Histologic Evaluation of Osteoarthritis in Mice

After surgery, mice that received different treatments were kept separately, and each treatment group had two cages (3 mice in a 400 square inch cage). At 8 weeks post-surgery, cartilage specimens were fixed in 4% paraformaldehyde for paraffin embedding. Each paraffin-embedded cartilage sample was sectioned at 5 μm, and every tenth section was stained with 0.1% safranin O solution and 0.001% Fast Green solution (Sigma‒Aldrich, St. Louis, MO, USA). For simple histologic scoring of OA in the mouse, we used an approved 0–6 subjective scoring system [22 (link)]. Histologic scores were evaluated in a blinded manner according to a grading scale (0 for normal cartilage, 0.5–4 for moderately degenerated cartilage, and 5–6 for severely degenerated cartilage).