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      Analytic Models of Oxygen and Nutrient Diffusion, Metabolism Dynamics, and Architecture Optimization in Three-Dimensional Tissue Constructs with Applications and Insights in Cerebral Organoids

      1 , 2

      Tissue Engineering Part C: Methods

      Mary Ann Liebert Inc

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          Brain energy metabolism: focus on astrocyte-neuron metabolic cooperation.

          The energy requirements of the brain are very high, and tight regulatory mechanisms operate to ensure adequate spatial and temporal delivery of energy substrates in register with neuronal activity. Astrocytes-a type of glial cell-have emerged as active players in brain energy delivery, production, utilization, and storage. Our understanding of neuroenergetics is rapidly evolving from a "neurocentric" view to a more integrated picture involving an intense cooperativity between astrocytes and neurons. This review focuses on the cellular aspects of brain energy metabolism, with a particular emphasis on the metabolic interactions between neurons and astrocytes. Copyright © 2011 Elsevier Inc. All rights reserved.
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            A nuclear factor induced by hypoxia via de novo protein synthesis binds to the human erythropoietin gene enhancer at a site required for transcriptional activation.

            We have identified a 50-nucleotide enhancer from the human erythropoietin gene 3'-flanking sequence which can mediate a sevenfold transcriptional induction in response to hypoxia when cloned 3' to a simian virus 40 promoter-chloramphenicol acetyltransferase reporter gene and transiently expressed in Hep3B cells. Nucleotides (nt) 1 to 33 of this sequence mediate sevenfold induction of reporter gene expression when present in two tandem copies compared with threefold induction when present in a single copy, suggesting that nt 34 to 50 bind a factor which amplifies the induction signal. DNase I footprinting demonstrated binding of a constitutive nuclear factor to nt 26 to 48. Mutagenesis studies revealed that nt 4 to 12 and 19 to 23 are essential for induction, as substitutions at either site eliminated hypoxia-induced expression. Electrophoretic mobility shift assays identified a nuclear factor which bound to a probe spanning nt 1 to 18 but not to a probe containing a mutation which eliminated enhancer function. Factor binding was induced by hypoxia, and its induction was sensitive to cycloheximide treatment. We have thus defined a functionally tripartite, 50-nt hypoxia-inducible enhancer which binds several nuclear factors, one of which is induced by hypoxia via de novo protein synthesis.
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              Low O2 tensions and the prevention of differentiation of hES cells.

              Early-stage mammalian embryos develop in a low O(2) environment (hypoxia). hES cells, however, are generally cultured under an atmosphere of 21% O(2) (normoxia), under which conditions they tend to differentiate spontaneously. Such conditions may not be the most suitable, therefore, for hES cell propagation. Here we have tested two hypotheses. The first hypothesis was that hES cells would grow as well under hypoxic as under normoxic conditions. The second hypothesis was that hypoxic culture would reduce the amount of spontaneous cell differentiation that occurs in hES colonies. Both hypotheses proved to be correct. Cells proliferated as well under 3% and 5% O(2) as they did under 21% O(2), and growth was only slightly reduced at 1% O(2). The appearance of differentiated regions as assessed morphologically, biochemically (by the production of human chorionic gonadotropin and progesterone), and immunohistochemically (by the loss of stage-specific embryonic antigen-4 and Oct-4 and gain of stage-specific embryonic antigen-1 marker expression) was markedly reduced under hypoxic conditions. In addition, hES cell growth under hypoxia provided enhanced formation of embryoid bodies. Hypoxic culture would appear to be necessary to maintain full pluripotency of hES cells.
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                Author and article information

                Journal
                Tissue Engineering Part C: Methods
                Tissue Engineering Part C: Methods
                Mary Ann Liebert Inc
                1937-3384
                1937-3392
                March 2016
                March 2016
                : 22
                : 3
                : 221-249
                Affiliations
                [1 ]Institute of Neural Regeneration & Tissue Engineering, Highland, Utah, United States.
                [2 ]Department of Engineering Science, Institute of Biomedical Engineering, University of Oxford, Oxford, United Kingdom.
                Article
                10.1089/ten.tec.2015.0375
                © 2016

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