Use of Hyperosmolar Stress to Measure Stress-Activated Protein Kinase Activation and Function in Human HTR Cells and Mouse Trophoblast Stem Cells

Zhong, Wenjing; Xie, Yahong; Wang, Yingchun; Lewis, Jennifer; Trostinskaia, Anna; Wang, Fangfei; Puscheck, Elizabeth E.; Rappolee, Daniel A.

doi:10.1177/1933719107307182

Cited by 33 publications

(59 citation statements)

References 48 publications

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“…Thus, we added 100mM sorbitol, which increases SAPK ~5 fold and induces differentiation at 20% O 2 despite FGF4 2, 3, 42, 54 . However, the added stress did not increase mRNA expression of any of the 3 terminal differentiation markers (PL1, Plf, and synTa) during hypoxia (Fig.…”

Section: Resultsmentioning

confidence: 99%

Hypoxic stress induces, but cannot sustain trophoblast stem cell differentiation to labyrinthine placenta due to mitochondrial insufficiency

et al. 2014

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Dysfunctional stem cell differentiation into placental lineages is associated with gestational diseases. Of the differentiated lineages available to trophoblast stem cells (TSC), elevated O2 and mitochondrial function are necessary to placental lineages at the maternal-placental surface and important in the etiology of preeclampsia. TSC lineage imbalance leads to embryonic failure during uterine implantation. Stress at implantation exacerbates stem cell depletion by decreasing proliferation and increasing differentiation. Implantation site O2 is normally ~2%. In culture, exposure to 2% O2 and fibroblast growth factor (FGF)4 enabled highest mouse TSC multipotency and proliferation. In contrast, hypoxic stress (0.5% O2) initiated the most TSC differentiation after 24 hr despite FGF4. However, hypoxic stress supported differentiation poorly after 4–7 days, despite FGF4 removal. At all tested O2 levels, FGF4 maintained Warburg metabolism; mitochondrial inactivity and aerobic glycolysis. However, hypoxic stress suppressed mitochondrial membrane potential, maintained low mitochondrial cytochrome c oxidase (oxidative phosphorylation/OxPhos), and high pyruvate kinase M2 (glycolysis) despite FGF4 removal. Inhibiting OxPhos inhibited differentiation at the differentiation optimum at 20% O2. Moreover, adding differentiation-inducing hyperosmolar stress failed to induce differentiation during hypoxia. Thus, differentiation depended on OxPhos at 20% O2; hypoxic and hyperosmolar stresses did not induce differentiation at 0.5% O2. Hypoxia-limited differentiation and mitochondrial inhibition and activation suggest that differentiation into two lineages of the labyrinthine placenta requires O2>0.5–2% and mitochondrial function. Stress-activated protein kinase increases an early lineage and suppresses later lineages in proportion to the deviation from optimal O2 for multipotency, thus it is the first enzyme reported to prioritize differentiation.

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Section: Resultsmentioning

confidence: 99%

Hypoxic stress induces, but cannot sustain trophoblast stem cell differentiation to labyrinthine placenta due to mitochondrial insufficiency

et al. 2014

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“…2; Zhong et al 2007, 2010, Xie et al 2008, 2010, Liu et al 2009. The stress enzymes our laboratory has studied most are stressactivated protein kinase protein (aka MAPK8/9, SAPK/ JNK1/2) and AMPK (aka AMPKa1/2, PRKAA1/2).…”

Section: Cellular Stress Signaling Pathwaysmentioning

confidence: 99%

Stress responses at the endometrial–placental interface regulate labyrinthine placental differentiation from trophoblast stem cells

Rappolee¹,

Zhou²,

Puscheck³

et al. 2013

Reproduction

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Development can happen in one of two ways. Cells performing a necessary function can differentiate from stem cells before the need for it arises and stress does not develop. Or need arises before function, stress develops and stress signals are part of the normal stimuli that regulate developmental mechanisms. These mechanisms adjust stem cell differentiation to produce function in a timely and proportional manner. In this review, we will interpret data from studies of null lethal mutants for placental stress genes that suggest the latter possibility. Acknowledged stress pathways participate in stress-induced and -regulated differentiation in two ways. These pathways manage the homeostatic response to maintain stem cells during the stress. Stress pathways also direct stem cell differentiation to increase the first essential lineage and suppress later lineages when stem cell accumulation is diminished. This stress-induced differentiation maintains the conceptus during stress. Pathogenic outcomes arise because population sizes of normal stem cells are first depleted by decreased accumulation. The fraction of stem cells is further decreased by differentiation that is induced to compensate for smaller stem cell populations. Analysis of placental lethal null mutant genes known to mediate stress responses suggests that the labyrinthine placenta develops during, and is regulated by, hypoxic stress.Reproduction (2013) 145 R139-R155

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“…For embryos, culture conditions cannot create the low SAPK activity level in age-matched embryos in vivo (Fig. 3; [Xie et al 2007; Zhong et al 2007; Zhou et al 2011]), although recent, improved culture media lower SAPK activity levels compared to culture media formulated 30–40 years ago [Wang et al 2005]. For cultured TSC, contradictory signals including nonphysiological hyperoxic O 2 levels activate AMPK by 3–5-fold more than levels more closely emulating the milieu of the stem cell niche in vivo [Zhou et al 2011].…”

Section: How To Further Improve Culture Stem Cell Testing To Better Ementioning

confidence: 99%

“…We previously published many reports that mapped a three dimensional stress enzyme contour map that reported a 1–10-fold upregulation of the stress enzyme SAPK [Xie et al 2008; Xie et al 2007; Zhong et al 2007]. But the ‘1-fold’ basal level is established using biological variables such serum and added factors such as FGF4 and heparin (Fig.…”

Section: How To Further Improve Culture Stem Cell Testing To Better Ementioning

confidence: 99%

“…Dioxin and BaP are more stressful than supoptimal media, but 400mM sorbitol and microgravity simulation(MGS; [Wang et al 2009]) or shear stress [Xie et al 2006b] cause maximal stress and stress effects (see text). For TSC, in descending order of stress, 400mM sorbitol ([Zhong et al 2007], 9x), 0% O 2 ([Zhou et al 2011], 4-fold), media with 200mM sorbitol or 50mM sorbitol (3.5-fold and 2-fold, respectively), and standard media is normalized to 1-fold. However standard media has fibroblast growth factor (FGF)4 to maintain potency and proliferation and 20% O 2 that is known to slow proliferation and increase differentiation.…”

Section: Figurementioning

confidence: 99%

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Toxic stress prioritizes and imbalances stem cell differentiation: implications for new biomarkers and in vitro toxicology tests

Rappolee

Xie

Slater

et al. 2012

Systems Biology in Reproductive Medicine

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This hypothesis and review provides rules of stem cell stress responses that provide biomarkers and alternative testing that replaces or reduces gestational tests using whole animals. These rules for the stress responses of cultured stem cells validate the organismal strategy of the stress response and show that it emulates what must happen if the conceptus implants during a response to stress in vivo. Specifically there is a profound threshold during a stress dose response where stem cells accumulate significantly less well. Below this threshold stress enzymes manage the stress response by converting catabolic to anabolic processes and by suppressing apoptosis, without affecting differentiation. However above this threshold the stem cell survival response converts to an organismal survival response where stress enzymes switch to new substrates and mediate loss of potency factors, gain of early essential differentiated lineages, and suppression of later essential lineages. Stressed stem cells ‘compensate’ for lower accumulation rates by differentiating a higher fraction of cells, and the organismal survival response further enhances adaptation by prioritizing the differentiation of early essential lineages. Thus compensatory and prioritized differentiation and the sets of markers produced are part of a response of cultured embryos and stem cells that emulate what must happen during implantation of a stressed gestation. Knowledge of these markers and use of stressed stem cell assays in culture should replace or reduce the number of animals needed for developmental toxicity and should produce biomarkers for stressed development in vitro and in vivo.

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Use of Hyperosmolar Stress to Measure Stress-Activated Protein Kinase Activation and Function in Human HTR Cells and Mouse Trophoblast Stem Cells

Cited by 33 publications

References 48 publications

Hypoxic stress induces, but cannot sustain trophoblast stem cell differentiation to labyrinthine placenta due to mitochondrial insufficiency

Hypoxic stress induces, but cannot sustain trophoblast stem cell differentiation to labyrinthine placenta due to mitochondrial insufficiency

Stress responses at the endometrial–placental interface regulate labyrinthine placental differentiation from trophoblast stem cells

Toxic stress prioritizes and imbalances stem cell differentiation: implications for new biomarkers and in vitro toxicology tests

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