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Embryonic stem (ES) cells were first discovered in mice in 1981 by Martin Evans. In 1998, James Thomson identified a method to isolate embryonic stem cells from humans. Human embryonic stem cells (hESCs) are obtained from 3-5 day old embryos that remain unused after an in vitro fertilization procedure.
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Profiling Individual Human Embryonic Stem Cells by Quantitative RT-PCR
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Visualization and quantification of dynamic intercellular coupling in human embryonic stem cells using single cell

Zhenzhen Fan1, Xufeng Xue2, Jianping Fu1,2

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Sonoporation enables precise dye loading and calcium imaging in human embryonic stem cells (hESCs). This technique visualizes gap junction intercellular communication (GJIC) and reveals heterogeneous cell connectivity in hESC colonies.

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Area of Science:

  • Cell Biology
  • Biophysics
  • Stem Cell Research

Background:

  • Gap junctions (GJs) are protein channels facilitating intercellular communication.
  • GJ intercellular communication (GJIC) is crucial for human embryonic stem cell (hESC) behavior, including proliferation and differentiation.

Purpose of the Study:

  • To develop and apply a novel sonoporation method for single-cell dye loading and GJIC visualization in hESC colonies.
  • To dynamically quantify GJIC and calcium (Ca2+) diffusion in hESCs.

Main Methods:

  • Utilized sonoporation with microbubbles to create transient pores for intracellular dye and Ca2+ loading in single hESCs.
  • Employed live imaging to monitor intercellular dye transfer and Ca2+ diffusion.
  • Applied mass transport models to quantify cell-cell permeability based on dye diffusion.

Main Results:

  • Successfully demonstrated single-cell intracellular dye loading and dynamic visualization of GJIC in hESC colonies.
  • Observed heterogeneous intercellular connectivity within hESC colonies.
  • Characterized diverse spatiotemporal patterns of intercellular Ca2+ waves initiated by single-cell sonoporation.

Conclusions:

  • Sonoporation is an effective technique for studying GJIC and Ca2+ dynamics in hESCs at the single-cell level.
  • The study highlights the complex and heterogeneous nature of intercellular communication in hESC colonies.