Airway Smooth Muscle Hyperplasia Assessment

Lung tissues were carefully removed and fixed in 10% formalin. Tissues were serially dehydrated in increasing concentrations of ethanol, cleared in xylene in a TP 1020 tissue processor (Leica Biosystems, Wetzlar, Germany), and embedded in paraffin using a Leica EG 1160 embedding machine (Leica Biosystems, Wetzlar, Germany). Transverse sections of 3 μm were cut with a Leica RM 2125 Microtome (Leica Biosystems, Wetzlar, Germany), deparaffinized, and hydrated with distilled water. Tissue sections were stained with H and E appropriately for airway integrity examination and observed under light microscope for assessment of smooth muscle hyperplasia of the airway.
Scoring was done according to Gibson–Corley [21 (link)]. Scoring adopted was 0—normal smooth muscle hyperplasia, 1—mild smooth muscle hyperplasia, 2—moderate smooth muscle hyperplasia, and 3—severe smooth muscle hyperplasia.