Lung histopathology
Mouse lungs were fixed with 10% formalin, and paraffin sections were
prepared by the Histology Core at Peking Union Medical College Hospital.
Hematoxylin and eosin staining was performed to examine inflammatory
infiltrates; Alcian blue-periodic acid-Schiff staining (Sigma-Aldrich)
was performed to evaluate mucus production. To determine the severity of
inflammatory cell infiltration, peribronchial and perivascular
inflammatory cell infiltration was blindly evaluated using a five-point
scoring system, as previously
described[17]: 0, no cells; 1, a few
cells; 2, a ring of cells with 1-cell depth; 3, a ring of cells with
2–4-cell depth; 4, a ring of cells with >4-cell depth. To
evaluate the severity of mucus production, goblet cell hyperplasia in
airway epithelium was evaluated semiquantitatively under a light
microscope (20× magnification) in a blinded manner, using a five-point
grading system described
previously[17]. Alcian blue-stained
goblet cells were counted and expressed as the percentage of the total
number of epithelial cells: 1, <25%; 2, 25%–50%; 3,
50%–75%; 4, >75%. The scoring of inflammatory cells and
goblet cells was performed in at least three different fields for each
lung section. Average scores were obtained from 10 mice.