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.
Copyright © 2022 Brookes, Thorpe, Rigby Evans, Keeling and Lee.

Cancer stem cells, sometimes referred to as tumor initiating cells, play pivotal roles in tumor initiation, progression, metastasis, resistance to therapy, and relapse. Understanding how these populations of cells expand in response to a host of conditions is critical in determining effective cancer therapeutics. A defining feature of cancer stem cells is the ability to switch between modes of quiescence and symmetric/asymmetric division to protect and conserve the population, this feature is traditionally reserved for normal adult stem cell populations. Understanding how the core cell cycle machinery responds to external cues to drive symmetric/asymmetric division vs. quiescence will reveal fundamental information about how cancer stem cell populations survive and expand. This chapter will describe methods to study the cell cycle dynamics in brain cancer stem cell populations and how they compare to the other populations in a tumor.

It has been very difficult, if not impossible, to establish mouse induced pluripotent stem cells (iPSCs) from differentiated cells, such as fibroblasts, without leukemia inhibitory factor (LIF). We have established and maintained LIF-independent iPSCs for longer than 120 days with modified Oct4 along with Sox2, Klf4, and c-Myc. The iPSCs will provide a novel tool to investigate the roles of the LIF-Stat3 signaling pathway in mouse pluripotent stem cells.
Copyright © 2015. Published by Elsevier B.V.