Tissue optical clearing, three-dimensional imaging, and computer morphometry in whole mouse lungs and human airways.
In whole adult mouse lung, full identification of airway nerves (or other cellular/subcellular objects) has not been possible due to patchy distribution and micron-scale size. Here we describe a method using tissue clearing to acquire the first complete image of three-dimensional (3D) innervation in the lung. We then created a method to pair analysis of nerve (or any other colabeled epitope) images with identification of 3D tissue compartments and airway morphometry by using fluorescent casting and morphometry software (which we designed and are making available as open-source). We then tested our method to quantify a sparse heterogeneous nerve population by examining visceral pleural nerves. Finally, we demonstrate the utility of our method in human tissue to image full thickness innervation in irregular 3D tissue compartments and to quantify sparse objects (intrinsic airway ganglia). Overall, this method can uniquely pair the advantages of whole tissue imaging and cellular/subcellular fluorescence microscopy.
Division of Pulmonary and Critical Care, Oregon Health and Sciences University, Portland, Oregon.
Gregory D
GD
Scott
Emily D
ED
Blum
Allison D
AD
Fryer
David B
DB
Jacoby
D000328:Adult; D000818:Animals; D006801:Humans; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008168:Lung / Q000033:anatomy & histology*; D051379:Mice; D008810:Mice, Inbred C57BL; D009420:Nervous System / Q000033:anatomy & histology*; D061848:Optical Imaging / Q000379:methods*; D012137:Respiratory System / Q000033:anatomy & histology*
clearing; modeling; morphometry; nerve; visceral pleura