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Pipette-based Method to Study Embryoid Body Formation Derived from Mouse and Human Pluripotent Stem Cells Partially Recapitulating Early Embryonic Development Under Simulated Microgravity Conditions

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Abstract

The in vitro differentiation of pluripotent stem cells partially recapitulates early in vivo embryonic development. More recently, embryonic development under the influence of microgravity has become a primary focus of space life sciences. In order to integrate the technique of pluripotent stem cell differentiation with simulated microgravity approaches, the 2-D clinostat compatible pipette-based method was experimentally investigated and adapted for investigating stem cell differentiation processes under simulated microgravity conditions. In order to keep residual accelerations as low as possible during clinorotation, while also guaranteeing enough material for further analysis, stem cells were exposed in 1-mL pipettes with a diameter of 3.5 mm. The differentiation of mouse and human pluripotent stem cells inside the pipettes resulted in the formation of embryoid bodies at normal gravity (1 g) after 24 h and 3 days. Differentiation of the mouse pluripotent stem cells on a 2-D pipette-clinostat for 3 days also resulted in the formation of embryoid bodies. Interestingly, the expression of myosin heavy chain was downregulated when cultivation was continued for an additional 7 days at normal gravity. This paper describes the techniques for culturing and differentiation of pluripotent stem cells and exposure to simulated microgravity during culturing or differentiation on a 2-D pipette clinostat. The implementation of these methodologies along with -omics technologies will contribute to understand the mechanisms regulating how microgravity influences early embryonic development.

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Acknowledgments

This work was funded by the European Space Agency, Noordwijk, Netherlands in the frame of the GBF Programm (4000110259 ESA-CORA-GBF)

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Authors and Affiliations

  1. Institute of Neurophysiology and Center for Molecular Medicine Cologne (CMMC), University of Cologne, Robert-Koch-Str. 39, 50931, Cologne, Germany

    Vaibhav Shinde, Jürgen Hescheler & Agapios Sachinidis

  2. German Aerospace Center (DLR), Institute of Aerospace Medicine, Gravitational Biology, Linder Hoehe, 51147, Cologne, Germany

    Sonja Brungs & Ruth Hemmersbach

Authors
  1. Vaibhav Shinde
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  2. Sonja Brungs
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  5. Agapios Sachinidis
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Correspondence to Agapios Sachinidis.

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Shinde, V., Brungs, S., Hescheler, J. et al. Pipette-based Method to Study Embryoid Body Formation Derived from Mouse and Human Pluripotent Stem Cells Partially Recapitulating Early Embryonic Development Under Simulated Microgravity Conditions. Microgravity Sci. Technol. 28, 287–295 (2016). https://doi.org/10.1007/s12217-015-9469-2

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