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Quantitative Pulmonary Neutrophil Dynamics Using Computer-Vision Stabilized Intravital Imaging.

Yoshikazu Tsukasaki1, Peter T Toth1,2, Esmaeil Davoodi-Bojd1

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American Journal of Respiratory Cell and Molecular Biology
|September 23, 2021
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Computer-vision stabilized intravital imaging overcomes respiratory motion artifacts in mouse lungs. This technique reveals dynamic pulmonary polymorphonuclear neutrophil (PMN) behavior and changes during endotoxemia, offering new insights into lung inflammation.

Keywords:
intravital imaginglung marginated poolneutrophil dynamicsneutrophil pathophysiologytwo-photon microscopy

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Area of Science:

  • Pulmonary physiology and immunology
  • In vivo imaging techniques
  • Computational biology

Background:

  • Intravital imaging of the lung is hindered by respiratory motion artifacts.
  • Understanding pulmonary polymorphonuclear neutrophil (PMN) dynamics is crucial for studying lung inflammation and injury.

Purpose of the Study:

  • To develop and validate a computer-vision stabilized intravital imaging method for precise lung motion correction.
  • To quantitatively analyze pulmonary PMN dynamics in real-time in mouse lungs.
  • To investigate PMN behavior in basal states and during endotoxemia-induced inflammation.

Main Methods:

  • Development of Computer-Vision Stabilized Intravital Imaging (CVSI) using a computer-vision stabilization algorithm.
  • Application of CVSI to live, respiring mouse models.
  • High-throughput quantitative analysis of PMN patrolling and recruitment in lung microvessels.
  • Observation of PMN morphology and migratory capabilities under basal and endotoxemic conditions.

Main Results:

  • CVSI achieved submicron precision in correcting lung movements and deformations.
  • Real-time dynamics of PMN patrolling in lung microvessels were quantified.
  • Marginated PMNs exhibited dynamic 'catch and release' behavior, deforming to enter smaller vessels.
  • Sequestered PMNs in endotoxemia lost migratory capacity and showed morphological changes.

Conclusions:

  • CVSI is a powerful tool for overcoming motion artifacts in lung intravital imaging.
  • Direct visualization reveals novel dynamic behaviors of pulmonary PMNs.
  • These findings enhance understanding of PMN physiology, pathophysiology, and their role in inflammatory lung injury.