Dynamics of single human embryonic stem cells and their pairs: a quantitative analysis

Wadkin, Laura E; Elliot, L. F.; Neganova, Irina; Parker, N. G.; Chichagova, Valeria; Swan, Georgina; Laude, Alex; Lako, Majlinda; Shukurov, Anvar

doi:10.1038/s41598-017-00648-0

Cited by 14 publications

(35 citation statements)

References 14 publications

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“…After these gaps are completely filled, we observe in large colonies the emergence of collective effects driven by the constant pushing and pulling of cells which drive the smallest and newly divided individuals to cluster in patches within the colony. Since the analysed colonies were grown on Matrigel TM , their migratory effects are large (Wadkin et al, 2017(Wadkin et al, , 2018 and this could be a relevant factor in the spatial organisation of the cells within the colony. The continuous re-organisation of the colonies implies that neighbours are interchanged continually and consequently the cell population is continuously mixed; this directly influences the level of clonality within the colonies and the outcome of community effects that will furthermore influence the pluripotency achieved by the population (Chang et al, 2019;Nemashkalo et al, 2017).…”

Section: Discussionmentioning

confidence: 99%

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Quantification of the morphological characteristics of hESC colonies

Orozco-Fuentes

Neganova

Wadkin

et al. 2019

Sci Rep

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The maintenance of the undifferentiated state in human embryonic stem cells (hESCs) is critical for further application in regenerative medicine, drug testing and studies of fundamental biology. Currently, the selection of the best quality cells and colonies for propagation is typically performed by eye, in terms of the displayed morphological features, such as prominent/abundant nucleoli and a colony with a tightly packed appearance and a well-defined edge. Using image analysis and computational tools, we precisely quantify these properties using phase-contrast images of hESC colonies of different sizes (0.1–1.1 ) during days 2, 3 and 4 after plating. Our analyses reveal noticeable differences in their structure influenced directly by the colony area . Large colonies (A > 0.6 mm2) have cells with smaller nuclei and a short intercellular distance when compared with small colonies (A < 0.2 mm2). The gaps between the cells, which are present in small and medium sized colonies with A ≤ 0.6 mm2, disappear in large colonies (A > 0.6 mm2) due to the proliferation of the cells in the bulk. This increases the colony density and the number of nearest neighbours. We also detect the self-organisation of cells in the colonies where newly divided (smallest) cells cluster together in patches, separated from larger cells at the final stages of the cell cycle. This might influence directly cell-to-cell interactions and the community effects within the colonies since the segregation induced by size differences allows the interchange of neighbours as the cells proliferate and the colony grows. Our findings are relevant to efforts to determine the quality of hESC colonies and establish colony characteristics database.

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Section: Discussionmentioning

confidence: 99%

“…Previous works within our group have demonstrated that isolated hESCs growing on Matrigel™ with mTESR1 media are highly motile (∼ 16.25 µm/h) and sensitive to the presence of nearby cells (Wadkin et al, 2017(Wadkin et al, , 2018. As these single cells proliferate and possibly aggregate with other cells, colonies are formed.…”

Section: Introductionmentioning

confidence: 93%

Quantification of the morphological characteristics of hESC colonies

Orozco-Fuentes

Neganova

Wadkin

et al. 2019

Sci Rep

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“…The bias in the Figure 12. (a) Single hESCs preferentially move along their elongation axis, at speed higher for a stronger elongation. (b) Cells separated by 70 µm or less move in a coordinated manner, whereas a wider separation implies independent biased random walk [33].…”

Section: Discussionmentioning

confidence: 99%

“…The lineage trees for these cells are provided in Ref. [33]. We define the time variable t as zero at the start of the image recording.…”

Section: Methodsmentioning

confidence: 99%

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Correlated random walks of human embryonic stem cells in vitro

et al. 2018

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We perform a detailed analysis of the migratory motion of human embryonic stem cells in two-dimensions, both when isolated and in close proximity to another cell, recorded with time-lapse microscopic imaging. We show that isolated cells tend to perform an unusual locally anisotropic walk, moving backwards and forwards along a preferred local direction correlated over a timescale of around 50 min and aligned with the axis of the cell elongation. Increasing elongation of the cell shape is associated with increased instantaneous migration speed. We also show that two cells in close proximity tend to move in the same direction, with the average separation of [Formula: see text]m or less and the correlation length of around 25 μm, a typical cell diameter. These results can be used as a basis for the mathematical modelling of the formation of clonal hESC colonies.

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Endothelial Differentiation G Protein-Coupled Receptor 5 Plays an Important Role in Induction and Maintenance of Pluripotency

Neganova

Cotts

Banks³

et al. 2019

Stem Cells

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Direct reprogramming of human somatic cells toward induced pluripotent stem cells holds great promise for regenerative medicine and basic biology. We used a high‐throughput small interfering RNA screening assay in the initiation phase of reprogramming for 784 genes belonging to kinase and phosphatase families and identified 68 repressors and 22 effectors. Six new candidates belonging to the family of the G protein‐coupled receptors (GPCRs) were identified, suggesting an important role for this key signaling pathway during somatic cell‐induced reprogramming. Downregulation of one of the key GPCR effectors, endothelial differentiation GPCR5 (EDG5), impacted the maintenance of pluripotency, actin cytoskeleton organization, colony integrity, and focal adhesions in human embryonic stem cells, which were associated with the alteration in the RhoA‐ROCK‐Cofilin‐PAXILLIN‐actin signaling pathway. Similarly, downregulation of EDG5 during the initiation stage of somatic cell‐induced reprogramming resulted in alteration of cytoskeleton, loss of human‐induced pluripotent stem cell colony integrity, and a significant reduction in partially and fully reprogrammed cells as well as the number of alkaline phosphatase positive colonies at the end of the reprogramming process. Together, these data point to an important role of EDG5 in the maintenance and acquisition of pluripotency. Stem Cells 2019;37:318–331

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Dynamics of single human embryonic stem cells and their pairs: a quantitative analysis

Cited by 14 publications

References 14 publications

Quantification of the morphological characteristics of hESC colonies

Quantification of the morphological characteristics of hESC colonies

Correlated random walks of human embryonic stem cells in vitro

Endothelial Differentiation G Protein-Coupled Receptor 5 Plays an Important Role in Induction and Maintenance of Pluripotency

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