cells, its physiological function(s) remain unknown, and PrP knockout mice do not exhibit overt and undisputed phenotypes. Here we showed that PrP is expressed on the surface of several bone marrow cell populations successively enriched in long-term (LT) hematopoietic stem cells (HSCs) using flow cytometry analysis. Affinity purification of the PrP-positive and -negative fractions from these populations, followed by competitive bone marrow reconstitution assays, shows that all LT HSCs express PrP. HSCs from PrP-null bone marrow exhibited impaired self-renewal in serial transplantation of lethally irradiated mouse recipients both in the presence and absence of competitors. When treated with a cell cycle-specific myelotoxic agent, the animals reconstituted with PrP-null HSCs exhibit increased sensitivity to hematopoietic cell depletion. Ectopic expression of PrP in PrP-null bone marrow cells by retroviral infection rescued the defective hematopoietic engraftment during serial transplantation. Therefore, PrP is a marker for HSCs and supports their self-renewal.

  9. Lepidopteran cell lines after long-term culture in alternative media: comparison of growth rates and baculovirus replication.

    Science.gov (United States)

    Lynn, Dwight E

    2006-01-01

    Three insect cell lines, IPLB-LdFB and IPLB-LdEIta from gypsy moth fat body and embryos and UFL-AG-286 from velvetbean caterpillar embryos, have been concurrently maintained for 1 to 12 yr on two media formulations, modified TC-100 containing 9% fetal bovine serum and Ex-cell 400, a commercial serum-free medium (SFM). Cells grown in each medium were tested for susceptibility to and productivity of various multiply embedded nucleopolyhedroviruses. The three lines chosen for these experiments fall into three categories of relative growth in SFM versus TC-100: LdFB cells grew similarly in each medium, LdEIta grew better in Ex-Cell than in TC-100, and AG-286 grew better in TC-100 than in Ex-Cell. The susceptibility of cells to infection also varies, although without any apparent correlation to which medium was best for supporting growth. Endpoint assays suggested that LdFB cells grown in serum-containing medium are more susceptible to virus infection than their SFM counterparts, while the opposite is true for LdEIta cells. Production of virus, based on numbers of occlusion bodies, showed fewer differences with only AcMNPV production with AG-286 in TC-100 being statistically higher than production of the same virus in Ex-cell 400. These studies suggest that long-term passage in alternative media may impact the ability of cells to support virus infection and replication, but the effects on each cell line and virus system need to be determined. PMID:16848634

  10. Maternal deprivation has sexually dimorphic long-term effects on hypothalamic cell-turnover, body weight and circulating hormone levels.

    Science.gov (United States)

    Viveros, María-Paz; Llorente, Ricardo; Díaz, Francisca; Romero-Zerbo, Silvana Y; Bermudez-Silva, Francisco J; Rodríguez de Fonseca, Fernando; Argente, Jesús; Chowen, Julie A

    2010-11-01

    Maternal deprivation (MD) has numerous outcomes, including modulation of neuroendocrine functions. We previously reported that circulating leptin levels are reduced and hypothalamic cell-turnover is affected during MD, with some of these effects being sexually dimorphic. As leptin modulates the development of hypothalamic circuits involved in metabolic control, we asked whether MD has long-term consequences on body weight, leptin levels and the expression of neuropeptides involved in metabolism. Rats were separated from their mother for 24h starting on postnatal day (PND) 9 and sacrificed at PNDs 13, 35 and 75. In both sexes MD reduced body weight, but only until puberty, while leptin levels were unchanged at PND 35 and significantly reduced at PND 75. Adiponectin levels were also reduced at PND 75 in females, while testosterone levels were reduced in males. At PND 13, MD modulated cell-turnover markers in the hypothalamus of males, but not females and increased nestin, a marker of immature neurons, in both sexes, with males having higher levels than females and a significantly greater rise in response to MD. There was no effect of MD on hypothalamic mRNA levels of the leptin receptor or metabolic neuropeptides or the mRNA levels of leptin and adiponectin in adipose tissue. Thus, MD has long-term effects on the levels of circulating hormones that are not correlated with changes in body weight. Furthermore, these endocrine outcomes are different between males and females, which could be due to the fact that MD may have sexually dimorphic effects on hypothalamic development.

  11. Regeneration-associated WNT Signaling Is Activated in Long-term Reconstituting AC133bright Acute Myeloid Leukemia Cells

    Directory of Open Access Journals (Sweden)

    Alessandro Beghini

    2012-12-01

    Full Text Available Acute myeloid leukemia (AML is a genetically heterogeneous clonal disorder characterized by two molecularly distinct self-renewing leukemic stem cell (LSC populations most closely related to normal progenitors and organized as a hierarchy. A requirement for WNT/β-catenin signaling in the pathogenesis of AML has recently been suggested by a mouse model. However, its relationship to a specific molecular function promoting retention of self-renewing leukemia-initiating cells (LICs in human remains elusive. To identify transcriptional programs involved in the maintenance of a self-renewing state in LICs, we performed the expression profiling in normal (n = 10 and leukemic (n = 33 human long-term reconstituting AC133+ cells, which represent an expanded cell population in most AML patients. This study reveals the ligand-dependent WNT pathway activation in AC133bright AML cells and shows a diffuse expression and release of WNT10B, a hematopoietic stem cell regenerative-associated molecule. The establishment of a primary AC133+ AML cell culture (A46 demonstrated that leukemia cells synthesize and secrete WNT ligands, increasing the levels of dephosphorylated β-catenin in vivo. We tested the LSC functional activity in AC133+ cells and found significant levels of engraftment upon transplantation of A46 cells into irradiated Rag2-/-γc-/- mice. Owing to the link between hematopoietic regeneration and developmental signaling, we transplanted A46 cells into developing zebrafish. This system revealed the formation of ectopic structures by activating dorsal organizer markers that act downstream of the WNT pathway. In conclusion, our findings suggest that AC133bright LSCs are promoted by misappropriating homeostatic WNT programs that control hematopoietic regeneration.

  12. Human tooth germ stem cells preserve neuro-protective effects after long-term cryo-preservation.

    Science.gov (United States)

    Yalvaç, Mehmet E; Ramazanoglu, Mustafa; Tekguc, Murat; Bayrak, Omer F; Shafigullina, Aygul K; Salafutdinov, Ilnur I; Blatt, Natalia L; Kiyasov, Andrey P; Sahin, Fikrettin; Palotás, András; Rizvanov, Albert A

    2010-02-01

    The use of mesenchymal stem cells (MSCs) has been shown to be promising in chronic disorders such as diabetes, Alzheimer's dementia, Parkinson's disease, spinal cord injury and brain ischemia. Recent studies revealed that human tooth germs (hTG) contain MSCs which can be easily isolated, expanded and cryo-preserved. In this report, we isolated human tooth germ stem cells (hTGSCs) with MSC characteristics from third molar tooth germs, cryo-preserved them at -80( degrees )C for 6 months, and evaluated for their surface antigens, expression of pluri-potency associated genes, differentiation capacity, karyotype, and proliferation rate. These characteristics were compared to their non-frozen counterparts. In addition, neuro-protective effects of cryo-preserved cells on neuro-blastoma SH-SY5Y cells were also assessed after exposure to stress conditions induced by hydrogen-peroxide (oxidative stress) and paclitaxel (microtubule stabilizing mitotic inhibitor). After long term cryo-preservation hTGSCs expressed surface antigens CD29, CD73, CD90, CD105, and CD166, but not CD34, CD45 or CD133, which was typical for non-frozen hTGSCs. Cryo-preserved hTGSCs were able to differentiate into osteo-, adipo- and neuro-genic cells. They also showed normal karyotype after high number of population doublings and unchanged proliferation rate. On the other hand, cryo-preserved cells demonstrated a tendency for lower level of pluri-potency associated gene expression (nanog, oct4, sox2, klf4, c-myc) than non-frozen hTGSCs. hTGSCs conditioned media increased survival of SH-SY5Y cells exposed to oxidative stress or paclitaxel. These findings confirm that hTGSCs preserve their major characteristics and exert neuro-protection after long-term cryo-preservation, suggesting that hTGSCs, harvested from young individuals and stored for possible use later as they grow old, might be employed in cellular therapy of age-related degenerative disorders. PMID:20158462

  13. Pairing of pre- and postsynaptic activities in cerebellar Purkinje cells induces long-term changes in synaptic efficacy in vitro.

    Science.gov (United States)

    Crepel, F; Jaillard, D

    1991-01-01

    1. An in vitro slice preparation of rat cerebellar cortex was used to analyse long-lasting modifications of synaptic transmission at parallel fibre (PF)-Purkinje cell (PC) synapses. These use-dependent changes were induced by pairing PF-mediated EPSPs evoked at low frequency (1 Hz) with different levels of membrane polarization (or bioelectrical activities) of PCs for 15 min. 2. Experiments were performed on forty-eight PCs recorded intracellularly in a conventional perfused chamber, and in fifty other cells maintained in a static chamber either in the presence (n = 21) or in the absence (n = 29) of 400 nM-phorbol 12,13-dibutyrate (PDBu). 3. In these three experimental conditions, PF-mediated EPSPs were always measured on PCs maintained at a holding potential of -75 mV, and further hyperpolarized by constant hyperpolarizing pulses. This allowed us both to test the input resistance of PCs and to avoid their firing during PF-mediated EPSPs. 4. In all cells retained for the present study, latencies of PF-mediated EPSPs evoked at 0.2 Hz were stable during the pre-pairing period, and the same was true for their amplitude and time course. 5. In the perfused chamber, pairing of PF-mediated EPSPs with the same hyperpolarization of PCs as that used for measurements of synaptic responses had no effect on these EPSPs in 30% of PCs. It induced long-term depression (LTD) and long-term potentiation (LTP) in 23 and 47% of the tested cells respectively (n = 17). 6. In the perfused chamber, pairing of PF-mediated EPSPs with moderate depolarization of PCs (n = 19) giving rise to a sustained firing of sodium spikes significantly favoured the appearance of LTP as compared to the previous pairing protocol. However, there were still 27 and 15% of cells which showed no modification and LTD respectively. 7. In contrast, pairing of PF-mediated EPSPs with calcium (Ca2+) spikes evoked by strong depolarization of PCs (n = 12) led to LTD of synaptic transmission in nearly half of the tested

  14. IGF-1-mediated osteoblastic niche expansion enhances long-term hematopoietic stem cell engraftment after murine bone marrow transplantation.

    Science.gov (United States)

    Caselli, Anna; Olson, Timothy S; Otsuru, Satoru; Chen, Xiaohua; Hofmann, Ted J; Nah, Hyun-Duck; Grisendi, Giulia; Paolucci, Paolo; Dominici, Massimo; Horwitz, Edwin M

    2013-10-01

    The efficiency of hematopoietic stem cell (HSC) engraftment after bone marrow (BM) transplantation depends largely on the capacity of the marrow microenvironment to accept the transplanted cells. While radioablation of BM damages osteoblastic stem cell niches, little is known about their restoration and mechanisms governing their receptivity to engraft transplanted HSCs. We previously reported rapid restoration and profound expansion of the marrow endosteal microenvironment in response to marrow radioablation. Here, we show that this reorganization represents proliferation of mature endosteal osteoblasts which seem to arise from a small subset of high-proliferative, relatively radio-resistant endosteal cells. Multiple layers of osteoblasts form along the endosteal surface within 48 hours after total body irradiation, concomitant with a peak in marrow cytokine expression. This niche reorganization fosters homing of the transplanted hematopoietic cells to the host marrow space and engraftment of long-term-HSC. Inhibition of insulin-like growth factor (IGF)-1-receptor tyrosine kinase signaling abrogates endosteal osteoblast proliferation and donor HSC engraftment, suggesting that the cytokine IGF-1 is a crucial mediator of endosteal niche reorganization and consequently donor HSC engraftment. Further understanding of this novel mechanism of IGF-1-dependent osteoblastic niche expansion and HSC engraftment may yield clinical applications for improving engraftment efficiency after clinical HSC transplantation.

  15. Isotropic Versus Bipolar Functionalized Biomimetic Artificial Basement Membranes and Their Evaluation in Long-Term Human Cell Co-Culture.

    Science.gov (United States)

    Rossi, Angela; Wistlich, Laura; Heffels, Karl-Heinz; Walles, Heike; Groll, Jürgen

    2016-08-01

    In addition to dividing tissues into compartments, basement membranes are crucial as cell substrates and to regulate cellular behavior. The development of artificial basement membranes is indispensable for the ultimate formation of functional engineered tissues; however, pose a challenge due to their complex structure. Herein, biodegradable electrospun polyester meshes are presented, exhibiting isotropic or bipolar bioactivation as a biomimetic and biofunctional model of the natural basement membrane. In a one-step preparation process, reactive star-shaped prepolymer additives, which generate a hydrophilic fiber surface, are electrospun with cell-adhesion-mediating peptides, derived from major components of the basement membrane. Human skin cells adhere to the functionalized meshes, and long-term co-culture experiments confirm that the artificial basement membranes recapitulate and preserve tissue specific functions. Several layers of immortalized human keratinocytes grow on the membranes, differentiating toward the surface and expressing typical epithelial markers. Fibroblasts migrate into the reticular lamina mimicking part of the mesh. Both cells types begin to produce extracellular matrix proteins and to remodel the initial membrane. It is shown at the example of skin that the artificial basement membrane design provokes biomimetic responses of different cell types and can thus be used as basis for the future development of basement membrane containing tissues. PMID:27283510

  16. Short and long-term safety of lenograstim administration in healthy peripheral haematopoietic progenitor cell donors: a single centre experience.

    Science.gov (United States)

    Martino, M; Console, G; Dattola, A; Callea, I; Messina, G; Moscato, T; Massara, E; Irrera, G; Fedele, R; Gervasi, A; Bresolin, G; Iacopino, P

    2009-08-01

    Healthy donors (HDs) who were mobilized using lenograstim (LENO) and who were undergoing peripheral haematopoietic progenitor cell collection with apheresis (HPC-A) were enrolled in a surveillance protocol. In all, 184 HDs have been assessed with a median follow-up of 62 months (range 2-155). HDs received LENO at a median dose of 10 microg/kg (range 5-15). Bone pain was reported as the most frequent short-term adverse event (71.2%). Other commonly observed short-term symptoms included fatigue (19.0%), fever (5.4%), headache (27.7%), nausea (12.0%) and insomnia (22.3%). Spleen size increased in 4.3% of the donors. No vascular disorders or cardiac disease occurred. Long-term follow-up included monitoring of adverse events, neoplastic disease or other pathologies. Transit ischaemic attack occurred in one donor (39 months post-donation). One autoimmune event was reported at 28 months post-recombinant human granulocyte (rhG)-CSF (ankylosing spondylitis); one donor with a history of chronic obstructive pulmonary disease developed secondary polyglobulia (50 months post-rhG-CSF). One donor was diagnosed with lung cancer at 19 months post-donation. No haematological disease was observed. In conclusion, the short-term safety appears to be verified, whereas, although the study identified no increased risks of malignancy among HDs who received rhG-CSF, long-term safety requires more complete data sets, especially a longer follow-up and a larger number of HDs. PMID:19182833

  17. Modification of fluorous substrates with oligo(ethylene glycol) via "click" chemistry for long-term resistance of cell adhesion.

    Science.gov (United States)

    Contreras-Caceres, Rafael; Santos, Catherine M; Li, Siheng; Kumar, Amit; Zhu, Zhiling; Kolar, Satya S; Casado-Rodriguez, Miguel A; Huang, Yongkai; McDermott, Alison; Lopez-Romero, Juan Manuel; Cai, Chengzhi

    2015-11-15

    In this work perfluorinated substrates fabricated from SiO2 glass slides are modified with oligo(ethylene glycol) (OEG) units for long-term resistance of cell adhesion purposes, based on fluorous interactions and click chemistry. Specifically, fluorous substrates, prepared by treatment of glass slides with 1H, 1H, 2H, 2H-perfluorodecyltrimethoxysilane (FAS17), were coated with ethynyl-OEG-C8F17, followed by covalent attachment of an azido-OEG via copper-catalyzed azide-alkyne cycloaddition (CuAAC) "click" reaction. We demonstrate that the resultant surface avoid fibrinogen adsorption and resisted cell adhesion for over 14days. X-ray photoemission spectroscopy (XPS) analysis and contact angle goniometry measurements confirm the presence of the OEG molecules on the fluorous substrates. Bright field optical images show total absence of 3T3 fibroblast cells on the OEG modified fluorinated substrate for 1 and 5days, and a remarkably decrease of cell adhesion at 14days. PMID:26210101

  18. Improvement of the long term stability in the high temperature solid oxide fuel cell using functional layers

    Energy Technology Data Exchange (ETDEWEB)

    Brueckner, B.; Guenther, C.; Ruckdaeschel, R. [Siemens AG, Erlangen (Germany)] [and others

    1996-12-31

    In the planar Siemens design of the solid oxide fuel cell a metallic interconnector is used to seperate the ceramic single cells. A disadvantage of the metallic bipolar plate which consists of a chromium alloy is the formation of high volatile Cr-oxides and hydroxides at the surface at the cathode side. The reaction products evaporate and are reduced at the cathode/electrolyte interface to form new crystalline phases. This process gives rise to long term cell degradation. Protective coatings might be successful in preventing the chromium oxide evaporation. The required properties of the protective layers are (I) high electrical conductivity, (II) similar coefficients of thermal expansion to the bipolar plate (III), chemical compatibility to the bipolar plate and cathode material, (IV) a low diffusion coefficient of Cr and (V) chemical stability up to 1223K under oxygen atmosphere. Furthermore, during operation at 1223K an electrical contact between the metallic plate and the electrodes has to be maintained. This problem could be solved using ceramic layer between the metallic plate and the single cells.

  19. Variations in gene and protein expression in canine chondrodystrophic nucleus pulposus cells following long-term three-dimensional culture.

    Directory of Open Access Journals (Sweden)

    Munetaka Iwata

    Full Text Available Intervertebral disc (IVD degeneration greatly affects quality of life. The nucleus pulposus (NP of chondrodystrophic dog breeds (CDBs is similar to the human NP, because the cells disappear with age and are replaced by fibrochondrocyte-like cells. However, because IVD develops as early as within the first year of life, we used canines as a model to investigate in vitro the mechanisms underlying IVD degeneration. Specifically, we evaluated the potential of a three-dimensional (3D culture of healthy NP as an in vitro model system to investigate the mechanisms of IVD degeneration. Agarose hydrogels were populated with healthy NP cells from beagles after performing magnetic resonance imaging, and mRNA expression profiles and pericellular extracellular matrix (ECM protein distribution were determined. After 25 days of 3D culture, there was a tendency for redifferentiation into the native NP phenotype, and mRNA levels of Col2A1, COMP, and CK18 were not significantly different from those of freshly isolated cells. Our findings suggest that long-term 3D culture promoted chondrodystrophic NP redifferentiation through reconstruction of the pericellular microenvironment. Further, lipopolysaccharide (LPS induced expression of TNF-α, MMP3, MMP13, VEGF, and PGES mRNA in the 3D cultures, creating a molecular milieu that mimics that of degenerated NP. These results suggest that this in vitro model represents a reliable and cost-effective tool for evaluating new therapies for disc degeneration.

  20. Enhancement of cancer stem-like and epithelial−mesenchymal transdifferentiation property in oral epithelial cells with long-term nicotine exposure: Reversal by targeting SNAIL

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Cheng-Chia [Institute of Oral Science, Chung Shan Medical University, Taichung, Taiwan (China); School of Dentistry, Chung Shan Medical University, Taichung, Taiwan (China); Department of Dentistry, Chung Shan Medical University Hospital, Taichung, Taiwan (China); Chang, Yu-Chao, E-mail: cyc@csmu.edu.tw [School of Dentistry, Chung Shan Medical University, Taichung, Taiwan (China); Department of Dentistry, Chung Shan Medical University Hospital, Taichung, Taiwan (China)

    2013-02-01

    Cigarette smoking is one of the major risk factors in the development and further progression of tumorigenesis, including oral squamous cell carcinoma (OSCC). Recent studies suggest that interplay cancer stem-like cells (CSCs) and epithelial−mesenchymal transdifferentiation (EMT) properties are responsible for the tumor maintenance and metastasis in OSCC. The aim of the present study was to investigate the effects of long-term exposure with nicotine, a major component in cigarette, on CSCs and EMT characteristics. The possible reversal regulators were further explored in nicotine-induced CSCs and EMT properties in human oral epithelial (OE) cells. Long-term exposure with nicotine was demonstrated to up-regulate ALDH1 population in normal gingival and primary OSCC OE cells dose-dependently. Moreover, long-term nicotine treatment was found to enhance the self-renewal sphere-forming ability and stemness gene signatures expression and EMT regulators in OE cells. The migration/cell invasiveness/anchorage independent growth and in vivo tumor growth by nude mice xenotransplantation assay was enhanced in long-term nicotine-stimulated OE cells. Knockdown of Snail in long-term nicotine-treated OE cells was found to reduce their CSCs properties. Therapeutic delivery of Si-Snail significantly blocked the xenograft tumorigenesis of long-term nicotine-treated OSCC cells and largely significantly improved the recipient survival. The present study demonstrated that the enrichment of CSCs coupled EMT property in oral epithelial cells induced by nicotine is critical for the development of OSCC tumorigenesis. Targeting Snail might offer a new strategy for the treatment of OSCC patients with smoking habit. -- Highlights: ► Sustained nicotine treatment induced CSCs properties of oral epithelial cells. ► Long-term nicotine treatment enhance EMT properties of oral epithelial cells. ► Long-term nicotine exposure increased tumorigenicity of oral epithelial cells. ► Si

  1. Enhancement of cancer stem-like and epithelial−mesenchymal transdifferentiation property in oral epithelial cells with long-term nicotine exposure: Reversal by targeting SNAIL

    International Nuclear Information System (INIS)

    Cigarette smoking is one of the major risk factors in the development and further progression of tumorigenesis, including oral squamous cell carcinoma (OSCC). Recent studies suggest that interplay cancer stem-like cells (CSCs) and epithelial−mesenchymal transdifferentiation (EMT) properties are responsible for the tumor maintenance and metastasis in OSCC. The aim of the present study was to investigate the effects of long-term exposure with nicotine, a major component in cigarette, on CSCs and EMT characteristics. The possible reversal regulators were further explored in nicotine-induced CSCs and EMT properties in human oral epithelial (OE) cells. Long-term exposure with nicotine was demonstrated to up-regulate ALDH1 population in normal gingival and primary OSCC OE cells dose-dependently. Moreover, long-term nicotine treatment was found to enhance the self-renewal sphere-forming ability and stemness gene signatures expression and EMT regulators in OE cells. The migration/cell invasiveness/anchorage independent growth and in vivo tumor growth by nude mice xenotransplantation assay was enhanced in long-term nicotine-stimulated OE cells. Knockdown of Snail in long-term nicotine-treated OE cells was found to reduce their CSCs properties. Therapeutic delivery of Si-Snail significantly blocked the xenograft tumorigenesis of long-term nicotine-treated OSCC cells and largely significantly improved the recipient survival. The present study demonstrated that the enrichment of CSCs coupled EMT property in oral epithelial cells induced by nicotine is critical for the development of OSCC tumorigenesis. Targeting Snail might offer a new strategy for the treatment of OSCC patients with smoking habit. -- Highlights: ► Sustained nicotine treatment induced CSCs properties of oral epithelial cells. ► Long-term nicotine treatment enhance EMT properties of oral epithelial cells. ► Long-term nicotine exposure increased tumorigenicity of oral epithelial cells. ► Si

  2. Expansion and long-term culture of human spermatogonial stem cells via the activation of SMAD3 and AKT pathways

    Science.gov (United States)

    Guo, Ying; Liu, Linhong; Sun, Min; Hai, Yanan; Li, Zheng

    2015-01-01

    Spermatogonial stem cells (SSCs) can differentiate into spermatids, reflecting that they could be used in reproductive medicine for treating male infertility. SSCs are able to become embryonic stem-like cells with the potentials of differentiating into numerous cell types of the three germ layers and they can transdifferentiate to mature and functional cells of other lineages, highlighting significant applications of human SSCs for treating human diseases. However, human SSCs are very rare and a long-term culture system of human SSCs has not yet established. This aim of study was to isolate, identify and culture human SSCs for a long period. We isolated GPR125-positive spermatogonia with high purity and viability from adult human testicular tissues utilizing the two-step enzymatic digestion and magnetic-activated cell sorting with antibody against GPR125. These freshly isolated cells expressed a number of markers for SSCs, including GPR125, PLZF, GFRA1, RET, THY1, UCHL1 and MAGEA4, but not the hallmarks for spermatocytes and spermatozoa, e.g. SYCP1, SYCP3, PRM1, and TNP1. The isolated human SSCs could be cultured for two months with a significant increase of cell number with the defined medium containing growth factors and hydrogel. Notably, the expression of numerous SSC markers was maintained during the cultivation of human SSCs. Furthermore, SMAD3 and AKT phosphorylation was enhanced during the culture of human SSCs. Collectively, these results suggest that human SSCs can be cultivated for a long period and expanded whilst retaining an undifferentiated status via the activation of SMAD3 and AKT pathways. This study could provide sufficient cells of SSCs for their basic research and clinic applications in reproductive and regenerative medicine. PMID:26088866

  3. Construction of a Defined Biomimetic Matrix for Long-Term Maintenance of Mouse Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Adnan, Nihad; Mie, Masayasu; Haque, Amranul; Hossain, Sharif; Mashimo, Yasumasa; Akaike, Toshihiro; Kobatake, Eiry

    2016-07-20

    The existing in vitro culture systems often use undefined and animal-derived components for the culture of pluripotent stem cells. Artificial bioengineered peptides have the potential to become alternatives to these components of extracellular matrix (ECM). Integrins and cadherins are two cell adhesion proteins important for stem cell self-renewal, differentiation, and phenotype stability. In the present study, we sought to mimic the physico-biochemical properties of natural ECMs that allow self-renewal of mouse induced pluripotent stem cells (iPSCs). We develop a genetically engineered ECM protein (ERE-CBP) that contains (i) an integrin binding peptide sequence (RGD/R), (ii) an E-/N-cadherin binding peptide sequence (SWELYYPLRANL/CBP), and (iii) 12 repeats of APGVGV elastin-like polypeptides (ELPs/E).While ELPs allow efficient coating by binding to nontreated hydrophobic tissue culture plates, RGD/R and CBP support integrin- and cadherin-dependent cell attachment, respectively. Mouse iPSCs on this composite matrix exhibit a more compact phenotype compared to cells on control gelatin substrate. We also demonstrated that the ERE-CBP supports proliferation and long-term self-renewal of mouse iPSCs for up to 17 passages without GSK3β (CHIR99021) and Erk (PD0325901) inhibitors. Overall, our engineered ECM protein, which is cost-effective to produce in prokaryotic origin and flexible to modify with other cell adhesion peptides or growth factors, provides a novel approach for expansion of mouse iPSCs in vitro. PMID:27269811

  4. Calcium Dynamics of Ex Vivo Long-Term Cultured CD8+ T Cells Are Regulated by Changes in Redox Metabolism

    Science.gov (United States)

    Gran, Margaret A.; Potnis, Anish; Hill, Abby; Lu, Hang

    2016-01-01

    T cells reach a state of replicative senescence characterized by a decreased ability to proliferate and respond to foreign antigens. Calcium release associated with TCR engagement is widely used as a surrogate measure of T cell response. Using an ex vivo culture model that partially replicates features of organismal aging, we observe that while the amplitude of Ca2+ signaling does not change with time in culture, older T cells exhibit faster Ca2+ rise and a faster decay. Gene expression analysis of Ca2+ channels and pumps expressed in T cells by RT-qPCR identified overexpression of the plasma membrane CRAC channel subunit ORAI1 and PMCA in older T cells. To test whether overexpression of the plasma membrane Ca2+ channel is sufficient to explain the kinetic information, we adapted a previously published computational model by Maurya and Subramaniam to include additional details on the store-operated calcium entry (SOCE) process to recapitulate Ca2+ dynamics after T cell receptor stimulation. Simulations demonstrated that upregulation of ORAI1 and PMCA channels is not sufficient to explain the observed alterations in Ca2+ signaling. Instead, modeling analysis identified kinetic parameters associated with the IP3R and STIM1 channels as potential causes for alterations in Ca2+ dynamics associated with the long term ex vivo culturing protocol. Due to these proteins having known cysteine residues susceptible to oxidation, we subsequently investigated and observed transcriptional remodeling of metabolic enzymes, a shift to more oxidized redox couples, and post-translational thiol oxidation of STIM1. The model-directed findings from this study highlight changes in the cellular redox environment that may ultimately lead to altered T cell calcium dynamics during immunosenescence or organismal aging. PMID:27526200

  5. Sniff-Like Patterned Input Results in Long-Term Plasticity at the Rat Olfactory Bulb Mitral and Tufted Cell to Granule Cell Synapse

    Science.gov (United States)

    Perez de los Cobos Pallares, Fernando; Loebel, Alex; Lukas, Michael

    2016-01-01

    During odor sensing the activity of principal neurons of the mammalian olfactory bulb, the mitral and tufted cells (MTCs), occurs in repetitive bursts that are synchronized to respiration, reminiscent of hippocampal theta-gamma coupling. Axonless granule cells (GCs) mediate self- and lateral inhibitory interactions between the excitatory MTCs via reciprocal dendrodendritic synapses. We have explored long-term plasticity at this synapse by using a theta burst stimulation (TBS) protocol and variations thereof. GCs were excited via glomerular stimulation in acute brain slices. We find that TBS induces exclusively long-term depression in the majority of experiments, whereas single bursts (“single-sniff paradigm”) can elicit both long-term potentiation and depression. Statistical analysis predicts that the mechanism underlying this bidirectional plasticity involves the proportional addition or removal of presynaptic release sites. Gamma stimulation with the same number of APs as in TBS was less efficient in inducing plasticity. Both TBS- and “single-sniff paradigm”-induced plasticity depend on NMDA receptor activation. Since the onset of plasticity is very rapid and requires little extra activity, we propose that these forms of plasticity might play a role already during an ongoing search for odor sources. Our results imply that components of both short-term and long-term olfactory memory may be encoded at this synapse. PMID:27747107

  6. Sniff-Like Patterned Input Results in Long-Term Plasticity at the Rat Olfactory Bulb Mitral and Tufted Cell to Granule Cell Synapse

    Directory of Open Access Journals (Sweden)

    Mahua Chatterjee

    2016-01-01

    Full Text Available During odor sensing the activity of principal neurons of the mammalian olfactory bulb, the mitral and tufted cells (MTCs, occurs in repetitive bursts that are synchronized to respiration, reminiscent of hippocampal theta-gamma coupling. Axonless granule cells (GCs mediate self- and lateral inhibitory interactions between the excitatory MTCs via reciprocal dendrodendritic synapses. We have explored long-term plasticity at this synapse by using a theta burst stimulation (TBS protocol and variations thereof. GCs were excited via glomerular stimulation in acute brain slices. We find that TBS induces exclusively long-term depression in the majority of experiments, whereas single bursts (“single-sniff paradigm” can elicit both long-term potentiation and depression. Statistical analysis predicts that the mechanism underlying this bidirectional plasticity involves the proportional addition or removal of presynaptic release sites. Gamma stimulation with the same number of APs as in TBS was less efficient in inducing plasticity. Both TBS- and “single-sniff paradigm”-induced plasticity depend on NMDA receptor activation. Since the onset of plasticity is very rapid and requires little extra activity, we propose that these forms of plasticity might play a role already during an ongoing search for odor sources. Our results imply that components of both short-term and long-term olfactory memory may be encoded at this synapse.

  7. Establishment of a long-term three-dimensional primary culture of mouse glandular stomach epithelial cells within the stem cell niche

    International Nuclear Information System (INIS)

    Highlights: ► We established a 3D culture system to allow long-term culture of stomach cells. ► In this culture system, gastric epithelial cells grew for about 3 months. ► The cultured cells differentiated into multi-units of the stomach. ► This culture method should be useful for elucidating the cause of gastric diseases. -- Abstract: Compared to the small intestine and colon, little is known about stem cells in the stomach because of a lack of specific stem cell markers and an in vitro system that allows long-term culture. Here we describe a long-term three-dimensional (3D) primary gastric culture system within the stem cell niche. Glandular stomach cells from neonatal mice cultured in collagen gel yielded expanding sphere-like structures for 3 months. The wall of the gastrospheres consisted of a highly polarized epithelial monolayer with an outer lining of myofibroblasts. The epithelial cells showed a tall columnar cell shape, basal round nuclei, and mucus-filled cytoplasm as well as expression of MUC5AC, indicating differentiation into gastric surface mucous cells. These cells demonstrated the features of fully differentiated gastric surface mucous cells such as microvilli, junctional complexes, and glycogen and secretory granules. Fewer than 1% of cultured epithelial cells differentiated into enteroendocrine cells. Active proliferation of the epithelial cells and many apoptotic cells in the inner lumen revealed the rapid cell turnover in gastrospheres in vitro. This method enables us to investigate the role of signaling between cell–cell and epithelial–mesenchymal interactions in an environment that is extremely similar to the in vivo environment

  8. Establishment of a long-term three-dimensional primary culture of mouse glandular stomach epithelial cells within the stem cell niche

    Energy Technology Data Exchange (ETDEWEB)

    Katano, Takahito [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Ootani, Akifumi [Department of Gastroenterology and GI Endoscopy Center, Shin-Kokura Hospital, Federation of National Public Service Personnel Mutual Aid Associations, 1-3-1 Kanada, Kokurakita-ku, Kitakyushu 803-0816 (Japan); Department of Internal Medicine, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga 849-8501 (Japan); Mizoshita, Tsutomu, E-mail: tmizoshi@med.nagoya-cu.ac.jp [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Tanida, Satoshi; Tsukamoto, Hironobu; Ozeki, Keiji; Ebi, Masahide; Mori, Yoshinori; Kataoka, Hiromi; Kamiya, Takeshi [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Toda, Shuji [Department of Pathology and Microbiology, Faculty of Medicine, Saga University, 5-1-1 Nabeshima, Saga 849-8501 (Japan); Joh, Takashi [Department of Gastroenterology and Metabolism, Nagoya City University Graduate School of Medical Sciences, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan)

    2013-03-22

    Highlights: ► We established a 3D culture system to allow long-term culture of stomach cells. ► In this culture system, gastric epithelial cells grew for about 3 months. ► The cultured cells differentiated into multi-units of the stomach. ► This culture method should be useful for elucidating the cause of gastric diseases. -- Abstract: Compared to the small intestine and colon, little is known about stem cells in the stomach because of a lack of specific stem cell markers and an in vitro system that allows long-term culture. Here we describe a long-term three-dimensional (3D) primary gastric culture system within the stem cell niche. Glandular stomach cells from neonatal mice cultured in collagen gel yielded expanding sphere-like structures for 3 months. The wall of the gastrospheres consisted of a highly polarized epithelial monolayer with an outer lining of myofibroblasts. The epithelial cells showed a tall columnar cell shape, basal round nuclei, and mucus-filled cytoplasm as well as expression of MUC5AC, indicating differentiation into gastric surface mucous cells. These cells demonstrated the features of fully differentiated gastric surface mucous cells such as microvilli, junctional complexes, and glycogen and secretory granules. Fewer than 1% of cultured epithelial cells differentiated into enteroendocrine cells. Active proliferation of the epithelial cells and many apoptotic cells in the inner lumen revealed the rapid cell turnover in gastrospheres in vitro. This method enables us to investigate the role of signaling between cell–cell and epithelial–mesenchymal interactions in an environment that is extremely similar to the in vivo environment.

  9. Effects of inhaled anesthetic isoflurane on long-term potentiation of CA3 pyramidal cell afferents in vivo

    Directory of Open Access Journals (Sweden)

    Ballesteros KA

    2012-11-01

    Full Text Available Kristen A Ballesteros,1 Angela Sikorski,2 James E Orfila,3 Joe L Martinez Jr41Department of Biology, The University of Texas at San Antonio, San Antonio, TX, USA; 2Texas A&M University Texarkana, Texarkana, TX, USA; 3University of Colorado in Denver, Denver, CO, USA; 4University of Illinois in Chicago, Chicago, IL, USAAbstract: Isoflurane is a preferred anesthetic, due to its properties that allow a precise concentration to be delivered continually during in vivo experimentation. The major mechanism of action of isoflurane is modulation of the γ-amino butyric acid (GABAA receptor-chloride channel, mediating inhibitory synaptic transmission. Animal studies have shown that isoflurane does not cause cell death, but it does inhibit cell growth and causes long-term hippocampal learning deficits. As there are no studies characterizing the effects of isoflurane on electrophysiological aspects of long-term potentiation (LTP in the hippocampus, it is important to determine whether isoflurane alters the characteristic responses of hippocampal afferents to cornu ammonis region 3 (CA3. We investigated the effects of isoflurane on adult male rats during in vivo induction of LTP, using the mossy fiber pathway, the lateral perforant pathway, the medial perforant pathway, and the commissural CA3 (cCA3 to CA3, with intracranial administration of Ringer’s solution, naloxone, RS-aminoindan-1, 5-dicarboxylic acid (AIDA, or 3-[(R-2-carboxypiperazin-4-yl]-propo-2-enyl-1-phosphonic acid (CPP. Then, we compared these responses to published electrophysiological data, using sodium pentobarbital as an anesthetic, under similar experimental conditions. Our results showed that LTP was exhibited in animals anesthetized with isoflurane under vehicle conditions. With the exception of AIDA in the lateral perforant pathway, the defining characteristics of the four pathways appeared to remain intact, except for the observation that LTP was markedly reduced in animals

  10. A long-term analysis of Pt counter electrodes for Dye-sensitized Solar Cells exploiting a microfluidic housing system

    International Nuclear Information System (INIS)

    The study of the degradation process occurring in Dye-sensitized Solar Cells (DSCs) is still a hot topic, in view of the final industrialization and application of this class of devices. Currently the long-term analysis of DSCs is carried out on the entire devices, while the monitoring of cell components cannot be performed in situ directly on the materials, but only through indirect methods. In this paper we report on the analysis of two different kinds of Pt counter electrodes through direct measurements performed under real operating conditions, thanks to the use of a home-made microfluidic housing system, which allows the opening and the investigation of the cell components. The counter electrode samples were studied through X-Ray Photoelectron Spectroscopy, Field Emission Scanning Electron Microscopy, Energy Dispersive X-ray Spectroscopy, UV–visible Spectroscopy and Electrochemical Impedance Spectroscopy for a period longer than 1 year. The results showed that the performances of both classes of Pt counter electrodes remained stable for all the investigation period, despite some slight variation of the morphology. DSCs fabricated employing aged counter electrodes exhibited the same photovoltaic performance behavior of reference cells using fresh-produced counter electrodes, thus demonstrating that both class of materials do not undergo degradation during normal operating conditions. - Highlights: • The analysis of Pt counter electrodes for Dye-sensitized Solar Cells was carried out. • Two families of counter electrodes were studied for a period longer than 1 year. • The analyzed samples were investigated in real operating condition. • A small detachment of the Pt clusters in the thermal samples was observed. • The charge transfer properties remained unchanged for all the investigation period

  11. Loading of red blood cells with an analyte-sensitive dye for development of a long-term monitoring technique

    Science.gov (United States)

    Ritter, Sarah C.; Meissner, Kenith E.

    2012-03-01

    Measurement of blood analytes, such as pH and glucose, provide crucial information about a patient's health. Some such analytes, such as glucose in the case of diabetes, require long-term or near-continuous monitoring for proper disease management. However, current monitoring techniques are far from ideal: multiple-per-day finger stick tests are inconvenient and painful for the patient; implantable sensors have short functional life spans (i.e., 3-7 days). Red blood cells serve as an attractive alternative for carriers of analyte sensors. Once reintroduced to the blood stream, these carriers may continue to live for the remainder of their life span (120 days for humans). They are also biodegradable and biocompatible, thereby eliminating the immune system response common for many implanted devices. The proposed carrier system takes advantage of the ability of the red blood cells to swell in response to a decrease in the osmolarity of the extracellular solution. Just before the membranes lyse, they develop small pores on the scale of tens of nanometers. Analyte-sensitive dyes in the extracellular solution may then diffuse into the perforated red blood cells and become entrapped upon restoration of physiological temperature and osmolarity. Because the membranes contain various analyte transporters, intracellular analyte levels rapidly equilibrate to those of the extracellular solution. A fluorescent dye has been loaded inside of red blood cells using a preswelling technique. Alterations in preparation parameters have been shown to affect characteristics of the resulting dye-loaded red blood cells (e.g., intensity of fluorescence).

  12. A long-term analysis of Pt counter electrodes for Dye-sensitized Solar Cells exploiting a microfluidic housing system

    Energy Technology Data Exchange (ETDEWEB)

    Sacco, Adriano, E-mail: adriano.sacco@iit.it [Center for Space Human Robotics @Polito, Istituto Italiano di Tecnologia, Corso Trento 21, 10129 Torino (Italy); Pugliese, Diego; Lamberti, Andrea [Applied Science and Technology Department, Politecnico di Torino, Corso Duca degli Abruzzi 24, 10129 Torino (Italy); Castellino, Micaela; Chiodoni, Angelica [Center for Space Human Robotics @Polito, Istituto Italiano di Tecnologia, Corso Trento 21, 10129 Torino (Italy); Virga, Alessandro [Applied Science and Technology Department, Politecnico di Torino, Corso Duca degli Abruzzi 24, 10129 Torino (Italy); Bianco, Stefano [Center for Space Human Robotics @Polito, Istituto Italiano di Tecnologia, Corso Trento 21, 10129 Torino (Italy); Applied Science and Technology Department, Politecnico di Torino, Corso Duca degli Abruzzi 24, 10129 Torino (Italy)

    2015-07-01

    The study of the degradation process occurring in Dye-sensitized Solar Cells (DSCs) is still a hot topic, in view of the final industrialization and application of this class of devices. Currently the long-term analysis of DSCs is carried out on the entire devices, while the monitoring of cell components cannot be performed in situ directly on the materials, but only through indirect methods. In this paper we report on the analysis of two different kinds of Pt counter electrodes through direct measurements performed under real operating conditions, thanks to the use of a home-made microfluidic housing system, which allows the opening and the investigation of the cell components. The counter electrode samples were studied through X-Ray Photoelectron Spectroscopy, Field Emission Scanning Electron Microscopy, Energy Dispersive X-ray Spectroscopy, UV–visible Spectroscopy and Electrochemical Impedance Spectroscopy for a period longer than 1 year. The results showed that the performances of both classes of Pt counter electrodes remained stable for all the investigation period, despite some slight variation of the morphology. DSCs fabricated employing aged counter electrodes exhibited the same photovoltaic performance behavior of reference cells using fresh-produced counter electrodes, thus demonstrating that both class of materials do not undergo degradation during normal operating conditions. - Highlights: • The analysis of Pt counter electrodes for Dye-sensitized Solar Cells was carried out. • Two families of counter electrodes were studied for a period longer than 1 year. • The analyzed samples were investigated in real operating condition. • A small detachment of the Pt clusters in the thermal samples was observed. • The charge transfer properties remained unchanged for all the investigation period.

  13. Different Immunological Phenotypes Associated with Preserved CD4+ T Cell Counts in HIV-Infected Controllers and Viremic Long Term Non-Progressors

    DEFF Research Database (Denmark)

    Gaardbo, Julie Christine; Hartling, Hans J; Ronit, Andreas;

    2013-01-01

    HIV-infected controllers control viral replication and maintain normal CD4+ T cell counts. Long Term Non-Progressors (LTNP) also maintain normal CD4+ T cell counts, but have on-going viral replication. We hypothesized that different immunological mechanisms are responsible for preserved CD4+ T cell...... counts in controllers and LTNP....

  14. Improved salinity tolerance of rice through cell type-specific expression of AtHKT1;1

    OpenAIRE

    Darren Plett; Gehan Safwat; Matthew Gilliham; Inge Skrumsager Møller; Stuart Roy; Neil Shirley; Andrew Jacobs; Alexander Johnson; Mark Tester

    2010-01-01

    Previously, cell type-specific expression of AtHKT1;1, a sodium transporter, improved sodium (Na(+)) exclusion and salinity tolerance in Arabidopsis. In the current work, AtHKT1;1, was expressed specifically in the root cortical and epidermal cells of an Arabidopsis GAL4-GFP enhancer trap line. These transgenic plants were found to have significantly improved Na(+) exclusion under conditions of salinity stress. The feasibility of a similar biotechnological approach in crop plants was explored...

  15. Long-term collections

    CERN Multimedia

    Collectes à long terme

    2007-01-01

    The Committee of the Long Term Collections (CLT) asks for your attention for the following message from a young Peruvian scientist, following the earthquake which devastated part of her country a month ago.

  16. Long-term potentiation promotes proliferation/survival and neuronal differentiation of neural stem/progenitor cells.

    Directory of Open Access Journals (Sweden)

    Taesup Cho

    Full Text Available Neural stem cell (NSC replacement therapy is considered a promising cell replacement therapy for various neurodegenerative diseases. However, the low rate of NSC survival and neurogenesis currently limits its clinical potential. Here, we examined if hippocampal long-term potentiation (LTP, one of the most well characterized forms of synaptic plasticity, promotes neurogenesis by facilitating proliferation/survival and neuronal differentiation of NSCs. We found that the induction of hippocampal LTP significantly facilitates proliferation/survival and neuronal differentiation of both endogenous neural progenitor cells (NPCs and exogenously transplanted NSCs in the hippocampus in rats. These effects were eliminated by preventing LTP induction by pharmacological blockade of the N-methyl-D-aspartate glutamate receptor (NMDAR via systemic application of the receptor antagonist, 3-[(R-2-carboxypiperazin-4-yl]-propyl-1-phosphonic acid (CPP. Moreover, using a NPC-neuron co-culture system, we were able to demonstrate that the LTP-promoted NPC neurogenesis is at least in part mediated by a LTP-increased neuronal release of brain-derived neurotrophic factor (BDNF and its consequent activation of tropomysosin receptor kinase B (TrkB receptors on NSCs. Our results indicate that LTP promotes the neurogenesis of both endogenous and exogenously transplanted NSCs in the brain. The study suggests that pre-conditioning of the host brain receiving area with a LTP-inducing deep brain stimulation protocol prior to NSC transplantation may increase the likelihood of success of using NSC transplantation as an effective cell therapy for various neurodegenerative diseases.

  17. Invariant TAD Boundaries Constrain Cell-Type-Specific Looping Interactions between Promoters and Distal Elements around the CFTR Locus.

    Science.gov (United States)

    Smith, Emily M; Lajoie, Bryan R; Jain, Gaurav; Dekker, Job

    2016-01-01

    Three-dimensional genome structure plays an important role in gene regulation. Globally, chromosomes are organized into active and inactive compartments while, at the gene level, looping interactions connect promoters to regulatory elements. Topologically associating domains (TADs), typically several hundred kilobases in size, form an intermediate level of organization. Major questions include how TADs are formed and how they are related to looping interactions between genes and regulatory elements. Here we performed a focused 5C analysis of a 2.8 Mb chromosome 7 region surrounding CFTR in a panel of cell types. We find that the same TAD boundaries are present in all cell types, indicating that TADs represent a universal chromosome architecture. Furthermore, we find that these TAD boundaries are present irrespective of the expression and looping of genes located between them. In contrast, looping interactions between promoters and regulatory elements are cell-type specific and occur mostly within TADs. This is exemplified by the CFTR promoter that in different cell types interacts with distinct sets of distal cell-type-specific regulatory elements that are all located within the same TAD. Finally, we find that long-range associations between loci located in different TADs are also detected, but these display much lower interaction frequencies than looping interactions within TADs. Interestingly, interactions between TADs are also highly cell-type-specific and often involve loci clustered around TAD boundaries. These data point to key roles of invariant TAD boundaries in constraining as well as mediating cell-type-specific long-range interactions and gene regulation. PMID:26748519

  18. Loss of growth inhibitory effects of retinoic acid in human breast cancer cells following long-term exposure to retinoic acid

    OpenAIRE

    Stephen, R; Darbre, P D

    2000-01-01

    Although retinoids are known to be inhibitory to breast cancer cell growth, a key remaining question is whether they would remain effective if administered long-term. We describe here the long-term effects of all-trans retinoic acid on two oestrogen-dependent human breast cancer cell lines MCF7 and ZR-75-1. Although both cell lines were growth inhibited by retinoic acid in the short-term in either the absence or the presence of oestradiol, prolonged culture with 1 μM all-trans retinoic acid r...

  19. Urea as a long-term stable alternative to guanidium thiocyanate additive in dye-sensitized solar cell

    Science.gov (United States)

    Kim, Mi-Jeong; Park, Nam-Gyu

    2012-09-01

    Photovoltaic performance of 0.05 M urea-contained redox electrolyte is compared to that of 0.05 M guanidinium thiocyanate (GSCN)-contained one in dye-sensitized solar cell. No significant difference in the initial photovoltaic performance is observed, which means that the role of urea additive is similar to that of GSCN. Initial solar-to-electrical conversion efficiency of the device containing GSCN shows 7% that is diminished to 5.8% after 40 days, whereas the device containing urea exhibits stable photovoltaic performance showing that initial efficiency of 7.2% is almost remained unchanged after 40 days (7.1%). The lowered efficiency of the GSCN-contained device is mainly due to the decreased photocurrent density, which is ascribed to the formation of needle-shaped crystals on TiO2 layer. Infrared spectroscopic study confirms that the crystals are dye analogue, which is indicative of dye desorption in the presence of GSCN. On the other hand, no crystals are formed in the urea-contained electrolyte, which implies that dye desorption is negligible. Urea additive is thus found to be less reactive in dye desorption than GSCN, leading to long-term stability.

  20. Long-term Disease Free and Successful Pregnancy in a Woman with Gonadal Dysgenesis and Malignant Germ Cell Tumor

    Directory of Open Access Journals (Sweden)

    Azamsadat Mousavi

    2012-06-01

    Full Text Available Objective: To report a case of long-term disease free and successful pregnancy after fertility sparing staging surgery with adjuvant chemotherapy in a 46,Xy gonadal dysgenetic with malignant germ cell tumor.Materials and methods: A case report from a university hospital about a 19-year-old female with 46,XY karyotype ( Swyer syndrome. The patient underwent bilateral gonadectomy and staging with uterus preservation. Six course adjuvant chemotherapy with VBP (Vinblastin, Bleomycin, Cisplatin was given. The case got pregnant through IVF- embryo donation. Disease free period and successful pregnancy is reported.Results: After treatment the patient is free of the disease after 11 years follow-up. She underwent in vitro fertilization treatment with oocyte donation and gave birth to a healthy ch.Conclusion: Improved multimodality treatment, allowance for consideration of fertility options for some women with gynecologic cancers. Since major concern in women with XY gonadal dysgenesis is ovarian malignancy, even with stage II dysgerminoma hysterectomy may not be required in some cases considering the opportunity for childbearing with the use of embryo transfer.

  1. Autologous hematopoietic cell transplantation following high-dose immunosuppressive therapy for advanced multiple sclerosis: long-term results.

    Science.gov (United States)

    Bowen, J D; Kraft, G H; Wundes, A; Guan, Q; Maravilla, K R; Gooley, T A; McSweeney, P A; Pavletic, S Z; Openshaw, H; Storb, R; Wener, M; McLaughlin, B A; Henstorf, G R; Nash, R A

    2012-07-01

    The purpose of the study was to determine the long-term safety and effectiveness of high-dose immunosuppressive therapy (HDIT) followed by autologous hematopoietic cell transplantation (AHCT) in advanced multiple sclerosis (MS). TBI, CY and antithymocyte globulin were followed by transplantation of autologous, CD34-selected PBSCs. Neurological examinations, brain magnetic resonance imaging and cerebrospinal fluid (CSF) for oligoclonal bands (OCB) were serially evaluated. Patients (n=26, mean Expanded Disability Status Scale (EDSS)=7.0, 17 secondary progressive, 8 primary progressive, 1 relapsing/remitting) were followed for a median of 48 months after HDIT followed by AHCT. The 72-month probability of worsening ≥1.0 EDSS point was 0.52 (95% confidence interval, 0.30-0.75). Five patients had an EDSS at baseline of ≤6.0; four of them had not failed treatment at last study visit. OCB in CSF persisted with minor changes in the banding pattern. Four new or enhancing lesions were seen on MRI, all within 13 months of treatment. In this population with high baseline EDSS, a significant proportion of patients with advanced MS remained stable for as long as 7 years after transplant. Non-inflammatory events may have contributed to neurological worsening after treatment. HDIT/AHCT may be more effective in patients with less advanced relapsing/remitting MS.

  2. Long term high temperature oxidation characteristics of La and Cu alloyed ferritic stainless steels for solid oxide fuel cell interconnects

    Science.gov (United States)

    Swaminathan, Srinivasan; Lee, Young-Su; Kim, Dong-Ik

    2016-09-01

    To ensure the best performance of solid oxide fuel cell metallic interconnects, the Fe-22 wt.% Cr ferritic stainless steels with various La contents (0.006-0.6 wt.%) and Cu addition (1.57 wt.%), are developed. Long-term isothermal oxidation behavior of these steels is investigated in air at 800 °C, for 2700 h. Chemistry, morphology, and microstructure of the thermally grown oxide scale are examined using XPS, SEM-EDX, and XRD techniques. Broadly, all the steels show a double layer consisting of an inner Cr2O3 and outer (Mn, Cr)3O4. Distinctly, in the La-added steels, binary oxides of Cr, Mn and Ti are found at the oxide scale surface together with (Mn, Cr)3O4. Furthermore, all La-varied steels possess the metallic Fe protrusions along with discontinuous (Mn, Cr)3O4 spinel zones at the oxide scale/metal interface and isolated precipitates of Ti-oxides in the underlying matrix. Increase of La content to 0.6 wt.% is detrimental to the oxidation resistance. For the Cu-added steel, Cu is found to segregate strongly at the oxide scale/metal interface which inhibits the ingress of oxygen thereby suppressing the subscale formation of (Mn, Cr)3O4. Thus, Cu addition to the Fe-22Cr ferritic stainless steels benefits the oxidation resistance.

  3. Long-Term Performance of Chemically and Physically Modified Activated Carbons in Air Cathodes of Microbial Fuel Cells

    KAUST Repository

    Zhang, Xiaoyuan

    2014-07-31

    © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim. Activated carbon (AC) is a low-cost and effective catalyst for oxygen reduction in air cathodes of microbial fuel cells (MFCs), but its performance must be maintained over time. AC was modified by three methods: 1)pyrolysis with iron ethylenediaminetetraacetic acid (AC-Fe), 2)heat treatment (AC-heat), and 3)mixing with carbon black (AC-CB). The maximum power densities after one month with these AC cathodes were 35% higher with AC-Fe (1410±50mW m-2) and AC-heat (1400±20mW m-2), and 16% higher with AC-CB (1210±30mW m-2) than for plain AC (1040±20mW m-2), versus 1270±50mW m-2 for a Pt control. After 16months, the Pt cathodes produced only 250±10mW m-2. However, the AC-heat and AC-CB cathodes still produced 960-970mW m-2, whereas plain AC produced 860±60mW m-2. The performance of the AC cathodes was restored to >85% of the initial maximum power densities by cleaning with a weak acid solution. Based on cost considerations among the AC materials, AC-CB appears to be the best choice for long-term performance.

  4. Long-term performance of activated carbon air cathodes with different diffusion layer porosities in microbial fuel cells

    KAUST Repository

    Zhang, Fang

    2011-08-01

    Activated carbon (AC) air-cathodes are inexpensive and useful alternatives to Pt-catalyzed electrodes in microbial fuel cells (MFCs), but information is needed on their long-term stability for oxygen reduction. AC cathodes were constructed with diffusion layers (DLs) with two different porosities (30% and 70%) to evaluate the effects of increased oxygen transfer on power. The 70% DL cathode initially produced a maximum power density of 1214±123mW/m 2 (cathode projected surface area; 35±4W/m 3 based on liquid volume), but it decreased by 40% after 1 year to 734±18mW/m 2. The 30% DL cathode initially produced less power than the 70% DL cathode, but it only decreased by 22% after 1 year (from 1014±2mW/m 2 to 789±68mW/m 2). Electrochemical tests were used to examine the reasons for the degraded performance. Diffusion resistance in the cathode was found to be the primary component of the internal resistance, and it increased over time. Replacing the cathode after 1 year completely restored the original power densities. These results suggest that the degradation in cathode performance was due to clogging of the AC micropores. These findings show that AC is a cost-effective material for oxygen reduction that can still produce ~750mW/m 2 after 1 year. © 2011 Elsevier B.V.

  5. Creep Behavior of Glass/Ceramic Sealant and its Effect on Long-term Performance of Solid Oxide Fuel Cells

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Wenning N.; Sun, Xin; Koeppel, Brian J.; Stephens, Elizabeth V.; Khaleel, Mohammad A.

    2009-10-14

    The creep behavior of glass or glass-ceramic sealant materials used in solid oxide fuel cells (SOFCs) becomes relevant under SOFC operating temperatures. In this paper, the creep of glass-ceramic sealants was experimentally examined, and a standard linear solid model was applied to capture the creep behavior of glass ceramic sealant materials developed for planar SOFCs at high temperatures. The parameters of this model were determined based on the creep test results. Furthermore, the creep model was incorporated into finite-element software programs SOFC-MP and Mentat-FC developed at Pacific Northwest National Laboratory for multi-physics simulation of SOFCs. The effect of creep of glass ceramic sealant materials on the long-term performance of SOFC stacks was investigated by studying the stability of the flow channels and the stress redistribution in the glass seal and on the various interfaces of the glass seal with other layers. Finite element analyses were performed to quantify the stresses in various parts. The stresses in glass seals were released because of creep behavior during operations.

  6. A review on durability issues and restoration techniques in long-term operations of direct methanol fuel cells

    Science.gov (United States)

    Mehmood, Asad; Scibioh, M. Aulice; Prabhuram, Joghee; An, Myung-Gi; Ha, Heung Yong

    2015-11-01

    Direct methanol fuel cells (DMFCs) remain attractive among advanced energy conversion technologies due to their high energy density and simple system configuration. Although they made an early market entry but failed to attain a large-scale commercialization mainly because of their inferior performance sustainment in lifetime operations and high production costs. There have been lots of R&D efforts made to upgrade the long-term durability of DMFCs to a commercially acceptable standard. These rigorous efforts have been useful in gaining insights about various degradation mechanisms and their origins. This review first briefly describes the recent progress in lifetime enhancement of DMFC technology reported by various groups in academia and industry. Then, it is followed by comprehensive discussions on the major performance degradation routes and associated physico-chemical origins, and influence of operational parameters, together with the methods which have been employed to alleviate and restore the performance losses. Finally, a brief summary of the presented literature survey is provided in conjunction with some possible future research directions.

  7. Amoeba-based computing for traveling salesman problem: long-term correlations between spatially separated individual cells of Physarum polycephalum.

    Science.gov (United States)

    Zhu, Liping; Aono, Masashi; Kim, Song-Ju; Hara, Masahiko

    2013-04-01

    A single-celled, multi-nucleated amoeboid organism, a plasmodium of the true slime mold Physarum polycephalum, can perform sophisticated computing by exhibiting complex spatiotemporal oscillatory dynamics while deforming its amorphous body. We previously devised an "amoeba-based computer (ABC)" to quantitatively evaluate the optimization capability of the amoeboid organism in searching for a solution to the traveling salesman problem (TSP) under optical feedback control. In ABC, the organism changes its shape to find a high quality solution (a relatively shorter TSP route) by alternately expanding and contracting its pseudopod-like branches that exhibit local photoavoidance behavior. The quality of the solution serves as a measure of the optimality of which the organism maximizes its global body area (nutrient absorption) while minimizing the risk of being illuminated (exposure to aversive stimuli). ABC found a high quality solution for the 8-city TSP with a high probability. However, it remains unclear whether intracellular communication among the branches of the organism is essential for computing. In this study, we conducted a series of control experiments using two individual cells (two single-celled organisms) to perform parallel searches in the absence of intercellular communication. We found that ABC drastically lost its ability to find a solution when it used two independent individuals. However, interestingly, when two individuals were prepared by dividing one individual, they found a solution for a few tens of minutes. That is, the two divided individuals remained correlated even though they were spatially separated. These results suggest the presence of a long-term memory in the intrinsic dynamics of this organism and its significance in performing sophisticated computing. PMID:23438635

  8. Input- and Cell-Type-Specific Endocannabinoid-Dependent LTD in the Striatum

    Directory of Open Access Journals (Sweden)

    Yu-Wei Wu

    2015-01-01

    Full Text Available Changes in basal ganglia plasticity at the corticostriatal and thalamostriatal levels are required for motor learning. Endocannabinoid-dependent long-term depression (eCB-LTD is known to be a dominant form of synaptic plasticity expressed at these glutamatergic inputs; however, whether eCB-LTD can be induced at all inputs on all striatal neurons is still debatable. Using region-specific Cre mouse lines combined with optogenetic techniques, we directly investigated and distinguished between corticostriatal and thalamostriatal projections. We found that eCB-LTD was successfully induced at corticostriatal synapses, independent of postsynaptic striatal spiny projection neuron (SPN subtype. Conversely, eCB-LTD was only nominally present at thalamostriatal synapses. This dichotomy was attributable to the minimal expression of cannabinoid type 1 (CB1 receptors on thalamostriatal terminals. Furthermore, coactivation of dopamine receptors on SPNs during LTD induction re-established SPN-subtype-dependent eCB-LTD. Altogether, our findings lay the groundwork for understanding corticostriatal and thalamostriatal synaptic plasticity and for striatal eCB-LTD in motor learning.

  9. CAST-ChIP Maps Cell-Type-Specific Chromatin States in the Drosophila Central Nervous System

    Directory of Open Access Journals (Sweden)

    Tamás Schauer

    2013-10-01

    Full Text Available Chromatin organization and gene activity are responsive to developmental and environmental cues. Although many genes are transcribed throughout development and across cell types, much of gene regulation is highly cell-type specific. To readily track chromatin features at the resolution of cell types within complex tissues, we developed and validated chromatin affinity purification from specific cell types by chromatin immunoprecipitation (CAST-ChIP, a broadly applicable biochemical procedure. RNA polymerase II (Pol II CAST-ChIP identifies ∼1,500 neuronal and glia-specific genes in differentiated cells within the adult Drosophila brain. In contrast, the histone H2A.Z is distributed similarly across cell types and throughout development, marking cell-type-invariant Pol II-bound regions. Our study identifies H2A.Z as an active chromatin signature that is refractory to changes across cell fates. Thus, CAST-ChIP powerfully identifies cell-type-specific as well as cell-type-invariant chromatin states, enabling the systematic dissection of chromatin structure and gene regulation within complex tissues such as the brain.

  10. Long-term survival of human neural stem cells in the ischemic rat brain upon transient immunosuppression.

    Directory of Open Access Journals (Sweden)

    Laura Rota Nodari

    Full Text Available Understanding the physiology of human neural stem cells (hNSCs in the context of cell therapy for neurodegenerative disorders is of paramount importance, yet large-scale studies are hampered by the slow-expansion rate of these cells. To overcome this issue, we previously established immortal, non-transformed, telencephalic-diencephalic hNSCs (IhNSCs from the fetal brain. Here, we investigated the fate of these IhNSC's immediate progeny (i.e. neural progenitors; IhNSC-Ps upon unilateral implantation into the corpus callosum or the hippocampal fissure of adult rat brain, 3 days after global ischemic injury. One month after grafting, approximately one fifth of the IhNSC-Ps had survived and migrated through the corpus callosum, into the cortex or throughout the dentate gyrus of the hippocampus. By the fourth month, they had reached the ipsilateral subventricular zone, CA1-3 hippocampal layers and the controlateral hemisphere. Notably, these results could be accomplished using transient immunosuppression, i.e administering cyclosporine for 15 days following the ischemic event. Furthermore, a concomitant reduction of reactive microglia (Iba1+ cells and of glial, GFAP+ cells was also observed in the ipsilateral hemisphere as compared to the controlateral one. IhNSC-Ps were not tumorigenic and, upon in vivo engraftment, underwent differentiation into GFAP+ astrocytes, and β-tubulinIII+ or MAP2+ neurons, which displayed GABAergic and GLUTAmatergic markers. Electron microscopy analysis pointed to the formation of mature synaptic contacts between host and donor-derived neurons, showing the full maturation of the IhNSC-P-derived neurons and their likely functional integration into the host tissue. Thus, IhNSC-Ps possess long-term survival and engraftment capacity upon transplantation into the globally injured ischemic brain, into which they can integrate and mature into neurons, even under mild, transient immunosuppressive conditions. Most notably

  11. Efficient long-term survival of cell grafts after myocardial infarction with thick viable cardiac tissue entirely from pluripotent stem cells.

    Science.gov (United States)

    Matsuo, Takehiko; Masumoto, Hidetoshi; Tajima, Shuhei; Ikuno, Takeshi; Katayama, Shiori; Minakata, Kenji; Ikeda, Tadashi; Yamamizu, Kohei; Tabata, Yasuhiko; Sakata, Ryuzo; Yamashita, Jun K

    2015-11-20

    Poor engraftment of cells after transplantation to the heart is a common and unresolved problem in the cardiac cell therapies. We previously generated cardiovascular cell sheets entirely from pluripotent stem cells with cardiomyocytes, endothelial cells and vascular mural cells. Though sheet transplantation showed a better engraftment and improved cardiac function after myocardial infarction, stacking limitation (up to 3 sheets) by hypoxia hampered larger structure formation and long-term survival of the grafts. Here we report an efficient method to overcome the stacking limitation. Insertion of gelatin hydrogel microspheres (GHMs) between each cardiovascular cell sheet broke the viable limitation via appropriate spacing and fluid impregnation with GHMs. Fifteen sheets with GHMs (15-GHM construct; >1 mm thickness) were stacked within several hours and viable after 1 week in vitro. Transplantation of 5-GHM constructs (≈2 × 10(6) of total cells) to a rat myocardial infarction model showed rapid and sustained functional improvements. The grafts were efficiently engrafted as multiple layered cardiovascular cells accompanied by functional capillary networks. Large engrafted cardiac tissues (0.8 mm thickness with 40 cell layers) successfully survived 3 months after TX. We developed an efficient method to generate thicker viable tissue structures and achieve long-term survival of the cell graft to the heart.

  12. Reassessment of long-term depression in cerebellar Purkinje cells in mice carrying mutated GluA2 C terminus.

    Science.gov (United States)

    Yamaguchi, Kazuhiko; Itohara, Shigeyoshi; Ito, Masao

    2016-09-01

    Long-term depression (LTD) of synaptic transmission from parallel fibers (PFs) to a Purkinje cell (PC) in the cerebellum has been considered to be a core mechanism of motor learning. Recently, however, discrepancies between LTD and motor learning have been reported in mice with a mutation that targeted the expression of PF-PC LTD by blocking AMPA-subtype glutamate receptor internalization regulated via the phosphorylation of AMPA receptors. In these mice, motor learning behavior was normal, but no PF-PC LTD was observed. We reexamined slices obtained from these GluA2 K882A and GluA2 Δ7 knockin mutants at 3-6 mo of age. The conventional protocols of stimulation did not induce LTD in these mutant mice, as previously reported, but surprisingly, LTD was induced using certain modified protocols. Such modifications involved increases in the number of PF stimulation (from one to two or five), replacement of climbing fiber stimulation with somatic depolarization (50 ms), filling a patch pipette with a Cs(+)-based solution, or extension of the duration of conjunction. We also found that intracellular infusion of a selective PKCα inhibitor (Gö6976) blocked LTD induction in the mutants, as in WT, suggesting that functional compensation occurred downstream of PKCα. The possibility that LTD in the mutants was caused by changes in membrane resistance, access resistance, or presynaptic property was excluded. The present results demonstrate that LTD is inducible by intensified conjunctive stimulations even in K882A and Δ7 mutants, indicating no contradiction against the LTD hypothesis of motor learning. PMID:27551099

  13. Early Significant Tumor Volume Reduction After Radiosurgery in Brain Metastases From Renal Cell Carcinoma Results in Long-Term Survival

    International Nuclear Information System (INIS)

    Purpose: To retrospectively evaluate survival of patients with brain metastasis from renal cell carcinoma (RCC) after radiosurgery. Patients and Methods: Between 1998 and 2010, 46 patients were treated with radiosurgery, and the total number of lesions was 99. The mean age was 58.9 years (range, 33–78 years). Twenty-six patients (56.5%) had a single brain metastasis. The mean tumor volume was 3.0 cm3 (range, 0.01–35.1 cm3), and the mean marginal dose prescribed was 20.8 Gy (range, 12–25 Gy) at the 50% isodose line. A patient was classified into the good-response group when the sum of the volume of the brain metastases decreased to less than 75% of the original volume at a 1-month follow-up evaluation using MRI. Results: As of December 28, 2010, 39 patients (84.8%) had died, and 7 (15.2%) survived. The overall median survival time was 10.0 ± 0.4 months (95% confidence interval, 9.1–10.8). After treatment, local tumor control was achieved in 72 (84.7%) of the 85 tumors assessed using MRI after radiosurgery. The good-response group survived significantly longer than the poor-response group (median survival times of 18.0 and 9.0 months, respectively; p = 0.025). In a multivariate analysis, classification in the good-response group was the only independent prognostic factor for longer survival (p = 0.037; hazard ratio = 0.447; 95% confidence interval, 0.209–0.953). Conclusions: Radiosurgery seems to be an effective treatment modality for patients with brain metastases from RCC. The early significant tumor volume reduction observed after radiosurgery seems to result in long-term survival in RCC patients with brain metastases.

  14. Carbazole dye with phosphonic acid anchoring groups for long-term heat stability of dye-sensitized solar cells

    International Nuclear Information System (INIS)

    In dye-sensitized solar cells (DSSCs), the binding strength between the dye and TiO2 surface may affect the heat stability of the DSSCs. Carboxylic acids have generally been used as the anchoring groups for the adsorption of dyes on TiO2 surfaces. However, a phosphonic acid anchor is expected to effect more stable bonding to a TiO2 surface because of its tridentate binding capability. In this study, we developed a new carbazole dye that employed a phosphonic acid anchor, MK-94. We studied the effect of the dye anchoring group on the heat stability of DSSCs by comparing MK-94, MK-2 (a carbazole dye with a carboxylic acid anchoring group), and Z-907 (a well-known, robust Ru complex dye with a carboxylic acid anchoring group). In the initial performance assessment, the DSSC with MK-94 exhibited a lower short circuit current (jsc) but a higher open circuit voltage (ΔVoc) than MK-2. With respect to heat stability, decreases in jsc and ΔVoc were suppressed with MK-94, and the drop of the power conversion efficiency (PCE) was kept within 6% after 504 h of heating. In addition, MK-94 demonstrated the longest electron lifetime and largest charge transfer resistance than the other dyes during the stability test. In contrast, the PCE decreased 27% from the initial value in the DSSC with MK-2. Thus, the phosphonic acid anchor was effective in extending the long-term heat stability of the DSSCs

  15. Breast Implant–Associated Anaplastic Large-Cell Lymphoma: Long-Term Follow-Up of 60 Patients

    Science.gov (United States)

    Miranda, Roberto N.; Aladily, Tariq N.; Prince, H. Miles; Kanagal-Shamanna, Rashmi; de Jong, Daphne; Fayad, Luis E.; Amin, Mitual B.; Haideri, Nisreen; Bhagat, Govind; Brooks, Glen S.; Shifrin, David A.; O'Malley, Dennis P.; Cheah, Chan Y.; Bacchi, Carlos E.; Gualco, Gabriela; Li, Shiyong; Keech, John A.; Hochberg, Ephram P.; Carty, Matthew J.; Hanson, Summer E.; Mustafa, Eid; Sanchez, Steven; Manning, John T.; Xu-Monette, Zijun Y.; Miranda, Alonso R.; Fox, Patricia; Bassett, Roland L.; Castillo, Jorge J.; Beltran, Brady E.; de Boer, Jan Paul; Chakhachiro, Zaher; Ye, Dongjiu; Clark, Douglas; Young, Ken H.; Medeiros, L. Jeffrey

    2014-01-01

    Purpose Breast implant–associated anaplastic large-cell lymphoma (ALCL) is a recently described clinicopathologic entity that usually presents as an effusion-associated fibrous capsule surrounding an implant. Less frequently, it presents as a mass. The natural history of this disease and long-term outcomes are unknown. Patients and Methods We reviewed the literature for all published cases of breast implant–associated ALCL from 1997 to December 2012 and contacted corresponding authors to update clinical follow-up. Results The median overall survival (OS) for 60 patients was 12 years (median follow-up, 2 years; range, 0-14 years). Capsulectomy and implant removal was performed on 56 of 60 patients (93%). Therapeutic data were available for 55 patients: 39 patients (78%) received systemic chemotherapy, and of the 16 patients (28%) who did not receive chemotherapy, 12 patients opted for watchful waiting and four patients received radiation therapy alone. Thirty-nine (93%) of 42 patients with disease confined by the fibrous capsule achieved complete remission, compared with complete remission in 13 (72%) of 18 patients with a tumor mass. Patients with a breast mass had worse OS and progression-free survival (PFS; P = .052 and P = .03, respectively). The OS or PFS were similar between patients who received and did not receive chemotherapy (P = .44 and P = .28, respectively). Conclusion Most patients with breast implant–associated ALCL who had disease confined within the fibrous capsule achieved complete remission. Proper management for these patients may be limited to capsulectomy and implant removal. Patients who present with a mass have a more aggressive clinical course that may be fatal, justifying cytotoxic chemotherapy in addition to removal of implants. PMID:24323027

  16. Long-term survival and T-cell kinetics in relapsed/refractory ALL patients who achieved MRD response after blinatumomab treatment.

    Science.gov (United States)

    Zugmaier, Gerhard; Gökbuget, Nicola; Klinger, Matthias; Viardot, Andreas; Stelljes, Matthias; Neumann, Svenja; Horst, Heinz-A; Marks, Reinhard; Faul, Christoph; Diedrich, Helmut; Reichle, Albrecht; Brüggemann, Monika; Holland, Chris; Schmidt, Margit; Einsele, Hermann; Bargou, Ralf C; Topp, Max S

    2015-12-10

    This long-term follow-up analysis evaluated overall survival (OS) and relapse-free survival (RFS) in a phase 2 study of the bispecific T-cell engager antibody construct blinatumomab in 36 adults with relapsed/refractory B-precursor acute lymphoblastic leukemia (ALL). In the primary analysis, 25 (69%) patients with relapsed/refractory ALL achieved complete remission with full (CR) or partial (CRh) hematologic recovery of peripheral blood counts within the first 2 cycles. Twenty-five patients (69%) had a minimal residual disease (MRD) response (bone marrow, and 1 patient with normocellular bone marrow but low peripheral counts. Ten of the 36 patients (28%) were long-term survivors (OS ≥30 months). Median OS was 13.0 months (median follow-up, 32.6 months). MRD response was associated with significantly longer OS (Mantel-Byar P = .009). All 10 long-term survivors had an MRD response. Median RFS was 8.8 months (median follow-up, 28.9 months). A plateau for RFS was reached after ∼18 months. Six of the 10 long-term survivors remained relapse-free, including 4 who received allogeneic stem cell transplantation (allo-SCT) as consolidation for blinatumomab and 2 who received 3 additional cycles of blinatumomab instead of allo-SCT. Three long-term survivors had neurologic events or cytokine release syndrome, resulting in temporary blinatumomab discontinuation; all restarted blinatumomab successfully. Long-term survivors had more pronounced T-cell expansion than patients with OS <30 months. PMID:26480933

  17. Older patients with inoperable non-small cell lung cancer. Long-term survival after concurrent chemoradiotherapy

    Energy Technology Data Exchange (ETDEWEB)

    Semrau, Sabine; Fietkau, Rainer [Friedrich-Alexander-University Erlangen-Nuernberg, Department of Radiation Oncology, Erlangen (Germany); Zettl, Heike [Rostock Cancer Registry University of Rostock, Rostock (Germany); Hildebrandt, Guido [University of Rostock, Department of Radiation Therapy, Rostock (Germany); Klautke, Gunther [Klinikum Chemnitz, Department of Radiation Therapy, Chemnitz (Germany)

    2014-12-15

    Considering the various comorbidities associated with aging, the feasibility and usefulness of concurrent chemoradiotherapy (CRT) in older patients with inoperable non-small cell lung cancer (NSCLC) is a controversial issue. Here, we compared the feasibility of CRT and the effects of various comorbidities on the prognosis of a minimally selected population of inoperable NSCLC patients aged 60-77 years. The study comprised 161 patients with inoperable NSCLC who received CRT with a target radiation dose greater than 60 Gy and platinum-based chemotherapy from 1998 to 2007. The total population included 69 patients aged 60-69 years and 53 aged 70-77 years. These two age cohorts were included in the study with a follow-up of a median 14.5 months. The two groups showed no differences in long-term survival, as reflected by the 5-year survival rates of 13.0 ± 4.1 % (60- to 69-year-olds) and 14.4 ± 4.9 % (70- to 77-year-olds). During the treatment phase, the groups were comparable in terms of toxicity and the feasibility of chemotherapy. Compared to patients in their 60s, the septuagenarians had more pulmonary comorbidities (p = 0.02), diabetes mellitus (p = 0.04), cardiac comorbidities (p = 0.08), and previous cancer disease (p = 0.08) that exerted a negative effect on survival. In patients without comorbidities, there were no differences between the age groups. Age is not a contraindication for concurrent CRT per se, because elderly patients do not have a worse long-term prognosis than younger seniors. However, ''elderly patients'' (≥ 70-77 years) have more concomitant diseases associated with shorter survival than ''moderately aged patients'' (≥ 60-69 years). (orig.) [German] Hinsichtlich der verschiedenen altersbedingten Komorbiditaeten werden die Durchfuehrbarkeit und der Nutzen einer simultanen Chemoradiotherapie (''concurrent chemoradiotherapy'', CRT) bei alten Patienten mit einem inoperablen nicht

  18. Long term morphological modelling

    DEFF Research Database (Denmark)

    Kristensen, Sten Esbjørn; Deigaard, Rolf; Taaning, Martin;

    2010-01-01

    A morphological modelling concept for long term nearshore morphology is proposed and examples of its application are presented and discussed. The model concept combines parameterised representations of the cross-shore morphology, with a 2DH area model for waves, currents and sediment transport in...

  19. Long-Term Collections

    CERN Multimedia

    Comité des collectes à long terme

    2011-01-01

    It is the time of the year when our fireman colleagues go around the laboratory for their traditional calendars sale. A part of the money of the sales will be donated in favour of the long-term collections. We hope that you will welcome them warmly.

  20. Gene expression profiles of hepatic cell-type specific marker genes in progression of liver fibrosis

    Institute of Scientific and Technical Information of China (English)

    Yoshiyuki Takahara; Mitsuo Takahashi; Hiroki Wagatsuma; Fumihiko Yokoya; Qing-Wei Zhang; Mutsuyo Yamaguchi; Hiroyuki Aburatani; Norifumi Kawada

    2006-01-01

    AIM: To determine the gene expression profile data for the whole liver during development of dimethylnitrosamine (DMN)-induced hepatic fibrosis.METHODS: Marker genes were identified for different types of hepatic cells, including hepatic stellate cells (HSCs), Kupffer cells (including other inflammatory cells),and hepatocytes, using independent temporal DNA microarray data obtained from isolated hepatic cells.RESULTS: The cell-type analysis of gene expression gave several key results and led to formation of three hypotheses: (1) changes in the expression of HSCspecific marker genes during fibrosis were similar to gene expression data in in vitro cultured HSCs, suggesting a major role of the self-activating characteristics of HSCs in formation of fibrosis; (2) expression of mast cell-specific marker genes reached a peak during liver fibrosis,suggesting a possible role of mast cells in formation of fibrosis; and (3) abnormal expression of hepatocytespecific marker genes was found across several metabolic pathways during fibrosis, including sulfur-containing amino acid metabolism, fatty acid metabolism, and drug metabolism, suggesting a mechanistic relationship between these abnormalities and symptoms of liver fibrosis.CONCLUSION: Analysis of marker genes for specific hepatic cell types can identify the key aspects of fibrogenesis. Sequential activation of inflammatory cells and the self-supporting properties of HSCs play an important role in development of fibrosis.

  1. Long-term follow-up of post hematopoietic stem cell transplantation for Hurler syndrome: Clinical, biochemical, and pathological improvements

    Directory of Open Access Journals (Sweden)

    Eriko Yasuda

    2015-03-01

    In conclusion, this long-term post-HSCT observation should shed light on a new aspect of therapeutic effect associated with skeletal pathology and GAG levels as a biomarker, indicating that HSCT is a primary choice at an early stage for not only CNS but also skeletal system in combination of appropriate surgical procedures.

  2. Defining cell-type specificity at the transcriptional level in human disease

    OpenAIRE

    Ju, Wenjun; Greene, Casey S; Eichinger, Felix; Nair, Viji; Hodgin, Jeffrey B.; Bitzer, Markus; Lee, Young-Suk; Zhu, Qian; Kehata, Masami; Li, Min; Jiang, Song; Rastaldi, Maria Pia; Cohen, Clemens D; Troyanskaya, Olga G.; Kretzler, Matthias

    2013-01-01

    Cell-lineage–specific transcripts are essential for differentiated tissue function, implicated in hereditary organ failure, and mediate acquired chronic diseases. However, experimental identification of cell-lineage–specific genes in a genome-scale manner is infeasible for most solid human tissues. We developed the first genome-scale method to identify genes with cell-lineage–specific expression, even in lineages not separable by experimental microdissection. Our machine-learning–based approa...

  3. Cell-type-specific neuroanatomy of cliques of autism-related genes in the mouse brain

    Directory of Open Access Journals (Sweden)

    Pascal eGrange

    2015-05-01

    Full Text Available Two cliques of genes identified computationally for their high co-expression in the mouse brain according to the Allen Brain Atlas, and for their enrichment in genes related to autism spectrum disorder, have recently been shown to be highly co-expressed in the cerebellar cortex, compared to what could be expected by chance. Moreover, the expression of these cliques of genes is not homogeneous across the cerebellar cortex, and it has been noted that their expression pattern seems to highlight the granular layer. However, this observation was only made by eye, and recent advances in computational neuroanatomy allow to rank cell types in the mouse brain (characterized by their transcriptome profiles according to the similarity between their spatial density profiles and the expression profiles of the cliques. We establish by Monte Carlo simulation that with probability at least 99%, the expression profiles of the two cliques are more similar to the density profile of granule cells than 99% of the expression of cliques containing the same number of genes (Purkinje cells also score above 99% in one of the cliques. Thresholding the expression profiles shows that the signal is more intense in the granular layer. Finally, we work out pairs of cell types whose combined expression profiles are more similar to the expression profiles of the cliquesthan any single cell type. These pairs predominantly consist of one cortical pyramidal cell and one cerebellar cell (whichcan be either a granule cell or a Purkinje cell.

  4. Cell-type-specific neuroanatomy of cliques of autism-related genes in the mouse brain.

    Science.gov (United States)

    Grange, Pascal; Menashe, Idan; Hawrylycz, Michael

    2015-01-01

    Two cliques of genes identified computationally for their high co-expression in the mouse brain according to the Allen Brain Atlas, and for their enrichment in genes related to autism spectrum disorder (ASD), have recently been shown to be highly co-expressed in the cerebellar cortex, compared to what could be expected by chance. Moreover, the expression of these cliques of genes is not homogeneous across the cerebellar cortex, and it has been noted that their expression pattern seems to highlight the granular layer. However, this observation was only made by eye, and recent advances in computational neuroanatomy allow to rank cell types in the mouse brain (characterized by their transcriptome profiles) according to the similarity between their spatial density profiles and the spatial expression profiles of the cliques. We establish by Monte Carlo simulation that with probability at least 99%, the expression profiles of the two cliques are more similar to the density profile of granule cells than 99% of the expression of cliques containing the same number of genes (Purkinje cells also score above 99% in one of the cliques). Thresholding the expression profiles shows that the signal is more intense in the granular layer. Finally, we work out pairs of cell types whose combined expression profiles are more similar to the expression profiles of the cliques than any single cell type. These pairs predominantly consist of one cortical pyramidal cell and one cerebellar cell (which can be either a granule cell or a Purkinje cell). PMID:26074809

  5. Assessment of Membrane Fluidity Fluctuations during Cellular Development Reveals Time and Cell Type Specificity

    KAUST Repository

    Noutsi, Pakiza

    2016-06-30

    Cell membrane is made up of a complex structure of lipids and proteins that diffuse laterally giving rise to what we call membrane fluidity. During cellular development, such as differentiation cell membranes undergo dramatic fluidity changes induced by proteins such as ARC and Cofilin among others. In this study we used the generalized polarization (GP) property of fluorescent probe Laurdan using two-photon microscopy to determine membrane fluidity as a function of time and for various cell lines. A low GP value corresponds to a higher fluidity and a higher GP value is associated with a more rigid membrane. Four different cell lines were monitored such as hN2, NIH3T3, HEK293 and L6 cells. Membrane fluidity was measured at 12h, 72h and 92 h. Our results show significant changes in membrane fluidity among all cell types at different time points. GP values tend to increase significantly within 92 h in hN2 cells and 72 h in NIH3T3 cells and only at 92 h in HEK293 cells. L6 showed a marked decrease in membrane fluidity at 72 h and starts to increase at 92 h. As expected, NIH3T3 cells have more rigid membrane at earlier time points. On the other hand, neurons tend to have the highest membrane fluidity at early time points emphasizing its correlation with plasticity and the need for this malleability during differentiation. This study sheds light on the involvement of membrane fluidity during neuronal differentiation and development of other cell lines.

  6. Assessment of Membrane Fluidity Fluctuations during Cellular Development Reveals Time and Cell Type Specificity.

    Science.gov (United States)

    Noutsi, Pakiza; Gratton, Enrico; Chaieb, Sahraoui

    2016-01-01

    Cell membrane is made up of a complex structure of lipids and proteins that diffuse laterally giving rise to what we call membrane fluidity. During cellular development, such as differentiation cell membranes undergo dramatic fluidity changes induced by proteins such as ARC and Cofilin among others. In this study we used the generalized polarization (GP) property of fluorescent probe Laurdan using two-photon microscopy to determine membrane fluidity as a function of time and for various cell lines. A low GP value corresponds to a higher fluidity and a higher GP value is associated with a more rigid membrane. Four different cell lines were monitored such as hN2, NIH3T3, HEK293 and L6 cells. Membrane fluidity was measured at 12h, 72h and 92 h. Our results show significant changes in membrane fluidity among all cell types at different time points. GP values tend to increase significantly within 92 h in hN2 cells and 72 h in NIH3T3 cells and only at 92 h in HEK293 cells. L6 showed a marked decrease in membrane fluidity at 72 h and starts to increase at 92 h. As expected, NIH3T3 cells have more rigid membrane at earlier time points. On the other hand, neurons tend to have the highest membrane fluidity at early time points emphasizing its correlation with plasticity and the need for this malleability during differentiation. This study sheds light on the involvement of membrane fluidity during neuronal differentiation and development of other cell lines. PMID:27362860

  7. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, Martha R.; Garbe, James C.

    2016-06-28

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  8. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, Martha R; Garbe, James C

    2015-02-24

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  9. Long-term follow-up of kidney allografts in patients with sickle cell hemoglobinopathy Transplante renal na anemia falciforme

    Directory of Open Access Journals (Sweden)

    João R. Friedrisch

    2003-06-01

    Full Text Available Although sickle cell anemia and sickle cell disease produce a variety of functional renal abnormalities they uncommonly cause end stage renal failure. Renal transplantation has been a successful alternative for the treatment of the rare terminal chronic renal failure with outcomes comparable with non-sickle recipients. This approach, however, has not been often described on patients with renal failure associated with SC hemoglobinopathy. Here we report the outcomes of two patients with chronic renal failure due to SC hemoglobinopathies who underwent renal transplantation. At the time of the transplantation they were both severely anemic and had frequent vasoocclosive pain crises. Both patients evolved with good allograft function, near normal hematological parameters, and very rare pain crisis, thirteen and eight years after transplant. These cases illustrate that terminal renal failure due to SC hemoglobinopathy can be successfully managed by renal transplantation and satisfactory long-term results are achievable not only in terms of renal allograft function but also of their hematological condition.Embora a anemia falciforme e as síndromes falciformes freqüentemente causem várias alterações funcionais renais, não é comum a insuficiência renal terminal. Nestes casos, o transplante renal é uma alternativa que se acompanha de resultados comparáveis aos obtidos em receptores sem hemoglobinopatias. Esta estratégia terapêutica tem sido, no entanto, pouco relatada para portadores de hemoglobinopatia SC. Este relato descreve a evolução de dois pacientes portadores de hemoglobinopatia SC que foram submetidos ao transplante renal. No momento do transplante ambos apresentavam severa anemia e crises dolorosas freqüentes. Os pacientes evoluíram com boa função do enxerto, parâmetros hematológicos quase normais e praticamente assintomáticos do ponto de vista da hemoglobinopatia, treze e oito anos após o transplante. Estes casos ilustram

  10. Gene Expression Programs in Response to Hypoxia: Cell Type Specificity and Prognostic Significance in Human Cancers.

    Directory of Open Access Journals (Sweden)

    2006-01-01

    Full Text Available BACKGROUND: Inadequate oxygen (hypoxia triggers a multifaceted cellular response that has important roles in normal physiology and in many human diseases. A transcription factor, hypoxia-inducible factor (HIF, plays a central role in the hypoxia response; its activity is regulated by the oxygen-dependent degradation of the HIF-1alpha protein. Despite the ubiquity and importance of hypoxia responses, little is known about the variation in the global transcriptional response to hypoxia among different cell types or how this variation might relate to tissue- and cell-specific diseases. METHODS AND FINDINGS: We analyzed the temporal changes in global transcript levels in response to hypoxia in primary renal proximal tubule epithelial cells, breast epithelial cells, smooth muscle cells, and endothelial cells with DNA microarrays. The extent of the transcriptional response to hypoxia was greatest in the renal tubule cells. This heightened response was associated with a uniquely high level of HIF-1alpha RNA in renal cells, and it could be diminished by reducing HIF-1alpha expression via RNA interference. A gene-expression signature of the hypoxia response, derived from our studies of cultured mammary and renal tubular epithelial cells, showed coordinated variation in several human cancers, and was a strong predictor of clinical outcomes in breast and ovarian cancers. In an analysis of a large, published gene-expression dataset from breast cancers, we found that the prognostic information in the hypoxia signature was virtually independent of that provided by the previously reported wound signature and more predictive of outcomes than any of the clinical parameters in current use. CONCLUSIONS: The transcriptional response to hypoxia varies among human cells. Some of this variation is traceable to variation in expression of the HIF1A gene. A gene-expression signature of the cellular response to hypoxia is associated with a significantly poorer prognosis

  11. Cell type specificity and structural determinants of IRES activity from the 5' leaders of different HIV-1 transcripts.

    Science.gov (United States)

    Plank, Terra-Dawn M; Whitehurst, James T; Kieft, Jeffrey S

    2013-07-01

    Internal ribosome entry site (IRES) RNAs are important regulators of gene expression, but their diverse molecular mechanisms remain partially understood. The HIV-1 gag transcript leader contains an IRES that may be a good model for understanding the function of many other IRESs. We investigated the possibility that this IRES' function is linked to both the structure of the RNA and its cellular environment. We find that in the context of a bicistronic reporter construct, HIV-1 gag IRES' activity is cell type-specific, with higher activity in T-cell culture systems that model the natural target cells for HIV-1 infection. This finding underscores how an IRES may be fine tuned to function in certain cells, perhaps owing to cell type-specific protein factors. Using RNA probing and mutagenesis, we demonstrate that the HIV-1 gag IRES does not use pre-folded RNA structure to drive function, a finding that gives insight into how conformationally dynamic IRESs operate. Furthermore, we find that a common exon drives IRES activity in a diverse set of alternatively spliced transcripts. We propose a mechanism in which a structurally plastic RNA element confers the ability to initiate translation internally, and activity from this common element is modulated by 3' nucleotides added by alternative splicing. PMID:23661682

  12. Cell type specificity and structural determinants of IRES activity from the 5′ leaders of different HIV-1 transcripts

    Science.gov (United States)

    Plank, Terra-Dawn M.; Whitehurst, James T.; Kieft, Jeffrey S.

    2013-01-01

    Internal ribosome entry site (IRES) RNAs are important regulators of gene expression, but their diverse molecular mechanisms remain partially understood. The HIV-1 gag transcript leader contains an IRES that may be a good model for understanding the function of many other IRESs. We investigated the possibility that this IRES’ function is linked to both the structure of the RNA and its cellular environment. We find that in the context of a bicistronic reporter construct, HIV-1 gag IRES’ activity is cell type-specific, with higher activity in T-cell culture systems that model the natural target cells for HIV-1 infection. This finding underscores how an IRES may be fine tuned to function in certain cells, perhaps owing to cell type-specific protein factors. Using RNA probing and mutagenesis, we demonstrate that the HIV-1 gag IRES does not use pre-folded RNA structure to drive function, a finding that gives insight into how conformationally dynamic IRESs operate. Furthermore, we find that a common exon drives IRES activity in a diverse set of alternatively spliced transcripts. We propose a mechanism in which a structurally plastic RNA element confers the ability to initiate translation internally, and activity from this common element is modulated by 3′ nucleotides added by alternative splicing. PMID:23661682

  13. Cell type-specific interactions of transcription factors with a housekeeping promoter in vivo.

    OpenAIRE

    Stapleton, G; Somma, M P; Lavia, P

    1993-01-01

    Mammalian housekeeping promoters represent a class of regulatory elements different from those of tissues-specific genes, lacking a TATA box and associated with CG-rich DNA. We have compared the organization of the housekeeping Htf9 promoter in different cell types by genomic footprinting. The sites of in vivo occupancy clearly reflected local combinations of tissue-specific and ubiquitous binding factors. The flexibility of the Htf9 promoter in acting as the target of cell-specific combinati...

  14. CD4+CD25+CD127 regulatory cells play multiple roles in maintaining HIV-1 p24 production in patients on long-term treatment: HIV-1 p24-producing cells and suppression of anti-HIV immunity

    Directory of Open Access Journals (Sweden)

    Yan-Mei Jiao

    2015-08-01

    Conclusions: CD4+CD25+CD127 regulatory cells play multiple roles in maintaining HIV-1 p24 production in long-term ART patients. Treg cells may be a target for eliminating the latent HIV reservoir after effective long-term ART.

  15. Unique cell type-specific junctional complexes in vascular endothelium of human and rat liver sinusoids.

    Directory of Open Access Journals (Sweden)

    Cyrill Géraud

    Full Text Available Liver sinusoidal endothelium is strategically positioned to control access of fluids, macromolecules and cells to the liver parenchyma and to serve clearance functions upstream of the hepatocytes. While clearance of macromolecular debris from the peripheral blood is performed by liver sinusoidal endothelial cells (LSECs using a delicate endocytic receptor system featuring stabilin-1 and -2, the mannose receptor and CD32b, vascular permeability and cell trafficking are controlled by transcellular pores, i.e. the fenestrae, and by intercellular junctional complexes. In contrast to blood vascular and lymphatic endothelial cells in other organs, the junctional complexes of LSECs have not yet been consistently characterized in molecular terms. In a comprehensive analysis, we here show that LSECs express the typical proteins found in endothelial adherens junctions (AJ, i.e. VE-cadherin as well as α-, β-, p120-catenin and plakoglobin. Tight junction (TJ transmembrane proteins typical of endothelial cells, i.e. claudin-5 and occludin, were not expressed by rat LSECs while heterogenous immunreactivity for claudin-5 was detected in human LSECs. In contrast, junctional molecules preferentially associating with TJ such as JAM-A, B and C and zonula occludens proteins ZO-1 and ZO-2 were readily detected in LSECs. Remarkably, among the JAMs JAM-C was considerably over-expressed in LSECs as compared to lung microvascular endothelial cells. In conclusion, we show here that LSECs form a special kind of mixed-type intercellular junctions characterized by co-occurrence of endothelial AJ proteins, and of ZO-1 and -2, and JAMs. The distinct molecular architecture of the intercellular junctional complexes of LSECs corroborates previous ultrastructural findings and provides the molecular basis for further analyses of the endothelial barrier function of liver sinusoids under pathologic conditions ranging from hepatic inflammation to formation of liver metastasis.

  16. Cocultivation of umbilical cord blood CD34+ cells with retro-transduced hMSCs leads to effective amplification of long-term culture-initiating cells

    Institute of Scientific and Technical Information of China (English)

    Chun-Gang Xie; Jin-Fu Wang; Ying Xiang; Li-Yan Qiu; Bing-Bing Jia; Li-Juan Wang; Guo-Zhong Wang; Guo-Ping Huang

    2006-01-01

    AIM: To establish a novel coculture system for ex vivo expansion of umbilical cord blood(UCB) hematopoietic progenitors using thrombopoietin (TPO)/Flt-3 ligand(FL)-transduced human marrow-derived mesenchymal stem cells (tfhMSCs) as feeder.METHODS: UCB CD34+ cells were isolated and cultured using four culture systems in serum-containing or serumfree medium. Suitable aliquots of cultured cells were used to monitor cell production, clonogenic activity,and long-term culture-initiating culture (LTC-IC) output.Finally, the severe-combined immunodeficient (SCID)mouse-repopulating cell (SRC) assay was performed to confirm ability of the cultured cells to reconstitute longterm hematopoiesis.RESULTS: There were no significant differences in the number of total nucleated cells among different culture systems in serum-containing medium during 21-d culture. However, on d 14, the outputs of CD34+ cells,CFU-C and CFU-GEMM in tfhMSCs coculture system were significantly enhanced. LTC-IC assay demonstrated that the tfhMSCs coculture system had the most powerful activity. The severe-combined immunodeficient (SCID)mouse repopulating cell (SRC) assay confirmed extensive ability of the expanded cells to reconstitute long-term hematopoiesis. Furthermore, PCR analysis demonstrated the presence of human hematopoietic cells in the bone marrow and peripheral blood cells of NOD/SCID mice.CONCLUSION: The TPO/FL-transduced hMSCs, in combination with additive cytokines, can effectively expand hematopoietic progenitors from UCB in vitro and the tfhMSCs coculture system may be a suitable system for ex vivo manipulation of primitive progenitor cells under contact culture conditions.

  17. Therapy-related leukemia as a result of long-term chemotherapy for non-small cell lung cancer

    International Nuclear Information System (INIS)

    Recent advances in chemotherapy have prolonged the survival of lung cancer patients. However, there have been reports of patients who suffer therapy-related leukemia after chemotherapy. A 64-year-old woman was given a diagnosis of adenocarcinoma (T4N0M0 stage IIIB) in 2004, and received chemotherapy with cisplatin and other drugs from 2004 to 2009, and radiotherapy for brain metastasis in 2008. In January 2009, she was admitted to our hospital with low-grade fever, fatigue, and prolonged neutropenia. A peripheral blood smear revealed a predominance of circulating myeloblasts. Examination of the bone marrow aspirate confirmed acute myelogenous leukemia. We think that therapy-related leukemia developed because of long-term chemotherapy and radiotherapy. Therapy-related leukemia can develop in lung cancer patients who undergo long-term therapy. (author)

  18. Cardiovascular protection of magnolol: cell-type specificity and dose-related effects

    Directory of Open Access Journals (Sweden)

    Ho Jennifer

    2012-07-01

    Full Text Available Abstract Magnolia officinalis has been widely used in traditional Chinese medicine. Magnolol, an active component isolated from Magnolia officinalis, is known to be a cardiovascular protector since 1994. The multiplex mechanisms of magnolol on cardiovascular protection depends on cell types and dosages, and will be reviewed and discussed in this article. Magnolol under low and moderate dosage possesses the ability to protect heart from ischemic/reperfusion injury, reduces atherosclerotic change, protects endothelial cell against apoptosis and inhibits neutrophil-endothelial adhesion. The moderate to high concentration of magnolol mainly acts on smooth muscle cells and platelets. Magnolol induces apoptosis in vascular smooth muscle cells at moderate concentration and inhibits proliferation at moderate and high concentration. High concentration of magnolol also abrogates platelet activation, aggregation and thrombus formation. Magnolol also serves as an smooth muscle relaxant only upon the high concentration. Oral intake of magnolol to reach the therapeutic level for cardiovascular protection is applicable, thus makes magnolol an agent of great potential for preventing cardiovascular diseases in high-risk patients.

  19. Long-term exposure of A549 cells to titanium dioxide nanoparticles induces DNA damage and sensitizes cells towards genotoxic agents.

    Science.gov (United States)

    Armand, Lucie; Tarantini, Adeline; Beal, David; Biola-Clier, Mathilde; Bobyk, Laure; Sorieul, Sephanie; Pernet-Gallay, Karin; Marie-Desvergne, Caroline; Lynch, Iseult; Herlin-Boime, Nathalie; Carriere, Marie

    2016-09-01

    Titanium dioxide nanoparticles (TiO2-NPs) are one of the most produced NPs in the world. Their toxicity has been studied for a decade using acute exposure scenarios, i.e. high exposure concentrations and short exposure times. In the present study, we evaluated their genotoxic impact using long-term and low concentration exposure conditions. A549 alveolar epithelial cells were continuously exposed to 1-50 μg/mL TiO2-NPs, 86% anatase/14% rutile, 24 ± 6 nm average primary diameter, for up to two months. Their cytotoxicity, oxidative potential and intracellular accumulation were evaluated using MTT assay and reactive oxygen species measurement, transmission electron microscopy observation, micro-particle-induced X-ray emission and inductively-coupled plasma mass spectroscopy. Genotoxic impact was assessed using alkaline and Fpg-modified comet assay, immunostaining of 53BP1 foci and the cytokinesis-blocked micronucleus assay. Finally, we evaluated the impact of a subsequent exposure of these cells to the alkylating agent methyl methanesulfonate. We demonstrate that long-term exposure to TiO2-NPs does not affect cell viability but causes DNA damage, particularly oxidative damage to DNA and increased 53BP1 foci counts, correlated with increased intracellular accumulation of NPs. In addition, exposure over 2 months causes cellular responses suggestive of adaptation, characterized by decreased proliferation rate and stabilization of TiO2-NP intracellular accumulation, as well as sensitization to MMS. Taken together, these data underline the genotoxic impact and sensitization effect of long-term exposure of lung alveolar epithelial cells to low levels of TiO2-NPs. PMID:26785166

  20. Long-term immune deficiency after allogeneic stem cell transplantation: B-cell deficiency is associated with late infections

    OpenAIRE

    Corre, Elise; Carmagnat, Maryvonnick; Busson, Marc; de Latour, Regis Peffault; Robin, Marie; Ribaud, Patricia; Toubert, Antoine; Rabian, Claire; Socié, Gerard

    2010-01-01

    Immune reconstitution was analyzed in 140 consecutive patients who were 2-year disease-free and who underwent myeloablative allogeneic transplantation. A CD4 and CD8 defect was observed involving naive, terminally differentiated, memory and competent cells and above limits values for activated subsets. Natural killer cells normalize at six months while we observed expansion of CD19+/CD5+ B cells after three months and a persisting defect of memory B cells. Chronic graft-versus-host disease di...

  1. Long-term in vivo provision of antigen-specific T cell immunity by programming hematopoietic stem cells

    Science.gov (United States)

    Yang, Lili; Baltimore, David

    2005-03-01

    A method to genetically program mouse hematopoietic stem cells to develop into functional CD8 or CD4 T cells of defined specificity in vivo is described. For this purpose, a bicistronic retroviral vector was engineered that efficiently delivers genes for both and chains of T cell receptor (TCR) to hematopoietic stem cells. When modified cell populations were used to reconstruct the hematopoietic lineages of recipient mice, significant percentages of antigen-specific CD8 or CD4 T cells were observed. These cells expressed normal surface markers and responded to peptide antigen stimulation by proliferation and cytokine production. Moreover, they could mature into memory cells after peptide stimulation. Using TCRs specific for a model tumor antigen, we found that the recipient mice were able to partially resist a challenge with tumor cells carrying the antigen. By combining cells modified with CD8- and CD4-specific TCRs, and boosting with dendritic cells pulsed with cognate peptides, complete suppression of tumor could be achieved and even tumors that had become established would regress and be eliminated after dendritic cell/peptide immunization. This methodology of "instructive immunotherapy" could be developed for controlling the growth of human tumors and attacking established pathogens.

  2. In vivo vascularization of cell sheets provided better long-term tissue survival than injection of cell suspension.

    Science.gov (United States)

    Takeuchi, Ryohei; Kuruma, Yosuke; Sekine, Hidekazu; Dobashi, Izumi; Yamato, Masayuki; Umezu, Mitsuo; Shimizu, Tatsuya; Okano, Teruo

    2016-08-01

    Cell sheets have shown a remarkable ability for repairing damaged myocardium in clinical and preclinical studies. Although they demonstrate a high degree of viability as engrafted cells in vivo, the reason behind their survivability is unclear. In this study, the survival and vascularization of rat cardiac cell sheets transplanted in the subcutaneous tissue of athymic rats were investigated temporally. The cell sheets showed significantly higher survival than cell suspensions for up to 12 months, using an in vivo bioluminescence imaging system to detect luciferase-positive transplanted cells. Terminal deoxynucleotidyl transferase dUTP nick-end labelling (TUNEL) assay also showed a smaller number of apoptotic cells in the cell sheets than in the cell suspensions at 1 day. Rapid vascular formation and maturation were observed inside the cell sheets using an in vivo imaging system. Leaky vessels appeared at 6 h, red blood cells flowing through functional vessels appeared at 12 h, and morphologically matured vessels appeared at 7 days. In addition, immunostaining of cell sheets with nerve/glial antigen-2 (NG2) showed that vessel maturity increased over time. Interestingly, these results correlated with the dynamics of cell sheet mRNA expression. Genes related to endothelial cells (ECs) proliferation, migration and vessel sprouting were highly expressed within 1 day, and genes related to pericyte recruitment and vessel maturation were highly expressed at 3 days or later. This suggested that the cell sheets could secrete appropriate angiogenic factors in a timely way after transplantation, and this ability might be a key reason for their high survival. Copyright © 2014 John Wiley & Sons, Ltd. PMID:24470393

  3. Innate immune response to pulmonary contusion: Identification of cell-type specific inflammatory responses

    OpenAIRE

    Hoth, J. Jason; Wells, Jonathan D.; Yoza, Barbara K.; McCall, Charles E.

    2012-01-01

    Lung injury from pulmonary contusion is a common traumatic injury, predominantly seen after blunt chest trauma such as in vehicular accidents. The local and systemic inflammatory response to injury includes activation of innate immune receptors, elaboration of a variety inflammatory mediators, and recruitment of inflammatory cells to the injured lung. Using a mouse model of pulmonary contusion, we had previously shown that innate immune Toll like receptors 2 and 4 (TLR2 and TLR4) mediate the ...

  4. Cell Type-Specific Manipulation with GFP-Dependent Cre Recombinase

    OpenAIRE

    Tang, Jonathan C. Y.; Rudolph, Stephanie; Dhande, Onkar S.; Abraira, Victoria E.; Choi, Seungwon; Lapan, Sylvain; Drew, Iain R.; Drokhlyansky, Eugene; Huberman, Andrew D.; Regehr, Wade G.; Cepko, Constance L.

    2015-01-01

    Summary There are many transgenic GFP reporter lines that allow visualization of specific populations of cells. Using such lines for functional studies requires a method that transforms GFP into a molecule that enables genetic manipulation. Here we report the creation of a method that exploits GFP for gene manipulation, Cre Recombinase Dependent on GFP (CRE-DOG), a split component system that uses GFP and its derivatives to directly induce Cre/loxP recombination. Using plasmid electroporation...

  5. Cell-Type Specific Determinants of NRAMP1 Expression in Professional Phagocytes

    Directory of Open Access Journals (Sweden)

    Mathieu F. M. Cellier

    2013-01-01

    Full Text Available The Natural resistance-associated macrophage protein 1 (Nramp1 or Solute carrier 11 member 1, Slc11a1 transports divalent metals across the membrane of late endosomes and lysosomes in professional phagocytes. Nramp1 represents an ancient eukaryotic cell-autonomous defense whereas the gene duplication that yielded Nramp1 and Nramp2 predated the origin of Sarcopterygians (lobe-finned fishes and tetrapods. SLC11A1 genetic polymorphisms associated with human resistance to tuberculosis consist of potential regulatory variants. Herein, current knowledge of the regulation of SLC11A1 gene expression is reviewed and comprehensive analysis of ENCODE data available for hematopoietic cell-types suggests a hypothesis for the regulation of SLC11A1 expression during myeloid development and phagocyte functional polarization. SLC11A1 is part of a 34.6 kb CTCF-insulated locus scattered with predicted regulatory elements: a 3' enhancer, a large 5' enhancer domain and four elements spread around the transcription start site (TSS, including several C/EBP and PU.1 sites. SLC11A1 locus ends appear mobilized by ETS-related factors early during myelopoiesis; activation of both 5' and 3' enhancers in myelo-monocytic cells correlate with transcription factor binding at the TSS. Characterizing the corresponding cis/trans determinants functionally will establish the mechanisms involved and possibly reveal genetic variation that impacts susceptibility to infectious or immune diseases.

  6. Chimeric Allografts Induced by Short-Term Treatment With Stem Cell Mobilizing Agents Result in Long-Term Kidney Transplant Survival Without Immunosuppression: II, Study in Miniature Swine.

    Science.gov (United States)

    Cameron, A M; Wesson, R N; Ahmadi, A R; Singer, A L; Hu, X; Okabayashi, T; Wang, Y; Shigoka, M; Fu, Y; Gao, W; Raccusen, L C; Montgomery, R A; Williams, G M; Sun, Z

    2016-07-01

    Transplantation is now lifesaving therapy for patients with end-stage organ failure but requires lifelong immunosuppression with resultant morbidity. Current immunosuppressive strategies inhibit T cell activation and prevent donor-recipient engagement. Therefore, it is not surprising that few host cells are demonstrated in donor grafts. However, our recent small animal studies found large numbers of recipient stem cells present after transplantation and pharmacological mobilization, resulting in a chimeric, repopulated organ. We now confirm these findings in a well-characterized large animal preclinical model. Here, we show that AMD3100 and FK506 mobilization of endogenous stem cells immediately post kidney transplantation combined with repeat therapy at 1, 2, and 3 months led to drug-free long-term survival in maximally immunologically mismatched swine. Three long-term recipients have stable chimeric transplants, preserved antidonor skin graft responses, and normal serum creatinine levels despite withdrawal of all medication for 3 years. PMID:26748958

  7. Long-term administration of scopolamine interferes with nerve cell proliferation, differentiation and migration in adult mouse hippocampal dentate gyrus, but it does not induce cell death

    Institute of Scientific and Technical Information of China (English)

    Bing Chun Yan; Yun Lyul Lee; Il-Jun Kang; Moo-Ho Won; Joon Ha Park; Bai Hui Chen; Jeong-Hwi Cho; In Hye Kim; Ji Hyeon Ahn; Jae-Chul Lee; In Koo Hwang; Jun Hwi Cho

    2014-01-01

    Long-term administration of scopolamine, a muscarinic receptor antagonist, can inhibit the survival of newly generated cells, but its effect on the proliferation, differentiation and migration of nerve cells in the adult mouse hippocampal dentate gyrus remain poorly understood. In this study, we used immunohistochemistry and western blot methods to weekly detect the biological behaviors of nerve cells in the hippocampal dentate gyrus of adult mice that received intraperito-neal administration of scopolamine for 4 weeks. Expression of neuronal nuclear antigen (NeuN;a neuronal marker) and Fluoro-Jade B (a marker for the localization of neuronal degeneration) was also detected. After scopolamine treatment, mouse hippocampal neurons did not die, and Ki-67 (a marker for proliferating cells)-immunoreactive cells were reduced in number and reac hed the lowest level at 4 weeks. Doublecortin (DCX; a marker for newly generated neurons)-im-munoreactive cells were gradually shortened in length and reduced in number with time. After scopolamine treatment for 4 weeks, nearly all of the 5-bromo-2′-deoxyuridine (BrdU)-labeled newly generated cells were located in the subgranular zone of the dentate gyrus, but they did not migrate into the granule cell layer. Few mature BrdU/NeuN double-labeled cells were seen in the subgranular zone of the dentate gyrus. These ifndings suggest that long-term administration of scopolamine interferes with the proliferation, differentiation and migration of nerve cells in the adult mouse hippocampal dentate gyrus, but it does not induce cell death.

  8. NOX1-induced accumulation of reactive oxygen species in abdominal fat-derived mesenchymal stromal cells impinges on long-term proliferation

    OpenAIRE

    Sela, M; Tirza, G; Ravid, O; Volovitz, I; Solodeev, I; Friedman, O; Zipori, D; E. Gur; Krelin, Y; Shani, N

    2015-01-01

    Mesenchymal stromal cells (MSCs) are multipotent and can be derived from different adult tissues including fat. Our repeated attempts to produce long-term proliferative cultures of rat abdominal adipose stem cells (aASCs) under normal oxygen concentration (21%) were unsuccessful. We set to examine the events controlling this cytostasis of aASCs and found that it resulted from overproduction of reactive oxygen species (ROS) that led to apoptosis. ROS overproduction in aASCs was accompanied by ...

  9. Long-term Smoking Mediated Down-regulation of Smad3 Induces Resistance to Carboplatin in Non-Small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Debangshu Samanta

    2012-07-01

    Full Text Available While numerous cell signaling pathways are known to play decisive roles in chemotherapeutic response, relatively little is known about the impact of the Smad-dependent transforming growth factor β pathway on the therapeutic outcome. Previous reports suggested that patients with lung cancer who continue to smoke while receiving chemotherapy have a poorer outcome than their nonsmoking counterparts do. In our previous study, we showed that long-term cigarette smoke condensate (CSC-mediated down-regulation of Smad3 induces tumorigenesis. The objective of this study was to determine the mechanism of function of Smad3 in chemoresistance induced by CSC in human lung cell lines, namely, A549 and HPL1A. Long-term CSC treatment increases the half-maximal inhibitory concentration (IC50 of carboplatin and makes cells resistant to carboplatin. The increase in IC50 of long-term CSC-treated cells is due to the reduced induction in apoptosis by carboplatin. The increase in IC50 and decrease in apoptosis in long-term CSC-treated cells is correlated with the expression of Bcl2. We have determined that Bcl2 is both necessary and sufficient to make the cells resistant to carboplatin. We have also shown that Smad3 acts upstream to regulate the expression of Bcl2 specifically and, thus, sensitivity of the cells to carboplatin. This is supported by the inverse correlation between the expressions of Smad3 and Bcl2 in human lung tumors. Collectively, these data suggest that loss of Smad3 expression in CSC-treated cells induces resistance to carboplatin by upregulating the expression of Bcl2. This study explains, at least in part, the higher chemoresistance rate observed in smokers.

  10. Radio(chemo)therapy for locally advanced squamous cell carcinoma of the esophagus. Long-term outcome

    Energy Technology Data Exchange (ETDEWEB)

    Ordu, Arif Deniz; Deymann, Lisa Felicia; Scherer, Vera; Combs, Stephanie E. [Technische Universitaet Muenchen, Department of Radiation Oncology, Klinikum rechts der Isar, Muenchen (Germany); Nieder, Carsten [University of Tromsoe, Department of Oncology and Palliative Medicine, Nordland Hospital Trust, Bodoe (Norway); Institute of Clinical Medicine, Faculty of Health Sciences, Tromsoe (Norway); Geinitz, Hans [Technische Universitaet Muenchen, Department of Radiation Oncology, Klinikum rechts der Isar, Muenchen (Germany); Krankenhaus der Barmherzigen Schwestern Linz, Department of Radiation Oncology, Linz (Austria); Kup, Philipp Guenther [Marien Hospital Herne, Universitaetsklinikum der Ruhr-Universitaet Bochum, Department of Radiation Oncology, Herne (Germany); Fakhrian, Khashayar [Technische Universitaet Muenchen, Department of Radiation Oncology, Klinikum rechts der Isar, Muenchen (Germany); Marien Hospital Herne, Universitaetsklinikum der Ruhr-Universitaet Bochum, Department of Radiation Oncology, Herne (Germany); Universitaetsklinikum der Ruhr-Universitaet Bochum, Department of Radiation Oncology, Sankt Josef Hospital Bochum, Bochum (Germany)

    2014-11-18

    The purpose of this work is to report the long-term outcomes of three-dimensional conformal radio(chemo)therapy in the curative management of esophageal squamous cell carcinoma (ESCC). A retrospective analysis of patients treated with radio(chemo)therapy between 1988 and 2011 at Klinikum rechts der Isar, Technische Universitaet Muenchen was performed. In all, 168 patients received radio(chemo)therapy for ESCC in curative intention. The median follow-up time was 91 months (range 1-212 months). There were 128 men and 40 women with a median age of 63 years. Selection criteria for radio(chemo)therapy were unfit for surgery and/or unresectable primary tumor (n = 146, 87 %) or patients' choice (n = 22, 13 %). The majority of the patients received a combination of cisplatin and 5-fluorouracil chemotherapy with 54 Gy in 30 fractions of radiotherapy. The median overall survival (OS) was 20 months (95 % confidence interval 17-23 months). The OS at 2 and 5 years for the whole cohort was 41 ± 4 % and 22 ± 3 %, respectively. Forty patients (24 %) suffered an in-field recurrence. The most common acute nonhematologic toxicity >grade 2 was dysphagia in 35 % of the patients. Acute hematologic toxicity > grade 2 was recorded in 14 % of the patients. There was no grade 5 toxicity observed during the study. Poor ECOG performance status (0-1 vs. 2-3, HR = 1.70, p = 0.002) and weight loss ≥ 10 % before the start of therapy (HR = 1.99, p = 0.001) were among the factors significantly associated with poor OS in multivariate analysis. Three-dimensional conformal definitive radio(chemo)therapy is well tolerated and leads to long-term survival in more than 20 % of patients with advanced disease and/or contraindication to surgery. However, 24 % in-field recurrence remains a major concern. Prospective trials are warranted to assess if a well-tailored conformal radiochemotherapy can improve the local control and obviate the need for surgical resection in patients with good general

  11. Cell-Type-Specific Sensorimotor Processing in Striatal Projection Neurons during Goal-Directed Behavior.

    Science.gov (United States)

    Sippy, Tanya; Lapray, Damien; Crochet, Sylvain; Petersen, Carl C H

    2015-10-21

    Goal-directed sensorimotor transformation drives important aspects of mammalian behavior. The striatum is thought to play a key role in reward-based learning and action selection, receiving glutamatergic sensorimotor signals and dopaminergic reward signals. Here, we obtain whole-cell membrane potential recordings from the dorsolateral striatum of mice trained to lick a reward spout after a whisker deflection. Striatal projection neurons showed strong task-related modulation, with more depolarization and action potential firing on hit trials compared to misses. Direct pathway striatonigral neurons, but not indirect pathway striatopallidal neurons, exhibited a prominent early sensory response. Optogenetic stimulation of direct pathway striatonigral neurons, but not indirect pathway striatopallidal neurons, readily substituted for whisker stimulation evoking a licking response. Our data are consistent with direct pathway striatonigral neurons contributing a "go" signal for goal-directed sensorimotor transformation leading to action initiation. VIDEO ABSTRACT.

  12. Investigation of Neuronal Cell Type-Specific Gene Expression of Ca2+/Calmodulin-dependent Protein Kinase II.

    Directory of Open Access Journals (Sweden)

    Mima Kazuko

    2002-01-01

    Full Text Available The promoter activity of the rat Ca2+/calmodulin-dependent protein kinase II gene was analyzed using the luciferase reporter gene in neuronal and non-neuronal cell lines. Neuronal cell type-specific promoter activity was found in the 5'-flanking region of &agr; and &bgr; isoform genes of the kinase. Silencer elements were also found further upstream of promoter regions. A brain-specific protein bound to the DNA sequence of the 5'-flanking region of the gene was found by gel mobility shift analysis in the nuclear extract of the rat brain, including the cerebellum, forebrain, and brainstem, but not in that of non-neuronal tissues, including liver, kidney and spleen. The luciferase expression system and gel shift analysis can be used as an additional and better index by which to monitor gene expression in most cell types.

  13. Rho-Signaling-Directed YAP/TAZ Activity Underlies the Long-Term Survival and Expansion of Human Embryonic Stem Cells.

    Science.gov (United States)

    Ohgushi, Masatoshi; Minaguchi, Maki; Sasai, Yoshiki

    2015-10-01

    Human embryonic stem cells (hESCs) can survive and proliferate for an extended period of time in culture, but unlike that of tumor-derived cells, this form of cellular immortality does not depend on genomic aberrations. In this study, we sought to elucidate the molecular basis of this long-term growth property of hESCs. We found that the survival of hESCs depends on the small GTPase Rho and its activator AKAP-Lbc. We show that AKAP-Lbc/Rho signaling sustains the nuclear function of the transcriptional cofactors YAP and TAZ by modulating actin microfilament organization. By inducing reprogramming and differentiation, we found that dependency on this Rho signaling pathway is associated with the pluripotent state. Thus, our findings show that the capacity of hESCs to undergo long-term expansion in vitro is intrinsically coupled to their cellular identity through interconnected molecular circuits that link cell survival to pluripotency. PMID:26321201

  14. Cell-type specific oxytocin gene expression from AAV delivered promoter deletion constructs into the rat supraoptic nucleus in vivo.

    Directory of Open Access Journals (Sweden)

    Raymond L Fields

    Full Text Available The magnocellular neurons (MCNs in the hypothalamus selectively express either oxytocin (OXT or vasopressin (AVP neuropeptide genes, a property that defines their phenotypes. Here we examine the molecular basis of this selectivity in the OXT MCNs by stereotaxic microinjections of adeno-associated virus (AAV vectors that contain various OXT gene promoter deletion constructs using EGFP as the reporter into the rat supraoptic nucleus (SON. Two weeks following injection of the AAVs, immunohistochemical assays of EGFP expression from these constructs were done to determine whether the EGFP reporter co-localizes with either the OXT- or AVP-immunoreactivity in the MCNs. The results show that the key elements in the OT gene promoter that regulate the cell-type specific expression the SON are located -216 to -100 bp upstream of the transcription start site. We hypothesize that within this 116 bp domain a repressor exists that inhibits expression specifically in AVP MCNs, thereby leading to the cell-type specific expression of the OXT gene only in the OXT MCNs.

  15. MOBE-ChIP: a large-scale chromatin immunoprecipitation assay for cell type-specific studies.

    Science.gov (United States)

    Lau, On Sun; Bergmann, Dominique C

    2015-10-01

    Cell type-specific transcriptional regulators play critical roles in the generation and maintenance of multicellularity. As they are often expressed at low levels, in vivo DNA-binding studies of these regulators by standard chromatin immunoprecipitation (ChIP) assays are technically challenging. We describe here an optimized ChIP protocol termed Maximized Objects for Better Enrichment (MOBE)-ChIP, which enhances the sensitivity of ChIP assays for detecting cell type-specific signals. The protocol, which is based on the disproportional increase of target signals over background at higher scales, uses substantially greater volume of starting materials than conventional ChIPs to achieve high signal enrichment. This technique can capture weak binding events that are ambiguous in standard ChIP assays, and is useful both in gene-specific and whole-genome analysis. This protocol has been optimized for Arabidopsis, but should be applicable to other model systems with minor modifications. The full procedure can be completed within 3 days.

  16. Long-term immune deficiency after allogeneic stem cell transplantation: B-cell deficiency is associated with late infections

    Science.gov (United States)

    Corre, Elise; Carmagnat, Maryvonnick; Busson, Marc; de Latour, Regis Peffault; Robin, Marie; Ribaud, Patricia; Toubert, Antoine; Rabian, Claire; Socié, Gerard

    2010-01-01

    Immune reconstitution was analyzed in 140 consecutive patients who were 2-year disease-free and who underwent myeloablative allogeneic transplantation. A CD4 and CD8 defect was observed involving naive, terminally differentiated, memory and competent cells and above limits values for activated subsets. Natural killer cells normalize at six months while we observed expansion of CD19+/CD5+ B cells after three months and a persisting defect of memory B cells. Chronic graft-versus-host disease did not influence significantly those parameters for CD8 subsets while the naïve and competent CD4 subsets were strongly affected. But the most profound impact of chronic graft-versus-host disease was on B-cell subsets, especially on the memory B population. The cumulative incidence of late severe infections was low (14% at four years). Using Cox’s models, only low B-cell counts at 12 (P=0.02) and 24 (P=0.001) months were associated with the hazard of developing late infection, in particular if patients did not develop chronic graft-versus-host disease. PMID:20133894

  17. A Single CD8+ T Cell Epitope Sets the Long-Term Latent Load of a Murid Herpesvirus

    OpenAIRE

    Sofia Marques; Marta Alenquer; Stevenson, Philip G.; J. Pedro Simas

    2008-01-01

    Author Summary Persistent viruses present a major challenge to the immune response. Gamma-herpesviruses are a prime example, and the archetypal family member, Epstein-Barr virus (EBV), has been studied for many years. A major unanswered question with EBV is why long-term virus loads—a key pathogenesis outcome—vary so widely between individuals. As most EBV studies are necessarily descriptive, the murid gamma-herpesvirus MuHV-4 provides an important focus of pathogenesis research. Here, we use...

  18. Effect of long-term exposure of SH-SY5Y cells to morphine: a whole cell proteomic analysis

    Directory of Open Access Journals (Sweden)

    Moulédous Lionel

    2006-12-01

    Full Text Available Abstract Background Opiate addiction reflects plastic changes that endurably alter synaptic transmission within relevant neuronal circuits. The biochemical mechanisms of these adaptations remain largely unknown and proteomics-based approaches could lead to a broad characterization of the molecular events underlying adaptations to chronic drug exposure. Results Thus, we have started proteomic analyses of the effects of chronic morphine exposure in a recombinant human neuroblastoma SH-SY5Y clone that stably overexpresses the μ-opioid receptor. Cells were treated with morphine for 6, 24 and 72 hours, the proteins were separated by 2-D gel electrophoresis and stained with Coomassie blue, and the protein map was compared with that obtained from untreated cells. Spots showing a statistically significant variation were selected for identification using mass spectrometric analyses. Conclusion A total of 45 proteins were identified, including proteins involved in cellular metabolism, cytoskeleton organization, vesicular trafficking, transcriptional and translational regulation, and cell signaling.

  19. Long-term detection of fluorescently labeled human mesenchymal stem cell in vitro and in vivo by semi-automated microscopy.

    Science.gov (United States)

    Polzer, Hans; Volkmer, Elias; Saller, Maximilian M; Prall, Wolf C; Haasters, Florian; Drosse, Inga; Anz, David; Mutschler, Wolf; Schieker, Matthias

    2012-02-01

    The use of seeded scaffolds in regenerative medicine is limited by the low survival of transplanted mesenchymal stem cells (MSC). Current approaches aim at improving cell viability but require an adequate long-term detection of the transplanted cells. Unfortunately, commonly performed labeling techniques have not been validated for this purpose, and studies often reveal inconclusive results. Consequently, we intended to identify the most suitable method for long-term detection of human MSC (hMSC) in vitro and in vivo. hMSC were labeled using the vital stainings PKH26 and carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) as well as enhanced green fluorescent protein (eGFP) transduction. Metabolic activity and relative fluorescence intensity (RFI) were quantified in vitro over 21 days at 8 time points using standardized semi-automated microscopy and flow cytometry. In vivo, cell seeded scaffolds were subcutaneously implanted in nude mice, and RFI was analyzed over 42 days at 5 time points. In vitro, PKH26 and CFDA-SE significantly reduced metabolic activity. RFI of both stainings significantly decreased after 1 day and further faded to RFI over the total period of 21 days. In vivo, RFI of eGFP labeled cells reached a plateau phase after 21 days and displayed a 3.8-fold higher RFI compared with PKH26 and CFDA-SE on day 42 evaluated in 280 field of views per scaffold using three scaffolds for each labeling technique and time point. We conclude that PKH26 and CFDA-SE are unsuitable for long-term detection of hMSC. eGFP transduction, in turn, allows long-term detection of hMSC in vitro and in vivo. Our results suggest that eGFP is currently the best option among the fluorescent labeling techniques to follow the fate of transplanted hMSC. PMID:21951128

  20. Long-Term Collections

    CERN Multimedia

    Staff Association

    2016-01-01

    45 years helping in developing countries! CERN personnel have been helping the least fortunate people on the planet since 1971. How? With the Long-Term Collections! Dear Colleagues, The Staff Association’s Long-Term Collections (LTC) Committee is delighted to share this important milestone in the life of our Laboratory with you. Indeed, whilst the name of CERN is known worldwide for scientific discoveries, it also shines in the many humanitarian projects which have been supported by the LTC since 1971. Several schools and clinics, far and wide, carry its logo... Over the past 45 years, 74 projects have been supported (9 of which are still ongoing). This all came from a group of colleagues who wanted to share a little of what life offered them here at CERN, in this haven of mutual understanding, peace and security, with those who were less fortunate elsewhere. Thus, the LTC were born... Since then, we have worked as a team to maintain the dream of these visionaries, with the help of regular donat...

  1. Long-Term Collection

    CERN Multimedia

    Staff Association

    2016-01-01

    Dear Colleagues, As previously announced in Echo (No. 254), your delegates took action to draw attention to the projects of the Long-Term Collections (LTC), the humanitarian body of the CERN Staff Association. On Tuesday, 11 October, at noon, small Z-Cards were widely distributed at the entrances of CERN restaurants and we thank you all for your interest. We hope to have achieved an important part of our goal, which was to inform you, convince you and find new supporters among you. We will find out in the next few days! An exhibition of the LTC was also set up in the Main Building for the entire week. The Staff Association wants to celebrate the occasion of the Long-Term Collection’s 45th anniversary at CERN because, ever since 1971, CERN personnel have showed great support in helping the least fortunate people on the planet in a variety of ways according to their needs. On a regular basis, joint fundraising appeals are made with the Directorate to help the victims of natural disasters around th...

  2. Collectes à long terme

    CERN Document Server

    Collectes à long terme

    2014-01-01

    En cette fin d’année 2014 qui approche à grands pas, le Comité des Collectes à Long Terme remercie chaleureusement ses fidèles donatrices et donateurs réguliers pour leurs contributions à nos actions en faveur des plus démunis de notre planète. C’est très important, pour notre Comité, de pouvoir compter sur l’appui assidu que vous nous apportez. Depuis plus de 40 ans maintenant, le modèle des CLT est basé principalement sur des actions à long terme (soit une aide pendant 4-5 ans par projet, mais plus parfois selon les circonstances), et sa planification demande une grande régularité de ses soutiens financiers. Grand MERCI à vous ! D’autres dons nous parviennent au cours de l’année, et ils sont aussi les bienvenus. En particulier, nous tenons à remercier...

  3. A quantitative comparison of cell-type-specific microarray gene expression profiling methods in the mouse brain.

    Directory of Open Access Journals (Sweden)

    Benjamin W Okaty

    Full Text Available Expression profiling of restricted neural populations using microarrays can facilitate neuronal classification and provide insight into the molecular bases of cellular phenotypes. Due to the formidable heterogeneity of intermixed cell types that make up the brain, isolating cell types prior to microarray processing poses steep technical challenges that have been met in various ways. These methodological differences have the potential to distort cell-type-specific gene expression profiles insofar as they may insufficiently filter out contaminating mRNAs or induce aberrant cellular responses not normally present in vivo. Thus we have compared the repeatability, susceptibility to contamination from off-target cell-types, and evidence for stress-responsive gene expression of five different purification methods--Laser Capture Microdissection (LCM, Translating Ribosome Affinity Purification (TRAP, Immunopanning (PAN, Fluorescence Activated Cell Sorting (FACS, and manual sorting of fluorescently labeled cells (Manual. We found that all methods obtained comparably high levels of repeatability, however, data from LCM and TRAP showed significantly higher levels of contamination than the other methods. While PAN samples showed higher activation of apoptosis-related, stress-related and immediate early genes, samples from FACS and Manual studies, which also require dissociated cells, did not. Given that TRAP targets actively translated mRNAs, whereas other methods target all transcribed mRNAs, observed differences may also reflect translational regulation.

  4. The cytoskeleton of Drosophila-derived Schneider line-1 and Kc23 cells undergoes significant changes during long-term culture

    Science.gov (United States)

    Schatten, H.; Hedrick, J.; Chakrabarti, A.

    1998-01-01

    Insect cell cultures derived from Drosophila melanogaster are increasingly being used as an alternative system to mammalian cell cultures, as they are amenable to genetic manipulation. Although Drosophila cells are an excellent tool for the study of genes and expression of proteins, culture conditions have to be considered in the interpretation of biochemical results. Our studies indicate that significant differences occur in cytoskeletal structure during the long-term culture of the Drosophila-derived cell lines Schneider Line-1 (S1) and Kc23. Scanning, transmission-electron, and immunofluorescence microscopy studies reveal that microfilaments, microtubules, and centrosomes become increasingly different during the culture of these cells from 24 h to 7-14 days. Significant cytoskeletal changes are observed at the cell surface where actin polymerizes into microfilaments, during the elongation of long microvilli. Additionally, long protrusions develop from the cell surface; these protrusions are microtubule-based and establish contact with neighboring cells. In contrast, the microtubule network in the interior of the cells becomes disrupted after four days of culture, resulting in altered transport of mitochondria. Microtubules and centrosomes are also affected in a small percent of cells during cell division, indicating an instability of centrosomes. Thus, the cytoskeletal network of microfilaments, microtubules, and centrosomes is affected in Drosophila cells during long-term culture. This implies that gene regulation and post-translational modifications are probably different under different culture conditions.

  5. Long-term results of a randomized phase III trial of TPF induction chemotherapy followed by surgery and radiation in locally advanced oral squamous cell carcinoma

    OpenAIRE

    Zhong, Lai-ping; Zhang, Chen-Ping; Ren, Guo-xin; Guo, Wei; William, William N.; Hong, Christopher S.; Sun, Jian; ZHU, HAN-GUANG; Tu, Wen-yong; Li, Jiang; Cai, Yi-li; Yin, Qiu-ming; WANG, LI-ZHEN; Wang, Zhong-he; Hu, Yong-jie

    2015-01-01

    Previously, we conducted a randomized phase III trial of TPF (docetaxel, cisplatin, and 5-fluorouracil) induction chemotherapy in surgically managed locally advanced oral squamous cell carcinoma (OSCC) and found no improvement in overall survival. This study reports long-term follow-up results from our initial trial. All patients had clinical stage III or IVA locally advanced OSCC. In the experimental group, patients received two cycles of TPF induction chemotherapy (75mg/m2 docetaxel d1, 75m...

  6. Purkinje cell long-term depression is prevented by T-588, a neuroprotective compound that reduces cytosolic calcium release from intracellular stores

    OpenAIRE

    Kimura, Tatsuo; Sugimori, Mutsuyuki; Llinás, Rodolfo R.

    2005-01-01

    Long-term depression (LTD) of the parallel-fiber (PF) Purkinje synapse induced by four different experimental paradigms could be prevented in rat cerebellar slices by T-588, a neuroprotective compound. The paradigms consisted of pairing PF activation with climbing-fiber activation, direct depolarization, glutamic iontophoretic depolarization, or caffeine. In all cases, LTD was determined by patch-clamp recording of PF excitatory postsynaptic currents at the Purkinje cell somata. T-588 at 1 μM...

  7. Tissue-culture light sheet fluorescence microscopy (TC-LSFM) allows long-term imaging of three-dimensional cell cultures under controlled conditions.

    Science.gov (United States)

    Pampaloni, Francesco; Berge, Ulrich; Marmaras, Anastasios; Horvath, Peter; Kroschewski, Ruth; Stelzer, Ernst H K

    2014-10-01

    Fluorescence long-term imaging of cellular processes in three-dimensional cultures requires the control of media supply, temperature, and pH, as well as minimal photodamage. We describe a system based on a light sheet fluorescence microscope (LSFM), which is optimized for long-term, multi-position imaging of three-dimensional in-gel cell cultures. The system integrates a stable culture condition control system in the optical path of the light-sheet microscope. A further essential element is a biocompatible agarose container suitable for the LSFM, in which any cell type can be cultured in different gel matrices. The TC-LSFM allows studying any in vitro cultured cell type reacting to, dividing in, or migrating through a three-dimensional extracellular matrix (ECM) gel. For this reason we called it "tissue culture-LSFM" (TC-LSFM). The TC-LSFM system allows fast imaging at multiple locations within a millimeter-sized ECM gel. This increases the number of analyzed events and allows testing population effects. As an example, we show the maturation of a cyst of MDCK (canine kidney epithelial) cells over a period of three days. Moreover, we imaged, tracked, and analyzed MDCK cells during the first five days of cell aggregate formation and discovered a remarkable heterogeneity in cell cycle lengths and an interesting cell death pattern. Thus, TC-LSFM allows performing new long-term assays assessing cellular behavior in three-dimensional ECM-gel cultures. For example migration, invasion or differentiation in epithelial cell systems, stem cells, as well as cancer cells can be investigated.

  8. Long-term Temozolomide Treatment Induces Marked Amino Metabolism Modifications and an Increase in TMZ Sensitivity in Hs683 Oligodendroglioma Cells

    Directory of Open Access Journals (Sweden)

    Delphine Lamoral-Theys

    2010-01-01

    Full Text Available Gliomas account for more than 50% of all primary brain tumors. The worst prognosis is associated with gliomas of astrocytic origin, whereas gliomas with an oligodendroglial origin offer higher sensitivity to chemotherapy, especially when oligodendroglioma cells display 1p19q deletions. Temozolomide (TMZ provides therapeutic benefits and is commonly used with radiotherapy in highly malignant astrocytic tumors, including glioblastomas. The actual benefits of TMZ during long-term treatment in oligodendroglioma patients have not yet been clearly defined. In this study, we have investigated the effects of such a long-term TMZ treatment in the unique Hs683 oligodendroglioma model. We have observed increased TMZ sensitivity of Hs683 orthotopic tumors that were previously treated in vitro with months of progressive exposure to increasing TMZ concentrations before being xenografted into the brains of immunocompromised mice. Whole-genome and proteomic analyses have revealed that this increased TMZ sensitivity of Hs683 oligodendroglioma cells previously treated for long periods with TMZ can be explained, at least partly, by a TMZ-induced p38-dependant dormancy state, which in turn resulted in changes in amino acid metabolism balance, in growth delay, and in a decrease in Hs683 oligodendroglioma cell-invasive properties. Thus, long-term TMZ treatment seems beneficial in this Hs683 oligodendroglioma model, which revealed itself unable to develop resistance against TMZ.

  9. Deferiprone versus deferoxamine in sickle cell disease: results from a 5-year long-term Italian multi-center randomized clinical trial.

    Science.gov (United States)

    Calvaruso, Giusi; Vitrano, Angela; Di Maggio, Rosario; Ballas, Samir; Steinberg, Martin H; Rigano, Paolo; Sacco, Massimiliano; Telfer, Paul; Renda, Disma; Barone, Rita; Maggio, Aurelio

    2014-12-01

    Blood transfusion and iron chelation currently represent a supportive therapy to manage anemia, vasculopathy and vaso-occlusion crises in Sickle-Cell-Disease. Here we describe the first 5-year long-term randomized clinical trial comparing Deferiprone versus Deferoxamine in patients with Sickle-Cell-Disease. The results of this study show that Deferiprone has the same effectiveness as Deferoxamine in decreasing body iron burden, measured as repeated measurements of serum ferritin concentrations on the same patient over 5-years and analyzed according to the linear mixed-effects model (LMM) (p=0.822). Both chelators are able to decrease, significantly, serum ferritin concentrations, during 5-years, without any effect on safety (p=0.005). Moreover, although the basal serum ferritin levels were higher in transfused compared with non-transfused group (p=0.031), the changes over time in serum ferritin levels were not statistically significantly different between transfused and non-transfused cohort of patients (p=0.389). Kaplan-Meier curve, during 5-years of study, suggests that Deferiprone does not alter survival in comparison with Deferoxamine (p=0.38). In conclusion, long-term iron chelation therapy with Deferiprone was associated with efficacy and safety similar to that of Deferoxamine. Therefore, in patients with Sickle-Cell-Disease, Deferiprone may represent an effective long-term treatment option. PMID:24814618

  10. Human brain derived cells respond in a type-specific manner after exposure to urban particulate matter (PM).

    Science.gov (United States)

    Campbell, Arezoo; Daher, Nancy; Solaimani, Parrisa; Mendoza, Kriscelle; Sioutas, Constantinos

    2014-10-01

    Exposure to particulate matter (PM), a component of urban air pollution, may cause adverse effects in the brain. Although the exact mechanisms involved are unknown, both oxidative and inflammatory responses have been reported. Since the main route of exposure to particulate matter is through inhalation, there is a potential for compounds to directly enter the brain and alter normal cellular function. Enhancement in both oxidative stress and neuroinflammatory markers has been observed in neurodegenerative disorders and PM-induced potentiation of these events may accelerate the disease process. The objective of this pilot study was to use normal human brain cells, a model system which has not been previously used, to assess cell-type-specific responses after exposure to ultrafine particles (UFP). Human microglia, neurons, and astrocytes were grown separately or as co-cultures and then exposed to aqueous UFP suspensions. Reactive Oxygen Species (ROS) formation and the proinflammatory cytokine tumor necrosis factor alpha (TNF-α) were measured as markers of oxidative stress or inflammation respectively. Our results revealed that after exposure to 2 μg/ml of particles, normal human neurons exhibit a decrease in ROS formation and an increase in TNF-α. The observed decrease in ROS formation persisted in the presence of glial cells, which contrasts previous studies done in rodent cells reporting that PM-induced microglial activation modulates neuronal responses. Our study indicates that human CNS cells may respond differently compared to rodent cells and that their use may be more predictive in risk assessment.

  11. Cell-type-specific tuning of Cav1.3 Ca2+-channels by a C-terminal automodulatory domain

    Science.gov (United States)

    Scharinger, Anja; Eckrich, Stephanie; Vandael, David H.; Schönig, Kai; Koschak, Alexandra; Hecker, Dietmar; Kaur, Gurjot; Lee, Amy; Sah, Anupam; Bartsch, Dusan; Benedetti, Bruno; Lieb, Andreas; Schick, Bernhard; Singewald, Nicolas; Sinnegger-Brauns, Martina J.; Carbone, Emilio; Engel, Jutta; Striessnig, Jörg

    2015-01-01

    Cav1.3 L-type Ca2+-channel function is regulated by a C-terminal automodulatory domain (CTM). It affects channel binding of calmodulin and thereby tunes channel activity by interfering with Ca2+- and voltage-dependent gating. Alternative splicing generates short C-terminal channel variants lacking the CTM resulting in enhanced Ca2+-dependent inactivation and stronger voltage-sensitivity upon heterologous expression. However, the role of this modulatory domain for channel function in its native environment is unkown. To determine its functional significance in vivo, we interrupted the CTM with a hemagglutinin tag in mutant mice (Cav1.3DCRDHA/HA). Using these mice we provide biochemical evidence for the existence of long (CTM-containing) and short (CTM-deficient) Cav1.3 α1-subunits in brain. The long (HA-labeled) Cav1.3 isoform was present in all ribbon synapses of cochlear inner hair cells. CTM-elimination impaired Ca2+-dependent inactivation of Ca2+-currents in hair cells but increased it in chromaffin cells, resulting in hyperpolarized resting potentials and reduced pacemaking. CTM disruption did not affect hearing thresholds. We show that the modulatory function of the CTM is affected by its native environment in different cells and thus occurs in a cell-type specific manner in vivo. It stabilizes gating properties of Cav1.3 channels required for normal electrical excitability. PMID:26379493

  12. Cell-type-specific tuning of Cav1.3 Ca2+-channels by a C-terminal automodulatory domain

    Directory of Open Access Journals (Sweden)

    Anja eScharinger

    2015-08-01

    Full Text Available Cav1.3 L-type Ca2+-channel function is regulated by a C-terminal automodulatory domain (CTM. It affects channel binding of calmodulin and thereby tunes channel activity by interfering with Ca2+- and voltage-dependent gating. Alternative splicing generates short C-terminal channel variants lacking the CTM resulting in enhanced Ca2+-dependent inactivation and stronger voltage-sensitivity upon heterologous expression. However, the role of this modulatory domain for channel function in its native environment is unkown. To determine its functional significance in vivo, we interrupted the CTM with a hemagglutinin tag in mutant mice (Cav1.3DCRDHA/HA. Using these mice we provide biochemical evidence for the existence of long (CTM-containing and short (CTM-deficient Cav1.3 α1-subunits in brain. The long (HA-labeled Cav1.3 isoform was present in all ribbon synapses of cochlear inner hair cells. CTM-elimination impaired Ca2+-dependent inactivation of Ca2+-currents in hair cells but increased it in chromaffin cells, resulting in hyperpolarized resting potentials and reduced pacemaking. CTM disruption did not affect hearing thresholds. We show that the modulatory function of the CTM is affected by its native environment in different cells and thus occurs in a cell-type specific manner in vivo. It is required to stabilize gating properties of Cav1.3 channels required for normal electrical excitability.

  13. Canonical symplectic particle-in-cell method for long-term large-scale simulations of the Vlasov–Maxwell equations

    Energy Technology Data Exchange (ETDEWEB)

    Qin, Hong; Liu, Jian; Xiao, Jianyuan; Zhang, Ruili; He, Yang; Wang, Yulei; Sun, Yajuan; Burby, Joshua W.; Ellison, Leland; Zhou, Yao

    2015-12-14

    Particle-in-cell (PIC) simulation is the most important numerical tool in plasma physics. However, its long-term accuracy has not been established. To overcome this difficulty, we developed a canonical symplectic PIC method for the Vlasov-Maxwell system by discretising its canonical Poisson bracket. A fast local algorithm to solve the symplectic implicit time advance is discovered without root searching or global matrix inversion, enabling applications of the proposed method to very large-scale plasma simulations with many, e.g. 10(9), degrees of freedom. The long-term accuracy and fidelity of the algorithm enables us to numerically confirm Mouhot and Villani's theory and conjecture on nonlinear Landau damping over several orders of magnitude using the PIC method, and to calculate the nonlinear evolution of the reflectivity during the mode conversion process from extraordinary waves to Bernstein waves.

  14. Long Term Culture of the A549 Cancer Cell Line Promotes Multilamellar Body Formation and Differentiation towards an Alveolar Type II Pneumocyte Phenotype

    Science.gov (United States)

    Cooper, James Ross; Abdullatif, Muhammad Bilal; Burnett, Edward C.; Kempsell, Karen E.; Conforti, Franco; Tolley, Howard; Collins, Jane E.; Davies, Donna E.

    2016-01-01

    Pulmonary research requires models that represent the physiology of alveolar epithelium but concerns with reproducibility, consistency and the technical and ethical challenges of using primary or stem cells has resulted in widespread use of continuous cancer or other immortalized cell lines. The A549 ‘alveolar’ cell line has been available for over four decades but there is an inconsistent view as to its suitability as an appropriate model for primary alveolar type II (ATII) cells. Since most work with A549 cells involves short term culture of proliferating cells, we postulated that culture conditions that reduced proliferation of the cancer cells would promote a more differentiated ATII cell phenotype. We examined A549 cell growth in different media over long term culture and then used microarray analysis to investigate temporal regulation of pathways involved in cell cycle and ATII differentiation; we also made comparisons with gene expression in freshly isolated human ATII cells. Analyses indicated that long term culture in Ham’s F12 resulted in substantial modulation of cell cycle genes to result in a quiescent population of cells with significant up-regulation of autophagic, differentiation and lipidogenic pathways. There were also increased numbers of up- and down-regulated genes shared with primary cells suggesting adoption of ATII characteristics and multilamellar body (MLB) development. Subsequent Oil Red-O staining and Transmission Electron Microscopy confirmed MLB expression in the differentiated A549 cells. This work defines a set of conditions for promoting ATII differentiation characteristics in A549 cells that may be advantageous for studies with this cell line. PMID:27792742

  15. Long-term MRI cell tracking after intraventricular delivery in a patient with global cerebral ischemia and prospects for magnetic navigation of stem cells within the CSF.

    Directory of Open Access Journals (Sweden)

    Miroslaw Janowski

    Full Text Available BACKGROUND: The purpose of the study was to evaluate the long-term clinical tracking of magnetically labeled stem cells after intracerebroventricular transplantation as well as to investigate in vitro feasibility for magnetic guidance of cell therapy within large fluid compartments. METHOD: After approval by our Institutional Review Board, an 18-month-old patient, diagnosed as being in a vegetative state due to global cerebral ischemia, underwent cell transplantation to the frontal horn of the lateral ventricle, with umbilical cord blood-derived stem cells labeled with superparamagnetic iron oxide (SPIO contrast agent. The patient was followed over 33 months with clinical examinations and MRI. To evaluate the forces governing the distribution of cells within the fluid compartment of the ventricular system in vivo, a gravity-driven sedimentation assay and a magnetic field-driven cell attraction assay were developed in vitro. RESULTS: Twenty-four hours post-transplantation, MR imaging (MRI was able to detect hypointense cells in the occipital horn of the lateral ventricle. The signal gradually decreased over 4 months and became undetectable at 33 months. In vitro, no significant difference in cell sedimentation between SPIO-labeled and unlabeled cells was observed (p = NS. An external magnet was effective in attracting cells over distances comparable to the size of human lateral ventricles. CONCLUSIONS: MR imaging of SPIO-labeled cells allows monitoring of cells within lateral ventricles. While the initial biodistribution is governed by gravity-driven sedimentation, an external magnetic field may possibly be applied to further direct the distribution of labeled cells within large fluid compartments such as the ventricular system.

  16. Long-term engraftment of p18INK4C-deficient hematopoietic stem cells is enhanced in the sublethally-irradiated recipients

    Institute of Scientific and Technical Information of China (English)

    SONG Xianmin; YUAN Youzhong; SHEN Hongmei; YU Hui; XU Feng; HUANG Paulina; SHIELDS Donna; WANG Jianmin; CHENG Tao

    2006-01-01

    Non-myeloablative regimens for host conditioning have been widely used in clinical hematopoietic stem cell transplantation due to their reduced toxicity on the recipients. But a milder conditioning regimen may require a higher engrafting ability of donor stem cells in competing with endogenous stem cells. Thus, new strategies for enhancing the competitiveness of donor stem cells in non-myeloablative recipients would have important implications for current clinical stem cell trans- plantation. It is known that the absence of p18INK4C (p18) gene can enhance the self-renewal potential of hematopoietic stem cells (HSCs). We applied the approach of competitive bone marrow trans- plantation to evaluate the impact of p18 gene deletion on long-term engraftment of HSCs in sub- lethally irradiated hosts. We found that p18-/- HSCs had a significant advantage over wild-type HSCs during long-term engraftment in the mouse recipients that received a sub-lethal irradiation (5-Gy). The engraftment efficiency of p18-/- HSCs in the sub-lethally irradiated recipients was similar to that in the lethally irradiated (10-Gy) recipients. Our current study demonstrates that enhanced engraftment of donor HSCs in the absence of p18 does not strictly depend on the dose of irradiation used for host conditioning. Therefore, p18 might serve as a potential drug target for increasing the efficacy of stem cell transplant in the patients that are preconditioned with either a myeloablative or non-myeloablative regimen.

  17. Use of autologous human mesenchymal stromal cell/fibrin clot constructs in upper limb non-unions: long-term assessment.

    Directory of Open Access Journals (Sweden)

    Stefano Giannotti

    Full Text Available BACKGROUND: Tissue engineering appears to be an attractive alternative to the traditional approach in the treatment of fracture non-unions. Mesenchymal stromal cells (MSCs are considered an appealing cell source for clinical intervention. However, ex vivo cell expansion and differentiation towards the osteogenic lineage, together with the design of a suitable scaffold have yet to be optimized. Major concerns exist about the safety of MSC-based therapies, including possible abnormal overgrowth and potential cancer evolution. AIMS: We examined the long-term efficacy and safety of ex vivo expanded bone marrow MSCs, embedded in autologous fibrin clots, for the healing of atrophic pseudarthrosis of the upper limb. Our research work relied on three main issues: use of an entirely autologous context (cells, serum for ex vivo cell culture, scaffold components, reduced ex vivo cell expansion, and short-term MSC osteoinduction before implantation. METHODS AND FINDINGS: Bone marrow MSCs isolated from 8 patients were expanded ex vivo until passage 1 and short-term osteo-differentiated in autologous-based culture conditions. Tissue-engineered constructs designed to embed MSCs in autologous fibrin clots were locally implanted with bone grafts, calibrating their number on the extension of bone damage. Radiographic healing was evaluated with short- and long-term follow-ups (range averages: 6.7 and 76.0 months, respectively. All patients recovered limb function, with no evidence of tissue overgrowth or tumor formation. CONCLUSIONS: Our study indicates that highly autologous treatment can be effective and safe in the long-term healing of bone non-unions. This tissue engineering approach resulted in successful clinical and functional outcomes for all patients.

  18. LONG TERM COLLECTIONS

    CERN Multimedia

    STAFF ASSOCIATION

    2010-01-01

    ACKNOWLEDGMENTS The Long-Term Collections (CLT) committee would like to warmly thank its faithful donors who, year after year, support our actions all over the world. Without you, all this would not be possible. We would like to thank, in particular, the CERN Firemen’s Association who donated 5000 CHF in the spring thanks to the sale of their traditional calendar, and the generosity of the CERN community. A huge thank you to the firemen for their devotion to our cause. And thank you to all those who have opened their door, their heart, and their purses! Similarly, we warmly thank the CERN Yoga Club once again for its wonderful donation of 2000 CHF we recently received. We would also like to tell you that all our projects are running well. Just to remind you, we are currently supporting the activities of the «Réflexe-Partage» Association in Mali; the training centre of «Education et Développement» in Abomey, Benin; and the orphanage and ...

  19. Long-Term Quantitative Biodistribution and Side Effects of Human Mesenchymal Stem Cells (hMSCs Engraftment in NOD/SCID Mice following Irradiation

    Directory of Open Access Journals (Sweden)

    Sabine François

    2014-01-01

    Full Text Available There is little information on the fate of infused mesenchymal stem cells (MSCs and long-term side effects after irradiation exposure. We addressed these questions using human MSCs (hMSCs intravenously infused to nonobese diabetic/severe combined immunodeficient (NOD/SCID mice submitted to total body irradiation (TBI or local irradiation (abdominal or leg irradiation. The animals were sacrificed 3 to 120 days after irradiation and the quantitative and spatial distribution of hMSCs were studied by polymerase chain reaction (PCR. Following their infusion into nonirradiated animals, hMSCs homed to various tissues. Engraftment depended on the dose of irradiation and the area exposed. Total body irradiation induced an increased hMSC engraftment level compared to nonirradiated mice, while local irradiations increased hMSC engraftment locally in the area of irradiation. Long-term engraftment of systemically administered hMSCs in NOD/SCID mice increased significantly in response to tissue injuries produced by local or total body irradiation until 2 weeks then slowly decreased depending on organs and the configuration of irradiation. In all cases, no tissue abnormality or abnormal hMSCs proliferation was observed at 120 days after irradiation. This work supports the safe and efficient use of MSCs by injection as an alternative approach in the short- and long-term treatment of severe complications after radiotherapy for patients refractory to conventional treatments.

  20. Balancing intestinal and systemic inflammation through cell type-specific expression of the aryl hydrocarbon receptor repressor.

    Science.gov (United States)

    Brandstätter, Olga; Schanz, Oliver; Vorac, Julia; König, Jessica; Mori, Tetsushi; Maruyama, Toru; Korkowski, Markus; Haarmann-Stemmann, Thomas; von Smolinski, Dorthe; Schultze, Joachim L; Abel, Josef; Esser, Charlotte; Takeyama, Haruko; Weighardt, Heike; Förster, Irmgard

    2016-01-01

    As a sensor of polyaromatic chemicals the aryl hydrocarbon receptor (AhR) exerts an important role in immune regulation besides its requirement for xenobiotic metabolism. Transcriptional activation of AhR target genes is counterregulated by the AhR repressor (AhRR) but the exact function of the AhRR in vivo is currently unknown. We here show that the AhRR is predominantly expressed in immune cells of the skin and intestine, different from other AhR target genes. Whereas AhRR antagonizes the anti-inflammatory function of the AhR in the context of systemic endotoxin shock, AhR and AhRR act in concert to dampen intestinal inflammation. Specifically, AhRR contributes to the maintenance of colonic intraepithelial lymphocytes and prevents excessive IL-1β production and Th17/Tc17 differentiation. In contrast, the AhRR enhances IFN-γ-production by effector T cells in the inflamed gut. Our findings highlight the physiologic importance of cell-type specific balancing of AhR/AhRR expression in response to microbial, nutritional and other environmental stimuli. PMID:27184933

  1. Attempts to predict the long-term decrease in lung function due to radiotherapy of non-small cell lung cancer

    International Nuclear Information System (INIS)

    Purpose: To obtain a model which can predict long-term decrease in lung function due to radiation damage from dose-volume data for patients with non-small cell lung cancer. Patients and methods: 27 patients were included, all long-term survivors after radical radiation therapy. For each patient a regression analysis was performed on a post-RT succession of measurements of FEV1 in order to estimate the decrease after 2 years and a standard error (SE) on this regression estimate. The modelling was based on dose-volume histograms (DVH) exported from the treatment planning system, and involved fits of threshold models, a mean lung dose model as well as more complex models based on the relative damaged volume (rdV). Results: Decreases after 2 years of up to 28% in FEV1 was measured (median 10%), with significant day-to-day variation in FEV1 for the individual patient. The threshold models predicted the long-term decrease in FEV1 well when the SE was interpreted as the uncertainty of the measured decrease. The best threshold value, marginally, was 30 Gy with an R2 of 0.46. The mean lung dose model did not perform so well. A complex model based on rdV performed better than any of the other models (R2 =0.52). Conclusion: The long-term decrease in FEV1 could be predicted from a simple dose-volume model when the SE was interpreted as the uncertainty of the measured decrease

  2. In Vivo Senescence in the Sbds-Deficient Murine Pancreas: Cell-Type Specific Consequences of Translation Insufficiency.

    Science.gov (United States)

    Tourlakis, Marina E; Zhang, Siyi; Ball, Heather L; Gandhi, Rikesh; Liu, Hongrui; Zhong, Jian; Yuan, Julie S; Guidos, Cynthia J; Durie, Peter R; Rommens, Johanna M

    2015-06-01

    Genetic models of ribosome dysfunction show selective organ failure, highlighting a gap in our understanding of cell-type specific responses to translation insufficiency. Translation defects underlie a growing list of inherited and acquired cancer-predisposition syndromes referred to as ribosomopathies. We sought to identify molecular mechanisms underlying organ failure in a recessive ribosomopathy, with particular emphasis on the pancreas, an organ with a high and reiterative requirement for protein synthesis. Biallelic loss of function mutations in SBDS are associated with the ribosomopathy Shwachman-Diamond syndrome, which is typified by pancreatic dysfunction, bone marrow failure, skeletal abnormalities and neurological phenotypes. Targeted disruption of Sbds in the murine pancreas resulted in p53 stabilization early in the postnatal period, specifically in acinar cells. Decreased Myc expression was observed and atrophy of the adult SDS pancreas could be explained by the senescence of acinar cells, characterized by induction of Tgfβ, p15(Ink4b) and components of the senescence-associated secretory program. This is the first report of senescence, a tumour suppression mechanism, in association with SDS or in response to a ribosomopathy. Genetic ablation of p53 largely resolved digestive enzyme synthesis and acinar compartment hypoplasia, but resulted in decreased cell size, a hallmark of decreased translation capacity. Moreover, p53 ablation resulted in expression of acinar dedifferentiation markers and extensive apoptosis. Our findings indicate a protective role for p53 and senescence in response to Sbds ablation in the pancreas. In contrast to the pancreas, the Tgfβ molecular signature was not detected in fetal bone marrow, liver or brain of mouse models with constitutive Sbds ablation. Nevertheless, as observed with the adult pancreas phenotype, disease phenotypes of embryonic tissues, including marked neuronal cell death due to apoptosis, were determined to

  3. Activity-dependent long-term plasticity of afferent synapses on grafted stem/progenitor cell-derived neurons.

    OpenAIRE

    Toft Sörensen, Andreas; Rogelius, Nina; Lundberg, Cecilia; Kokaia, Merab

    2011-01-01

    Stem cell-based cell replacement therapies aiming at restoring injured or diseased brain function ultimately rely on the capability of transplanted cells to promote functional recovery. The mechanisms by which stem cell-based therapies for neurological conditions can lead to functional recovery are uncertain, but structural and functional repair appears to depend on integration of transplanted cell-derived neurons into neuronal circuitries. The nature by which stem/progenitor cell-derived neu...

  4. Combined gene/cell therapies provide long-term and pervasive rescue of multiple pathological symptoms in a murine model of globoid cell leukodystrophy.

    Science.gov (United States)

    Ricca, Alessandra; Rufo, Nicole; Ungari, Silvia; Morena, Francesco; Martino, Sabata; Kulik, Wilem; Alberizzi, Valeria; Bolino, Alessandra; Bianchi, Francesca; Del Carro, Ubaldo; Biffi, Alessandra; Gritti, Angela

    2015-06-15

    Globoid cell leukodystrophy (GLD) is a lysosomal storage disease caused by deficient activity of β-galactocerebrosidase (GALC). The infantile forms manifest with rapid and progressive central and peripheral demyelination, which represent a major hurdle for any treatment approach. We demonstrate here that neonatal lentiviral vector-mediated intracerebral gene therapy (IC GT) or transplantation of GALC-overexpressing neural stem cells (NSC) synergize with bone marrow transplant (BMT) providing dramatic extension of lifespan and global clinical-pathological rescue in a relevant GLD murine model. We show that timely and long-lasting delivery of functional GALC in affected tissues ensured by the exclusive complementary mode of action of the treatments underlies the outstanding benefit. In particular, the contribution of neural stem cell transplantation and IC GT during the early asymptomatic stage of the disease is instrumental to enhance long-term advantage upon BMT. We clarify the input of central nervous system, peripheral nervous system and periphery to the disease, and the relative contribution of treatments to the final therapeutic outcome, with important implications for treatment strategies to be tried in human patients. This study gives proof-of-concept of efficacy, tolerability and clinical relevance of the combined gene/cell therapies proposed here, which may constitute a feasible and effective therapeutic opportunity for children affected by GLD. PMID:25749991

  5. Mouse Pancreas Tissue Slice Culture Facilitates Long-Term Studies of Exocrine and Endocrine Cell Physiology in situ

    OpenAIRE

    Speier, Stephan; Marciniak, Anja; Selck, Claudia; Friedrich, Betty

    2013-01-01

    Studies on pancreatic cell physiology rely on the investigation of exocrine and endocrine cells in vitro. Particularly, in the case of the exocrine tissue these studies have suffered from a reduced functional viability of acinar cells in culture. As a result not only investigations on dispersed acinar cells and isolated acini were limited in their potential, but also prolonged studies on pancreatic exocrine and endocrine cells in an intact pancreatic tissue environment were unfeasible. To ove...

  6. Long-term low-dose α-particle enhanced the potential of malignant transformation in human bronchial epithelial cells through MAPK/Akt pathway

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Weili; Xiao, Linlin; Dong, Chen; He, Mingyuan; Pan, Yan; Xie, Yuexia; Tu, Wenzhi; Fu, Jiamei; Shao, Chunlin, E-mail: clshao@shmu.edu.cn

    2014-05-09

    Highlights: • Multi-exposures of 25 mGy α-ray enhanced cell proliferation, adhesion, and invasion. • MAPK/Akt but not JNK/P66 was positively correlated with cell invasive phenotypes. • LDR of α-irradiation triggers cell malignant transformation through MAPK/Akt. - Abstract: Since the wide usage of ionizing radiation, the cancer risk of low dose radiation (LDR) (<0.1 Gy) has become attractive for a long time. However, most results are derived from epidemiologic studies on atomic-bomb survivors and nuclear accidents surrounding population, and the molecular mechanism of this risk is elusive. To explore the potential of a long-term LDR-induced malignant transformation, human bronchial epithelial cells Beas-2B were fractionally irradiated with 0.025 Gy α-particles for 8 times in total and then further cultured for 1–2 months. It was found that the cell proliferation, the abilities of adhesion and invasion, and the protein expressions of p-ERK, p-Akt, especially p-P38 were not only increased in the multiply-irradiated cells but also in their offspring 1–2 months after the final exposure, indicating high potentiality of cell malignant transformation. On opposite, the expressions of p-JNK and p-P66 were diminished in the subcultures of irradiated cells and thus may play a role of negative regulation in canceration. When the cells were transferred with p38 siRNA, the LDR-induced enhancements of cell adhesion and invasion were significantly reduced. These findings suggest that long-term LDR of α-particles could enhance the potential of malignant transformation incidence in human bronchial epithelial cells through MAPK/Akt pathway.

  7. As-prepared MoS2 quantum dot as a facile fluorescent probe for long-term tracing of live cells.

    Science.gov (United States)

    Zhou, Kai; Zhang, Yue; Xia, Zhining; Wei, Weili

    2016-07-01

    Recently, the newly emerged two-dimensional nanomaterials, layered transition metal dichalcogenide (e.g. MoS2) nanosheets, have drawn tremendous attentions due to their extraordinary electronic and optical properties, and MoS2 quantum dots (MoS2 QDs) with lateral sizes less than 10 nm have been found to be highly luminescent. In the present study, a facile approach for large-scale preparation of MoS2 QDs by Na intercalation reaction without using any toxic organic reagents is proposed. MoS2 QDs were carefully characterized by various techniques including transmission electron microscopy, atomic force microscopy, dynamic light scattering, spectroscopy, in vitro cytotoxicology, and capillary electrophoresis. The as-prepared MoS2 QDs were strongly fluorescent, highly photo-stable, low in cytotoxicity, and readily reactive to thiols. These inherent properties of MoS2 QDs make them excellent fluorescent probes for long-term live cell tracing. The results of live cells imaging indicated that MoS2 QD stained cells remained highly fluorescent after long-term culture, and could be easily traced from other co-cultured cell lines.

  8. As-prepared MoS2 quantum dot as a facile fluorescent probe for long-term tracing of live cells

    Science.gov (United States)

    Zhou, Kai; Zhang, Yue; Xia, Zhining; Wei, Weili

    2016-07-01

    Recently, the newly emerged two-dimensional nanomaterials, layered transition metal dichalcogenide (e.g. MoS2) nanosheets, have drawn tremendous attentions due to their extraordinary electronic and optical properties, and MoS2 quantum dots (MoS2 QDs) with lateral sizes less than 10 nm have been found to be highly luminescent. In the present study, a facile approach for large-scale preparation of MoS2 QDs by Na intercalation reaction without using any toxic organic reagents is proposed. MoS2 QDs were carefully characterized by various techniques including transmission electron microscopy, atomic force microscopy, dynamic light scattering, spectroscopy, in vitro cytotoxicology, and capillary electrophoresis. The as-prepared MoS2 QDs were strongly fluorescent, highly photo-stable, low in cytotoxicity, and readily reactive to thiols. These inherent properties of MoS2 QDs make them excellent fluorescent probes for long-term live cell tracing. The results of live cells imaging indicated that MoS2 QD stained cells remained highly fluorescent after long-term culture, and could be easily traced from other co-cultured cell lines.

  9. Long-Term In Vitro Expansion of Salivary Gland Stem Cells Driven by Wnt Signals

    OpenAIRE

    Maimets, Martti; Rocchi, Cecilia; Bron, Reinier; Pringle, Sarah; Kuipers, Jeroen; Giepmans, Ben N.G.; Robert G.J. Vries; Clevers, Hans; de Haan, Gerald; van Os, Ronald; Coppes, Robert P.

    2015-01-01

    Summary Adult stem cells are the ultimate source for replenishment of salivary gland (SG) tissue. Self-renewal ability of stem cells is dependent on extrinsic niche signals that have not been unraveled for the SG. The ductal compartment in SG has been identified as the location harboring stem cells. Here, we report that rare SG ductal EpCAM+ cells express nuclear β-catenin, indicating active Wnt signaling. In cell culture experiments, EpCAMhigh cells respond potently to Wnt signals stimulatin...

  10. A Two-Component DNA-Prime/Protein-Boost Vaccination Strategy for Eliciting Long-Term, Protective T Cell Immunity against Trypanosoma cruzi

    OpenAIRE

    Shivali Gupta; Garg, Nisha J.

    2015-01-01

    In this study, we evaluated the long-term efficacy of a two-component subunit vaccine against Trypanosoma cruzi infection. C57BL/6 mice were immunized with TcG2/TcG4 vaccine delivered by a DNA-prime/Protein-boost (D/P) approach and challenged with T. cruzi at 120 or 180 days post-vaccination (dpv). We examined whether vaccine-primed T cell immunity was capable of rapid expansion and intercepting the infecting T. cruzi. Our data showed that D/P vaccine elicited CD4+ (30-38%) and CD8+ (22-42%) ...

  11. Autonomous isolation, long-term culture and differentiation potential of adult salivary gland-derived stem/progenitor cells.

    Science.gov (United States)

    Baek, Hyunjung; Noh, Yoo Hun; Lee, Joo Hee; Yeon, Soo-In; Jeong, Jaemin; Kwon, Heechung

    2014-09-01

    Salivary gland stem/progenitor cells belong to the endodermal lineage and may serve as good candidates to replace their dysfunctional counterparts. The objective of this study was to isolate large numbers of salivary gland tissue-derived stem cells (SGSCs) from adult rats in order to develop a clinically applicable method that does not involve sorting or stem cell induction by duct ligation. We analysed SGSCs isolated from normal rat salivary glands to determine whether they retained the major characteristics of stem cells, self-renewal and multipotency, especially with respect to the various endodermal cell types. SGSCs expressed high levels of integrin α6β1 and c-kit, which are surface markers of SGSCs. In particular, the integrin α6β1(+) /c-kit(+) salivary gland cells maintained the morphology, proliferation activity and multipotency of stem cells for up to 92 passages in 12 months. Furthermore, we analysed the capacity of SGSCs to differentiate into endoderm lineage cell types, such as acinar-like and insulin-secreting cells. When cultured on growth factor reduced matrigel, the morphology of progenitor cells changed to acinar-like structures and these cells expressed the acinar cell-specific marker, α-amylase, and tight junction markers. Moreover, reverse transcription-polymerase chain reaction (RT-PCR) data showed increased expression of pancreatic cell markers, including insulin, Pdx1, pan polypeptide and neurogenin-3, when these cells formed pancreatic clusters in the presence of activin A, exendin-4 and retinoic acid. These data demonstrate that adult salivary stem/progenitor cells may serve as a potential source for cell therapy in salivary gland hypofunction and diabetes.

  12. Reversal of morphine-induced cell-type-specific synaptic plasticity in the nucleus accumbens shell blocks reinstatement.

    Science.gov (United States)

    Hearing, Matthew C; Jedynak, Jakub; Ebner, Stephanie R; Ingebretson, Anna; Asp, Anders J; Fischer, Rachel A; Schmidt, Clare; Larson, Erin B; Thomas, Mark John

    2016-01-19

    Drug-evoked plasticity at excitatory synapses on medium spiny neurons (MSNs) of the nucleus accumbens (NAc) drives behavioral adaptations in addiction. MSNs expressing dopamine D1 (D1R-MSN) vs. D2 receptors (D2R-MSN) can exert antagonistic effects in drug-related behaviors, and display distinct alterations in glutamate signaling following repeated exposure to psychostimulants; however, little is known of cell-type-specific plasticity induced by opiates. Here, we find that repeated morphine potentiates excitatory transmission and increases GluA2-lacking AMPA receptor expression in D1R-MSNs, while reducing signaling in D2-MSNs following 10-14 d of forced abstinence. In vivo reversal of this pathophysiology with optogenetic stimulation of infralimbic cortex-accumbens shell (ILC-NAc shell) inputs or treatment with the antibiotic, ceftriaxone, blocked reinstatement of morphine-evoked conditioned place preference. These findings confirm the presence of overlapping and distinct plasticity produced by classes of abused drugs within subpopulations of MSNs that may provide targetable molecular mechanisms for future pharmacotherapies. PMID:26739562

  13. Cell type-specific genes show striking and distinct patterns of spatial expression in the mouse brain.

    Science.gov (United States)

    Ko, Younhee; Ament, Seth A; Eddy, James A; Caballero, Juan; Earls, John C; Hood, Leroy; Price, Nathan D

    2013-02-19

    To characterize gene expression patterns in the regional subdivisions of the mammalian brain, we integrated spatial gene expression patterns from the Allen Brain Atlas for the adult mouse with panels of cell type-specific genes for neurons, astrocytes, and oligodendrocytes from previously published transcriptome profiling experiments. We found that the combined spatial expression patterns of 170 neuron-specific transcripts revealed strikingly clear and symmetrical signatures for most of the brain's major subdivisions. Moreover, the brain expression spatial signatures correspond to anatomical structures and may even reflect developmental ontogeny. Spatial expression profiles of astrocyte- and oligodendrocyte-specific genes also revealed regional differences; these defined fewer regions and were less distinct but still symmetrical in the coronal plane. Follow-up analysis suggested that region-based clustering of neuron-specific genes was related to (i) a combination of individual genes with restricted expression patterns, (ii) region-specific differences in the relative expression of functional groups of genes, and (iii) regional differences in neuronal density. Products from some of these neuron-specific genes are present in peripheral blood, raising the possibility that they could reflect the activities of disease- or injury-perturbed networks and collectively function as biomarkers for clinical disease diagnostics.

  14. DETECTION OF E6, E7 AND CELL-TYPE SPECIFIC ENHANCER OF HUMAN PAPILLOMAVIRUS TYPE 16 IN BREAST CARCINOMA

    Institute of Scientific and Technical Information of China (English)

    HE Qian; CHU Yong-lie; JIA Xiao-li; ZHANG Shu-qun; LIU Wen-kang

    2008-01-01

    Objective To detect HPV16 E6, E7 genes and cell-type specific enhancer (CTSE) of long control region (LCR) in breast carcinoma (BC).Methods HPV16 E6,E7 genes and CTSE were detected in 40 BCs and 20 normal breast tissue (NBT) using polymerase chain reaction (PCR).Results The positive rates of HPV16 E6, E7genes and CTSE were 60% (24/40),55% (22/40) and 67.5%(27/40)respectively in BCs, whereas only 5% (1/20), 5%(1/20) and 15% (3/20) in NBTs (P<0.05). There exited significant correlation between E6 gene and CTSE in BCs (P<0.05), as well as E7 gene and CTSE. The infection of HPV16 E6, E7 and CTSE had no statistic relationship with pathological features.Conclusion There were HPV16 E6, E7 genes and CTSE together in BCs and CTSE may play an important role in pathogenesis of BC.

  15. Long-term T cell memory requires the surface expression of self-peptide/major histocompatibility complex molecules

    OpenAIRE

    Markiewicz, Mary A.; Girao, Cristina; Opferman, Joseph T.; Sun, Jiling; Hu, Qinghui; Agulnik, Alexander A.; Bishop, Colin E.; Thompson, Craig B.; Ashton-Rickardt, Philip G.

    1998-01-01

    How memory T cells are maintained in vivo is poorly understood. To address this problem, a male-specific peptide (H-Y) was identified and used to activate female anti-H-Y T cells in vitro. Anti-H-Y T cells survived in vivo for at least 70 days in the absence of antigen. This persistence was not because of the intrinsic ability of memory T cells to survive in vivo. Instead, the survival and function of adoptively transferred memory cells was found to require transporter of antigen protein 1-de...

  16. The age-dependent epigenetic and physiological changes in an Arabidopsis T87 cell suspension culture during long-term cultivation

    Energy Technology Data Exchange (ETDEWEB)

    Kwiatkowska, Aleksandra, E-mail: A.Kwiatkows@gmail.com [Department of Botany, University of Rzeszow, Kolbuszowa (Poland); Zebrowski, Jacek [Department of Plant Physiology, University of Rzeszow, Kolbuszowa (Poland); Oklejewicz, Bernadetta [Department of Genetics, University of Rzeszow, Kolbuszowa (Poland); Czarnik, Justyna [Department of Botany, University of Rzeszow, Kolbuszowa (Poland); Halibart-Puzio, Joanna [Department of Plant Physiology, University of Rzeszow, Kolbuszowa (Poland); Wnuk, Maciej [Department of Genetics, University of Rzeszow, Kolbuszowa (Poland)

    2014-05-02

    Highlights: • A decrease in proliferation rate during long-term cultivation of Arabidopsis cells. • Age-dependent increase in senescence-associated gene expression in Arabidopsis cells. • Age-related increase in DNA methylation, H3K9me2, and H3K27me3 in Arabidopsis cells. • High potential of photosynthetic efficiency of long-term cultured Arabidopsis cells. - Abstract: Plant cell suspension cultures represent good model systems applicable for both basic research and biotechnological purposes. Nevertheless, it is widely known that a prolonged in vitro cultivation of plant cells is associated with genetic and epigenetic instabilities, which may limit the usefulness of plant lines. In this study, the age-dependent epigenetic and physiological changes in an asynchronous Arabidopsis T87 cell culture were examined. A prolonged cultivation period was found to be correlated with a decrease in the proliferation rate and a simultaneous increase in the expression of senescence-associated genes, indicating that the aging process started at the late growth phase of the culture. In addition, increases in the heterochromatin-specific epigenetic markers, i.e., global DNA methylation, H3K9 dimethylation, and H3K27 trimethylation, were observed, suggesting the onset of chromatin condensation, a hallmark of the early stages of plant senescence. Although the number of live cells decreased with an increase in the age of the culture, the remaining viable cells retained a high potential to efficiently perform photosynthesis and did not exhibit any symptoms of photosystem II damage.

  17. Role of morphology and crystallinity of nanorod and planar electron transport layers on the performance and long term durability of perovskite solar cells

    Science.gov (United States)

    Fakharuddin, Azhar; Di Giacomo, Francesco; Ahmed, Irfan; Wali, Qamar; Brown, Thomas M.; Jose, Rajan

    2015-06-01

    High efficiency is routinely reported in CH3NH3PbI3-xClx sensitized mesoscopic solar cells (PSCs) employing planar and scaffold architectures; however, a systematic comparison of their photovoltaic performance under similar experimental conditions and their long term stability have so far not been discussed. In this paper, we compare the performance and durability of PSCs employing these two device configurations and conclude that although a planar architecture routinely provides high initial photoconversion efficiency (PCE), particularly high open-circuit voltage (VOC), a scaffold is crucial to achieve long term durable performance of such devices. In a comparative study of scaffold (rutile nanorods, NRs) vs. planar devices, the efficiency in latter dropped off by one order of magnitude in ∼300 h despite their similar initial PCE of ∼12%. We compared the performance and the durability of two types of scaffolds, i.e., pristine and TiCl4 treated NRs, and observed that the pristine NRs showed >10% improvement in the PCE after ∼1300 h whereas the cells employing post-treated NR scaffold retained ∼60% of initial value. We address the origin of the different photovoltaic performance of planar and scaffold devices in the context of photoanode morphology and its possible effect on the cell durability.

  18. Long-term clinical results of autologous bone marrow CD 133+ cell transplantation in patients with ST-elevation myocardial infarction

    Science.gov (United States)

    Kirgizova, M. A.; Suslova, T. E.; Markov, V. A.; Karpov, R. S.; Ryabov, V. V.

    2015-11-01

    The aim of the study was investigate the long-term results of autologous bone marrow CD 133+ cell transplantation in patients with primary ST-Elevation Myocardial Infarction (STEMI). Methods and results: From 2006 to 2007, 26 patients with primary STEMI were included in an open randomized study. Patients were randomized to two groups: 1st - included patients underwent PCI and transplantation of autologous bone marrow CD 133+ cell (n = 10); 2nd - patients with only PCI (n = 16). Follow-up study was performed 7.70±0.42 years after STEMI and consisted in physical examination, 6-min walking test, Echo exam. Total and cardiovascular mortality in group 1 was lower (20% (n = 2) vs. 44% (n = 7), p = 0.1 and 22% (n = 2) vs. 25% (n = 4), (p=0.53), respectively). Analysis of cardiac volumetric parameters shows significant differences between groups: EDV of 100.7 ± 50.2 mL vs. 144.40±42.7 mL, ESV of 56.3 ± 37.8 mL vs. 89.7 ± 38.7 mL in 1st and 2nd groups, respectively. Data of the study showed positive effects of autologous bone marrow CD 133+ cell transplantation on the long-term survival of patients and structural status of the heart.

  19. Effects of Long-term exposure of Gelatinated and Non-gelatinated Cadmium Telluride Quantum Dots on Differentiated PC12 cells

    LENUS (Irish Health Repository)

    Prasad, Babu R

    2012-01-20

    Abstract Background The inherent toxicity of unmodified Quantum Dots (QDs) is a major hindrance to their use in biological applications. To make them more potent as neuroprosthetic and neurotherapeutic agents, thioglycolic acid (TGA) capped CdTe QDs, were coated with a gelatine layer and investigated in this study with differentiated pheochromocytoma 12 (PC12) cells. The QD - cell interactions were investigated after incubation periods of up to 17 days by MTT and APOTOX-Glo Triplex assays along with using confocal microscopy. Results Long term exposure (up to 17 days) to gelatinated TGA-capped CdTe QDs of PC12 cells in the course of differentiation and after neurites were grown resulted in dramatically reduced cytotoxicity compared to non-gelatinated TGA-capped CdTe QDs. Conclusion The toxicity mechanism of QDs was identified as caspase-mediated apoptosis as a result of cadmium leaking from the core of QDs. It was therefore concluded that the gelatine capping on the surface of QDs acts as a barrier towards the leaking of toxic ions from the core QDs in the long term (up to 17 days).

  20. High-throughput sequencing reveals miRNA effects on the primary and secondary production properties in long-term subcultured Taxus cells.

    Science.gov (United States)

    Zhang, Meng; Dong, Yanshan; Nie, Lin; Lu, Mingbo; Fu, Chunhua; Yu, Longjiang

    2015-01-01

    Plant-cell culture technology is a promising alternative for production of high-value secondary metabolites but is limited by the decreased metabolite production after long-term subculture. The goal of this study was to determine the effects of miRNAs on altered gene expression profiles during long-term subculture. Two Taxus cell lines, CA (subcultured for 10 years) and NA (subcultured for 6 months), were high-throughput sequenced at the mRNA and miRNA levels. A total of 265 known (78.87% of 336) and 221 novel (79.78% of 277) miRNAs were differentially expressed. Furthermore, 67.17% of the known differentially expressed (DE) miRNAs (178) and 60.63% of the novel DE-miRNAs (134) were upregulated in NA. A total of 275 inverse-related miRNA/mRNA modules were identified by target prediction analysis. Functional annotation of the targets revealed that the high-ranking miRNA targets were those implicated in primary metabolism and abiotic or biotic signal transduction. For example, various genes for starch metabolism and oxidative phosphorylation were inversely related to the miRNA levels, thereby indicating that miRNAs have important roles in these pathways. Interestingly, only a few genes for secondary metabolism were inversely related to miRNA, thereby indicating that factors other than miRNA are present in the regulatory system. Moreover, miR8154 and miR5298b were upregulated miRNAs that targeted a mass of DE genes. The overexpression of these miRNAs in CA increased the genes of taxol, phenylpropanoid, and flavonoid biosynthesis, thereby suggesting their function as crucial factors that regulate the entire metabolic network during long-term subculture. Our current studies indicated that a positive conversion of production properties from secondary metabolism to primary metabolism occurred in long-term subcultured cells. miRNAs are important regulators in the upregulation of primary metabolism.

  1. Comparative Analysis of KnockOut™ Serum with Fetal Bovine Serum for the In Vitro Long-Term Culture of Human Limbal Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Shaokun Zhang

    2016-01-01

    Full Text Available The limbal epithelial cells can be maintained on 3T3 feeder layer with fetal bovine serum supplemented culture medium, and these cells have been used to successfully treat limbal stem cell deficiency. However, fetal bovine serum contains unknown components and displays quantitative and qualitative lot-to-lot variations. To improve the culture condition, the defined KnockOut serum replacement was investigated to replace fetal bovine serum for culturing human limbal epithelial cell. Human primary limbal epithelial cells were cultured in KnockOut serum and fetal bovine serum supplemented medium, respectively. The cell growth rate, gene expression, and maintenance of limbal epithelial stem cells were studied and compared between these two groups. Human primary limbal epithelial cells were isolated and successfully serially cultivated in this novel KnockOut serum supplemented medium; the cell proliferation and stem cell maintenance were similar to those of cells grown in fetal bovine serum supplemented medium. These data suggests that this KnockOut serum supplemented medium is an efficient replacement to traditional fetal bovine serum supplemented medium for limbal epithelial cell culture, and this medium has great potential for long term maintenance of limbal epithelial cells, limbal epithelial stem cells transplantation, and tissue regeneration.

  2. The differential short- and long-term effects of HIV-1 latency-reversing agents on T cell function.

    Science.gov (United States)

    Clutton, G; Xu, Y; Baldoni, P L; Mollan, K R; Kirchherr, J; Newhard, W; Cox, Kara; Kuruc, J D; Kashuba, A; Barnard, R; Archin, N; Gay, C L; Hudgens, M G; Margolis, D M; Goonetilleke, N

    2016-01-01

    Despite the extraordinary success of HIV-1 antiretroviral therapy in prolonging life, infected individuals face lifelong therapy because of a reservoir of latently-infected cells that harbor replication competent virus. Recently, compounds have been identified that can reverse HIV-1 latency in vivo. These latency- reversing agents (LRAs) could make latently-infected cells vulnerable to clearance by immune cells, including cytolytic CD8+ T cells. We investigated the effects of two leading LRA classes on CD8+ T cell phenotype and function: the histone deacetylase inhibitors (HDACis) and protein kinase C modulators (PKCms). We observed that relative to HDACis, the PKCms induced much stronger T cell activation coupled with non-specific cytokine production and T cell proliferation. When examining antigen-specific CD8+ T cell function, all the LRAs except the HDACi Vorinostat reduced, but did not abolish, one or more measurements of CD8+ T cell function. Importantly, the extent and timing of these effects differed between LRAs. Panobinostat had detrimental effects within 10 hours of drug treatment, whereas the effects of the other LRAs were observed between 48 hours and 5 days. These observations suggest that scheduling of LRA and CD8+ T cell immunotherapy regimens may be critical for optimal clearance of the HIV-1 reservoir. PMID:27480951

  3. [{sup 89}Zr]Oxinate{sub 4} for long-term in vivo cell tracking by positron emission tomography

    Energy Technology Data Exchange (ETDEWEB)

    Charoenphun, Putthiporn; Meszaros, Levente K.; Chuamsaamarkkee, Krisanat; Sharif-Paghaleh, Ehsan; Ballinger, James R.; Mullen, Gregory E.D. [St Thomas' Hospital, King' s College London, Division of Imaging Sciences and Biomedical Engineering, London (United Kingdom); Ferris, Trevor J.; Went, Michael J. [University of Kent, School of Physical Sciences, Canterbury (United Kingdom); Blower, Philip J. [St Thomas' Hospital, King' s College London, Division of Imaging Sciences and Biomedical Engineering, London (United Kingdom); King' s College London, Division of Chemistry, London (United Kingdom)

    2014-10-31

    {sup 111}In (typically as [{sup 111}In]oxinate{sub 3}) is a gold standard radiolabel for cell tracking in humans by scintigraphy. A long half-life positron-emitting radiolabel to serve the same purpose using positron emission tomography (PET) has long been sought. We aimed to develop an {sup 89}Zr PET tracer for cell labelling and compare it with [{sup 111}In]oxinate{sub 3} single photon emission computed tomography (SPECT). [{sup 89}Zr]Oxinate{sub 4} was synthesised and its uptake and efflux were measured in vitro in three cell lines and in human leukocytes. The in vivo biodistribution of eGFP-5T33 murine myeloma cells labelled using [{sup 89}Zr]oxinate{sub 4} or [{sup 111}In]oxinate{sub 3} was monitored for up to 14 days. {sup 89}Zr retention by living radiolabelled eGFP-positive cells in vivo was monitored by FACS sorting of liver, spleen and bone marrow cells followed by gamma counting. Zr labelling was effective in all cell types with yields comparable with {sup 111}In labelling. Retention of {sup 89}Zr in cells in vitro after 24 h was significantly better (range 71 to >90 %) than {sup 111}In (43-52 %). eGFP-5T33 cells in vivo showed the same early biodistribution whether labelled with {sup 111}In or {sup 89}Zr (initial pulmonary accumulation followed by migration to liver, spleen and bone marrow), but later translocation of radioactivity to kidneys was much greater for {sup 111}In. In liver, spleen and bone marrow at least 92 % of {sup 89}Zr remained associated with eGFP-positive cells after 7 days in vivo. [{sup 89}Zr]Oxinate{sub 4} offers a potential solution to the emerging need for a long half-life PET tracer for cell tracking in vivo and deserves further evaluation of its effects on survival and behaviour of different cell types. (orig.)

  4. TLR-2 Activation Induces Regulatory T Cells and Long-Term Suppression of Asthma Manifestations in Mice

    OpenAIRE

    Nawijn, Martijn C.; Motta, Alexandre C; Renée Gras; Soheila Shirinbak; Hadi Maazi; van Oosterhout, Antoon J. M.

    2013-01-01

    Asthma is a chronic inflammatory disease of the airways characterized by variable airway obstruction and airway hyperresponsiveness (AHR). The T regulatory (Treg) cell subset is critically important for the regulation of immune responses. Adoptive transfer of Treg cells has been shown to be sufficient for the suppression of airway inflammation in experimental allergic asthma. Intervention strategies aimed at expanding the Treg cell population locally in the airways of sensitized individuals a...

  5. Hematopoietic microenvironment. Origin, lineage, and transplantability of the stromal cells in long-term bone marrow cultures from chimeric mice.

    OpenAIRE

    Perkins, S; Fleischman, R A

    1988-01-01

    Studies of bone marrow transplant patients have suggested that the stromal cells of the in vitro hematopoietic microenvironment are transplantable into conditioned recipients. Moreover, in patients with myeloproliferative disorders, all of the stromal cells, which include presumptive endothelial cells, appear to be derived from hematopoietic precursors. To confirm these findings, we have constructed two chimeric mouse models: (a) traditional radiation chimeras, and (b) fetal chimeras, produce...

  6. Long-term outcomes among older patients following nonmyeloablative conditioning and allogeneic hematopoietic cell transplantation for advanced hematologic malignancies

    DEFF Research Database (Denmark)

    Sorror, Mohamed L; Sandmaier, Brenda M; Storer, Barry E;

    2011-01-01

    A minimally toxic nonmyeloablative regimen was developed for allogeneic hematopoietic cell transplantation (HCT) to treat patients with advanced hematologic malignancies who are older or have comorbid conditions....

  7. Bovine colostrum supports the serum-free proliferation of epithelial cells but not of fibroblasts in long-term culture

    OpenAIRE

    Klagsbrun, M

    1980-01-01

    Medium lacking serum but supplemented with milk will support the growth of sparse cells in culture. Milk obtained within 8 h after the birth of a calf (day 1 colostrum) is the most effective in supporting proliferation. In mixed cultures of early-passage bovine embryonic kidney (BEK) or early-passage calf kidney (CK) cells, both epithelial cells and fibroblasts grow in Dulbecco’s modified eagle’s medium (DMEM) supplemented with serum. However, only cells that appear to be epithelial-like grow...

  8. Long-term in-vivo tumorigenic assessment of human culture-expanded adipose stromal/stem cells

    International Nuclear Information System (INIS)

    After more than a decade of extensive experimentation, the promise of stem cells to revolutionize the field of medicine has negotiated their entry into clinical trial. Adipose tissue specifically holds potential as an attainable and abundant source of stem cells. Currently undergoing investigation are adipose stem cell (ASC) therapies for diabetes and critical limb ischemia, among others. In the enthusiastic pursuit of regenerative therapies, however, questions remain regarding ASC persistence and migration, and, importantly, their safety and potential for neoplasia. To date, assays of in vivo ASC activity have been limited by early end points. We hypothesized that with time, ASCs injected subcutaneously undergo removal by normal tissue turnover and homeostasis, and by the host's immune system. In this study, a high dose of culture expanded ASCs was formulated and implanted as multicellular aggregates into immunocompromised mice, which were maintained for over one year. Animals were monitored for toxicity, and surviving cells quantified at study endpoint. No difference in growth/weight or lifespan was found between cell-treated and vehicle treated animals, and no malignancies were detected in treated animals. Moreover, real-time PCR for a human specific sequence, ERV-3, detected no persistent ASCs. With the advent of clinical application, clarification of currently enigmatic stem cell properties has become imperative. Our study represents the longest duration determination of stem cell activity in vivo, and contributes strong evidence in support of the safety of adipose derived stem cell applications. -- Highlights: ► Adipose stem cells promise novel clinical therapies. ► Before clinical translation, safety profiles must be further elucidated. ► Subcutaneously injected non-autologous adipose stem cells do not form tumors. ► Subcutaneously injected non-autologous adipose stem cells undergo complete removal by one year.

  9. Long-term in-vivo tumorigenic assessment of human culture-expanded adipose stromal/stem cells

    Energy Technology Data Exchange (ETDEWEB)

    MacIsaac, Zoe Marie, E-mail: zmm4a@virgina.edu [University of Virginia (United States); Shang, Hulan, E-mail: shanghulan@gmail.com [Department of Plastic Surgery, University of Virginia (United States); Agrawal, Hitesh, E-mail: hiteshdos@hotmail.com [Department of Plastic Surgery, University of Virginia (United States); Yang, Ning, E-mail: ny6u@virgina.edu [Department of Plastic Surgery, University of Virginia (United States); Parker, Anna, E-mail: amp4v@virginia.edu [Department of Surgery, University of Virginia (United States); Katz, Adam J., E-mail: ajk2f@virginia.edu [Department of Plastic Surgery, University of Virginia (United States)

    2012-02-15

    After more than a decade of extensive experimentation, the promise of stem cells to revolutionize the field of medicine has negotiated their entry into clinical trial. Adipose tissue specifically holds potential as an attainable and abundant source of stem cells. Currently undergoing investigation are adipose stem cell (ASC) therapies for diabetes and critical limb ischemia, among others. In the enthusiastic pursuit of regenerative therapies, however, questions remain regarding ASC persistence and migration, and, importantly, their safety and potential for neoplasia. To date, assays of in vivo ASC activity have been limited by early end points. We hypothesized that with time, ASCs injected subcutaneously undergo removal by normal tissue turnover and homeostasis, and by the host's immune system. In this study, a high dose of culture expanded ASCs was formulated and implanted as multicellular aggregates into immunocompromised mice, which were maintained for over one year. Animals were monitored for toxicity, and surviving cells quantified at study endpoint. No difference in growth/weight or lifespan was found between cell-treated and vehicle treated animals, and no malignancies were detected in treated animals. Moreover, real-time PCR for a human specific sequence, ERV-3, detected no persistent ASCs. With the advent of clinical application, clarification of currently enigmatic stem cell properties has become imperative. Our study represents the longest duration determination of stem cell activity in vivo, and contributes strong evidence in support of the safety of adipose derived stem cell applications. -- Highlights: Black-Right-Pointing-Pointer Adipose stem cells promise novel clinical therapies. Black-Right-Pointing-Pointer Before clinical translation, safety profiles must be further elucidated. Black-Right-Pointing-Pointer Subcutaneously injected non-autologous adipose stem cells do not form tumors. Black-Right-Pointing-Pointer Subcutaneously injected non

  10. The predominant mechanism of intercellular calcium wave propagation changes during long-term culture of human osteoblast-like cells

    DEFF Research Database (Denmark)

    Henriksen, Zanne; Hiken, Jeffrey F; Steinberg, Thomas H;

    2006-01-01

    , but as cells differentiate in culture, gap-junction-mediated ICW become more prominent. These results suggest that P2Y receptor-mediated and gap junction-mediated mechanisms of intercellular calcium signaling may play different roles during differentiation of bone-forming cells....

  11. TLR-2 Activation Induces Regulatory T Cells and Long-Term Suppression of Asthma Manifestations in Mice

    NARCIS (Netherlands)

    Nawijn, Martijn C.; Motta, Alexandre C.; Gras, Renee; Shirinbak, Soheila; Maazi, Hadi; van Oosterhout, Antoon J. M.

    2013-01-01

    Asthma is a chronic inflammatory disease of the airways characterized by variable airway obstruction and airway hyperresponsiveness (AHR). The T regulatory (Treg) cell subset is critically important for the regulation of immune responses. Adoptive transfer of Treg cells has been shown to be sufficie

  12. Bovine colostrum supports the serum-free proliferation of epithelial cells but not of fibroblasts in long-term culture

    Science.gov (United States)

    Klagsbrun, M

    1980-01-01

    Medium lacking serum but supplemented with milk will support the growth of sparse cells in culture. Milk obtained within 8 h after the birth of a calf (day 1 colostrum) is the most effective in supporting proliferation. In mixed cultures of early-passage bovine embryonic kidney (BEK) or early-passage calf kidney (CK) cells, both epithelial cells and fibroblasts grow in Dulbecco’s modified eagle’s medium (DMEM) supplemented with serum. However, only cells that appear to be epithelial-like grow in DMEM supplemented with colostrum. Sparse cultures of early-passage human and rat fibroblasts that grow readily in DMEM supplemented with serum do not grow in DMEM supplemented with colostrum. Canine kidney epithelial cells (MDCK), when plated sparsely, grow exponentially in DMEM supplemented with day 1 bovine colostrum. The generation time is 26 h, the same growth rate as in DMEM supplemented with calf serum. The MDCK cells can be subcultured and regrown to confluence repeatedly in colostrum-supplemented DMEM. Growth in DMEM supplemented with colostrum does not alter the morphological characteristics of the MDCK cells, which are polygonal, contain microvilli at the apical surface, and are connected by tight junctions and desmosomes. MDCK cells do not proliferate in DMEM supplemented with milk obtained 1 wk after the birth of a calf. PMID:7358799

  13. Thin-Film Photovoltaic Cells: Long-Term Metal(loid) Leaching at Their End-of-Life

    NARCIS (Netherlands)

    Zimmermann, Y.S.; Schäffer, A.; Corvini, P.F.X.; Lenz, M.

    2013-01-01

    The photovoltaic effect of thin-film copper indium gallium selenide cells (CIGS) is conferred by the latter elements. Organic photovoltaic cells (OPV), relying on organic light-absorbing molecules, also contain a variety of metals (e.g., Zn, Al, In, Sn, Ag). The environmental impact of such technolo

  14. Effects of long-term administration of cancer-promoting substances on oral subepithelial mast cells in the rat.

    Science.gov (United States)

    Sand, L; Hilliges, M; Larsson, P A; Wallstrom, M; Hirsch, J M

    2002-01-01

    The role of oral subepithelial mast cells in the defence against tumours is a matter of controversy. The effect of established and suggested carcinogens, such as the carcinogen 4-nitroquinoline-N-oxide (4-NQO) and Herpes simplex virus type 1 (HSV-1), in combination with oral snuff on lower lip subepithelial mast cells (MC) was studied in rats. The rats were exposed to prolonged use of oral snuff. The test substances were administered in a surgically created canal in the lower lip of the rats. There were 15 rats in each test group and 10 rats in the control group. The amount of countable subepithelial mast cells decreased significantly when the rat oral mucosa was exposed to the oral carcinogen 4-NQO but the effect of oral snuff and HSV-1 infection was weak. Our findings suggest that mast cells play a role in immunological cell defence against chemical carcinogens. Further studies are needed to clarify the mechanisms. PMID:12529973

  15. Quantitative analysis of the acute and long-term CD4(+) T-cell response to a persistent gammaherpesvirus

    DEFF Research Database (Denmark)

    Christensen, Jan Pravsgaard; Doherty, P C

    1999-01-01

    The murine gammaherpesvirus 68 (MHV-68) replicates in respiratory epithelial cells, where it establishes a persistent, latent infection limited predominantly to B lymphocytes. The virus-specific CD4(+) T-cell response in C57BL/6 mice challenged intranasally with MHV-68 is detected first in the me......The murine gammaherpesvirus 68 (MHV-68) replicates in respiratory epithelial cells, where it establishes a persistent, latent infection limited predominantly to B lymphocytes. The virus-specific CD4(+) T-cell response in C57BL/6 mice challenged intranasally with MHV-68 is detected first...... were initially CD62Llow, with >80% maintaining that phenotype for the next 14 months. The overall conclusion is that MHV-68-specific CD4(+) T cells remain activated (CD62Llow) and at a stable frequency in the face of persistent infection....

  16. Long term culture of mesenchymal stem cells in hypoxia promotes a genetic program maintaining their undifferentiated and multipotent status

    Directory of Open Access Journals (Sweden)

    de Carvalho Marcelo

    2011-03-01

    Full Text Available Abstract Background In the bone marrow, hematopietic and mesenchymal stem cells form a unique niche in which the oxygen tension is low. Hypoxia may have a role in maintaining stem cell fate, self renewal and multipotency. However, whereas most studies addressed the effect of transient in vitro exposure of MSC to hypoxia, permanent culture under hypoxia should reflect the better physiological conditions. Results Morphologic studies, differentiation and transcriptional profiling experiments were performed on MSC cultured in normoxia (21% O2 versus hypoxia (5% O2 for up to passage 2. Cells at passage 0 and at passage 2 were compared, and those at passage 0 in hypoxia generated fewer and smaller colonies than in normoxia. In parallel, MSC displayed (>4 fold inhibition of genes involved in DNA metabolism, cell cycle progression and chromosome cohesion whereas transcripts involved in adhesion and metabolism (CD93, ESAM, VWF, PLVAP, ANGPT2, LEP, TCF1 were stimulated. Compared to normoxic cells, hypoxic cells were morphologically undifferentiated and contained less mitochondrias. After this lag phase, cells at passage 2 in hypoxia outgrew the cells cultured in normoxia and displayed an enhanced expression of genes (4-60 fold involved in extracellular matrix assembly (SMOC2, neural and muscle development (NOG, GPR56, SNTG2, LAMA and epithelial development (DMKN. This group described herein for the first time was assigned by the Gene Ontology program to "plasticity". Conclusion The duration of hypoxemia is a critical parameter in the differentiation capacity of MSC. Even in growth promoting conditions, hypoxia enhanced a genetic program that maintained the cells undifferentiated and multipotent. This condition may better reflect the in vivo gene signature of MSC, with potential implications in regenerative medicine.

  17. Expression of Gast, Cckbr, Reg1α genes in rat duodenal epithelial cells upon long-term gastric hypoacidity and after a multiprobiotic administration

    Directory of Open Access Journals (Sweden)

    Dranitsina A. S.

    2014-11-01

    Full Text Available Aim. Determination of the Cckbr, Gast and Reg1α genes expression in rat duodenal epithelial cells upon long- term hypoacidity and with the administration of the multiprobiotic Symbiter. Methods. The experiments were carried out on white non-strain male rats. The hypoacidic state was induced through intraperitoneal injection of omeprazole for 28 days. The level of genes expression was determined by semi-quantitative analysis with RT-PCR Results. The elevation of mRNA levels of the Cckbr and Gast genes in rat duodenal villus and crypt epitheliocytes, the increased expression of the Reg1A gene in crypt epithelial cells were shown as well as the appearance of the Reg1a gene expression in villus epitheliocytes upon hypoacidic conditions were shown. The content of mRNAs of the above mentioned genes decreased or remained at the control level upon the treatment of hypoacidic rats with the multiprobiotic Symbiter. Conclusions. Long-term gastric hypoacidity is accompanied by the changes in expression of the Cckbr, Gast and Reg1a genes in rat duodenum, whereas upon administration of the multiprobiotic Symbiter the pattern of studied gene expression did not changed in the most cases.

  18. Long-term delivery of nerve growth factor by encapsulated cell biodelivery in the Göttingen minipig basal forebrain

    DEFF Research Database (Denmark)

    Fjord-Larsen, L; Kusk, P; Tornøe, Jens;

    2010-01-01

    Nerve growth factor (NGF) prevents cholinergic degeneration in Alzheimer's disease (AD) and improves memory in AD animal models. In humans, the safe delivery of therapeutic doses of NGF is challenging. For clinical use, we have therefore developed an encapsulated cell (EC) biodelivery device......, capable of local delivery of NGF. The clinical device, named NsG0202, houses an NGF-secreting cell line (NGC-0295), which is derived from a human retinal pigment epithelial (RPE) cell line, stably genetically modified to secrete NGF. Bioactivity and correct processing of NGF was confirmed in vitro. Ns...

  19. Long-term MRI tracking of dual-labeled adipose-derived stem cells homing into mouse carotid artery injury

    Directory of Open Access Journals (Sweden)

    Qin JB

    2012-10-01

    Full Text Available Jin-Bao Qin,1,5,* Kang-An Li,2,* Xiang-Xiang Li,1,5 Qing-Song Xie,3 Jia-Ying Lin,4 Kai-Chuang Ye,1,5 Mi-Er Jiang,1,5 Gui-Xiang Zhang,2 Xin-Wu Lu1,51Department of Vascular Surgery, Shanghai Ninth People's Hospital Affiliated to Shanghai Jiao Tong University, School of Medicine, 2Department of Radiology, Shanghai First People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 3Department of Neurosurgery, Cixi Municipal People's Hospital, Zhejiang Province, China; 4Clinic for Gynecology, Charite-Universitatsmedizin Berlin, Berlin, Germany; 5Vascular Center, Shanghai Jiao Tong University, Shanghai, China*These two authors contributed equally to this workBackground: Stem cell therapy has shown great promise for regenerative repair of injured or diseased tissues. Adipose-derived stem cells (ADSCs have become increasingly attractive candidates for cellular therapy. Magnetic resonance imaging has been proven to be effective in tracking magnetic-labeled cells and evaluating their clinical relevance after cell transplantation. This study investigated the feasibility of imaging green fluorescent protein-expressing ADSCs (GFP-ADSCs labeled with superparamagnetic iron oxide particles, and tracked them in vivo with noninvasive magnetic resonance imaging after cell transplantation in a model of mouse carotid artery injury.Methods: GFP-ADSCs were isolated from the adipose tissues of GFP mice and labeled with superparamagnetic iron oxide particles. Intracellular stability, proliferation, and viability of the labeled cells were evaluated in vitro. Next, the cells were transplanted into a mouse carotid artery injury model. Clinical 3 T magnetic resonance imaging was performed immediately before and 1, 3, 7, 14, 21, and 30 days after cell transplantation. Prussian blue staining and histological analysis were performed 7 and 30 days after transplantation.Results: GFP-ADSCs were found to be efficiently labeled with superparamagnetic iron oxide

  20. LONG-TERM (OF MANY DAYS NORMOTHERMAL LIMBAL GRAFTS PRESERVATION AS A METHOD OF QUANTITY AND ACTIVITY INCREASE OF MMSC-LIKE LIMBAL CELLS

    Directory of Open Access Journals (Sweden)

    S. A. Borzenok

    2012-01-01

    Full Text Available The reliability increase of corneal transplants engraftment may be achieved by combined transplantation with preserved limbal grafts (LG. We studied two methods of LG preservation: normothermal preservation in Borzenok–Moroz medium or standard culture medium for 35 days and cryopreservation at –80 °C in Borzenok– Moroz medium with/without DMSO addition for 1 month. We found that long-term normothermal preservation (for 28 days and cryopreservation maintain LG cells viability and their capacity to maintain cytokine balance as well as to keep cytokine balance and to retain phenotype of MMSC (multipotent mesenchymal stromal cells and to activate tolerogenic properties of limbal cells the latter improving quality and increasing capacity of corneal transplant engraftment reliability without systemic immunosupression. 

  1. Long-Term Fructose Intake Increases Adipogenic Potential: Evidence of Direct Effects of Fructose on Adipocyte Precursor Cells

    OpenAIRE

    Zubiría, María Guillermina; Alzamendi, Ana; Moreno, Griselda; Rey, María Amanda; Spinedi, Eduardo; Giovambattista, Andrés

    2016-01-01

    We have previously addressed that fructose rich diet (FRD) intake for three weeks increases the adipogenic potential of stromal vascular fraction cells from the retroperitoneal adipose tissue (RPAT). We have now evaluated the effect of prolonged FRD intake (eight weeks) on metabolic parameters, number of adipocyte precursor cells (APCs) and in vitro adipogenic potential from control (CTR) and FRD adult male rats. Additionally, we have examined the direct fructose effects on the adipogenic cap...

  2. Effects of long-term hypothyroidism in the morphology and synaptic organization of cerebellar ectopic granule cells

    OpenAIRE

    Madeira, M. D.; Azevedo, F.P.; Paula-Barbosa, M M

    1988-01-01

    Abundant ectopic granule cells scattered in the cerebellar molecular layer have been observed in 30- day-old hypothyroid rats. Their morphological features indicate that they must be regarded as mature heterotopic cells arrested during their migration towards the granular layer. As their impoverished dendritic trees are identical to those seen in controls, it is unlikely that the lack of thyroid hormones played a major role in the deficient dendritic outgrowth....

  3. Multilayer graphene for long-term corrosion protection of stainless steel bipolar plates for polymer electrolyte membrane fuel cell

    DEFF Research Database (Denmark)

    Stoot, Adam Carsten; Camilli, Luca; Spiegelhauer, Susie Ann;

    2015-01-01

    Abstract Motivated by similar investigations recently published (Pu et al., 2015), we report a comparative corrosion study of three sets of samples relevant as bipolar plates for polymer electrolyte fuel cells: stainless steel, stainless steel with a nickel seed layer (Ni/SS) and stainless steel...... graphene film is still intact with unchanged defect density. Our results show that even non-perfect multilayer graphene films can considerably increase the lifetime of future-generation bipolar plates for fuel cells....

  4. Efficient and stable MGMT-mediated selection of long-term repopulating stem cells in nonhuman primates

    OpenAIRE

    Brian C. Beard; Trobridge, Grant D.; Ironside, Christina; McCune, Jeannine S.; Adair, Jennifer E.; Kiem, Hans-Peter

    2010-01-01

    HSC transplantation using genetically modified autologous cells is a promising therapeutic strategy for various genetic diseases, cancer, and HIV. However, for many of these conditions, the current efficiency of gene transfer to HSCs is not sufficient for clinical use. The ability to increase the percentage of gene-modified cells following transplantation is critical to overcoming this obstacle. In vivo selection with mutant methylguanine methyltransferase (MGMTP140K) has been proposed to ove...

  5. Phenotypic malignant changes and untargeted lipidomic analysis of long-term exposed prostate cancer cells to endocrine disruptors

    Energy Technology Data Exchange (ETDEWEB)

    Bedia, Carmen, E-mail: carmen.bedia@idaea.csic.es; Dalmau, Núria, E-mail: nuria.dalmau@idaea.csic.es; Jaumot, Joaquim, E-mail: joaquim.jaumot@idaea.csic.es; Tauler, Romà, E-mail: roma.tauler@idaea.csic.es

    2015-07-15

    Endocrine disruptors (EDs) are a class of environmental toxic molecules able to interfere with the normal hormone metabolism. Numerous studies involve EDs exposure to initiation and development of cancers, including prostate cancer. In this work, three different EDs (aldrin, aroclor 1254 and chlorpyrifos (CPF)) were investigated as potential inducers of a malignant phenotype in DU145 prostate cancer cells after a chronic exposure. Epithelial to mesenchymal transition (EMT) induction, proliferation, migration, colony formation and release of metalloproteinase 2 (MMP-2) were analyzed in 50-day exposed cells to the selected EDs. As a result, aldrin and CPF exposure led to an EMT induction (loss of 16% and 14% of E-cadherin levels, respectively, compared to the unexposed cells). Aroclor and CPF presented an increased migration (134% and 126%, respectively), colony formation (204% and 144%, respectively) and MMP-2 release (137% in both cases) compared to the unexposed cells. An untargeted lipidomic analysis was performed to decipher the lipids involved in the observed transformations. As general results, aldrin exposure showed a global decrease in phospholipids and sphingolipids, and aroclor and CPF showed an increase of certain phospholipids, glycosphingolipids as well as a remarkable increase of some cardiolipin species. Furthermore, the three exposures resulted in an increase of some triglyceride species. In conclusion, some significant changes in lipids were identified and thus we postulate that some lipid compounds and lipid metabolic pathways could be involved in the acquisition of the malignant phenotype in exposed prostate cancer cells to the selected EDs. - Highlights: • Aldrin, aroclor and chlorpyrifos induced an aggressive phenotype in DU145 cells. • An untargeted lipidomic analysis has been performed on chronic exposed cells. • Lipidomic results showed changes in specific lipid species under chronic exposure. • These lipids may have a role in the

  6. Drug and cell type-specific regulation of genes with different classes of estrogen receptor beta-selective agonists.

    Directory of Open Access Journals (Sweden)

    Sreenivasan Paruthiyil

    Full Text Available Estrogens produce biological effects by interacting with two estrogen receptors, ERalpha and ERbeta. Drugs that selectively target ERalpha or ERbeta might be safer for conditions that have been traditionally treated with non-selective estrogens. Several synthetic and natural ERbeta-selective compounds have been identified. One class of ERbeta-selective agonists is represented by ERB-041 (WAY-202041 which binds to ERbeta much greater than ERalpha. A second class of ERbeta-selective agonists derived from plants include MF101, nyasol and liquiritigenin that bind similarly to both ERs, but only activate transcription with ERbeta. Diarylpropionitrile represents a third class of ERbeta-selective compounds because its selectivity is due to a combination of greater binding to ERbeta and transcriptional activity. However, it is unclear if these three classes of ERbeta-selective compounds produce similar biological activities. The goals of these studies were to determine the relative ERbeta selectivity and pattern of gene expression of these three classes of ERbeta-selective compounds compared to estradiol (E(2, which is a non-selective ER agonist. U2OS cells stably transfected with ERalpha or ERbeta were treated with E(2 or the ERbeta-selective compounds for 6 h. Microarray data demonstrated that ERB-041, MF101 and liquiritigenin were the most ERbeta-selective agonists compared to estradiol, followed by nyasol and then diarylpropionitrile. FRET analysis showed that all compounds induced a similar conformation of ERbeta, which is consistent with the finding that most genes regulated by the ERbeta-selective compounds were similar to each other and E(2. However, there were some classes of genes differentially regulated by the ERbeta agonists and E(2. Two ERbeta-selective compounds, MF101 and liquiritigenin had cell type-specific effects as they regulated different genes in HeLa, Caco-2 and Ishikawa cell lines expressing ERbeta. Our gene profiling studies

  7. Cell type specificity of female lung cancer associated with sulfur dioxide from air pollutants in Taiwan: An ecological study

    Directory of Open Access Journals (Sweden)

    Tseng Ching-Yu

    2012-01-01

    Full Text Available Abstract Background Many studies have examined the association between air pollutants (including sulfur dioxide [SO2], carbon monoxide [CO], nitrogen dioxide [NO2], nitric oxide [NO], ozone [O3], and particulate matter 10] and lung cancer. However, data from previous studies on pathological cell types were limited, especially for SO2 exposure. We aimed to explore the association between SO2 exposure from outdoor air pollutants and female lung cancer incidence by cell type specificity. Methods We conducted an ecological study and calculated annual average concentration of 6 air pollutants (SO2, CO, NO2, NO, O3, and PM10 using data from Taiwan Environmental Protection Administration air quality monitoring stations. The Poisson regression models were used to evaluate the association between SO2 and age-standardized incidence rate of female lung cancer by two major pathological types (adenocarcinoma [AC] and squamous cell carcinoma [SCC]. In order to understand whether there is a dose-response relationship between SO2 and two major pathological types, we analyzed 4 levels of exposure based on quartiles of concentration of SO2. Results The Poisson regression results showed that with the first quartile of SO2 concentration as the baseline, the relative risks for AC/SCC type cancer among females were 1.20 (95% confidence interval [CI], 1.04-1.37/1.39 (95% CI, 0.96-2.01 for the second, 1.22 (95% CI, 1.04-1.43/1.58 (95% CI, 1.06-2.37 for the third, and 1.27 (95% CI, 1.06-1.52/1.80 (95% CI, 1.15-2.84 for the fourth quartile of SO2 concentration. The tests for trend were statistically significant for both AC and SCC at P = 0.0272 and 0.0145, respectively. Conclusion The current study suggests that SO2 exposure as an air pollutant may increase female lung cancer incidence and the associations with female lung cancer is much stronger for SCC than for AC. The findings of this study warrant further investigation on the role of SO2 in the etiology of SCC.

  8. Rate of primary refractory disease in B and T-cell non-Hodgkin's lymphoma: correlation with long-term survival.

    Directory of Open Access Journals (Sweden)

    Corrado Tarella

    Full Text Available BACKGROUND: Primary refractory disease is a main challenge in the management of non-Hodgkin's Lymphoma (NHL. This survey was performed to define the rate of refractory disease to first-line therapy in B and T-cell NHL subtypes and the long-term survival of primary refractory compared to primary responsive patients. METHODS: Medical records were reviewed of 3,106 patients who had undergone primary treatment for NHL between 1982 and 2012, at the Hematology Centers of Torino and Bergamo, Italy. Primary treatment included CHOP or CHOP-like regimens (63.2%, intensive therapy with autograft (16.9%, or other therapies (19.9%. Among B-cell NHL, 1,356 (47.8% received first-line chemotherapy with rituximab. Refractory disease was defined as stable/progressive disease, or transient response with disease progression within six months. RESULTS: Overall, 690 (22.2% patients showed primary refractory disease, with a higher incidence amongst T-cell compared to B-cell NHL (41.9% vs. 20.5%, respectively, p<0.001. Several other clinico-pathological factors at presentation were variably associated with refractory disease, including histological aggressive disease, unfavorable clinical presentation, Bone Marrow involvement, low lymphocyte/monocyte ration and male gender. Amongst B-cell NHL, the addition of rituximab was associated with a marked reduction of refractory disease (13.6% vs. 26.7% for non-supplemented chemotherapy, p<0.001. Overall, primary responsive patients had a median survival of 19.8 years, compared to 1.3 yr. for refractory patients. A prolonged survival was consistently observed in all primary responsive patients regardless of the histology. The long life expectancy of primary responsive patients was documented in both series managed before and after 2.000. Response to first line therapy resulted by far the most predictive factor for long-term outcome (HR for primary refractory disease: 16.52, p<0.001. CONCLUSION: Chemosensitivity to primary

  9. The impact of long-term haemofiltration (continuous veno-venous haemofiltration) on cell-mediated immunity during endotoxaemia

    DEFF Research Database (Denmark)

    Toft, P; Nilsen, B U; Bollen, P;

    2007-01-01

    BACKGROUND: Increased survival with high-volume continuous veno-venous haemofiltration (CVVH) has been demonstrated in critically ill patients. This may be the result of intensified blood purification or an effect on the immune system. We hypothesized that CVVH modifies the cell-mediated immunity...

  10. Very Long Term Stability of Mixed Chimerism after Allogeneic Hematopoietic Stem Cell Transplantation in Patients with Hematologic Malignancies

    Directory of Open Access Journals (Sweden)

    Emmanuel Levrat

    2015-01-01

    Full Text Available The objective of this study is to analyze the evolution of chimerism of all patients transplanted for hematologic malignancies in our unit during a 20-year period, alive without relapse at 1 year after allogeneic hematopoietic stem cell transplantation (HSCT. Chimerism was tested using short tandem repeat polymorphisms after separation into mononuclear cells and granulocytes by Ficoll density gradient centrifugation. Of 155 patients studied, 89 had full chimerism (FC, 36 mononuclear cells mixed chimerism (MNC-MC, and 30 granulocytic MC with or without mononuclear cells MC (Gran-MC. Survival was significantly better in MNC-MC than in Gran-MC patients, with FC patients being intermediate. There was more disease relapse in the Gran-MC group but not in the MNC-MC group as compared to FC. MC was stable up to 21 years in the MNC-MC group and up to 19 years in the Gran-MC group. Of MC patients alive at 10 years, MC persisted in 83% in the MNC-MC and 57% in the Gran-MC groups. In conclusion, mixed chimerism may remain stable over a very long time period. In survivors without relapse at 1 year after HSCT, determining lineage specific chimerism may be useful as outcome differs, MNC-MC being associated with better outcome than Gran-MC.

  11. Long-Term Excess Mortality for Survivors of Non-small Cell Lung Cancer in the Netherlands

    NARCIS (Netherlands)

    Janssen-Heijnen, Maryska L.; van Steenbergen, Liza N.; Steyerberg, Ewout; Visser, Otto; De Ruysscher, Dirk K.; Groen, Harry J.

    2012-01-01

    Introduction: Most patients diagnosed with non-small cell lung cancer (NSCLC) die within the first few years after diagnosis. However, only little is known about those who have survived these first years. We aimed to study conditional 5-year relative survival rates for NSCLC patients during long-ter

  12. Blood Vessel Matrix Seeded with Cells: A Better Alternative for Abdominal Wall Reconstruction—A Long-Term Study

    Directory of Open Access Journals (Sweden)

    Maciej Nowacki

    2015-01-01

    Full Text Available Purpose. The aim of this study was to present abdominal wall reconstruction using a porcine vascular graft seeded with MSC (mesenchymal stem cells on rat model. Material and Methods. Abdominal wall defect was prepared in 21 Wistar rats. Acellular porcine-vascular grafts taken from aorta and prepared with Triton X were used. 14 aortic grafts were implanted in place, of which 7 grafts were seeded with rat MSC cells (Group I, and 7 were acellular grafts (Group II. As a control, 7 standard polypropylene meshes were used for defect augmentation (Group III. The assessment method was performed by HE and CD31 staining after 6 months. The mechanical properties have been investigated by Zwick&Roell Z0.5. Results. The strongest angiogenesis and lowest inflammatory response were observed in Group I. Average capillaries density was 2.75, 0.75, and 1.53 and inflammatory effect was 0.29, 1.39, and 2.72 for Groups I, II, and III, respectively. The means of mechanical properties were 12.74±1.48, 7.27±1.56, and 14.4±3.7 N/cm in Groups I and II and control, respectively. Conclusions. Cell-seeded grafts have better mechanical properties than acellular grafts but worse than polypropylene mesh. Cells improved mechanical and physiological properties of decellularized natural scaffolds.

  13. Induction of multiple drug resistance in HMEC-1 endothelial cells after long-term exposure to sunitinib

    Directory of Open Access Journals (Sweden)

    Huang LM

    2014-12-01

    Full Text Available Limin Huang,1,* Chaoquan Hu,2,* Mélanie Di Benedetto,3 Rémi Varin,4 Jielin Liu,2,3 Li Wang,3 Jean-Pierre Vannier,4 Jian Jin,3,5 Anne Janin,3,6,7 He Lu,3,6 Hong Li4 1Department of Oncology, People’s Hospital of Guizhou Province, Guiyang, People’s Republic of China; 2Department of Surgery, Affiliated Hospital, Guiyang Medical University, Guiyang, People’s Republic of China; 3INSERM UMR-S 1165, Paris, France; 4MERCI (EA 3829, Faculté de Médecine et de Pharmacie, Université de Rouen, Rouen, France; 5School of Medicine and Pharmaceutics, Jiangnan University, Wuxi, People’s Republic of China; 6Université Paris Diderot, Laboratoire de Pathologie, Paris, France; 7AP-HP-Hôpital Saint-Louis, Laboratoire de Pathologie, Paris, France *These authors contributed equally to this studyAbstract: Multiple drug resistance is still an unsolved problem in cancer therapy. Our previous study demonstrated that the chemotherapeutic drug doxorubicin (Dox induced upregulation of P-glycoprotein (P-gp in endothelial cells, resulting in a 20-fold increase in drug resistance and reduced efficiency of Dox treatment in a mice tumor model. In this study, we exposed human microvascular endothelial cells (HMEC-1 to sunitinib, a tyrosine kinase receptor inhibitor, to induce drug resistance. The results show that sunitinib treatment induced multiple drug resistance in these cells. They became resistant not only to sunitinib but also to Dox, paclitaxel, and vinblastine. Significant increases in P-gp (9.3-fold, ABCG2 (breast cancer resistance protein, 1.9-fold, and multidrug resistance-associated protein 1 (2.7-fold gene transcription were found by quantitative polymerase chain reaction quantification, and their protein expression was confirmed by Western blot. These increases gave rise to an approximately five-fold increase in half maximal inhibitory concentration in these cells in response to sunitinib treatment in vitro. The inhibitors of adenosine triphosphate

  14. Fluorescent CdSe/ZnS Nanocrystal-Peptide Conjugates for Long-term, Nontoxic Imaging and Nuclear Targeting in Living Cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, F; Gerion, D

    2004-06-08

    One of the biggest challenges in cell biology is the imaging of living cells. For this purpose, the most commonly used visualization tool is fluorescent markers. However, conventional labels, such as organic fluorescent dyes or green fluorescent proteins (GFP), lack the photostability to allow the tracking of cellular events that happen over minutes to days. In addition, they are either toxic to cells (dyes), or difficult to construct and manipulate (GFP). We report here the use of a new class of fluorescent labels, silanized CdSe/ZnS nanocrystal-peptide conjugates, for imaging the nuclei of living cells. CdSe/ZnS nanocrystals, or so called quantum dots (qdots), are extremely photostable, and have been used extensively in cellular imaging of fixed cells. Most of the studies about living cells so far have been concerned only with particle entry into the cytoplasm or the localization of receptors on the cell membrane. Specific targeting of qdots to the nucleus of living cells has not been reported in previous studies, due to the lack of a targeting mechanism and proper particle size. Here we demonstrate for the first time the construction of a CdSe/ZnS nanocrystal-peptide conjugate that carries the SV40 large T antigen nuclear localization signal (NLS), and the transfection of the complex into living cells. By a novel adaptation of a commonly used cell transfection technique for qdots, we were able to introduce and retain the NLS-qdot conjugates in living cells for up to a week without detectable negative cellular effects. Moreover, we can visualize the movement of the CdSe/ZnS qdot-peptide conjugates from cytoplasm to the nucleus, and the accumulation of the complex in the cell nucleus, over a long observation time period. This report opens the door for using qdots to visualize long-term biological events that happen in the cell nucleus, and provides a new nontoxic, long-term imaging platform for nuclear trafficking mechanisms and cell nuclear processes.

  15. Fluorescent CdSe/ZnS nanocrystal-peptide conjugates for long-term, nontoxic imaging and nuclear targeting in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Fanqing; Gerion, Daniele

    2004-06-14

    One of the biggest challenges in cell biology is the imaging of living cells. For this purpose, the most commonly used visualization tool is fluorescent markers. However, conventional labels, such as organic fluorescent dyes or green fluorescent proteins (GFP), lack the photostability to allow the tracking of cellular events that happen over minutes to days. In addition, they are either toxic to cells (dyes), or difficult to construct and manipulate (GFP). We report here the use of a new class of fluorescent labels, silanized CdSe/ZnS nanocrystal-peptide conjugates, for imaging the nuclei of living cells. CdSe/ZnS nanocrystals, or so called quantum dots (qdots), are extremely photostable, and have been used extensively in cellular imaging of fixed cells. However, most of the studies about living cells so far have been concerned only with particle entry into the cytoplasm or the localization of receptors on the cell membrane. Specific targeting of qdots to the nucleus of living cells ha s not been reported in previous studies, due to the lack of a targeting mechanism and proper particle size. Here we demonstrate for the first time the construction of a CdSe/ZnS nanocrystal-peptide conjugate that carries the SV40 large T antigen nuclear localization signal (NLS), and the transfection of the complex into living cells. By a novel adaptation of commonly used cell transfection techniques for qdots, we were able to introduce and retain the NLS-qdots conjugate in living cells for up to a week without detectable negative cellular effects. Moreover, we can visualize the movement of the CdSe/ZnS nanocrystal-peptide conjugates from cytoplasm to the nucleus, and the accumulation of the complex in the cell nucleus, over a long observation time period. This report opens the door for using qdots to visualize long-term biological events that happen in the cell nucleus, and provides a new nontoxic, long-term imaging platform for cell nuclear processes.

  16. Fluorescent CdSe/ZnS nanocrystal-peptide conjugates for long-term, nontoxic imaging and nuclear targeting in living cells

    International Nuclear Information System (INIS)

    One of the biggest challenges in cell biology is the imaging of living cells. For this purpose, the most commonly used visualization tool is fluorescent markers. However, conventional labels, such as organic fluorescent dyes or green fluorescent proteins (GFP), lack the photostability to allow the tracking of cellular events that happen over minutes to days. In addition, they are either toxic to cells (dyes), or difficult to construct and manipulate (GFP). We report here the use of a new class of fluorescent labels, silanized CdSe/ZnS nanocrystal-peptide conjugates, for imaging the nuclei of living cells. CdSe/ZnS nanocrystals, or so called quantum dots (qdots), are extremely photostable, and have been used extensively in cellular imaging of fixed cells. However, most of the studies about living cells so far have been concerned only with particle entry into the cytoplasm or the localization of receptors on the cell membrane. Specific targeting of qdots to the nucleus of living cells ha s not been reported in previous studies, due to the lack of a targeting mechanism and proper particle size. Here we demonstrate for the first time the construction of a CdSe/ZnS nanocrystal-peptide conjugate that carries the SV40 large T antigen nuclear localization signal (NLS), and the transfection of the complex into living cells. By a novel adaptation of commonly used cell transfection techniques for qdots, we were able to introduce and retain the NLS-qdots conjugate in living cells for up to a week without detectable negative cellular effects. Moreover, we can visualize the movement of the CdSe/ZnS nanocrystal-peptide conjugates from cytoplasm to the nucleus, and the accumulation of the complex in the cell nucleus, over a long observation time period. This report opens the door for using qdots to visualize long-term biological events that happen in the cell nucleus, and provides a new nontoxic, long-term imaging platform for cell nuclear processes

  17. Long-term effects of bariatric surgery on meal disposal and β-cell function in diabetic and nondiabetic patients

    DEFF Research Database (Denmark)

    Camastra, Stefania; Muscelli, Elza; Gastaldelli, Amalia;

    2013-01-01

    Gastric bypass surgery leads to marked improvements in glucose tolerance and insulin sensitivity in obese type 2 diabetes (T2D); the impact on glucose fluxes in response to a physiological stimulus, such as a mixed meal test (MTT), has not been determined. We administered an MTT to 12 obese T2D...... patients and 15 obese nondiabetic (ND) subjects before and 1 year after surgery (10 T2D and 11 ND) using the double-tracer technique and modeling of β-cell function. In both groups postsurgery, tracer-derived appearance of oral glucose was biphasic, a rapid increase followed by a sharp drop, a pattern...... that was mirrored by postprandial glucose levels and insulin secretion. In diabetic patients, surgery lowered fasting and postprandial glucose levels, peripheral insulin sensitivity increased in proportion to weight loss (~30%), and β-cell glucose sensitivity doubled but did not normalize (compared with 21...

  18. Improving the long-term stability of PBDTTPD polymer solar cells through material purification aimed at removing organic impurities

    KAUST Repository

    Mateker, William R.

    2013-01-01

    While bulk heterojunction (BHJ) solar cells fabricated from high M n PBDTTPD achieve power conversion efficiencies (PCE) as high as 7.3%, the short-circuit current density (JSC) of these devices can drop by 20% after seven days of storage in the dark and under inert conditions. This degradation is characterized by the appearance of S-shape features in the reverse bias region of current-voltage (J-V) curves that increase in amplitude over time. Conversely, BHJ solar cells fabricated from low Mn PBDTTPD do not develop S-shaped J-V curves. However, S-shapes identical to those observed in high Mn PBDTTPD solar cells can be induced in low M n devices through intentional contamination with the TPD monomer. Furthermore, when high Mn PBDTTPD is purified via size exclusion chromatography (SEC) to reduce the content of low molecular weight species, the JSC of polymer devices is significantly more stable over time. After 111 days of storage in the dark under inert conditions, the J-V curves do not develop S-shapes and the JSC degrades by only 6%. The S-shape degradation feature, symptomatic of low device lifetimes, appears to be linked to the presence of low molecular weight contaminants, which may be trapped within samples of high Mn polymer that have not been purified by SEC. Although these impurities do not affect initial device PCE, they significantly reduce device lifetime, and solar cell stability is improved by increasing the purity of the polymer materials. © 2013 The Royal Society of Chemistry.

  19. Bright and stable CdSe/CdS@SiO2 nanoparticles suitable for long term cell labeling

    OpenAIRE

    Aubert, Tangi; Wassmuth, Daniel; Soenen, Stefaan; Braeckmans, Kevin; Hens, Zeger

    2014-01-01

    Semiconductor quantum dots (QDs) constitute very promising candidates as light emitters for numerous applications in the field of biotechnology, including cell labeling, in vivo imaging and diagnostics.[1] For such applications, semiconductor QDs represent an attractive alternative to classic organic fluorophores as they exhibit a higher brightness thanks to their large absorption cross-sections and high photoluminescence quantum yields. Nevertheless, QDs usually suffer from higly oxidative e...

  20. Retinol and retinyl esters in parenchymal and nonparenchymal rat liver cell fractions after long-term administration of ethanol

    Energy Technology Data Exchange (ETDEWEB)

    Rasmussen, M.; Blomhoff, R.; Helgerud, P.; Solberg, L.A.; Berg, T.; Norum, K.R.

    1985-09-01

    Chronic ethanol consumption reduces the liver retinoid store in man and rat. We have studied the effect of ethanol on some aspects of retinoid metabolism in parenchymal and nonparenchymal liver cells. Rats fed 36% of total energy intake as ethanol for 5-6 weeks had the liver retinoid concentration reduced to about one-third, as compared to pair-fed controls. The reduction in liver retinoid affected both the parenchymal and the nonparenchymal cell fractions. Plasma retinol level was normal. Liver uptake of injected chylomicron (3H)retinyl ester was similar in the experimental and control group. The transport of retinoid from the parenchymal to the nonparenchymal cells was not found to be significantly retarded in the ethanol-fed rats. Despite the reduction in total retinoid level in liver, the concentrations of unesterified retinol and retinyl oleate were increased in the ethanol fed rats. Hepatic retinol esterification was not significantly affected in the ethanol-fed rats. Since our study has demonstrated that liver uptake of chylomicron retinyl ester is not impaired in the ethanol-fed rat, we suggest that liver retinoid metabolism may be increased.

  1. Brain metastasis of ALK positive anaplastic large cell lymphoma after a long-term disease free survival in an old adult

    Science.gov (United States)

    Wang, Cai-Xia; Wang, Hai; Li, Jie; Ma, Heng-Hui; Yu, Bo; Shi, Shan-Shan; Zhou, Xiao-Jun; Shi, Qun-Li

    2014-01-01

    Anaplastic large cell lymphoma (ALCL) is a subtype of non-Hodgkin lymphoma composed of CD30-positive cells and now recognized as three different entities: primary cutaneous ALCL, primary systemic anaplastic lymphoma kinase (ALK)-positive (ALK+) ALCL and primary ALK-negative (ALK-) ALCL. ALK+ ALCL is supposed to have a better prognosis than ALK- ALCL. It is rarely metastasized to other sites, especially to the central nervous system (CNS). Herein, we present a rare case of systemic ALK+ ALCL which metastasized to the brain after a long-term disease free survival in an adult. Neuroimaging revealed a well-enhanced mass in the left frontal lobe. And it was completely resected. The results of the pathological and immunohistochemical studies were consistent with the metastasized ALK+ ALCL. The clinical findings, pathologic characteristics and treatment are described. PMID:24696735

  2. OPTIMIZATION OF THE CATHODE LONG-TERM STABILITY IN MOLTEN CARBONATE FUEL CELLS: EXPERIMENTAL STUDY AND MATHEMATICAL MODELING

    Energy Technology Data Exchange (ETDEWEB)

    Hector Colonmer; Prabhu Ganesan; Nalini Subramanian; Dr. Bala Haran; Dr. Ralph E. White; Dr. Branko N. Popov

    2002-09-01

    This project focused on addressing the two main problems associated with state of art Molten Carbonate Fuel Cells, namely loss of cathode active material and stainless steel current collector deterioration due to corrosion. We followed a dual approach where in the first case we developed novel materials to replace the cathode and current collector currently used in molten carbonate fuel cells. In the second case we improved the performance of conventional cathode and current collectors through surface modification. States of art NiO cathode in MCFC undergo dissolution in the cathode melt thereby limiting the lifetime of the cell. To prevent this we deposited cobalt using an electroless deposition process. We also coated perovskite (La{sub 0.8}Sr{sub 0.2}CoO{sub 3}) in NiO thorough a sol-gel process. The electrochemical oxidation behavior of Co and perovskites coated electrodes is similar to that of the bare NiO cathode. Co and perovskite coatings on the surface decrease the dissolution of Ni into the melt and thereby stabilize the cathode. Both, cobalt and provskites coated nickel oxide, show a higher polarization compared to that of nickel oxide, which could be due to the reduced surface area. Cobalt substituted lithium nickel oxide (LiNi{sub 0.8}Co{sub 0.2}O{sub 2}) and lithium cobalt oxide were also studied. LiNi{sub x}Co{sub 1-x}O{sub 2} was synthesized by solid-state reaction procedure using lithium nitrate, nickel hydroxide and cobalt oxalate precursor. LiNi{sub x}Co{sub 1-x}O{sub 2} showed smaller dissolution of nickel than state of art nickel oxide cathode. The performance was comparable to that of nickel oxide. The corrosion of the current collector in the cathode side was also studied. The corrosion characteristics of both SS304 and SS304 coated with Co-Ni alloy were studied. This study confirms that surface modification of SS304 leads to the formation of complex scales with better barrier properties and better electronic conductivity at 650 C. A three

  3. Curcumin as a double-edged sword for stem cells: dose, time and cell type-specific responses to curcumin

    OpenAIRE

    Attari, Fatemeh; Zahmatkesh, Maryam; Aligholi, Hadi; Mehr, Shahram Ejtemaei; Sharifzadeh, Mohammad; Gorji, Ali; Mokhtari, Tahmineh; Khaksarian, Mojtaba; Hassanzadeh, Gholamreza

    2015-01-01

    Background The beneficial effects of curcumin which includes its antioxidant, anti-inflammatory and cancer chemo-preventive properties have been identified. Little information is available regarding the optimal dose and treatment periods of curcumin on the proliferation rate of different sources of stem cells. Methods In this study, the effect of various concentrations of curcumin on the survival and proliferation of two types of outstanding stem cells which includes bone marrow stem cells (B...

  4. Long-term environmental stewardship.

    Energy Technology Data Exchange (ETDEWEB)

    Nagy, Michael David

    2010-08-01

    The purpose of this Supplemental Information Source Document is to effectively describe Long-Term Environmental Stewardship (LTES) at Sandia National Laboratories/New Mexico (SNL/NM). More specifically, this document describes the LTES and Long-Term Stewardship (LTS) Programs, distinguishes between the LTES and LTS Programs, and summarizes the current status of the Environmental Restoration (ER) Project.

  5. Pairing-specific long-term depression of Purkinje cell excitatory postsynaptic potentials results from a classical conditioning procedure in the rabbit cerebellar slice.

    Science.gov (United States)

    Schreurs, B G; Oh, M M; Alkon, D L

    1996-03-01

    1. Using a rabbit cerebellar slice preparation, we stimulated a classical conditioning procedure by stimulating parallel fiber inputs to Purkinje cells with the use of a brief, high-frequency train of eight constant-current pulses 80 ms before climbing fiber inputs to the same Purkinje cell were stimulated with the use of a brief, lower frequency train of three constant-current pulses. In all experiments, we assessed the effects of stimulation by measuring the peak amplitude of Purkinje cell excitatory postsynaptic potentials (EPSPs) to single parallel fiber test pulses. 2. Intradendritically recorded Purkinje cell EPSPs underwent a long-term (> 20 min) reduction in peak amplitude (30%) after paired stimulation of the parallel and climbing fibers but not after unpaired or parallel fiber alone stimulation. We call this phenomenon pairing-specific long-term depression (PSD). 3. Facilitation of the peak amplitude of a second EPSP elicited by a parallel fiber train occurred both before and after paired stimulation suggesting that the locus of depression was not presynaptic. Depression of the peak amplitude of a depolarizing response to focal application of glutamate following pairings of parallel and climbing fiber stimulation added support to a suggested postsynaptic locus of the PSD effect. 4. The application of aniracetam potentiated EPSP peak amplitude by 40%, but these values returned to baseline as a result of pairings. With the removal of aniracetam from the bath 20 min after pairings, normal levels of pairing-specific EPSP depression were observed, indicating that the effect did not result from direct desensitization of alpha-amino-3-hydroxy-5-methyl-4-isoxazole-proprionic acid (AMPA) receptors. 5. Incubation of slices in the protein kinase inhibitor H-7 potentiated EPSP peak amplitudes slightly (9%), but peak amplitudes returned to baseline levels after pairings. The net reduction in EPSP peak amplitude of classical conditioning.

  6. Potassium Current Is Not Affected by Long-Term Exposure to Ghrelin or GHRP-6 in Somatotropes GC Cells

    Directory of Open Access Journals (Sweden)

    Belisario Domínguez Mancera

    2013-01-01

    Full Text Available Ghrelin is a growth hormone (GH secretagogue (GHS and GHRP-6 is a synthetic peptide analogue; both act through the GHS receptor. GH secretion depends directly on the intracellular concentration of Ca2+; this is determined from the intracellular reserves and by the entrance of Ca2+ through the voltage-dependent calcium channels, which are activated by the membrane depolarization. Membrane potential is mainly determined by K+ channels. In the present work, we investigated the effect of ghrelin (10 nM or GHRP-6 (100 nM for 96 h on functional expression of voltage-dependent K+ channels in rat somatotropes: GC cell line. Physiological patch-clamp whole-cell recording was used to register the K+ currents. With Cd2+ (1 mM and tetrodotoxin (1 μm in the bath solution recording, three types of currents were characterized on the basis of their biophysical and pharmacological properties. GC cells showed a K+ current with a transitory component sensitive to 4-aminopyridine, which represents ~40% of the total outgoing current; a sustained component named delayed rectifier , sensitive to tetraethylammonium; and a third type of K+ current was recorded at potentials more negative than −80 mV, permitting the entrance of K+ named inward rectifier (KIR. Chronic treatment with ghrelin or GHRP-6 did not modify the functional expression of K+ channels, without significant changes ( in the amplitudes of the three currents observed; in addition, there were no modifications in their biophysical properties and kinetic activation or inactivation.

  7. A Low Ethanol Dose Affects all Types of Cells in Mixed Long-Term Embryonic Cultures of the Cerebellum

    DEFF Research Database (Denmark)

    Pickering, Chris; Wicher, Grzegorz; Rosendahl, Sofi;

    2010-01-01

    . We exposed a primary culture of rat cerebellum from embryonic day 17 (corresponding to second trimester in humans) to ethanol at a concentration of 17.6 mM which is roughly equivalent to one glass of wine. Acutely, there was no change in cell viability after 5 or 8 days of exposure relative to...... of this ethanol dose, cultures were exposed for 30 days. After this period, virtually no neurons or myelinating oligodendrocytes were present in the ethanol-treated cultures. In conclusion, chronic exposure to ethanol, even at small doses, dramatically and persistently affects normal development....

  8. Long Term Stability Investigation of Solid Oxide Electrolysis Cell with Infiltrated Porous YSZ Air Electrode Under High Current

    DEFF Research Database (Denmark)

    Veltzé, Sune; Ovtar, Simona; Simonsen, Søren Bredmose;

    2015-01-01

    The increased interest in stable and low cost electrodes for solid oxide cells (SOC) has driven the research of electrode preparation to infiltration of catalyst material into porous backbone material. The infiltration method enables a reduction of amount of catalyst material and increases its...... stabilised zirconia (YSZ) backbone air electrode and Ni/YSZ cermet fuel electrode. The SOC was tested at electrolysis conditions under high current (up to -1 A/cm2). The porous YSZ electrodes was infiltrated with gadolinium-doped ceria oxide (CGO), to act as a barrier layer between the catalyst...

  9. Long-Term Fructose Intake Increases Adipogenic Potential: Evidence of Direct Effects of Fructose on Adipocyte Precursor Cells.

    Science.gov (United States)

    Zubiría, María Guillermina; Alzamendi, Ana; Moreno, Griselda; Rey, María Amanda; Spinedi, Eduardo; Giovambattista, Andrés

    2016-01-01

    We have previously addressed that fructose rich diet (FRD) intake for three weeks increases the adipogenic potential of stromal vascular fraction cells from the retroperitoneal adipose tissue (RPAT). We have now evaluated the effect of prolonged FRD intake (eight weeks) on metabolic parameters, number of adipocyte precursor cells (APCs) and in vitro adipogenic potential from control (CTR) and FRD adult male rats. Additionally, we have examined the direct fructose effects on the adipogenic capacity of normal APCs. FRD fed rats had increased plasma levels of insulin, triglyceride and leptin, and RPAT mass and adipocyte size. FACS studies showed higher APCs number and adipogenic potential in FRD RPAT pads; data is supported by high mRNA levels of competency markers: PPARγ2 and Zfp423. Complementary in vitro experiments indicate that fructose-exposed normal APCs displayed an overall increased adipogenic capacity. We conclude that the RPAT mass expansion observed in eight week-FRD fed rats depends on combined accelerated adipogenesis and adipocyte hypertrophy, partially due to a direct effect of fructose on APCs. PMID:27049396

  10. Long-Term Fructose Intake Increases Adipogenic Potential: Evidence of Direct Effects of Fructose on Adipocyte Precursor Cells

    Directory of Open Access Journals (Sweden)

    María Guillermina Zubiría

    2016-04-01

    Full Text Available We have previously addressed that fructose rich diet (FRD intake for three weeks increases the adipogenic potential of stromal vascular fraction cells from the retroperitoneal adipose tissue (RPAT. We have now evaluated the effect of prolonged FRD intake (eight weeks on metabolic parameters, number of adipocyte precursor cells (APCs and in vitro adipogenic potential from control (CTR and FRD adult male rats. Additionally, we have examined the direct fructose effects on the adipogenic capacity of normal APCs. FRD fed rats had increased plasma levels of insulin, triglyceride and leptin, and RPAT mass and adipocyte size. FACS studies showed higher APCs number and adipogenic potential in FRD RPAT pads; data is supported by high mRNA levels of competency markers: PPARγ2 and Zfp423. Complementary in vitro experiments indicate that fructose-exposed normal APCs displayed an overall increased adipogenic capacity. We conclude that the RPAT mass expansion observed in eight week-FRD fed rats depends on combined accelerated adipogenesis and adipocyte hypertrophy, partially due to a direct effect of fructose on APCs.

  11. Multilayer graphene for long-term corrosion protection of stainless steel bipolar plates for polymer electrolyte membrane fuel cell

    Science.gov (United States)

    Stoot, Adam C.; Camilli, Luca; Spiegelhauer, Susie-Ann; Yu, Feng; Bøggild, Peter

    2015-10-01

    Motivated by similar investigations recently published (Pu et al., 2015), we report a comparative corrosion study of three sets of samples relevant as bipolar plates for polymer electrolyte fuel cells: stainless steel, stainless steel with a nickel seed layer (Ni/SS) and stainless steel with Ni seed layer coated by a multi-layered graphene thin film (G/Ni/SS). The graphene film, synthesized by chemical vapour deposition (CVD), has a moderate amount of defects according to Raman spectroscopy. Short/medium-term corrosion test shows no significant advantage of using G/Ni/SS rather than Ni/SS, both samples exhibiting a similar trend, thus questioning the short-term positive effect of graphene coatings. However, partial immersion in boiling seawater for three weeks reveals a clear superiority of the graphene coating with respect to steel just protected by Ni. After the test, the graphene film is still intact with unchanged defect density. Our results show that even non-perfect multilayer graphene films can considerably increase the lifetime of future-generation bipolar plates for fuel cells.

  12. Long-term culture of chicken primordial germ cells isolated from embryonic blood and production of germline chimaeric chickens.

    Science.gov (United States)

    Naito, Mitsuru; Harumi, Takashi; Kuwana, Takashi

    2015-02-01

    Production of germline chimaeric chickens by the transfer of cultured primordial germ cells (PGC) is a useful system for germline manipulation. A novel culture system was developed for chicken PGC isolated from embryonic blood. The isolated PGC were cultured on feeder cells derived from chicken embryonic fibroblast. The cultured PGC formed colonies and they proliferated about 300-times during the first 30 days. The cultured PGC retained the ability to migrate to recipient gonads and were also chicken VASA homologue (CVH)-positive. Female PGC were present in the mixed-sex PGC populations cultured for more than 90 days and gave rise to viable offspring efficiently via germline chimaeric chickens. Male cultured PGC were transferred to recipient embryos and produced putative chimaeric chickens. The DNA derived from the cultured PGC was detected in the sperm samples of male putative chimaeric chickens, but no donor derived offspring were obtained. Donor-derived offspring were also obtained from germline chimaeric chickens by the transfer of frozen-thawed cultured PGC. The culture method for PGC developed in the present study is useful for manipulation of the germline in chickens, such as preservation of genetic resources and gene transfer.

  13. Long-term activity of covalent grafted biocatalysts during intermittent use of a glucose/O{sub 2} biofuel cell

    Energy Technology Data Exchange (ETDEWEB)

    Merle, G. [Institut Europeen des Membranes, UMR 5635, Place Eugene Bataillon, CC 047, 34095 Montpellier (France); Habrioux, A.; Servat, K. [LACCO ' Equipe Electrocatalyse' , UMR 6503, CNRS-Universite de Poitiers, 40 av. du Recteur Pineau, 86022 Poitiers (France); Rolland, M.; Innocent, C. [Institut Europeen des Membranes, UMR 5635, Place Eugene Bataillon, CC 047, 34095 Montpellier (France); Kokoh, K.B. [LACCO ' Equipe Electrocatalyse' , UMR 6503, CNRS-Universite de Poitiers, 40 av. du Recteur Pineau, 86022 Poitiers (France); Tingry, S. [Institut Europeen des Membranes, UMR 5635, Place Eugene Bataillon, CC 047, 34095 Montpellier (France)], E-mail: sophie.tingry@iemm.univ-montp2.fr

    2009-04-15

    The operational stability of enzymes in a concentric glucose/O{sub 2} biofuel cell has been significantly improved with the synthesis of grafted enzyme electrodes compared to entrapped enzyme electrodes. The concentric device combined glucose electro-oxidation by glucose oxidase at the anode and oxygen electro-reduction by bilirubin oxidase at the cathode. The entrapped enzyme electrodes were prepared from physical immobilization of the enzymes by a polypyrrole polymer onto the electrode surface. The grafted enzyme electrodes were synthesized by grafting the enzymes via alkyl spacer arms to a poly(aminopropylpyrrole) film onto the electrode surface. From spectrophotometric and electrochemical analyses, it was demonstrated that the spacer arms increased the operational stability and enzyme mobility that favoured electron transfer from their active sites to the electrode. The maximum power output of the assembled biofuel cell was 20 {mu}W cm{sup -2}, at 0.20 V with 10 mM glucose in phosphate buffer pH 7.4. The grafted enzyme electrodes presented an unprecedented operational stability as the maximum of power density of the BFC remains constant after intermittent use over a 45-day period. This was a remarkable improvement compared to electrodes with entrapped enzymes, which lost 74% of their initial power density after intermittent use over a 17-day period.

  14. Significance of preoperative C-reactive protein as a parameter of the perioperative course and long-term prognosis in squamous cell carcinoma and adenocarcinoma of the oesophagus

    Institute of Scientific and Technical Information of China (English)

    Ines Gockel; Kathrin Dirksen; Claudia M Messow; Theodor Junginger

    2006-01-01

    AIM: C-reactive protein (CRP) is an acute-phase reactant and a known indicator of the malignant potential of the tumour. The aim of this study was to investigate the significance of preoperative CRP as a parameter of the perioperative course and long-term prognosis in patients with squamous cell carcinoma and adenocarcinoma of the oesophagus.METHODS: Serum CRP was determined preoperatively in 291 of 371 patients undergoing oesophagectomy for cancer from December 1989 to March 2004. Median patient age was 59 (28-79) year,82.5% of patients were males. Squamous cell carcinoma was diagnosed in 151(51.9%) and adenocarcinoma in 122 patients. Transhiatal oesophagectomy was done in 151 (51.9%) patients and 134 (46.0%) patients underwent the abdominothoracic procedure.RESULTS: In 127 (43.6%) patients the preoperative serum CRP concentration was within the normal range (<5 mg/dL), elevated CRP levels were measured in 164 (56.4%) patients. Tumour extension (P<0.0005)and the number of lymph nodes affected by metastatic spread (P=0.015) were significantly increased in the group with elevated CRP levels. Among the perioperative parameters both the number of blood transfusions (P =0.006) and the general complication rate (P=0.002)were higher in patients with elevated preoperative CRP levels. The long-term survival rate of 13.6 (0-109.8)mo was poorer in the group with elevated CRP levels compared to 18.9 (0-155.4) mo in the group with normal CRP levels (log-rank test:P=0.107). Multivariate analysis with backward variables selection identified preoperative CRP as an independent prognostic factor of the long-term prognosis in patients with oesophageal carcinoma, with a hazard ratio of 1.182 (95% confidence interval: 1.030-1.356).CONCLUSION: The preoperative serum CRP-level is an easily determined independent prognostic marker in patients with squamous cell carcinoma and adenocarcinoma of the oesophagus.

  15. Quality of life, social challenges, and psychosocial support for long-term survivors after allogeneic hematopoietic stem-cell transplantation.

    Science.gov (United States)

    Norkin, Maxim; Hsu, Jack W; Wingard, John R

    2012-01-01

    Over the last two decades quality of life (QoL) and the social challenges of allogeneic hematopoietic stem cell transplant (allo-HSCT) survivors have been emerging as subjects of extensive research and are now considered as very important aspects in the pretransplant evaluation and management of allo-HSCT recipients. Recognition of QoL challenges in allo-HSCT survivors allows timely interventions leading to improvement of post-transplant outcomes. It needs to be recognized that long-lasting life changes associated with survivorship after allo-HSCT also significantly affect QoL of partners of allo-HSCT survivors. Currently, resources should be focused on how research findings can be used by patients, their partners, and physicians to optimize QoL and psychosocial adjustment.

  16. Long-term palliative treatment of patient with signet ring cell gastric cancer using endoscopic photodynamic therapy

    Directory of Open Access Journals (Sweden)

    V. V. Sokolov

    2014-01-01

    Full Text Available A case of multiple course of photodynamic therapy (PDT in patient with gastric cancer T1N0M0. Morpholological diagnosis in this patient was signet ring cell cancer. For 8 years the patient underwent endoscopic organ-sparing treatment: PDT with Photohem (17 courses, electrocoagulation of tumor (3 sessions. The drug Photohem was introduced intravenously at dose of 3.0 mg/kg body weight for 48 h before PDT. The treatment result was only partial regression of gastric tumor, the maximal follow-up period with no endoscopic and morphological signs of tumor growth accounted for 8 months. However besides incomplete removal of gastric tumor and morphological type, for check-up examination 8 years after the onset of endoscopic treatment there were no features of regional and distant metastases according to chest and abdominal CT and US. 

  17. Microbial community analysis in a long-term membrane-less microbial electrolysis cell with hydrogen and methane production.

    Science.gov (United States)

    Rago, Laura; Ruiz, Yolanda; Baeza, Juan A; Guisasola, Albert; Cortés, Pilar

    2015-12-01

    A single-chamber microbial electrolysis cell (MEC) aiming at hydrogen production with acetate as sole carbon source failed due to methanogenesis build-up despite the significant amount of 2-bromoethanesulfonate (BES) dosage, 50 mM. Specific batch experiments and a thorough microbial community analysis, pyrosequencing and qPCR, of cathode, anode and medium were performed to understand these observations. The experimental data rebuts different hypothesis and shows that methanogenesis at high BES concentration was likely due to the capacity of some Archaea (hydrogen-oxidizing genus Methanobrevibacter) to resist high BES concentration up to 200 mM. Methanobrevibacter, of the Methanobacteriales order, represented almost the 98% of the total Archaea in the cathode whereas Geobacter was highly abundant in the anode (72% of bacteria). Moreover, at higher BES concentration (up to 200 mM), methanogenesis activity decreased resulting in an increase of homoacetogenic activity, which challenged the performance of the MEC for H2 production.

  18. The neural cell adhesion molecule-derived peptide FGL facilitates long-term plasticity in the dentate gyrus in vivo

    DEFF Research Database (Denmark)

    Dallérac, Glenn; Zerwas, Meike; Novikova, Tatiana;

    2011-01-01

    The neural cell adhesion molecule (NCAM) is known to play a role in developmental and structural processes but also in synaptic plasticity and memory of the adult animal. Recently, FGL, a NCAM mimetic peptide that binds to the Fibroblast Growth Factor Receptor 1 (FGFR-1), has been shown to have...... a beneficial impact on normal memory functioning, as well as to rescue some pathological cognitive impairments. Whether its facilitating impact may be mediated through promoting neuronal plasticity is not known. The present study was therefore designed to test whether FGL modulates the induction......-frequency stimulation (HFS) to the medial perforant path. The results suggest that although FGL did not alter basal synaptic transmission, it facilitated both the induction and maintenance of LTP. Interestingly, FGL also modified the heterosynaptic plasticity observed at the neighboring lateral perforant path synapses...

  19. Modeling Cu Migration in CdTe Solar Cells Under Device-Processing and Long-Term Stability Conditions: Preprint

    Energy Technology Data Exchange (ETDEWEB)

    Teeter, G.; Asher, S.

    2008-05-01

    An impurity migration model for systems with material interfaces is applied to Cu migration in CdTe solar cells. In the model, diffusion fluxes are calculated from the Cu chemical potential gradient. Inputs to the model include Cu diffusivities, solubilities, and segregation enthalpies in CdTe, CdS and contact materials. The model yields transient and equilibrium Cu distributions in CdTe devices during device processing and under field-deployed conditions. Preliminary results for Cu migration in CdTe photovoltaic devices using available diffusivity and solubility data from the literature show that Cu segregates in the CdS, a phenomenon that is commonly observed in devices after back-contact processing and/or stress conditions.

  20. Improving the Long-Term Stability of Perovskite Solar Cells with a Porous Al 2 O 3 Buffer Layer

    KAUST Repository

    Guarnera, Simone

    2015-02-05

    © 2015 American Chemical Society. Hybrid perovskites represent a new paradigm for photovoltaics, which have the potential to overcome the performance limits of current technologies and achieve low cost and high versatility. However, an efficiency drop is often observed within the first few hundred hours of device operation, which could become an important issue. Here, we demonstrate that the electrode\\'s metal migrating through the hole transporting material (HTM) layer and eventually contacting the perovskite is in part responsible for this early device degradation. We show that depositing the HTM within an insulating mesoporous "buffer layer" comprised of Al2O3 nanoparticles prevents the metal electrode migration while allowing for precise control of the HTM thickness. This enables an improvement in the solar cell fill factor and prevents degradation of the device after 350 h of operation. (Graph Presented).

  1. Long-term assessment of best cathode position to maximise microbial fuel cell performance in horizontal subsurface flow constructed wetlands.

    Science.gov (United States)

    Corbella, Clara; Garfí, Marianna; Puigagut, Jaume

    2016-09-01

    The cathode of microbial fuel cells (MFCs) implemented in constructed wetlands (CWs) is generally set in close contact with water surface to provide a rich oxygen environment. However, water level variations caused by plants evapotranspiration in CWs might decrease MFC performance by limiting oxygen transfer to the cathode. Main objective of this work was to quantify the effect of water level variation on MFC performance implemented in HSSF CW. For the purpose of this work two MFCs were implemented within a HSSF CW pilot plant fed with primary treated domestic wastewater. Cell voltage (Ecell) and the relative distance between the cathode and the water level were recorded for one year. Results showed that Ecell was greatly influenced by the relative distance between the cathode and the water level, giving an optimal cathode position of about 1 to 2cm above water level. Both water level variation and Ecell were daily and seasonal dependent, showing a pronounced day/night variation during warm periods and showing almost no daily variation during cold periods. Energy production under pronounced daily water level variation was 40% lower (80±56mWh/m(2)·day) than under low water level variation (131±61mWh/m(2)·day). Main conclusion of the present work is that of the performance of MFC implemented in HSSF CW is highly dependent on plants evapotranspiration. Therefore, MFC that are to be implemented in CWs shall be designed to be able to cope with pronounced water level variations. PMID:27151501

  2. Long-term complete remission in a patient with intravascular large B-cell lymphoma with central nervous system involvement

    Directory of Open Access Journals (Sweden)

    Sawada T

    2014-11-01

    Full Text Available Takeshi Sawada,1 Yasushi Omuro,1 Takeshi Kobayashi,2 Tunekazu Hishima,3 Fumiaki Koizumi,4 Yusuke Kanemasa,1 Tatsu Shimoyama,1 Eisaku Sasaki,1 Yoshiharu Maeda1 1Department of Chemotherapy, 2Department of Hematology, 3Department of Pathology, 4Department of Laboratory Medicine, Tokyo Metropolitan Cancer and Infectious Diseases Center, Komagome Hospital, Bunkyo-ku, Tokyo, Japan Abstract: This report describes a patient with intravascular large B-cell lymphoma (IVLBCL with central nervous system involvement at the time of diagnosis who achieved complete remission for over 5 years in response to therapy. The patient, a 71 year-old woman, was previously healthy with the exception of taking verapamil for paroxysmal supraventricular tachycardia. She had presented with pyrexia and gradually progressive anemia. Brain magnetic resonance imaging revealed an infarct-like lesion in the pons, although no paralysis was observed. She was diagnosed with IVLBCL on the basis of random skin biopsy. After eight cycles of rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone therapy, abnormal laboratory data had normalized, and no pontine lesion was evident on magnetic resonance imaging without receiving any intrathecal chemotherapy. IVLBCL is associated with poor prognosis, particularly in patients with central nervous system involvement. Early initiation of rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone therapy and drug interactions between anticancer agents and verapamil as a p-glycoprotein inhibitor were considered the possible reasons for favorable outcome in the present case. Keywords: intravascular large B-cell lymphoma, random skin biopsy, CNS involvement, rituximab, verapamil, blood–brain barrier

  3. Long-term environmental exposure to metals (Cu, Cd, Pb, Zn) activates the immune cell stress response in the common European sea star (Asterias rubens).

    Science.gov (United States)

    Matranga, V; Pinsino, A; Randazzo, D; Giallongo, A; Dubois, P

    2012-05-01

    The common sea star Asterias rubens represents a key-species of the North-Eastern Atlantic macro benthic community. The cells of their immune system, known as coelomocytes, are the first line of defence against environmental hazards. Here, we report the results of investigations on the immune cells response of sea stars exposed to marine environmental pollution for long periods. We show that levels of the heat shock cognate protein 70 (HSC70) in coelomocytes from A. rubens, which were collected during a field study in the Sǿrfjord (North Sea, SW coast of Norway) along a contamination gradient, are directly associated with the long-term accumulation of Cd, Cu heavy metals exclusively in the tegument. Conversely, Pb and Zn accumulation in the tegument did not relate to HSC70 levels and none of the metals were found accumulated in the pyloric coeca. In addition the coelomocytes from A. rubens, collected in high and low metal impacted stations were examined by a proteomic approach using two-dimensional electrophoresis (2DE). By comparison of the proteomic maps, we observed that 31 protein spots differed in their relative abundance, indicating a gene expression response to the metal mixture exposure. All together, our results confirm that the echinoderm immune cells are a suitable model for the assessment of long-term exposure to environmental pollution, moreover that the increased level of HSC70 can be considered a signal of an acquired tolerance within a large spectrum of protein profile changes occurring in response to metal contamination. PMID:22000270

  4. Adipose tissue-derived mesenchymal stem cells in long-term dialysis patients display downregulation of PCAF expression and poor angiogenesis activation.

    Directory of Open Access Journals (Sweden)

    Shuichiro Yamanaka

    Full Text Available We previously demonstrated that mesenchymal stem cells (MSCs differentiate into functional kidney cells capable of urine and erythropoietin production, indicating that they may be used for kidney regeneration. However, the viability of MSCs from dialysis patients may be affected under uremic conditions. In this study, we isolated MSCs from the adipose tissues of end-stage kidney disease (ESKD patients undergoing long-term dialysis (KD-MSCs; mean: 72.3 months and from healthy controls (HC-MSCs to compare their viability. KD-MSCs and HC-MSCs were assessed for their proliferation potential, senescence, and differentiation capacities into adipocytes, osteoblasts, and chondrocytes. Gene expression of stem cell-specific transcription factors was analyzed by PCR array and confirmed by western blot analysis at the protein level. No significant differences of proliferation potential, senescence, or differentiation capacity were observed between KD-MSCs and HC-MSCs. However, gene and protein expression of p300/CBP-associated factor (PCAF was significantly suppressed in KD-MSCs. Because PCAF is a histone acetyltransferase that mediates regulation of hypoxia-inducible factor-1α (HIF-1α, we examined the hypoxic response in MSCs. HC-MSCs but not KD-MSCs showed upregulation of PCAF protein expression under hypoxia. Similarly, HIF-1α and vascular endothelial growth factor (VEGF expression did not increase under hypoxia in KD-MSCs but did so in HC-MSCs. Additionally, a directed in vivo angiogenesis assay revealed a decrease in angiogenesis activation of KD-MSCs. In conclusion, long-term uremia leads to persistent and systematic downregulation of PCAF gene and protein expression and poor angiogenesis activation of MSCs from patients with ESKD. Furthermore, PCAF, HIF-1α, and VEGF expression were not upregulated by hypoxic stimulation of KD-MSCs. These results suggest that the hypoxic response may be blunted in MSCs from ESKD patients.

  5. Long-term improvement during tadalafil therapy in a patient with pulmonary hypertension secondary to pulmonary Langerhans cell histiocytosis.

    Science.gov (United States)

    Nemoto, Kenji; Oh-Ishi, Shuji; Inui, Toshihide; Nakazawa, Mariko; Hyodo, Kentaro; Nakajima, Masayuki; Kanazawa, Jun; Miura, Yukiko; Takaku, Takio; Minami, Yuko; Hayashihara, Kenji; Saito, Takefumi; Kawabata, Yoshinori

    2016-01-01

    Pulmonary arterial hypertension (PAH) secondary to pulmonary Langerhans cell histiocytosis (PLCH) is known to be a relatively common complication and is associated with a poor prognosis. However, the optimal therapeutic approach for these cases remains to be established. A 57-year-old man visited our hospital because of a progressive dry cough. A thoracic computed tomography examination showed a combination of diffuse thick-walled cysts and reticulonodular shadows that were predominant in bilateral upper lobes of the lungs. He was diagnosed as having PLCH based on the results of video-assisted thoracoscopic lung biopsies. During a 3-year clinical course, his condition deteriorated despite smoking cessation. A systemic evaluation demonstrated precapillary PAH caused by PLCH (PAH-PLCH), and treatment with tadalafil, a phosphodiesterase-5 inhibitor, was started. During a 50-month period of treatment with tadalafil, improvements in his dyspnea, 6-min walking distance, and hemodynamics were maintained without either overt hypoxemia or pulmonary edema. We considered that tadalafil therapy may be a useful option in the treatment of patients with PAH-PLCH. PMID:27330952

  6. Coordinated cell type-specific epigenetic remodeling in prefrontal cortex begins before birth and continues into early adulthood.

    Directory of Open Access Journals (Sweden)

    Hennady P Shulha

    2013-04-01

    Full Text Available Development of prefrontal and other higher-order association cortices is associated with widespread changes in the cortical transcriptome, particularly during the transitions from prenatal to postnatal development, and from early infancy to later stages of childhood and early adulthood. However, the timing and longitudinal trajectories of neuronal gene expression programs during these periods remain unclear in part because of confounding effects of concomitantly occurring shifts in neuron-to-glia ratios. Here, we used cell type-specific chromatin sorting techniques for genome-wide profiling of a histone mark associated with transcriptional regulation--H3 with trimethylated lysine 4 (H3K4me3--in neuronal chromatin from 31 subjects from the late gestational period to 80 years of age. H3K4me3 landscapes of prefrontal neurons were developmentally regulated at 1,157 loci, including 768 loci that were proximal to transcription start sites. Multiple algorithms consistently revealed that the overwhelming majority and perhaps all of developmentally regulated H3K4me3 peaks were on a unidirectional trajectory defined by either rapid gain or loss of histone methylation during the late prenatal period and the first year after birth, followed by similar changes but with progressively slower kinetics during early and later childhood and only minimal changes later in life. Developmentally downregulated H3K4me3 peaks in prefrontal neurons were enriched for Paired box (Pax and multiple Signal Transducer and Activator of Transcription (STAT motifs, which are known to promote glial differentiation. In contrast, H3K4me3 peaks subject to a progressive increase in maturing prefrontal neurons were enriched for activating protein-1 (AP-1 recognition elements that are commonly associated with activity-dependent regulation of neuronal gene expression. We uncovered a developmental program governing the remodeling of neuronal histone methylation landscapes in the prefrontal

  7. Long-Term Survival of a Patient with Giant Cell Glioblastoma: Case Report and Review of the Literature

    Directory of Open Access Journals (Sweden)

    E. Naydenov

    2009-07-01

    Full Text Available Glioblastoma multiforme (GBM is the most common glial tumor of the central nervous system. Overall survival is less than a year in most of the cases in spite of multimodal treatment approaches. A 45-year-old female with histologically confirmed giant cell GBM was treated at our institution. Subtotal excision of the lesion situated in the right precentral area was performed during the initial stay in August 2005. The patient improved after the procedure with no hypertension and additional neurological deficit. Radiotherapy plus concomitant and adjuvant temozolomide was performed. The patient was symptom-free for 35 months after initial surgery. From July 2008 the patient developed partial motor seizures in the left side of the body and progressive hemiparesis. Local tumor progression was demonstrated on the neuroimaging studies. In December 2008, a second operative intervention was performed with subtotal excision of the tumor. Forty-five months after the initial diagnosis the patient is still alive with moderate neurological deficit. Microarray analysis of the tumor found the following numeric chromosomal aberrations: monosomy 8, 10, 13, 22, and trisomy 21, as well as amplifications in 4q34.1, 4q28.2, 6q16.3, 7q36.1, 7p21.3, and deletions in 1q42.12, 1q32.2, 1q25.2, 1p33, 2q37.2, 18q22.3, 19p13.2, Xq28, and Xq27.3. GBMs seem to be a heterogeneous group of glial tumors with different clinical course and therapeutic response. Microarray analysis is a useful method to establish a number of possible molecular predictors.

  8. Differential microRNA expression signatures and cell type-specific association with Taxol resistance in ovarian cancer cells.

    Science.gov (United States)

    Kim, Yong-Wan; Kim, Eun Young; Jeon, Doin; Liu, Juinn-Lin; Kim, Helena Suhyun; Choi, Jin Woo; Ahn, Woong Shick

    2014-01-01

    Paclitaxel (Taxol) resistance remains a major obstacle for the successful treatment of ovarian cancer. MicroRNAs (miRNAs) have oncogenic and tumor suppressor activity and are associated with poor prognosis phenotypes. miRNA screenings for this drug resistance are needed to estimate the prognosis of the disease and find better drug targets. miRNAs that were differentially expressed in Taxol-resistant ovarian cancer cells, compared with Taxol-sensitive cells, were screened by Illumina Human MicroRNA Expression BeadChips. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to identify target genes of selected miRNAs. Kaplan-Meier survival analysis was applied to identify dysregulated miRNAs in ovarian cancer patients using data from The Cancer Genome Atlas. A total of 82 miRNAs were identified in ovarian carcinoma cells compared to normal ovarian cells. miR-141, miR-106a, miR-200c, miR-96, and miR-378 were overexpressed, and miR-411, miR-432, miR-494, miR-409-3p, and miR-655 were underexpressed in ovarian cancer cells. Seventeen miRNAs were overexpressed in Taxol-resistant cells, including miR-663, miR-622, and HS_188. Underexpressed miRNAs in Taxol-sensitive cells included miR-497, miR-187, miR-195, and miR-107. We further showed miR-663 and miR-622 as significant prognosis markers of the chemo-resistant patient group. In particular, the downregulation of the two miRNAs was associated with better survival, perhaps increasing the sensitivity of cancer cells to Taxol. In the chemo-sensitive patient group, only miR-647 could be a prognosis marker. These miRNAs inhibit several interacting genes of p53 networks, especially in TUOS-3 and TUOS-4, and showed cell line-specific inhibition effects. Taken together, the data indicate that the three miRNAs are closely associated with Taxol resistance and potentially better prognosis factors. Our results suggest that these miRNAs were successfully and reliably identified and would be used in the

  9. Cell-type-specific profiling of protein-DNA interactions without cell isolation using targeted DamID with next-generation sequencing.

    Science.gov (United States)

    Marshall, Owen J; Southall, Tony D; Cheetham, Seth W; Brand, Andrea H

    2016-09-01

    This protocol is an extension to: Nat. Protoc. 2, 1467-1478 (2007); doi:10.1038/nprot.2007.148; published online 7 June 2007The ability to profile transcription and chromatin binding in a cell-type-specific manner is a powerful aid to understanding cell-fate specification and cellular function in multicellular organisms. We recently developed targeted DamID (TaDa) to enable genome-wide, cell-type-specific profiling of DNA- and chromatin-binding proteins in vivo without cell isolation. As a protocol extension, this article describes substantial modifications to an existing protocol, and it offers additional applications. TaDa builds upon DamID, a technique for detecting genome-wide DNA-binding profiles of proteins, by coupling it with the GAL4 system in Drosophila to enable both temporal and spatial resolution. TaDa ensures that Dam-fusion proteins are expressed at very low levels, thus avoiding toxicity and potential artifacts from overexpression. The modifications to the core DamID technique presented here also increase the speed of sample processing and throughput, and adapt the method to next-generation sequencing technology. TaDa is robust, reproducible and highly sensitive. Compared with other methods for cell-type-specific profiling, the technique requires no cell-sorting, cross-linking or antisera, and binding profiles can be generated from as few as 10,000 total induced cells. By profiling the genome-wide binding of RNA polymerase II (Pol II), TaDa can also identify transcribed genes in a cell-type-specific manner. Here we describe a detailed protocol for carrying out TaDa experiments and preparing the material for next-generation sequencing. Although we developed TaDa in Drosophila, it should be easily adapted to other organisms with an inducible expression system. Once transgenic animals are obtained, the entire experimental procedure-from collecting tissue samples to generating sequencing libraries-can be accomplished within 5 d. PMID:27490632

  10. Long-term results of a randomized controlled trial evaluating preoperative chemotherapy in resectable non-small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Chen ZW

    2013-06-01

    Full Text Available Zhiwei Chen,* Qingquan Luo,* Hong Jian, Zhen Zhou, Baijun Cheng, Shun Lu, Meilin LiaoShanghai Lung Tumor Clinical Medical Center, Shanghai Chest Hospital, Shanghai Jiao Tong University, Shanghai, People’s Republic of China *These authors contributed equallyObjective: We aimed to evaluate whether preoperative chemotherapy provides benefits in the survival and prognosis of patients with non-small cell lung cancer (NSCLC in resectable stages I to IIIA, except T1N0. Methods: In this randomized, controlled trial, 356 patients with stage I (except for T1N0, II and IIIA NSCLC were assigned to either the preoperative chemotherapy plus surgery arm (179 patients or the primary surgery arm (177 patients. Both treatments were followed by adjuvant chemotherapy. The end point of this study included overall survival (OS, progression-free survival (PFS, and survival rate associated with clinical remission. Results: Statistical survival difference was found between the preoperative chemotherapy plus surgery arm and the surgery-alone arm. However, the median survival time (MST in the preoperative chemotherapy arm was lower than that of surgery-alone arm (MST, 45.42 months vs 57.59 months (P = 0.016. When comparing the effect of preoperative chemotherapy at each stage of NSCLC, a statistical survival difference was found in stage II NSCLC but not in stage I and IIIA (MST 40.86 months vs 80.81 months (P = 0.044. However, no statistically significant difference in PFS was noticed between the two arms, except for stage I NSCLC (hazard radio [HR] = 0.87; 95% CI, 0.561−1.629; P = 0.027. The survival rate was higher for patients who had clinical remission after preoperative chemotherapy, but the differences did not reach statistical significance (MST 42.10 months vs 35.33 months (P = 0.630. Conclusion: Preoperative chemotherapy did not show benefits in OS and PFS for stage I-IIIA NSCLC patients. Keywords: NSCLC, neoadjuvent, mitomycin, cisplatin, vindesine

  11. Diffuse large B-cell lymphoma associated with the use of biologic and other investigational agents: the importance of long-term post-marketing safety surveillance.

    Science.gov (United States)

    Goddard, Allison; Borovicka, Judy H; West, Dennis P; Evens, Andrew M; Laumann, Anne

    2011-01-01

    This case report describes a patient who developed diffuse large B-cell lymphoma (DLBCL) after receiving courses of two investigational biologic agents and cyclosporine followed by more than four years of subcutaneous efalizumab for the treatment of extensive chronic plaque psoriasis. Three years later, the patient remains free of lymphoma and his psoriasis is well controlled with thrice-weekly narrow-band ultraviolet phototherapy. This case emphasizes the importance of continued long-term post-marketing safety surveillance and the early reporting of all possible serious side effects, including cancers, related to the use of any newly available product. In particular, surveillance should focus on the immunomodulating biologic agents in order to identify possible dangerous sequelae.

  12. Long-term survival after gemcitabine and cisplatin in patients with locally advanced transitional cell carcinoma of the bladder: focus on supplementary treatment strategies

    DEFF Research Database (Denmark)

    Als, Anne Birgitte; Sengelov, Lisa; von der Maase, Hans

    2007-01-01

    radiotherapy as supplementary treatment. CONCLUSIONS: In patients with locally advanced bladder cancer, NED following chemotherapy alone or chemotherapy plus supplementary cystectomy or radiotherapy is essential to achieve long-term survival. Patients with a partial response should be offered radical......OBJECTIVE: The objective was to evaluate response and survival, as well as efficacy of subsequent supplementary treatment and follow-up strategy in patients with locally advanced transitional cell carcinoma of the bladder following combination chemotherapy with gemcitabine and cisplatin (GC......: A total of 25 patients (29.8%) with complete response to chemotherapy were followed by close surveillance. This group achieved a median overall survival of 47.6 mo. Another 25 patients had partial response to chemotherapy. Of these patients, 16 had supplementary treatment, with 10 achieving "no evidence...

  13. Long-term effects of ionizing radiation

    International Nuclear Information System (INIS)

    This paper approaches the long-term effects of ionizing radiation considering the common thought that killing of cells is the basis for deterministic effects and that the subtle changes in genetic information are important in the development of radiation-induced cancer, or genetic effects if these changes are induced in germ cells

  14. A long-term degradation study of power generation characteristics of anode-supported solid oxide fuel cells using LaNi(Fe)O{sub 3} electrode

    Energy Technology Data Exchange (ETDEWEB)

    Komatsu, Takeshi; Watanabe, Kimitaka; Arakawa, Masayasu; Arai, Hajime [NTT Corporation, NTT Energy and Environment Systems Laboratories, Morinosato-Wakamiya 3-1, Atsugi-shi, Kanagawa 243-0198 (Japan)

    2009-09-05

    The long-term operation of an anode-supported solid oxide fuel cell was examined to study the degradation factor. The cell was constructed using LaNi{sub 0.6}Fe{sub 0.4}O{sub 3} (LNF), alumina-doped scandia stabilized zirconia (SASZ), and NiO-SASZ as the cathode, electrolyte, and anode respectively. The cell had Pt current collectors and was operated for 6500 h. The test was carried out at 1073 K with a constant load of 0.4 A cm{sup -2} and included thermal cycling. The cell voltage degradation rate was below 0.86%/1000 h when the cell was operated for up to 5200 h. Changes in the resistance of the cells during the experiments were analyzed by impedance spectroscopy. The cathode polarization resistance and ohmic resistance increased with time. The elements (Si and B) contained in the water condensed from the cathode exhaust gas were identified using inductively coupled plasma (ICP). (author)

  15. Layered long-term co-culture of hepatocytes and endothelial cells on a transwell membrane: toward engineering the liver sinusoid

    International Nuclear Information System (INIS)

    This paper presents a novel liver model that mimics the liver sinusoid where most liver activities occur. A key aspect of our current liver model is a layered co-culture of primary rat hepatocytes (PRHs) and primary rat liver sinusoidal endothelial cells (LSECs) or bovine aortic endothelial cells (BAECs) on a transwell membrane. When a layered co-culture was attempted with a thin Matrigel layer placed between hepatocytes and endothelial cells to mimic the space of Disse, the cells did not form completely separated monolayers. However, when hepatocytes and endothelial cells were cultured on the opposite sides of a transwell membrane, PRHs co-cultured with LSECs or BAECs maintained their viability and normal morphology for 39 and 57 days, respectively. We assessed the presence of hepatocyte-specific differentiation markers to verify that PRHs remained differentiated in the long-term co-culture and analyzed hepatocyte function by monitoring urea synthesis. We also noted that the expression of cytochrome P-450 remained similar in the co-cultured system from day 1 to day 48. Thus, our novel liver model system demonstrated that primary hepatocytes can be cultured for extended times and retain their hepatocyte-specific functions when layered with endothelial cells. (paper)

  16. Long-term upregulation of inflammation and suppression of cell proliferation in the brain of adult rats exposed to traumatic brain injury using the controlled cortical impact model.

    Directory of Open Access Journals (Sweden)

    Sandra A Acosta

    Full Text Available The long-term consequences of traumatic brain injury (TBI, specifically the detrimental effects of inflammation on the neurogenic niches, are not very well understood. In the present in vivo study, we examined the prolonged pathological outcomes of experimental TBI in different parts of the rat brain with special emphasis on inflammation and neurogenesis. Sixty days after moderate controlled cortical impact injury, adult Sprague-Dawley male rats were euthanized and brain tissues harvested. Antibodies against the activated microglial marker, OX6, the cell cycle-regulating protein marker, Ki67, and the immature neuronal marker, doublecortin, DCX, were used to estimate microglial activation, cell proliferation, and neuronal differentiation, respectively, in the subventricular zone (SVZ, subgranular zone (SGZ, striatum, thalamus, and cerebral peduncle. Stereology-based analyses revealed significant exacerbation of OX6-positive activated microglial cells in the striatum, thalamus, and cerebral peduncle. In parallel, significant decrements in Ki67-positive proliferating cells in SVZ and SGZ, but only trends of reduced DCX-positive immature neuronal cells in SVZ and SGZ were detected relative to sham control group. These results indicate a progressive deterioration of the TBI brain over time characterized by elevated inflammation and suppressed neurogenesis. Therapeutic intervention at the chronic stage of TBI may confer abrogation of these deleterious cell death processes.

  17. Aqueous exposure to Aroclor 1254 modulates the mitogenic response of Atlantic salmon anterior kidney T-cells: Indications of short- and long-term immunomodulation

    International Nuclear Information System (INIS)

    Polychlorinated biphenyls (PCBs) exist as persistent organic pollutants in numerous river systems in the United States. Unfortunately, some of these rivers are sites of active Atlantic salmon restoration programs, and polychlorinated biphenyls have been implicated as ancillary factors contributing to failed salmon restoration. Here, we investigate the immediate and chronic effects of intermediate duration aqueous PCB exposure (1 or 10 μg L-1 Aroclor 1254) on the mitogen-stimulated lymphoproliferative response of Atlantic salmon anterior kidney leukocytes (AKLs). A short-term study was designed to examine immunomodulation in Atlantic salmon smolts immediately following 21 days of aqueous exposure, while a long-term study evaluated chronic impacts in the mitogen response in parr 15 months post-exposure as larvae. The proliferative response of AKLs to the mitogens concanavalin A (CON A), phytohemaglutinnin-P (PHA-P), pokeweed mitogen (PWM), and lipopolysaccharide were used as an indice of immunomodulation. The proliferative response to the T-cell mitogens CON A and PHA-P was significantly increased in the 10 μg L-1 group (n = 10; P = 0.043 and 0.002, respectively) immediately following exposure of smolts. Additionally, The PHA-P response was significantly increased in the 1 μg L-1 exposure group (n = 10, P = 0.036). In fish treated as larvae and tested 15 months later, the PHA-P sensitive populations exhibited elevated proliferation in the 1 and 10 μg L-1 groups (n = 12, P -1 treated groups. These results demonstrate an immunomodulatory effect of PCBs on T-cell mitogen sensitive populations of lymphocytes in Atlantic salmon as well as long-term immunomodulation in PHA-P and PWM sensitive populations

  18. Docosahexaenoic acid signaling modulates cell survival in experimental ischemic stroke penumbra and initiates long-term repair in young and aged rats.

    Directory of Open Access Journals (Sweden)

    Tiffany N Eady

    Full Text Available BACKGROUND: Docosahexaenoic acid, a major omega-3 essential fatty acid family member, improves behavioral deficit and reduces infarct volume and edema after experimental focal cerebral ischemia. We hypothesize that DHA elicits neuroprotection by inducing AKT/p70S6K phosphorylation, which in turn leads to cell survival and protects against ischemic stroke in young and aged rats. METHODS AND RESULTS: Rats underwent 2 h of middle cerebral artery occlusion (MCAo. DHA, neuroprotectin D1 (NPD1 or vehicle (saline was administered 3 h after onset of stroke. Neurological function was evaluated on days 1, 2, 3, and 7. DHA treatment improved functional recovery and reduced cortical, subcortical and total infarct volumes 7 days after stroke. DHA also reduced microglia infiltration and increased the number of astrocytes and neurons when compared to vehicle on days 1 and 7. Increases in p473 AKT and p308 AKT phosphorylation/activation were observed in animals treated with DHA 4 h after MCAo. Activation of other members of the AKT signaling pathway were also observed in DHA treated animals including increases in pS6 at 4 h and pGSK at 24 h. DHA or NPD1 remarkably reduced total and cortical infarct in aged rats. Moreover, we show that in young and aged rats DHA treatment after MCAo potentiates NPD1 biosynthesis. The phosphorylation of p308 AKT or pGSK was not different between groups in aged rats. However, pS6 expression was increased with DHA or NPD1 treatment when compared to vehicle. CONCLUSIONS: We suggest that DHA induces cell survival, modulates the neuroinflammatory response and triggers long term restoration of synaptic circuits. Both DHA and NPD1 elicited remarkable protection in aged animals. Accordingly, activation of DHA signaling might provide benefits in the management of ischemic stroke both acutely as well as long term to limit ensuing disabilities.

  19. Stem cell mobilisation by granulocyte-colony stimulating factor in patients with acute myocardial infarction. Long-term results of the REVIVAL-2 trial.

    Science.gov (United States)

    Steppich, Birgit; Hadamitzky, Martin; Ibrahim, Tareq; Groha, Philip; Schunkert, Heribert; Laugwitz, Karl-Ludwig; Kastrati, Adnan; Ott, Ilka

    2016-04-01

    Treatment with granulocyte-colony stimulating factor (G-CSF) mobilises cells from the bone marrow to the peripheral blood. Previous preclinical and early clinical trials may suggest that treatment with G-CSF leads to improved myocardial perfusion and function in acute or chronic ischaemic heart disease. In the REVIVAL-2 study we found that stem cell mobilisation by G-CSF does not influence infarct size, left ventricular function and coronary restenosis in patients with acute myocardial infarction (MI) that underwent successful percutaneous coronary intervention. The objective of the present analysis was to assess the impact of G-CSF treatment on seven-year clinical outcomes from the REVIVAL-2 trial. In the randomized, double-blind, placebo-controlled REVIVAL-2 study, 114 patients with the diagnosis of acute myocardial infarction were enrolled five days after successful reperfusion by percutaneous coronary intervention. Patients were assigned to receive 10 µg/kg G-CSF (n=56) or placebo (n=58) for five days. The primary endpoint for this long-term outcome analysis was the composite of death, myocardial infarction or stroke seven years after randomisation. The endpoint occurred in 14.3 % of patients in the G-CSF group versus 17.2 % assigned to placebo (p=0.67). The combined incidence of death or myocardial infarction occurred in 14.3 % of the patients assigned to G-CSF and 15.5 % of the patients assigned to placebo (p=0.85). In conclusion, these long-term follow-up data show that G-CSF does not improve clinical outcomes of patients with acute myocardial infarction.

  20. Differential microRNA expression signatures and cell type-specific association with Taxol resistance in ovarian cancer cells

    Directory of Open Access Journals (Sweden)

    Kim YW

    2014-02-01

    Full Text Available Yong-Wan Kim,1 Eun Young Kim,1 Doin Jeon,1 Juinn-Lin Liu,2 Helena Suhyun Kim,3 Jin Woo Choi,4 Woong Shick Ahn5 1Cancer Research Institute of Medical Science, The Catholic University of Korea, Seoul, Republic of Korea; 2Brain Tumor Center, Department of Neuro-Oncology, The University of Texas MD Anderson Cancer Center, TX, USA; 3Cancer Rehab Laboratory, RH Healthcare Systems Inc, TX, USA; 4Harvard Medical School and Wellman Center for Photomedicine, Cambridge, MA, USA; 5Department of Obstetrics and Gynecology, The Catholic University of Korea, Seoul, Republic of Korea Abstract: Paclitaxel (Taxol resistance remains a major obstacle for the successful treatment of ovarian cancer. MicroRNAs (miRNAs have oncogenic and tumor suppressor activity and are associated with poor prognosis phenotypes. miRNA screenings for this drug resistance are needed to estimate the prognosis of the disease and find better drug targets. miRNAs that were differentially expressed in Taxol-resistant ovarian cancer cells, compared with Taxol-sensitive cells, were screened by Illumina Human MicroRNA Expression BeadChips. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR was used to identify target genes of selected miRNAs. Kaplan–Meier survival analysis was applied to identify dysregulated miRNAs in ovarian cancer patients using data from The Cancer Genome Atlas. A total of 82 miRNAs were identified in ovarian carcinoma cells compared to normal ovarian cells. miR-141, miR-106a, miR-200c, miR-96, and miR-378 were overexpressed, and miR-411, miR-432, miR-494, miR-409-3p, and miR-655 were underexpressed in ovarian cancer cells. Seventeen miRNAs were overexpressed in Taxol-resistant cells, including miR-663, miR-622, and HS_188. Underexpressed miRNAs in Taxol-sensitive cells included miR-497, miR-187, miR-195, and miR-107. We further showed miR-663 and miR-622 as significant prognosis markers of the chemo-resistant patient group. In particular, the

  1. Characterisation of CD4 T cells in healthy and diseased koalas (Phascolarctos cinereus) using cell-type-specific monoclonal antibodies.

    Science.gov (United States)

    Mangar, Chandan; Armitage, Charles W; Timms, Peter; Corcoran, Lynn M; Beagley, Kenneth W

    2016-07-01

    The koala (Phascolarctos cinereus) is an arboreal herbivorous marsupial that is an Australian icon. Koalas in many parts of Australia are under multiple threats including habitat destruction, dog attacks, vehicular accidents, and infectious diseases such as Chlamydia spp. and the koala retrovirus (KoRV), which may contribute to the incidence of lymphoma and leukaemia in this species. Due to a lack of koala-specific immune reagents and assays there is currently no way to adequately analyse the immune response in healthy, diseased or vaccinated animals. This paper reports the production and characterisation of the first anti-koala CD4 monoclonal antibody (mAb). The koala CD4 gene was identified and used to develop recombinant proteins for mAb production. Fluorochrome-conjugated anti-CD4 mAb was used to measure the levels of CD4(+) lymphocytes collected from koala spleens (41.1%, range 20-45.1%) lymph nodes (36.3%, range 19-55.9%) and peripheral blood (23.8%, range 17.3-35%) by flow cytometry. Biotin-conjugated anti-CD4 mAb was used for western blot to determine an approximate size of 52 kDa for the koala CD4 molecule and used in immunohistochemistry to identify CD4(+) cells in the paracortical region and germinal centres of spleen and lymph nodes. Using the anti-CD4 mab we showed that CD4 cells from vaccinated, but not control, koalas proliferated following in vitro stimulation with UV-inactivated Chlamydia pecorum and recombinant chlamydial antigens. Since CD4(+) T cells have been shown to play a pivotal role in clearing chlamydial infection in both human and mouse infections, using this novel antibody will help determine the role CD4(+) T cells play in protection against chlamydial infection in koalas and also enhance our knowledge of how KoRV affects the koala immune system. PMID:26905635

  2. Long term results of postoperative Intensity-Modulated Radiation Therapy (IMRT) in the treatment of Squamous Cell Carcinoma (SCC) located in the oropharynx or oral cavity

    International Nuclear Information System (INIS)

    To report our long-term results with postoperative intensity-modulated radiation therapy (IMRT) in patients suffering from squamous-cell carcinoma (SCC) of the oral cavity or oropharynx. Seventy five patients were retrospectively analyzed. Median age was 58 years and 84 % were male. 76 % of the primaries were located in the oropharynx. Surgery resulted in negative margins (R0) in 64 % of the patients while 36 % suffered from positive margins (R1). Postoperative stages were as follows: stage1:4 %, stage2:9 %, stage3:17 %, stage4a:69 % with positive nodes in 84 %. Perineural invasion (Pn+) and extracapsular extension (ECE) were present in 7 % and 29 %, respectively. All patients received IMRT using the step-and-shoot approach with a simultaneously integrated boost (SIB) in 84 %. Concurrent systemic therapy was applied to 53 patients, mainly cisplatin weekly. Median follow-up was 55 months (5–150). 13 patients showed locoregional failures (4 isolated local, 4 isolated neck, 5 combined) transferring into 5-year-LRC rates of 85 %. Number of positive lymph nodes (n > 2) and presence of ECE were significantly associated with decreased LRC in univariate analysis, but only the number of nodes remained significant in multivariate analysis. Overall treatment failures occurred in 20 patients (9 locoregional only, 7 distant only, 4 combined), transferring into 3-and 5-year-FFTF rates of 77 % and 75 %, respectively. The 3-and 5-year-OS rates were 80 % and 72 %, respectively. High clinical stage, high N stage, number of positive nodes (n > 2), ECE and Pn1 were significantly associated with worse FFTF and OS in univariate analysis, but only number of nodes remained significant for FFTF in multivariate analysis. Maximum acute toxicity was grade 3 in 64 % and grade 4 in 1 %, mainly hematological or mucositis/dysphagia. Maximum late toxicity was grade 3 in 23 % of the patients, mainly long-term tube feeding dependency. Postoperative IMRT achieved excellent LRC and good OS with

  3. Prognostic model for long-term survival of locally advanced non-small-cell lung cancer patients after neoadjuvant radiochemotherapy and resection integrating clinical and histopathologic factors

    International Nuclear Information System (INIS)

    Outcome of consecutive patients with locally advanced non-small cell lung cancer and histopathologically proven mediastional lymph node metastases treated with induction chemotherapy, neoadjuvant radiochemotherapy and thoracotomy at the West German Cancer Center between 08/2000 and 06/2012 was analysed. A clinico-pathological prognostic model for survival was built including partial or complete response according to computed tomography imaging (CT) as clinical parameters as well as pathologic complete remission (pCR) and mediastinal nodal clearance (MNC) as histopathologic factors. Proportional hazard analysis (PHA) and recursive partitioning analysis (RPA) were used to identify prognostic factors for survival. Long-term survival was defined as survival ≥ 36 months. A total of 157 patients were treated, median follow-up was 97 months. Among these patients, pCR and MNC were observed in 41 and 85 patients, respectively. Overall survival was 56 ± 4% and 36 ± 4% at 24 and 60 months, respectively. Sensitivities of pCR and MNC to detect long-term survivors were 38% and 61%, specificities were 84% and 52%, respectively. Multivariable survival analysis revealed pCR, cN3 category, and gender, as prognostic factors at a level of α < 0.05. Considering only preoperative available parameters, CT response became significant. Classifying patients with a predicted hazard above the median as high risk group and the remaining as low risk patients yielded better separation of the survival curves by the inclusion of histopathologic factors than by preoperative factors alone (p < 0.0001, log rank test). Using RPA, pCR was identified as the top prognostic factor above clinical factors (p = 0.0006). No long term survivors were observed in patients with cT3-4 cN3 tumors without pCR. pCR is the dominant histopathologic response parameter and improves prognostic classifiers, based on clinical parameters. The validated prognostic model can be used to estimate individual prognosis and

  4. Dose-dependent and cell type-specific cell death and proliferation following in vitro exposure to radial extracorporeal shock waves.

    Science.gov (United States)

    Hochstrasser, Tanja; Frank, Hans-Georg; Schmitz, Christoph

    2016-01-01

    Radial extracorporeal shock wave (rESW) therapy is widely used in musculoskeletal disorders and wound repair. However, the mechanisms of action are still largely unknown. The current study compared the effects of rESWs on two cell types. Human fetal foreskin fibroblasts (HFFF2) and human placental choriocarcinoma cell line JEG-3 were exposed to 0, 100, 200, 500 or 5000 rESWs generated with a Swiss DolorClast device (2.5 bar, 1 Hz). FACS analysis immediately after rESW exposure showed that initially, rESWs rather induced mechanical cell destruction than regulated or programmed cell death. Cell damage was nearly negated by reducing cavitation. Furthermore, cell viability decreased progressively with higher numbers of rESWs. Exposure to rESWs had no impact on growth potential of JEG-3 cells, but dose-dependently increased growth potential of HFFF2 cells. Cultivation of cells that were initially exposed to sham-rESWs in conditioned media increased the growth potential of HFFF2 cells, nevertheless, an even stronger effect was achieved by direct exposure to rESWs. Additionally, cell cycle distribution analysis demonstrated a shift in proportion from G0/G1 to G2/M phase in HFFF2 cells, but not in JEG-3 cells. These data demonstrate that rESWs leads to initial and subsequent dose-dependent and cell type-specific effects in vitro. PMID:27477873

  5. Comparative Analysis of Short- and Long-Term Changes in Gene Expression Caused by Low Water Potential in Potato (Solanum tuberosum) Cell-Suspension Cultures.

    Science.gov (United States)

    Leone, A.; Costa, A.; Tucci, M.; Grillo, S.

    1994-10-01

    To dissect the cellular response to water stress and compare changes induced as a generalized response with those involved in tolerance/acclimation mechanisms, we analyzed changes in two-dimensional electrophoretic patterns of in vivo [35S]methionine-labeled polypeptides of cultured potato (Solanum tuberosum) cells after gradual and long exposure to polyethylene glycol (PEG)- mediated low water potential versus those induced in cells abruptly exposed to the same stress intensity. Protein synthesis was not inhibited by gradual stress imposition, and the expression of 17 proteins was induced in adapted cells. Some polypeptides were inducible under mild stress conditions (5% PEG) and accumulated further when cells were exposed to a higher stress intensity (10 and 20% PEG). The synthesis of another set of polypeptides was up-regulated only when more severe water-stress conditions were applied, suggesting that plant cells were able to monitor different levels of stress intensity and modulate gene expression accordingly. In contrast, in potato cells abruptly exposed to 20% PEG, protein synthesis was strongly inhibited. Nevertheless, a large set of polypeptides was identified whose expression was increased. Most of these polypeptides were not induced in adapted cells, but many of them were common to those observed in abscisic acid (ABA)-treated cells. These data, along with the finding that cellular ABA content increased in PEG-shocked cells but not in PEG-adapted cells, suggested that this hormone is mainly involved in the rapid response to stress rather than long-term adaptation. A further group of proteins included those induced after long exposure to both water stress and shock. Western blot analysis revealed that osmotin was one protein belonging to this common group. This class may represent induced proteins that accumulate specifically in response to low water potential and that are putatively involved in the maintenance of cellular homeostasis under prolonged

  6. Integrative modeling of eQTLs and cis-regulatory elements suggests mechanisms underlying cell type specificity of eQTLs.

    Directory of Open Access Journals (Sweden)

    Christopher D Brown

    Full Text Available Genetic variants in cis-regulatory elements or trans-acting regulators frequently influence the quantity and spatiotemporal distribution of gene transcription. Recent interest in expression quantitative trait locus (eQTL mapping has paralleled the adoption of genome-wide association studies (GWAS for the analysis of complex traits and disease in humans. Under the hypothesis that many GWAS associations tag non-coding SNPs with small effects, and that these SNPs exert phenotypic control by modifying gene expression, it has become common to interpret GWAS associations using eQTL data. To fully exploit the mechanistic interpretability of eQTL-GWAS comparisons, an improved understanding of the genetic architecture and causal mechanisms of cell type specificity of eQTLs is required. We address this need by performing an eQTL analysis in three parts: first we identified eQTLs from eleven studies on seven cell types; then we integrated eQTL data with cis-regulatory element (CRE data from the ENCODE project; finally we built a set of classifiers to predict the cell type specificity of eQTLs. The cell type specificity of eQTLs is associated with eQTL SNP overlap with hundreds of cell type specific CRE classes, including enhancer, promoter, and repressive chromatin marks, regions of open chromatin, and many classes of DNA binding proteins. These associations provide insight into the molecular mechanisms generating the cell type specificity of eQTLs and the mode of regulation of corresponding eQTLs. Using a random forest classifier with cell specific CRE-SNP overlap as features, we demonstrate the feasibility of predicting the cell type specificity of eQTLs. We then demonstrate that CREs from a trait-associated cell type can be used to annotate GWAS associations in the absence of eQTL data for that cell type. We anticipate that such integrative, predictive modeling of cell specificity will improve our ability to understand the mechanistic basis of human

  7. Long-term results of low dose daily cisplatin chemotherapy used concurrently with modestly accelerated radiotherapy in locally advanced squamous cell carcinomas of the head neck cancer region

    Directory of Open Access Journals (Sweden)

    Pramod Kumar Gupta

    2014-01-01

    Full Text Available Introduction: Concurrent single agent cisplatin (CDDP with radiotherapy (RT improves outcomes in locally advanced squamous cell carcinomas of the head neck (LA-SCCHN. CDDP at 100 mg/m 2 at 3 weekly intervals raise compliance, hospitalization, and supportive care issues. Low dose daily CDDP was delivered with RT to evaluate its compliance, long-term safety and efficacy. Patients and Methods: During the period of month between November 2005 and May 2007, 52 patients of stage III/IV LA-SCCHN were given with conventional RT in a phased manner (dose-70 Gy/35 fractions/6 weeks along with daily CDDP (6 mg/m 2 ; capped 10 mg-30 cycles over 6 weeks. No hospitalization or antiemetic cover was planned. Compliance, acute and late toxicity were recorded as per Radiation Therapy Oncology Group/European Organization for Research and Treatment of Cancer grading system and survival outcomes were evaluated. Results : The median follow-up was 63 months. 43 (83% cases complied with RT schedule and >28 cycles of CDDP was administered in 38 (73% cases. Confluent mucositis was seen in 65%, Grade III/IV dysphagia in 67%; 77% required enteral feed and hospitalization in 15%. There were four treatment related deaths. At 5 years, the loco-regional control was 25% (median-11 months and the overall survival was 31% (median-11 months. The 5 years actuarial rates of late Grade III/IV toxicity was 24%. Late swallowing difficulty/aspiration were seen in 17%; xerostomia-40%; ototoxicity-6%; nephrotoxicity-4%; and no second malignancy. Conclusion: Low dose cisplatin with moderately accelerated RT schedule appears feasible and logistically suitable "out-patient" option without increasing long-term toxicity in LA-SCCHN cancer region.

  8. Long-term results of a randomized phase III trial of TPF induction chemotherapy followed by surgery and radiation in locally advanced oral squamous cell carcinoma.

    Science.gov (United States)

    Zhong, Lai-ping; Zhang, Chen-ping; Ren, Guo-xin; Guo, Wei; William, William N; Hong, Christopher S; Sun, Jian; Zhu, Han-guang; Tu, Wen-yong; Li, Jiang; Cai, Yi-li; Yin, Qiu-ming; Wang, Li-zhen; Wang, Zhong-he; Hu, Yong-jie; Ji, Tong; Yang, Wen-jun; Ye, Wei-min; Li, Jun; He, Yue; Wang, Yan-an; Xu, Li-qun; Zhuang, Zhengping; Lee, J Jack; Myers, Jeffrey N; Zhang, Zhi-yuan

    2015-07-30

    Previously, we conducted a randomized phase III trial of TPF (docetaxel, cisplatin, and 5-fluorouracil) induction chemotherapy in surgically managed locally advanced oral squamous cell carcinoma (OSCC) and found no improvement in overall survival. This study reports long-term follow-up results from our initial trial. All patients had clinical stage III or IVA locally advanced OSCC. In the experimental group, patients received two cycles of TPF induction chemotherapy (75mg/m2 docetaxel d1, 75mg/m2 cisplatin d1, and 750mg/m2/day 5-fluorouracil d1-5) followed by radical surgery and post-operative radiotherapy; in the control group, patients received upfront radical surgery and post-operative radiotherapy. The primary endpoint was overall survival. Among 256 enrolled patients with a median follow-up of 70 months, estimated 5-year overall survival, disease-free survival, locoregional recurrence-free survival, and distant metastasis-free survival rates were 61.1%, 52.7%, 55.2%, and 60.4%, respectively. There were no significant differences in survival rates between experimental and control groups. However, patients with favorable pathologic responses had improved outcomes compared to those with unfavorable pathologic responses and to those in the control group. Although TPF induction chemotherapy did not improve long-term survival compared to surgery upfront in patients with stage III and IVA OSCC, a favorable pathologic response after induction chemotherapy may be used as a major endpoint and prognosticator in future studies. Furthermore, the negative results observed in this trial may be represent type II error from an underpowered study. Future larger scale phase III trials are warranted to investigate whether a significant benefit exists for TPF induction chemotherapy in surgically managed OSCC. PMID:26124084

  9. Regulation of insulin-like growth factor-binding protein-1 synthesis and secretion by progestin and relaxin in long term cultures of human endometrial stromal cells

    International Nuclear Information System (INIS)

    The decidualized endometrium during the first trimester of pregnancy synthesizes and secretes a 32-kDa insulin-like growth factor-binding protein (termed hIGFBP-1) at high levels. IGFBP-1 is the major soluble protein product of this tissue and is principally localized to the differentiated endometrial stromal cell, the decidual cell. In the present study long term culture of stromal cells from the nonpregnant endometrium have been employed to elucidate the hormonal requirements for IGFBP-1 production. Immunoreactive IGFBP-1 was undetectable in control cultures. However, inclusion of medroxyprogesterone acetate (MPA) induced rates of 0.35 +/- 0.09 microgram/0.1 mg cell DNA.day after 20-30 days. In these cultures cells exhibited morphological changes consistent with decidual cell differentiation. In all cultures removal of MPA after exposure for 10-16 days, with or without subsequent inclusion of relaxin (RLX), increased production of IGFBP-1 450- to 4600-fold to rates of 150-710 micrograms/0.1 mg cell DNA.day or 26-131 micrograms/10(6) cells.day on days 24-26. The rates tended to be higher with the inclusion of RLX and were sustained in contrast to cultures without RLX, where rates fell by day 30. Individual cultures responded differently to RLX when added from the initiation of culture, with either a response similar to MPA alone or a cyclical change in production, achieving maximal rates of 190-290 micrograms/0.1 mg cell DNA.day. Cultures in which RLX alone induced high IGFBP-1 high production were obtained from endometrium during the progesterone-dominated luteal phase. In cultures exhibiting high rates of immunoreactive IGFBP-1 production, the protein represented their major secretory protein product. This was confirmed by [35S]methionine incorporation and the presence of IGFBP-1 as the predominant protein in serum-free culture medium

  10. Regulation of insulin-like growth factor-binding protein-1 synthesis and secretion by progestin and relaxin in long term cultures of human endometrial stromal cells

    Energy Technology Data Exchange (ETDEWEB)

    Bell, S.C.; Jackson, J.A.; Ashmore, J.; Zhu, H.H.; Tseng, L. (Univ. of Leicester, (United Kingdom))

    1991-05-01

    The decidualized endometrium during the first trimester of pregnancy synthesizes and secretes a 32-kDa insulin-like growth factor-binding protein (termed hIGFBP-1) at high levels. IGFBP-1 is the major soluble protein product of this tissue and is principally localized to the differentiated endometrial stromal cell, the decidual cell. In the present study long term culture of stromal cells from the nonpregnant endometrium have been employed to elucidate the hormonal requirements for IGFBP-1 production. Immunoreactive IGFBP-1 was undetectable in control cultures. However, inclusion of medroxyprogesterone acetate (MPA) induced rates of 0.35 +/- 0.09 microgram/0.1 mg cell DNA.day after 20-30 days. In these cultures cells exhibited morphological changes consistent with decidual cell differentiation. In all cultures removal of MPA after exposure for 10-16 days, with or without subsequent inclusion of relaxin (RLX), increased production of IGFBP-1 450- to 4600-fold to rates of 150-710 micrograms/0.1 mg cell DNA.day or 26-131 micrograms/10(6) cells.day on days 24-26. The rates tended to be higher with the inclusion of RLX and were sustained in contrast to cultures without RLX, where rates fell by day 30. Individual cultures responded differently to RLX when added from the initiation of culture, with either a response similar to MPA alone or a cyclical change in production, achieving maximal rates of 190-290 micrograms/0.1 mg cell DNA.day. Cultures in which RLX alone induced high IGFBP-1 high production were obtained from endometrium during the progesterone-dominated luteal phase. In cultures exhibiting high rates of immunoreactive IGFBP-1 production, the protein represented their major secretory protein product. This was confirmed by ({sup 35}S)methionine incorporation and the presence of IGFBP-1 as the predominant protein in serum-free culture medium.

  11. Long-Term Safety Issues of iPSC-Based Cell Therapy in a Spinal Cord Injury Model: Oncogenic Transformation with Epithelial-Mesenchymal Transition

    Directory of Open Access Journals (Sweden)

    Satoshi Nori

    2015-03-01

    Full Text Available Previously, we described the safety and therapeutic potential of neurospheres (NSs derived from a human induced pluripotent stem cell (iPSC clone, 201B7, in a spinal cord injury (SCI mouse model. However, several safety issues concerning iPSC-based cell therapy remain unresolved. Here, we investigated another iPSC clone, 253G1, that we established by transducing OCT4, SOX2, and KLF4 into adult human dermal fibroblasts collected from the same donor who provided the 201B7 clone. The grafted 253G1-NSs survived, differentiated into three neural lineages, and promoted functional recovery accompanied by stimulated synapse formation 47 days after transplantation. However, long-term observation (for up to 103 days revealed deteriorated motor function accompanied by tumor formation. The tumors consisted of Nestin+ undifferentiated neural cells and exhibited activation of the OCT4 transgene. Transcriptome analysis revealed that a heightened mesenchymal transition may have contributed to the progression of tumors derived from grafted cells.

  12. Continuous Fermentation of Clostridium tyrobutyricum with Partial Cell Recycle as a Long-Term Strategy for Butyric Acid Production

    Directory of Open Access Journals (Sweden)

    Edgar C. Clausen

    2012-08-01

    Full Text Available In making alternative fuels from biomass feedstocks, the production of butyric acid is a key intermediate in the two-step production of butanol. The fermentation of glucose via Clostridium tyrobutyricum to butyric acid produces undesirable byproducts, including lactic acid and acetic acid, which significantly affect the butyric acid yield and productivity. This paper focuses on the production of butyric acid using Clostridium tyrobutyricum in a partial cell recycle mode to improve fermenter yield and productivity. Experiments with fermentation in batch, continuous culture and continuous culture with partial cell recycle by ultrafiltration were conducted. The results show that a continuous fermentation can be sustained for more than 120 days, which is the first reported long-term production of butyric acid in a continuous operation. Further, the results also show that partial cell recycle via membrane ultrafiltration has a great influence on the selectivity and productivity of butyric acid, with an increase in selectivity from ≈9% to 95% butyric acid with productivities as high as 1.13 g/Lh. Continuous fermentation with low dilution rate and high cell recycle ratio has been found to be desirable for optimum productivity and selectivity toward butyric acid and a comprehensive model explaining this phenomenon is given.

  13. Magnetic resonance hypointensive signal primarily originates from extracellular iron particles in the long-term tracking of mesenchymal stem cells transplanted in the infarcted myocardium

    Directory of Open Access Journals (Sweden)

    Huang Z

    2015-03-01

    Full Text Available Zheyong Huang,1,* Chenguang Li,1,* Shan Yang,2 Jianfeng Xu,1 Yunli Shen,3 Xinxing Xie,4 Yuxiang Dai,1 Hao Lu,1 Hui Gong,5 Aijun Sun,1 Juying Qian,1 Junbo Ge1 1Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University, Shanghai, People’s Republic of China; 2Department of Radiology, Zhongshan Hospital, Fudan University, Shanghai, People’s Republic of China; 3Department of Cardiology, Shanghai East Hospital, Tongji University, Shanghai, People’s Republic of China; 4Department of Cardiology, Qianfoshan Hospital, Shandong University, Jinan, Shandong Province, People’s Republic of China; 5Institute of Biomedical Science, Fudan University, Shanghai, People’s Republic of China *These authors contributed equally to this work Purpose: The long-lasting hypointensities in cardiac magnetic resonance (CMR were believed to originate from superparamagnetic iron oxide (SPIO-engulfed macrophages during long-term stem cell tracking. However, the iron clearance capacity of the ischemic heart was limited. Therefore, we speculated that the extracellular SPIO particles may also be involved in the generation of false-positive signals.Methods and results: Male swine mesenchymal stem cells (MSCs were incubated with SPIO for 24 hours, and SPIO labeling had no significant effects on either cell viability or differentiation. In vitro studies showed that magnetic resonance failed to distinguish SPIO from living SPIO-MSCs or dead SPIO-MSCs. Two hours after the establishment of the female swine acute myocardial infarction model, 2×107 male SPIO-labeled MSCs (n=5 or unlabeled MSCs (n=5 were transextracardially injected into the infarcted myocardium at ten distinct sites. In vivo CMR with T2 star weighted imaging-flash-2D sequence revealed a signal void corresponding to the initial SPIO-MSC injection sites. At 6 months after transplantation, CMR identified 32 (64% of the 50 injection sites, where massive Prussian blue-positive iron

  14. Long-Term Fate Mapping Using Conditional Lentiviral Vectors Reveals a Continuous Contribution of Radial Glia-Like Cells to Adult Hippocampal Neurogenesis in Mice.

    Directory of Open Access Journals (Sweden)

    Sarah-Ann Aelvoet

    Full Text Available Newborn neurons are generated throughout life in two neurogenic regions, the subventricular zone and the hippocampal dentate gyrus. Stimulation of adult neurogenesis is considered as an attractive endogenous repair mechanism to treat different neurological disorders. Although tremendous progress has been made in our understanding of adult hippocampal neurogenesis, important questions remain unanswered, regarding the identity and the behavior of neural stem cells in the dentate gyrus. We previously showed that conditional Cre-Flex lentiviral vectors can be used to label neural stem cells in the subventricular zone and to track the migration of their progeny with non-invasive bioluminescence imaging. Here, we applied these Cre-Flex lentiviral vectors to study neurogenesis in the dentate gyrus with bioluminescence imaging and histological techniques. Stereotactic injection of the Cre-Flex vectors into the dentate gyrus of transgenic Nestin-Cre mice resulted in specific labeling of the nestin-positive neural stem cells. The labeled cell population could be detected with bioluminescence imaging until 9 months post injection, but no significant increase in the number of labeled cells over time was observed with this imaging technique. Nevertheless, the specific labeling of the nestin-positive neural stem cells, combined with histological analysis at different time points, allowed detailed analysis of their neurogenic potential. This long-term fate mapping revealed that a stable pool of labeled nestin-positive neural stem cells continuously contributes to the generation of newborn neurons in the mouse brain until 9 months post injection. In conclusion, the Cre-Flex technology is a valuable tool to address remaining questions regarding neural stem cell identity and behavior in the dentate gyrus.

  15. The Effect of Long-Term Exercise on the Production of Osteoclastogenic and Antiosteoclastogenic Cytokines by Peripheral Blood Mononuclear Cells and on Serum Markers of Bone Metabolism

    Directory of Open Access Journals (Sweden)

    J. Kelly Smith

    2016-01-01

    Full Text Available Although it is recognized that the mechanical stresses associated with physical activity augment bone mineral density and improve bone quality, our understanding of how exercise modulates bone homeostasis at the molecular level is lacking. In a before and after trial involving 43 healthy adults, we measured the effect of six months of supervised exercise training on the spontaneous and phytohemagglutinin-induced production of osteoclastogenic cytokines (interleukin-1α, tumor necrosis factor-α, antiosteoclastogenic cytokines (transforming growth factor-β1 and interleukins 4 and 10, pleiotropic cytokines with variable effects on osteoclastogenesis (interferon-γ, interleukin-6, and T cell growth and differentiation factors (interleukins 2 and 12 by peripheral blood mononuclear cells. We also measured lymphocyte phenotypes and serum markers of bone formation (osteocalcin, bone resorption (C-terminal telopeptides of Type I collagen, and bone homeostasis (25 (OH vitamin D, estradiol, testosterone, parathyroid hormone, and insulin-like growth factor 1. A combination of aerobic, resistance, and flexibility exercises done on average of 2.5 hours a week attenuated the production of osteoclastogenic cytokines and enhanced the production of antiosteoclastogenic cytokines. These changes were accompanied by a 16% reduction in collagen degradation products and a 9.8% increase in osteocalcin levels. We conclude that long-term moderate intensity exercise exerts a favorable effect on bone resorption by changing the balance between blood mononuclear cells producing osteoclastogenic cytokines and those producing antiosteoclastogenic cytokines. This trial is registered with Clinical Trials.gov Identifier: NCT02765945.

  16. Prenatal stress induces long-term effects in cell turnover in the hippocampus-hypothalamus-pituitary axis in adult male rats.

    Directory of Open Access Journals (Sweden)

    Eva Baquedano

    Full Text Available Subchronic gestational stress leads to permanent modifications in the hippocampus-hypothalamus-pituitary-adrenal axis of offspring probably due to the increase in circulating glucocorticoids known to affect prenatal programming. The aim of this study was to investigate whether cell turnover is affected in the hippocampus-hypothalamus-pituitary axis by subchronic prenatal stress and the intracellular mechanisms involved. Restraint stress was performed in pregnant rats during the last week of gestation (45 minutes; 3 times/day. Only male offspring were used for this study and were sacrificed at 6 months of age. In prenatally stressed adults a decrease in markers of cell death and proliferation was observed in the hippocampus, hypothalamus and pituitary. This was associated with an increase in insulin-like growth factor-I mRNA levels, phosphorylation of CREB and calpastatin levels and inhibition of calpain -2 and caspase -8 activation. Levels of the anti-apoptotic protein Bcl-2 were increased and levels of the pro-apoptotic factor p53 were reduced. In conclusion, prenatal restraint stress induces a long-term decrease in cell turnover in the hippocampus-hypothalamus-pituitary axis that might be at least partly mediated by an autocrine-paracrine IGF-I effect. These changes could condition the response of this axis to future physiological and pathophysiological situations.

  17. Prenatal Stress Induces Long-Term Effects in Cell Turnover in the Hippocampus-Hypothalamus-Pituitary Axis in Adult Male Rats

    Science.gov (United States)

    Baquedano, Eva; García-Cáceres, Cristina; Diz-Chaves, Yolanda; Lagunas, Natalia; Calmarza-Font, Isabel; Azcoitia, Iñigo; Garcia-Segura, Luis M.; Argente, Jesús; Chowen, Julie A.; Frago, Laura M.

    2011-01-01

    Subchronic gestational stress leads to permanent modifications in the hippocampus-hypothalamus-pituitary-adrenal axis of offspring probably due to the increase in circulating glucocorticoids known to affect prenatal programming. The aim of this study was to investigate whether cell turnover is affected in the hippocampus-hypothalamus-pituitary axis by subchronic prenatal stress and the intracellular mechanisms involved. Restraint stress was performed in pregnant rats during the last week of gestation (45 minutes; 3 times/day). Only male offspring were used for this study and were sacrificed at 6 months of age. In prenatally stressed adults a decrease in markers of cell death and proliferation was observed in the hippocampus, hypothalamus and pituitary. This was associated with an increase in insulin-like growth factor-I mRNA levels, phosphorylation of CREB and calpastatin levels and inhibition of calpain -2 and caspase -8 activation. Levels of the anti-apoptotic protein Bcl-2 were increased and levels of the pro-apoptotic factor p53 were reduced. In conclusion, prenatal restraint stress induces a long-term decrease in cell turnover in the hippocampus-hypothalamus-pituitary axis that might be at least partly mediated by an autocrine-paracrine IGF-I effect. These changes could condition the response of this axis to future physiological and pathophysiological situations. PMID:22096592

  18. Long term effects of the implantation of Wharton's jelly-derived mesenchymal stem cells from the umbilical cord for newly-onset type 1 diabetes mellitus.

    Science.gov (United States)

    Hu, Jianxia; Yu, Xiaolong; Wang, Zhongchao; Wang, Fang; Wang, Li; Gao, Hong; Chen, Ying; Zhao, Wenjuan; Jia, Zhaotong; Yan, Shengli; Wang, Yangang

    2013-01-01

    Type 1 diabetes mellitus (T1DM) is an autoimmune disorder resulted from T cell-mediated destruction of pancreatic β-cells, how to regenerate β-cells and prevent the autoimmune destruction of remnant and neogenetic β-cells is a tough problem. Immunomodulatory propertity of mesenchymal stem cell make it illuminated to overcome it. We assessed the long-term effects of the implantation of Wharton's jelly-derived mesenchymal stem cells (WJ-MSCs) from the umbilical cord for Newly-onset T1DM. Twenty-nine patients with newly onset T1DM were randomly divided into two groups, patients in group I were treated with WJ-MSCs and patients in group II were treated with normal saline based on insulin intensive therapy. Patients were followed-up after the operation at monthly intervals for the first 3 months and thereafter every 3 months for the next 21 months, the occurrence of any side effects and results of laboratory examinations were evaluated. There were no reported acute or chronic side effects in group I compared with group II, both the HbA1c and C peptide in group I patients were significantly better than either pretherapy values or group II patients during the follow-up period. These data suggested that the implantation of WJ-MSCs for the treatment of newly-onset T1DM is safe and effective. This therapy can restore the function of islet β cells in a longer time, although precise mechanisms are unknown, the implantation of WJ-MSCs is expected to be an effective strategy for treatment of type1 diabetes.

  19. Long-term exposure to high levels of decabrominated diphenyl ether inhibits CD4 T-cell functions in C57Bl/6 mice.

    Science.gov (United States)

    Feng, Yan; Zeng, Weihong; Wang, Ying; Shen, Hao; Wang, Yan

    2016-09-01

    In recent years, the adverse health effects of decabrominated diphenyl ether (BDE-209) have raised more concerns as a growing number of studies reported its persistence in the environment and abundance in the human population, especially in occupational environmental compartments and exposed personnel. This study applies our previous animal model simulating occupational exposure to BDE-209 to investigate its potential adverse effects on CD4 T cells. Female C57Bl/6 mice were orally gavaged with BDE-209 at a dose of 800 mg kg(-1) every 2 days for 10 months and the blood of each mouse was collected for analysis. Kinetic changes of the peripheral immune system were investigated from 1 to 5 months of exposure. The chronic effects on cytokine production, proliferation and the antigen-specific responses of CD4 T cells were evaluated at 7, 9 and 10 months, respectively. The results have shown that impaired proliferation and cytokine (IFN-γ, IL-2 or TNF-α) production of CD4 T cells were observed in BDE-209-exposed mice, accompanied by increased T regulatory cells in the blood. BDE-209 exposure in vitro also suppressed the reactivity of CD4 T cells at concentrations of 0.01, 0.1, 1 and 10 μM. Furthermore, we observed weaker antigen-specific CD4 T-cell responses to Listeria monocytogenes infection in the mice exposed to BDE-209, suggesting decreased resistance to exogenous pathogens. Taken together, these observations indicate an impaired cellular immunity after long-term and relative high-dose exposure to BDE-209 in adult mice. Copyright © 2015 John Wiley & Sons, Ltd. PMID:26682527

  20. miR-155 Drives Metabolic Reprogramming of ER+ Breast Cancer Cells Following Long-Term Estrogen Deprivation and Predicts Clinical Response to Aromatase Inhibitors.

    Science.gov (United States)

    Bacci, Marina; Giannoni, Elisa; Fearns, Antony; Ribas, Ricardo; Gao, Qiong; Taddei, Maria Letizia; Pintus, Gianfranco; Dowsett, Mitch; Isacke, Clare M; Martin, Lesley-Ann; Chiarugi, Paola; Morandi, Andrea

    2016-03-15

    Aromatase inhibitors (AI) have become the first-line endocrine treatment of choice for postmenopausal estrogen receptor-positive (ER(+)) breast cancer patients, but resistance remains a major challenge. Metabolic reprogramming is a hallmark of cancer and may contribute to drug resistance. Here, we investigated the link between altered breast cancer metabolism and AI resistance using AI-resistant and sensitive breast cancer cells, patient tumor samples, and AI-sensitive human xenografts. We found that long-term estrogen deprivation (LTED), a model of AI resistance, was associated with increased glycolysis dependency. Targeting the glycolysis-priming enzyme hexokinase-2 (HK2) in combination with the AI, letrozole, synergistically reduced cell viability in AI-sensitive models. Conversely, MCF7-LTED cells, which displayed a high degree of metabolic plasticity, switched to oxidative phosphorylation when glycolysis was impaired. This effect was ER dependent as breast cancer cells with undetectable levels of ER failed to exhibit metabolic plasticity. MCF7-LTED cells were also more motile than their parental counterparts and assumed amoeboid-like invasive abilities upon glycolysis inhibition with 2-deoxyglucose (2-DG). Mechanistic investigations further revealed an important role for miR-155 in metabolic reprogramming. Suppression of miR-155 resulted in sensitization of MCF7-LTED cells to metformin treatment and impairment of 2-DG-induced motility. Notably, high baseline miR-155 expression correlated with poor response to AI therapy in a cohort of ER(+) breast cancers treated with neoadjuvant anastrozole. These findings suggest that miR-155 represents a biomarker potentially capable of identifying the subset of breast cancers most likely to adapt to and relapse on AI therapy. PMID:26795347

  1. Adverse Fat Depots and Marrow Adiposity Are Associated With Skeletal Deficits and Insulin Resistance in Long-Term Survivors of Pediatric Hematopoietic Stem Cell Transplantation.

    Science.gov (United States)

    Mostoufi-Moab, Sogol; Magland, Jeremy; Isaacoff, Elizabeth J; Sun, Wenli; Rajapakse, Chamith S; Zemel, Babette; Wehrli, Felix; Shekdar, Karuna; Baker, Joshua; Long, Jin; Leonard, Mary B

    2015-09-01

    Allogeneic hematopoietic stem-cell transplantation (alloHSCT) survivors treated with total body irradiation (TBI) exhibit bone deficits and excess adiposity, potentially related to altered mesenchymal stem cell differentiation into osteoblasts or adipocytes. We examined associations among fat distribution, bone microarchitecture, and insulin resistance in alloHSCT survivors after TBI. This was a cross-sectional observational study of 25 alloHSCT survivors (aged 12 to 25 years) a median of 9.7 (4.3 to 19.3) years after alloHSCT compared to 25 age-, race-, and sex-matched healthy controls. Vertebral MR spectroscopic imaging and tibia micro-MRI were used to quantify marrow adipose tissue (MAT) and trabecular microarchitecture. Additional measures included DXA whole-body fat mass (WB-FM), leg lean mass (Leg-LM), trunk visceral adipose tissue (VAT), and CT calf muscle density. Insulin resistance in alloHSCT survivors was estimated by HOMA-IR. AlloHSCT survivors had lower Leg-LM (p treatment-related morbidity and mortality in alloHSCT recipients after TBI. Trabecular deterioration was associated with marrow and visceral adiposity. Furthermore, long-term survivors demonstrated sarcopenic obesity, insulin resistance, and vertebral deformities. Future studies are needed to identify strategies to prevent and treat metabolic and skeletal complications in this growing population of childhood alloHSCT survivors. PMID:25801428

  2. Porcine adipose tissue-derived mesenchymal stem cells retain their proliferative characteristics, senescence, karyotype and plasticity after long-term cryopreservation.

    Directory of Open Access Journals (Sweden)

    Rafael Dariolli

    Full Text Available We and others have provided evidence that adipose tissue-derived mesenchymal stem cells (ASCs can mitigate rat cardiac functional deterioration after myocardial ischemia, even though the mechanism of action or the relevance of these findings to human conditions remains elusive. In this regard, the porcine model is a key translational step, because it displays heart anatomic-physiological features that are similar to those found in the human heart. Towards this end, we wanted to establish the cultural characteristics of porcine ASCs (pASCs with or without long-term cryostorage, considering that allogeneic transplantation may also be a future option. Compared to fresh pASCs, thawed cells displayed 90-95% viability and no changes in morphological characteristics or in the expression of surface markers (being pASCs characterized by positive markers CD29(+; CD90(+; CD44(+; CD140b(+; CD105(+; and negative markers CD31(-; CD34(-; CD45(- and SLA-DR(-; n = 3. Mean population doubling time was also comparable (64.26±15.11 hours to thawed cells vs. 62.74±18.07 hours to fresh cells and cumulative population doubling increased constantly until Passage 10 (P10 in the entire cell population, with a small and gradual increase in senescence (P5, 3.25%±0.26 vs. 3.47%±0.32 and P10, 9.6%±0.29 vs. 10.67%±1.25, thawed vs. fresh; SA-β-Gal staining. Chromosomal aberrations were not observed. In addition, under both conditions pASCs responded to adipogenic and osteogenic chemical cues in vitro. In conclusion, we have demonstrated the growth characteristics, senescence, and the capacity of pASCs to respond to chemical cues in vitro and have provided evidence that these properties are not influenced by cryostorage in 10% DMSO solution.

  3. Brief embryonic cadmium exposure induces a stress response and cell death in the developing olfactory system followed by long-term olfactory deficits in juvenile zebrafish

    International Nuclear Information System (INIS)

    The toxic effects of cadmium and other metals have been well established. A primary target of these metals is known to be the olfactory system, and fish exposed to a number of different waterborne metals display deficiencies in olfaction. Importantly, exposure over embryonic/larval development periods can cause deficits in chemosensory function in juvenile fish, but the specific cell types affected are unknown. We have previously characterized a transgenic zebrafish strain expressing the green fluorescent protein (eGFP) gene linked to the hsp70 gene promoter, and shown it to be a useful tool for examining cell-specific toxicity in living embryos and larvae. Here we show that the hsp70/eGFP transgene is strongly and specifically upregulated within the olfactory sensory neurons (OSNs) of transgenic zebrafish larvae following a brief 3-h exposure to water-borne cadmium. This molecular response was closely correlated to an endpoint for tissue damage within the olfactory placode, namely cell death. Furthermore, cadmium-induced olfactory cytotoxicity in zebrafish larvae gives rise to more permanent effects. Juvenile zebrafish briefly exposed to cadmium during early larval development display deficits in olfactory-dependent predator avoidance behaviors 4-6 weeks after a return to clean water. Lateral line neuromasts of exposed zebrafish larvae also activate both the endogenous hsp70 gene and the hsp70/eGFP transgene. The data reveal that even a very brief exposure period that gives rise to cell death within the developing olfactory placode results in long-term deficits in olfaction, and that hsp70/eGFP may serve as an effective indicator of sublethal cadmium exposure in sensory cells

  4. Promising long-term results with attenuated adverse effects by methotrexate-containing sequential chemoradiation therapy in locally advanced head and neck squamous cell carcinoma

    International Nuclear Information System (INIS)

    To reduce severe acute and late toxicities without compromising organ preservation survival in patients with locoregionally advanced head and neck squamous cell carcinoma, we performed three-drug induction methotrexate-cisplatin-fluorouracil with weekly cisplatin-fluorouracil concurrent chemoradiation. Two induction courses of methotrexate (40 mg/m2/day, days 1, 8 and 15), cisplatin and 5-fluorouracil (25 and 750 mg/m2/day, days 1-4) were given in new diagnoses of patients with non-nasopharyngeal locoregionally advanced head and neck squamous cell carcinoma. Responders received concurrent chemoradiation with weekly cisplatin (20 mg/m2/day) and 5-fluorouracil (400 mg/m2/day) on day 1. Among 57 patients (58% with Stage IV and hypopharyngeal cancer), the rates of Grade 3-4 toxicity were 30 and 74% during induction and concurrent chemoradiotherapy (CCRT), respectively. A total of 49 patients completed induction and began concurrent chemoradiation; 47 (96%) completed all planned treatment. With a median follow-up of 62 months (range 19-83 months) for the current survivors, the 3-year overall and disease-specific survival estimates were 50 and 58%, respectively. The 3-year organ preservation survival was 74% in patients who achieved complete remission after concurrent chemoradiation, and 96% of current survivors are tracheotomy and feeding tube-free. No patient without local/regional failure suffered from distant metastasis. Methotrexate-cisplatin-fluorouracil induction chemotherapy followed by weekly cisplatin-fluorouracil concurrent chemoradiation is an acute and late toxicity-acceptable protocol without attenuating organ preservation survival in patients with locoregionally advanced head and neck squamous cell carcinoma. In this patient cohort with advanced head and neck squamous cell carcinoma, overall and organ preservation survivals were encouraging, and provided promising long-term benefits of this approach. (author)

  5. Long-term survival and late effects among one-year survivors of second allogeneic hematopoietic cell transplantation for relapsed acute leukemia and myelodysplastic syndromes.

    Science.gov (United States)

    Duncan, Christine N; Majhail, Navneet S; Brazauskas, Ruta; Wang, Zhiwei; Cahn, Jean-Yves; Frangoul, Haydar A; Hayashi, Robert J; Hsu, Jack W; Kamble, Rammurti T; Kasow, Kimberly A; Khera, Nandita; Lazarus, Hillard M; Loren, Alison W; Marks, David I; Maziarz, Richard T; Mehta, Paulette; Myers, Kasiani C; Norkin, Maxim; Pidala, Joseph A; Porter, David L; Reddy, Vijay; Saber, Wael; Savani, Bipin N; Schouten, Harry C; Steinberg, Amir; Wall, Donna A; Warwick, Anne B; Wood, William A; Yu, Lolie C; Jacobsohn, David A; Sorror, Mohamed L

    2015-01-01

    We analyzed the outcomes of patients who survived disease-free for 1 year or more after a second allogeneic hematopoietic cell transplantation (HCT) for relapsed acute leukemia or myelodysplastic syndromes between 1980 and 2009. A total of 1285 patients received a second allogeneic transplant after disease relapse; among these, 325 were relapse free at 1 year after the second HCT. The median time from first to second HCT was 17 and 24 months for children and adults, respectively. A myeloablative preparative regimen was used in the second transplantation in 62% of children and 45% of adult patients. The overall 10-year conditional survival rates after second transplantation in this cohort of patients who had survived disease-free for at least 1 year was 55% in children and 39% in adults. Relapse was the leading cause of mortality (77% and 54% of deaths in children and adults, respectively). In multivariate analyses, only disease status before second HCT was significantly associated with higher risk for overall mortality (hazard ratio, 1.71 for patients with disease not in complete remission before second HCT, P < .01). Chronic graft-versus-host disease (GVHD) developed in 43% and 75% of children and adults after second transplantation. Chronic GVHD was the leading cause of nonrelapse mortality, followed by organ failure and infection. The cumulative incidence of developing at least 1 of the studied late effects within 10 years after second HCT was 63% in children and 55% in adults. The most frequent late effects in children were growth disturbance (10-year cumulative incidence, 22%) and cataracts (20%); in adults they were cataracts (20%) and avascular necrosis (13%). Among patients with acute leukemia and myelodysplastic syndromes who receive a second allogeneic HCT for relapse and survive disease free for at least 1 year, many can be expected to survive long term. However, they continue to be at risk for relapse and nonrelapse morbidity and mortality. Novel

  6. Long-term Survival and Late Effects among 1-year Survivors of Second Allogeneic Hematopoietic Cell Transplantation for Relapsed Acute Leukemia and Myelodysplastic Syndromes

    Science.gov (United States)

    Duncan, Christine N.; Majhail, Navneet S.; Brazauskas, Ruta; Wang, Zhiwei; Cahn, Jean-Yves; Frangoul, Haydar A.; Hayashi, Robert J.; Hsu, Jack W.; Kamble, Rammurti T.; Kasow, Kimberly A.; Khera, Nandita; Lazarus, Hillard M.; Loren, Alison W.; Marks, David I.; Maziarz, Richard T.; Mehta, Paulette; Myers, Kasiani C.; Norkin, Maxim; Pidala, Joseph A.; Porter, David L.; Reddy, Vijay; Saber, Wael; Savani, Bipin N.; Schouten, Harry C.; Steinberg, Amir; Wall, Donna A.; Warwick, Anne B.; Wood, William A.; Yu, Lolie C.; Jacobsohn, David A.; Sorror, Mohamed L.

    2014-01-01

    We analyzed the outcomes of patients who survived disease-free for 1-year or more following second allogeneic hematopoietic cell transplantation (HCT) for relapsed acute leukemia or myelodysplastic syndromes between 1980 and 2009. A total of 1285 patients received a second allogeneic transplant following disease relapse; among these 325 survived relapse-free at 1-year after the second HCT. The median time from first to second HCT was 17 and 24 months for children and adults, respectively. A myeloablative preparative regimen was used in the second transplant in 62% of children and 45% of adult patients. The overall 10-year conditional survival rates after second transplantation in this cohort of patients who had survived disease-free for at least one year were 55% in children and 39% in adults. Relapse was the leading cause of mortality (77% and 54% of deaths in children and adults, respectively). In multivariate analyses, only disease status prior to second HCT was significantly associated with higher risk for overall mortality (HR 1.71 for patients with disease not in complete remission prior to second HCT, P<0.01). Chronic graft-versus-host disease (GVHD) developed in 43% and 75% of children and adults following second transplant. Chronic GVHD was the leading cause of non-relapse mortality followed by organ failure and infection. The cumulative incidence of developing at least one of the studied late effects at 10-years after second HCT was 63% in children and 55% in adults. The most frequent late effects in children were growth disturbance (10-year cumulative incidence 22%) and cataracts (20%), and in adults were cataracts (20%) and avascular necrosis (13%). Among patients with acute leukemia and myelodysplastic syndromes who receive a second allogeneic HCT for relapse and survive disease-free for at least 1-year, many can be expected to survive long term. However, they continue to be at risk for relapse and non-relapse morbidity and mortality. Novel approaches

  7. Long-term outcome of irradiation with or without chemotherapy for esophageal squamous cell carcinoma: a final report on a prospective trial

    International Nuclear Information System (INIS)

    To investigate the long-term outcome of esophageal squamous cell carcinoma (SCC) treated by irradiation with or without concurrent chemotherapy. A prospective clinical trial was carried out from 1998 to 2000. One hundred and eleven patients were randomly enrolled to receive either late course accelerated hyperfractionated irradiation (LCAF) or LCAF with concurrent chemotherapy (LCAF + CT). For LCAF, 41.4 Gy in 23 fractions was first delivered at five fractions per week, followed by 27 Gy in 18 fractions at two 1.5 Gy fractions a day. Concurrent chemotherapy of cis-platinum and 5-fluorouracil was administered for four cycles. Overall survival (OS), locoregional recurrence and distant metastasis were observed. Late toxicity was scored by RTOG criteria, and quality of life (QOL) was also evaluated. The median follow-up time was 24 months for all patients and 138 months for 17 living patients. Median survival time was 25 months and 32 months in LCAF and LCAF + CT (p = 0.653), respectively. For an entire group of patients, overall survivals were 34%, 27% and 22%; locoregional recurrence rates were 30%, 36% and 41%; and distant metastasis rates were 26%, 28% and 29% at 5-yr, 8-yr and 10-yr, respectively. Incidences of ≥ Grade 3 late toxicity were 29% at 10-yr. There were no statistically significant differences between LCAF and LCAF + CT with respect to the parameters mentioned above. Cumulative incidence of late toxicities of ≥ Grade 3 increased sharply after the attained age of 70 years. Eighty-eight percent of patients lived with good KPS (≥ 90) and 94% could eat regular or soft diet. The long-term outcome of esophageal SCC patients who received LCAF or LCAF + CT was good. The locoregional and distant failures occurred more often in the first three years after treatment, but could continuously occur up to 10 years. The late toxicity was acceptable. Late toxicities ≥ Grade 3 were more likely to occur in elderly patients. QOL was good in living patients

  8. Long-term outcomes after proton therapy, with concurrent chemotherapy, for stage II–III inoperable non-small cell lung cancer

    International Nuclear Information System (INIS)

    Purpose: We report long-term disease control, survival, and toxicity for patients with locally advanced non-small cell lung cancer prospectively treated with concurrent proton therapy and chemotherapy on a nonrandomized case-only observational study. Methods: All patients received passive-scatter proton therapy, planned with 4D-CT–based simulation; all received proton therapy concurrent with weekly chemotherapy. Endpoints were local and distant control, disease-free survival (DFS), and overall survival (OS). Results: The 134 patients (21 stage II, 113 stage III; median age 69 years) had a median gross tumor volume (GTV) of 70 cm3 (range, 5–753 cm3); 77 patients (57%) received 74 Gy(RBE), and 57 (42%) received 60–72 Gy(RBE) (range, 60–74.1 Gy(RBE)). At a median follow-up time of 4.7 years, median OS times were 40.4 months (stage II) and 30.4 months (stage III). Five-year DFS rates were 17.3% (stage II) and 18.0% (stage III). OS, DFS, and local and distant control rates at 5 years did not differ by disease stage. Age and GTV were related to OS and DFS. Toxicity was tolerable, with 1 grade 4 esophagitis and 16 grade 3 events (2 pneumonitis, 6 esophagitis, 8 dermatitis). Conclusion: This report of outcomes after proton therapy for 134 patients indicated that this regimen produced excellent OS with tolerable toxicity

  9. Influence of (La,Sr)MnO3+δ cathode composition on cathode/electrolyte interfacial structure during long-term operation of solid oxide fuel cells

    Science.gov (United States)

    Matsui, Toshiaki; Mikami, Yuichi; Muroyama, Hiroki; Eguchi, Koichi

    2013-11-01

    Time-dependent events during operation of SOFCs, i.e., performance enhancement and/or deterioration, can be readily observed for the cell composed of strontium-doped lanthanum manganite (LSM) cathode and yttria-stabilized zirconia (YSZ) electrolyte, concomitant with the change in interfacial structure of LSM/YSZ. The influence of LSM composition on the electrochemical properties and microstructure of LSM/YSZ interface during prolonged operation was investigated. Four different LSM cathodes were used and the change in microstructure, especially TPB-length, was evaluated quantitatively by a focused ion beam-scanning electron microscope (FIB-SEM). For LSM cathodes with A-site deficient compositions, the change in TPB-length had a minor contribution to the performance enhancement after 20 h of galvanostatic operation. On the other hand, for 100 h duration an increase in cathode overpotential was confirmed, accompanied with the formation of thin layer of LSM over YSZ electrolyte. A series of phenomena were triggered by the change in oxygen nonstoichiometry of LSM under polarized states. The mechanism for microstructural change was proposed and the long-term stability of LSM/YSZ interface was discussed.

  10. Long-Term Survival in a Patient with Multiple Brain Metastases from Small-Cell Lung Cancer Treated with Gamma Knife Radiosurgery on Four Occasions: A Case Report

    Directory of Open Access Journals (Sweden)

    Ameer L. Elaimy

    2012-01-01

    Full Text Available Brain metastases are the most common cancerous neoplasm in the brain. The treatment of these lesions is challenging and often includes a multimodality management approach with whole-brain radiation therapy, stereotactic radiosurgery, and neurosurgery options. Although advances in biomedical imaging technologies and the treatment of extracranial cancer have led to the overall increase in the survival of brain metastases patients, the finding that select patients survive several years remains puzzling. For this reason, we present the case of a 70-year-old patient who was diagnosed with multiple brain metastases from small-cell lung cancer five years ago and is currently alive following treatment with chemotherapy for the primary cancer and whole-brain radiation therapy and Gamma Knife radiosurgery on four separate occasions for the neurological cancer. Since the diagnosis of brain metastases five years ago, the patient’s primary cancer has remained controlled. Furthermore, multiple repeat GKRS procedures provided this patient with high levels of local tumor control, which in combination with a stable primary cancer led to an extended period of survival and a highly functional life. Further analysis and clinical research will be valuable in assessing the durability of multiple GKRS for brain metastases patients who experience long-term survival.

  11. Correction of the equilibrium temperature caused by slight evaporation of water in protein crystal growth cells during long-term space experiments at International Space Station.

    Science.gov (United States)

    Fujiwara, Takahisa; Suzuki, Yoshihisa; Yoshizaki, Izumi; Tsukamoto, Katsuo; Murayama, Kenta; Fukuyama, Seijiro; Hosokawa, Kouhei; Oshi, Kentaro; Ito, Daisuke; Yamazaki, Tomoya; Tachibana, Masaru; Miura, Hitoshi

    2015-08-01

    The normal growth rates of the {110} faces of tetragonal hen egg-white lysozyme crystals, R, were measured as a function of the supersaturation σ parameter using a reflection type interferometer under μG at the International Space Station (NanoStep Project). Since water slightly evaporated from in situ observation cells during a long-term space station experiment for several months, equilibrium temperature T(e) changed, and the actual σ, however, significantly increased mainly due to the increase in salt concentration C(s). To correct σ, the actual C(s) and protein concentration C(p), which correctly represent the measured T(e) value in space, were first calculated. Second, a new solubility curve with the corrected C(s) was plotted. Finally, the revised σ was obtained from the new solubility curve. This correction method successfully revealed that the 2.8% water was evaporated from the solution, leading to 2.8% increase in the C(s) and C(p) of the solution.

  12. Motor learning in common marmosets: vestibulo-ocular reflex adaptation and its sensitivity to inhibitors of Purkinje cell long-term depression.

    Science.gov (United States)

    Anzai, Mari; Nagao, Soich

    2014-06-01

    Adaptation of the horizontal vestibulo-ocular reflex (HVOR) provides an experimental model for cerebellum-dependent motor learning. We developed an eye movement measuring system and a paradigm for induction of HVOR adaptation for the common marmoset. The HVOR gain in dark measured by 10° (peak-to-peak amplitude) and 0.11-0.5Hz turntable oscillation was around unity. The gain-up and gain-down HVOR adaptation was induced by 1h of sustained out-of-phase and in-phase 10°-0.33Hz combined turntable-screen oscillation in the light, respectively. To examine the role of long-term depression (LTD) of parallel fiber-Purkinje cell synapses, we intraperitonially applied T-588 or nimesulide, which block the induction of LTD in vitro or in vivo preparations, 1h before the test of HVOR adaptation. T-588 (3 and 5mg/kg bod