Histological Analysis of Glomerular Injury in Mice

For histological analysis, mouse renal tissues were fixed in 10% formalin, followed by 70% ethanol dehydration, embedded in paraffin and sectioned at 4-µm or 2-µm thickness for periodic acid-Schiff (PAS), periodic acid-methenamine silver (PAM) and Masson-trichrome staining. Glomerular injury score was assessed as described previously [15 (link)]. More than 100 PAS-stained random glomeruli per mouse (n = 5–6 mice) were examined, and scored from 0 to 4 (0, no lesion; 1, expansion of mesangial area; 2, expansion of Bowman’s epithelial cells and adhesion of glomeruli and Bowman’s capsule; 3, sclerotic area in 50–75% of glomerulus; 4, sclerotic area in 75–100% of glomerulus). Scoring was performed using Bio-Revo imaging and analysis software (Keyence, Japan). Values were computed and presented in a graph as percentage.