Oxygen-Controlled Automated Neural Differentiation of Mouse Embryonic Stem Cells

Mondragón-Terán, Paul; Tostoes, Rui; Mason, Chris; Lye, Gary J.; Veraitch, Farlan

doi:10.2217/rme.13.12

Cited by 3 publications

(3 citation statements)

References 51 publications

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“…3F). A decline in glucose consumption has been observed in mouse PSCs and mouse naïve PSC neural differentiation has been charted every 48 h (Fernandes et al, 2010;Mondragon-Teran et al, 2013); however, neither study contrasted mitochondrial metabolism with glycolytic metabolism, and both altered the medium base for neural inductions. Our findings establish the acquisition of a more quiescent metabolism upon PSC exit from pluripotency, in which carbohydrate utilization is halved by the NPC state.…”

Section: Glucose-derived Carbon Utilization Decreases With Nascent Ecmentioning

confidence: 99%

“…Despite the clear profile of increasing quiescence during hPSC ectoderm differentiation, a sharp peak in metabolic parameters, including ROS, and a trough in pluripotency markers occurred within the first 24 h of differentiation. This metabolic exit event has been missed by other metabolic assessments of differentiation that only assess every 48 h or longer (Cho et al, 2006;Fernandes et al, 2010;Prigione and Adjaye, 2010;Varum et al, 2011;Mondragon-Teran et al, 2013). Similar to the waves of transcriptional and proteomic remodeling that take place during reprogramming to iPSCs (Hansson et al, 2012;Polo et al, 2012), this metabolic exit event may be necessary to shift PSCs out of their stable primed state (Enver et al, 2009), thus marking the onset of differentiation.…”

Section: Npc Induction Reduces Mitochondrial Activity and Investmentmentioning

confidence: 99%

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Mitochondrial and glycolytic remodeling during nascent neural differentiation of human pluripotent stem cells

Lees

Harvey

2018

Development

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As human pluripotent stem cells (hPSCs) exit pluripotency, they reportedly switch from glycolytic energy production to primarily mitochondrial metabolism. Here, we show that upon ectoderm differentiation to neural precursor cells (NPCs), hPSCs increase glycolytic rate, ultimately producing more carbon as lactate than is consumed as glucose. However, glucose, lactate and pyruvate utilization decrease to half their PSC levels by the NPC stage, establishing a more quiescent metabolic state. Furthermore, we characterize a metabolic exit event within the first 24 h of differentiation, plausibly necessary to transition hPSCs out of the pluripotent state. Contrary to current thinking, mitochondrial mass does not increase during NPC induction. Instead, mitochondrial DNA copies and mitochondrial activity decrease, suggesting that mitochondrial metabolism either requires suppression, or is not required, for nascent ectoderm differentiation. Our work, therefore, contrasts with the dogma that the hPSC state is primarily glycolytic, transitioning to an oxidative metabolism upon the loss of the pluripotent state. Instead, we show that heightened glycolytic metabolism is acquired, indicating that metabolic modulation of both glycolysis and mitochondrial metabolism occurs during exit from pluripotency in hPSCs.

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Section: Glucose-derived Carbon Utilization Decreases With Nascent Ecmentioning

confidence: 99%

Section: Npc Induction Reduces Mitochondrial Activity and Investmentmentioning

confidence: 99%

Mitochondrial and glycolytic remodeling during nascent neural differentiation of human pluripotent stem cells

Lees

Harvey

2018

Development

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“…In contrast, others revealed that hypoxia promoted differentiation of human ESCs into cardiomyocytes [48] and chondrocytes [49]. Hypoxia also promoted mouse ESCs to differentiate to neurons [50], endothelial cells, and hematopoietic stem cells [51]. These apparently controversial reports on the effect of hypoxia on differentiation may be explained by the stage of stemness at which the hypoxia was introduced, and the duration and degree of hypoxia.…”

Section: Hypoxia and Hifs In Embryonic Stem Cells And Induced Pluripomentioning

confidence: 99%

Hypoxia Signaling Pathway in Stem Cell Regulation: Good and Evil

et al. 2018

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Purpose of Review This review summarizes the role of hypoxia and hypoxia-inducible factors (HIFs) in the regulation of stem cell biology, specifically focusing on maintenance, differentiation, and stress responses in the context of several stem cell systems. Stem cells for different lineages/tissues reside in distinct niches, and are exposed to diverse oxygen concentrations. Recent studies have revealed the importance of the hypoxia signaling pathway for stem cell functions. Recent Findings Hypoxia and HIFs contribute to maintenance of embryonic stem cells, generation of induced pluripotent stem cells, functionality of hematopoietic stem cells, and survival of leukemia stem cells. Harvest and collection of mouse bone marrow and human cord blood cells in ambient air results in fewer hematopoietic stem cells recovered due to the phenomenon of Extra PHysiologic Oxygen Shock/Stress (EPHOSS). Summary Oxygen is an important factor in the stem cell microenvironment. Hypoxia signaling and HIFs play important roles in modeling cellular metabolism in both stem cells and niches to regulate stem cell biology, and represent an additional dimension that allows stem cells to maintain an undifferentiated status and multilineage differentiation potential.

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The Differential Effects of 2% Oxygen Preconditioning on the Subsequent Differentiation of Mouse and Human Pluripotent Stem Cells

Fynes

Tostoes

Ruban

et al. 2014

Stem Cells and Development

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A major challenge facing the development of effective cell therapies is the efficient differentiation of pluripotent stem cells (PSCs) into pure populations. Lowering oxygen tension to physiological levels can affect both the expansion and differentiation stages. However, to date, there are no studies investigating the knock-on effect of culturing PSCs under low oxygen conditions on subsequent lineage commitment at ambient oxygen levels. PSCs were passaged three times at 2% O2 before allowing cells to spontaneously differentiate as embryoid bodies (EBs) in high oxygen (20% O2) conditions. Maintenance of mouse PSCs in low oxygen was associated with a significant increase in the expression of early differentiation markers FGF5 and Eomes, while conversely we observed decreased expression of these genes in human PSCs. Low oxygen preconditioning primed mouse PSCs for their subsequent differentiation into mesodermal and endodermal lineages, as confirmed by increased gene expression of Eomes, Goosecoid, Brachyury, AFP, Sox17, FoxA2, and protein expression of Brachyury, Eomes, Sox17, FoxA2, relative to high oxygen cultures. The effects extended to the subsequent formation of more mature mesodermal lineages. We observed significant upregulation of cardiomyocyte marker Nkx2.5, and critically a decrease in the number of contaminant pluripotent cells after 12 days using a directed cardiomyocyte protocol. However, the impact of low oxygen preconditioning was to prime human cells for ectodermal lineage commitment during subsequent EB differentiation, with significant upregulation of Nestin and β3-tubulin. Our research demonstrates the importance of oxygen tension control during cell maintenance on the subsequent differentiation of both mouse and human PSCs, and highlights the differential effects.

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Oxygen-Controlled Automated Neural Differentiation of Mouse Embryonic Stem Cells

Cited by 3 publications

References 51 publications

Mitochondrial and glycolytic remodeling during nascent neural differentiation of human pluripotent stem cells

Mitochondrial and glycolytic remodeling during nascent neural differentiation of human pluripotent stem cells

Hypoxia Signaling Pathway in Stem Cell Regulation: Good and Evil

The Differential Effects of 2% Oxygen Preconditioning on the Subsequent Differentiation of Mouse and Human Pluripotent Stem Cells

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