MIA (Sigma-Aldrich, St. Louis, MO, USA) was dissolved in phosphate-buffered saline (PBS). 0.1 mg MIA in 50 μl PBS was injected once into the right knee and PBS was injected into the left knee with a 27-gauge needle in a 1.0 ml syringe through the lateral infrapatellar area toward the inter condylar space of the femur in a deep knee flexed position (Fig. 2a). Seven days after the injection, rats were forced to run 15 km (20 m/min for 50 min, 5 days a week) over 21 days and were then evaluated. Rats were also kept without running for 21 days and evaluated (Fig. 2b).Fig. 2Outline of the study. a Schema to show the grouping of knee injection and running. b Protocol for injection of mono-iodoacetate (MIA), running, and evaluation