Osteoarthritis (OA) is a multifactorial disease, associated with articular cartilage degeneration and eventually joint destruction. The phases of the disease have been described in detail, and mechanical factors play an important role in the initiation of OA, but many questions remain about its etiology. Swelling of cartilage, one of the earliest signs of damage, is proportional to the amount of collagen damage. This strongly suggests that damage to the collagen network is an early event in cartilage degeneration. The goal of this study was to determine the mechanical cause of early collagen damage in articular cartilage after mechanical overloading. Both the shear strain along the fibrils and the maximum fibril strains were evaluated as possible candidates for causing collagen damage. This evaluation was done by comparing the locations of maximum shear and tensile strains with the locations of initial collagen damage after mechanical overloading in bovine explants as found using antibodies directed against denatured type II collagen (Col2-3/4M). Collagen damage could be initiated by excessive shear strains along the collagen fibrils, and by excessive fibrils strains. The locations of collagen damage after mechanical overloading were highly dependent on the cartilage thickness, with thinner cartilage being more susceptible to damage than thicker samples.