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Stepwise pluripotency transitions in mouse stem cells.

ABSTRACT: Pluripotent cells in mouse embryos, which first emerge in the inner cell mass of the blastocyst, undergo gradual transition marked by changes in gene expression, developmental potential, polarity, and morphology as they develop from the pre-implantation until post-implantation gastrula stage. Recent studies of cultured mouse pluripotent stem cells (PSCs) have clarified the presence of intermediate pluripotent stages between the naïve pluripotent state represented by embryonic stem cells (ESCs-equivalent to the pre-implantation epiblast) and the primed pluripotent state represented by epiblast stem cells (EpiSCs-equivalent to the late post-implantation gastrula epiblast). In this review, we discuss these recent findings in light of our knowledge on peri-implantation mouse development and consider the implications of these new PSCs to understand their temporal sequence and the feasibility of using them as model system for pluripotency.

SUBMITTER: Endoh M

PROVIDER: S-EPMC9442314 | biostudies-literature | 2022 Sep

REPOSITORIES: biostudies-literature

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Stepwise pluripotency transitions in mouse stem cells.

Endoh Mitsuhiro M Niwa Hitoshi H

EMBO reports 20220729 9

Pluripotent cells in mouse embryos, which first emerge in the inner cell mass of the blastocyst, undergo gradual transition marked by changes in gene expression, developmental potential, polarity, and morphology as they develop from the pre-implantation until post-implantation gastrula stage. Recent studies of cultured mouse pluripotent stem cells (PSCs) have clarified the presence of intermediate pluripotent stages between the naïve pluripotent state represented by embryonic stem cells (ESCs-eq ...[more]

PMID: 35903955

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Project description:BackgroundPluripotent embryonic stem cells grown under standard conditions (ESC) have a markedly glycolytic profile, which is shared with many different types of cancer cells. Thus, some therapeutic strategies suggest that pharmacologically shifting cancer cells towards an oxidative phenotype, using glycolysis inhibitors, may reduce cancer aggressiveness. Given the metabolic parallels between cancer and stemness would chemotherapeutical agents have an effect on pluripotency, and could a strategy involving these agents be envisioned to modulate stem cell fate in an accessible manner? In this manuscript we attempted to determine the effects of 3-bromopyruvate (3BrP) in pluripotency. Although it has other intracellular targets, this compound is a potent inhibitor of glycolysis enzymes thought to be important to maintain a glycolytic profile. The goal was also to determine if we could contribute towards a pharmacologically accessible metabolic strategy to influence cell differentiation.Methodology/principal findingsMouse embryonic stem cells (mESC) grown under standard pluripotency conditions (in the presence of Leukemia Inducing Factor- LIF) were treated with 3BrP. As a positive control for differentiation other mESCs were grown without LIF. Overall our results demonstrate that 3BrP negatively affects pluripotency, forcing cells to become less glycolytic and with more active mitochondria. These changes in metabolism are correlated with increased differentiation, even under pluripotency conditions (i.e. in the presence of LIF). However, 3BrP also significantly impaired cell function, and may have other roles besides affecting the metabolic profile of mESCs.Conclusions/findingsTreatment of mESCs with 3BrP triggered a metabolic switch and loss of pluripotency, even in the presence of LIF. Interestingly, the positive control for differentiation allowed for a distinction between 3BrP effects and changes associated with spontaneous differentiation/loss of pluripotency in the absence of LIF. Additionally, there was a slight differentiation bias towards mesoderm in the presence of 3BrP. However, the side effects on cellular function suggest that the use of this drug is probably not adequate to efficiently push cells towards specific differentiation fates.

Project description:Pluripotent cells are subject to much interest as a source of differentiated cellular material for research models, regenerative medical therapies and novel applications such as lab-cultured meat. Greater understanding of the pluripotent state and control over its differentiation is therefore desirable. The role of biomechanical properties in directing cell fate and cell behavior has been increasingly well described in recent years. However, many of the mechanisms which control cell morphology and mechanical properties in somatic cells are absent from pluripotent cells. We leveraged naturally occurring variation in biomechanical properties and expression of pluripotency genes in murine ESCs to investigate the relationship between these parameters. We observed considerable variation in a Rex1-GFP expression reporter line and found that this variation showed no apparent correlation to cell spreading morphology as determined by circularity, Feret ratio, phase contrast brightness or cell spread area, either on a parameter-by-parameter basis, or when evaluated using a combined metric derived by principal component analysis from the four individual criteria. We further confirmed that cell volume does not co-vary with Rex1-GFP expression. Interestingly, we did find that a subpopulation of cells that were readily detached by gentle agitation collectively exhibited higher expression of Nanog, and reduced LmnA expression, suggesting that elevated pluripotency gene expression may correlate with reduced adhesion to the substrate. Furthermore, atomic force microscopy and quantitative fluorescent imaging revealed a connection between cell stiffness and Rex1-GFP reporter expression. Cells expressing high levels of Rex1-GFP are consistently of a relatively low stiffness, while cells with low levels of Rex1-GFP tend toward higher stiffness values. These observations indicate some interaction between pluripotency gene expression and biomechanical properties, but also support a strong role for other interactions between the cell culture regime and cellular biomechanical properties, occurring independently of the core transcriptional network that supports pluripotency.

Dataset Information

Stepwise pluripotency transitions in mouse stem cells.

Publications

Stepwise pluripotency transitions in mouse stem cells.

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