Effect of Oxygen Tension on the Amino Acid Utilisation of Human Embryonic Stem Cells

Christensen, David R.; Calder, Philip C.; Houghton, Franchesca D.

doi:10.1159/000356665

Cited by 20 publications

(23 citation statements)

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“…Glutamine is the highest consumed amino acid in hESCs (Christensen et al, 2014) and it improves the generation of embryos after in vitro fertilization (Devreker et al, 1998). Moreover, the expression of SLC1A5 and GLS2 , respectively the major glutamine uptake transporter and mitochondrial glutaminase involved in converting glutamine into glutamate (Hensley et al, 2013), decreases during spontaneous differentiation (Figure S1A).…”

Section: Resultsmentioning

confidence: 99%

Glutamine Metabolism Regulates the Pluripotency Transcription Factor OCT4

et al. 2016

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SUMMARY The molecular mechanisms underlying the regulation of pluripotency by cellular metabolism in human embryonic stem cells (hESCs) are not fully understood. We found that high levels of glutamine metabolism are essential to prevent degradation of OCT4, a key transcription factor regulating hESC pluripotency. Glutamine withdrawal depletes the endogenous anti-oxidant glutathione, which results in the oxidation of OCT4 cysteine residues required for its DNA binding and enhanced OCT4 degradation. The emergence of the OCT4lo cell population following glutamine withdrawal did not result in greater propensity for cell death. Instead, glutamine withdrawal during vascular differentiation of hESCs generated cells with greater angiogenic capacity, thus indicating that modulating glutamine metabolism enhances the differentiation and functional maturation of cells. These findings demonstrate that the pluripotency transcription factor OCT4 can serve as a metabolic-redox sensor in hESCs and that metabolic cues can act in concert with growth factor signaling to orchestrate stem cell differentiation.

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Section: Resultsmentioning

confidence: 99%

Glutamine Metabolism Regulates the Pluripotency Transcription Factor OCT4

et al. 2016

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“…Amino acids were eluted using an elution gradient. Buffer A comprised 15 ml tetrahydrofuran (Fisher Scientific), 200 ml HPLC grade methanol and 800 ml sodium acetate (83 mM, pH 5.9) and buffer B 200 ml sodium acetate (83 mM, pH 5.9; Fisher Scientific) and 800 ml HPLC grade methanol (Christensen et al , 2014). This method allowed the separation and analysis of 18 amino acids; including essential amino acids; histidine (His), glutamine (Gln), arginine (Arg), threonine (Thr), tyrosine (Tyr), methionine (Met), valine (Val), tryptophan (Trp), phenylalanine (Phe), isoleucine (Iso), leucine (Leu), and lysine (Lys); and non-essential amino acids; aspartic acid (Asp), glutamate (Glu), asparagine (Asn), serine (Ser), glycine (Gly), and alanine (Ala).…”

Section: Methodsmentioning

confidence: 99%

Amino acid composition of human uterine fluid: association with age, lifestyle and gynaecological pathology

et al. 2015

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STUDY QUESTIONDo the amino acid levels of human uterine fluid vary with age, BMI, phase of menstrual cycle, benign pathology or diet?SUMMARY ANSWERThe levels of 18 amino acids in human uterine fluid were shown to be affected only by maternal diet.WHAT IS KNOWN ALREADYMurine, bovine and ovine uterine amino acid content has been reported, but no reliable data on the human exist. Murine studies have demonstrated that the intrauterine periconceptional nutritional environment is affected by maternal diet.STUDY DESIGN, SIZE, DURATIONUterine secretions were aspirated from 56 women aged 18–45 years. The women were recruited preoperatively from gynaecological theatre operating schedules or hysterosalpingo-contrast-sonography (HyCoSy) lists. A proportion of these women had proven fertility; however, the majority were being investigated for subfertility. The BMI, gynaecological history and dietary pattern of these women were also assessed.PARTICIPANTS/MATERIALS, SETTING, METHODSReverse phase high performance liquid chromatography was used to analyse the concentrations of 18 amino acids within the uterine fluid and blood serum. The results were analysed against the women's stage of cycle, age, BMI and diet.MAIN RESULTS AND THE ROLE OF CHANCEThe profile of 18 amino acids in uterine fluid was described. In total, human uterine fluid was observed to contain an amino acid concentration of 3.54 mM (interquartile range: 2.27–6.24 mM). The relative concentrations of 18 amino acids were not significantly altered by age, BMI, cycle phase or the presence of specific benign gynaecological pathologies. However, a diet identified by a validated scoring system as being less healthy was associated with higher concentrations of asparagine (P = 0.018), histidine (P = 0.011), serine (P = 0.033), glutamine (P = 0.049), valine (P = 0.025), phenylalanine (P = 0.019), isoleucine (P = 0.025) and leucine (P = 0.043) in the uterine fluid compared with a healthier diet, defined as one with a higher intake of fresh vegetables, fruit, whole-grain products and fish and a low intake of red and processed meat and high fat dairy products. There were no significant correlations between serum amino acid concentrations and those in the uterine fluid.LIMITATIONS, REASONS FOR CAUTIONOur results enabled us to detect the effect of diet on the concentrations of amino acids in human uterine fluid; however, the study may not have had sufficient numbers to detect mild effects of BMI or age.WIDER IMPLICATIONS OF THE FINDINGSThese findings increase our understanding of the nutritional environment encountered by the preimplantation embryo, and indicate how periconceptional diet may alter this. Given the importance of early embryo environment for programming of development and future health, this information may aid in the development of nutritional interventions aimed at optimizing the preimplantation phase of human embryo development in vivo.STUDY FUNDING/COMPETING INTEREST(S)This work was funded by the NIHR, the Medical Research Council (G0701153) and the University o...

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“…The effect of oxygen on preimplantation embryos, from which human embryonic stem (ES) cells are derived, has been comprehensively examined in several species, including the human. While embryos are capable of developing under 20% oxygen, studies have demonstrated compromised embryo development and viability under these conditions (Thompson et al 1990;Batt et al 1991;Catt and Henman 2000;Nanassy et al 2010;Harvey et al 2012;Christensen et al 2014). The use of atmospheric oxygen for embryo culture has also been associated with increased DNA fragmentation (Goto et al 1993;Kwon et al 1999;Kitagawa et al 2004), altered genomic (Harvey et al 2004;Rinaudo et al 2006) and proteomic profiles (Katz-Jaffe et al 2005) and perturbed metabolic activity Wale and Gardner 2012).…”

Section: Introductionmentioning

confidence: 99%

“…More recently, Forristal et al (2013) described enhanced glucose uptake and lactate production in human ES cells cultured under physiological oxygen conditions, which impacted on the expression of self-renewal markers, a response mediated through Hypoxia Inducible Factor 2 alpha (HIF2A). Characterisation of amino acid profiles in human ES cells in response to oxygen have shown changes in the use of specific amino acids, but these changes were not consistently seen across the two lines examined (Christensen et al 2014). The methodology used by Forristal et al (2013) and Christensen et al (2014), however, required a medium change to facilitate metabolic measurement.…”

Section: Introductionmentioning

confidence: 99%

Oxygen modulates human embryonic stem cell metabolism in the absence of changes in self-renewal

Harvey

Rathjen

2016

Reprod. Fertil. Dev.

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Human embryonic stem (ES) cells are routinely cultured under atmospheric oxygen (~20%), a concentration that is known to impair embryo development in vitro and is likely to be suboptimal for maintaining human ES cells compared with physiological (~5%) oxygen conditions. Conflicting reports exist on the effect of oxygen during human ES cell culture and studies have been largely limited to characterisation of typical stem cell markers or analysis of global expression changes. This study aimed to identify physiological markers that could be used to evaluate the metabolic impact of oxygen on the MEL-2 human ES cell line after adaptation to either 5% or 20% oxygen in extended culture. ES cells cultured under atmospheric oxygen displayed decreased glucose consumption and lactate production when compared with those cultured under 5% oxygen, indicating an overall higher flux of glucose through glycolysis under physiological conditions. Higher glucose utilisation at 5% oxygen was accompanied by significantly increased expression of all glycolytic genes analysed. Analysis of amino acid turnover highlighted differences in the consumption of glutamine and threonine and in the production of proline. The expression of pluripotency and differentiation markers was, however, unaltered by oxygen and no observable difference in proliferation between cells cultured in 5% and 20% oxygen was seen. Apoptosis was elevated under 5% oxygen conditions. Collectively these data suggest that culture conditions, including oxygen concentration, can significantly alter human ES cell physiology with coordinated changes in gene expression, in the absence of detectable alterations in undifferentiated marker expression.

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Effect of Oxygen Tension on the Amino Acid Utilisation of Human Embryonic Stem Cells

Cited by 20 publications

References 51 publications

Glutamine Metabolism Regulates the Pluripotency Transcription Factor OCT4

Glutamine Metabolism Regulates the Pluripotency Transcription Factor OCT4

Amino acid composition of human uterine fluid: association with age, lifestyle and gynaecological pathology

Oxygen modulates human embryonic stem cell metabolism in the absence of changes in self-renewal

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