Abstract
Numerous biological approaches are available to characterise the mechanisms which govern the formation of human embryonic stem cell (hESC) colonies. To understand how the kinematics of single and pairs of hESCs impact colony formation, we study their mobility characteristics using time-lapse imaging. We perform a detailed statistical analysis of their speed, survival, directionality, distance travelled and diffusivity. We confirm that single and pairs of cells migrate as a diffusive random walk for at least 7 hours of evolution. We show that the presence of Cell Tracer significantly reduces hESC mobility. Our results open the path to employ the theoretical framework of the diffusive random walk for the prognostic modelling and optimisation of the growth of hESC colonies. Indeed, we employ this random walk model to estimate the seeding density required to minimise the occurrence of hESC colonies arising from more than one founder cell and the minimal cell number needed for successful colony formation. Our prognostic model can be extended to investigate the kinematic behaviour of somatic cells emerging from hESC differentiation and to enable its wide application in phenotyping of pluripotent stem cells for large scale stem cell culture expansion and differentiation platforms.
Highlights
Human pluripotent stem cells (encompassing both human embryonic stem cell (hESC) and the human induced pluripotent stem cells) hold great potential for advancement of cellular therapies, disease modelling and drug discovery
The cells that are closer to each other move in a more systematic, directed manner, and display a higher ability to form colonies arising from more than one founder cell, suggesting that the separation distance of hESCs at the start of the colony formation from a few cells and their migration parameters are important for clonal expansion and have an impact on pluripotent phenotype and status of a colony as a whole[4]
Mechanisms that regulate the heterogeneous phenotype of a hESC colony and its relation to the functional properties of a single cell are poorly studied
Summary
Human pluripotent stem cells (encompassing both hESCs and the human induced pluripotent stem cells (hiPSCs)) hold great potential for advancement of cellular therapies, disease modelling and drug discovery. Our aim is to develop a quantitative model and understanding which would facilitate prediction of the behaviour of individual cells during the colony formation as well as within the colony This would allow a non-invasive characterization of hESC colonies, identification of cell fate history and their interaction with neighbours. Such a model would provide a useful platform for testing impacts of small molecules, Cell Tracers, differentiation agents and culture media, and enable molecular studies of mutual cell interactions To approach this major task systematically, we should first understand the behaviour of single hESCs, cell pairs, small clumps and growing colonies. In establishing the diffusive migration of cells and cell pairs, our work opens the possibility to use the well-developed, powerful mathematical theory of random walks for non-invasive prognostic modelling of the behaviour of single hESCs, with obvious implications for large scale expansion and differentiation assays
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