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Super-Resolution Imaging of the Actin Cytoskeleton in Living Cells Using TIRF-SIM.

Torsten Wöllert1, George M Langford2

  • 1Biology Department, Syracuse University, Syracuse, NY, USA.

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|September 20, 2021
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Summary

Super-resolution imaging reveals dynamic actin cytoskeleton remodeling in pancreatic beta cells during insulin secretion. This study utilized TIRF-SIM to observe real-time changes in microvilli and actin networks, crucial for understanding cellular function.

Keywords:
AiryscanCLSMCortical actin cytoskeletonFIJIImage processing and analysisLive cell imagingMicrovilliPancreatic beta cellsSuper-resolution microscopyTIRF-SIM

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Area of Science:

  • Cell Biology
  • Microscopy
  • Biophysics

Background:

  • Super-resolution (SR) microscopy enables visualization beyond the diffraction limit.
  • Studying dynamic cellular processes in living cells requires gentle imaging techniques with minimal photodamage.
  • The actin cytoskeleton plays a critical role in pancreatic beta cell function, particularly during insulin secretion.

Purpose of the Study:

  • To investigate the dynamic remodeling of the actin cytoskeleton in living pancreatic beta cells during insulin secretion using a gentle SR technique.
  • To demonstrate the utility of TIRF-SIM for real-time observation of rapid cellular events.
  • To quantify changes in actin organization and cell surface structures.

Main Methods:

  • Total Internal Reflection Fluorescence-Structured Illumination Microscopy (TIRF-SIM) was employed as the primary SR imaging technique.
  • Live pancreatic beta cells were imaged to capture dynamic actin cytoskeleton rearrangements.
  • FIJI plugins were utilized for image processing and quantitative analysis of TIRF-SIM data.

Main Results:

  • TIRF-SIM successfully visualized dynamic actin cytoskeleton remodeling in living pancreatic beta cells.
  • Microvilli on pancreatic beta cells were observed to translocate within the plasma membrane.
  • The cortical actin network exhibited significant reorganization upon stimulation of insulin secretion.

Conclusions:

  • TIRF-SIM is a suitable gentle SR technique for studying dynamic cellular processes in living cells.
  • Dynamic remodeling of the actin cytoskeleton, including microvilli translocation and cortical network reorganization, is associated with insulin secretion in pancreatic beta cells.
  • Quantitative analysis of TIRF-SIM data provides valuable insights into the mechanisms underlying insulin secretion.