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Imaging lung regeneration by light sheet microscopy.

Isabelle Salwig1,2,3, Birgit Spitznagel1, Marion Wiesnet1

  • 1Department of Cardiac Development and Remodeling, Max-Planck-Institute for Heart and Lung Research, Ludwigstr. 43, 61231, Bad Nauheim, Germany.

Histochemistry and Cell Biology
|July 21, 2020
PubMed
Summary

This study reveals how lung airway epithelial cells regenerate after injury using advanced 3D imaging. It identifies specific stem cell niches that initiate the repair process, highlighting the lung

Keywords:
Club cellsLight sheet microscopyLung regenerationOptical clearingRegenerative fociStem cell niches

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Area of Science:

  • Pulmonary Medicine
  • Regenerative Biology
  • Microscopy Techniques

Background:

  • Visualizing cellular dynamics in large biological specimens like whole lungs is crucial for understanding tissue regeneration.
  • Existing methods often struggle to preserve fluorescent signals in cleared tissues, limiting deep imaging capabilities.
  • Identifying stem cell niches is key to understanding organ repair mechanisms.

Purpose of the Study:

  • To investigate the dynamics of airway regeneration in cleared whole lung preparations using light sheet microscopy.
  • To optimize an optical clearing procedure for preserving genetically encoded fluorophores in mouse lung tissue.
  • To map the spatial and temporal course of bronchiolar epithelial repair following naphthalene-induced injury.

Main Methods:

  • Development and application of an optimized optical clearing protocol using pH-adjusted tert-butanol and ethyl cinnamate.
  • Utilizing a Scgb1a1-mCherry knock-in mouse model for specific labeling of bronchiolar Club cells.
  • Employing light sheet microscopy for high-resolution 3D imaging of cleared whole lung specimens at various time points post-injury.

Main Results:

  • The optimized clearing method effectively preserved mCherry fluorescence and minimized background noise.
  • 3D imaging revealed near-complete loss of Club cells by 3 days post-naphthalene (dpn), followed by regeneration foci at 7 dpn.
  • Regeneration initiated at airway bifurcations and distal terminal bronchioles, with significant expansion of new Club cells by 21 dpn.

Conclusions:

  • Regional stem cell niches serve as critical starting points for respiratory epithelial recovery.
  • The study underscores the significant regenerative potential of the lung epithelium.
  • Whole lung 3D imaging is a powerful tool for assessing pulmonary damage and repair.