Osteoarthritis (OA) is a degenerative disease characterized by injury of all joint tissues. Our previous study showed that in experimental osteoporosis, chiropractic manipulation (CM) exerts protective effects on bone. We here assessed whether CM might ameliorate OA by improving subchondral bone sclerosis, cartilage integrity and synovitis. Male New-Zealand rabbits underwent knee surgery to induce OA by anterior cruciate ligament injury. CM was performed using the chiropractic instrument ActivatorV 3 times/week for 8 weeks as follows: force 2 setting was applied to the tibial tubercle of the rabbit right hind limb (TM-OA), whereas the corresponding left hind limb received a false manipulation (FM-OA) consisting of ActivatorV firing in the air and slightly touching the tibial tubercle. After sacrifice, subchondral bone integrity was assessed in the tibiae by microCT and histology. Cartilage damage and synovitis were estimated by Mankin's and Krenn's scores, respectively, and histological techniques. Bone mineral density and content in both cortical and trabecular compartments of subchondral bone decreased in OA rabbits compared to controls, but partially reversed in the TM-OA group. Trabecular bone parameters in the latter group also showed a significant improvement compared to FM-OA group. Moreover RANKL, OPG, ALP and TRAP protein expression in subchondral bone significantly decreased in TM-OA rabbits with respect to FM-OA group. CM was associated with lower Mankin's and Krenn's scores and macrophage infiltrate together with a decreased protein expression of pro-inflammatory, fibrotic and angiogenic factors, in TM-OA rabbits with respect to FM-OA. Our results suggest that CM may mitigate OA progression by improving subchondral bone as well as cartilage and synovial membrane status. Osteoarthritis (OA) is one of the most common chronic diseases affecting all anatomical structures of the joint, namely cartilage, subchondral bone and synovial membrane 1. This disease affects about 15% of the population aged 25-75 years, and its prevalence significantly increases with age, affecting 70% of the population over 65 years 2. Although OA has been described as a cartilage disorder, changes in the underlying (subchondral) bone also occur in this disease 3. In this sense, different molecular alterations associated with the latter bone remodeling, e.g., in expression of nuclear factor ligand receptor kappa B (RANKL) and osteoprotegerin (OPG), have been described in OA 4-7. Preclinical and clinical studies point to the observed alterations in subchondral bone as an important OA pathogenic factor 8. In fact, studies in animal models of combined osteoporosis (OP) and OA (OPOA) demonstrate that OP induces cartilage damage 9. In this setting, the observed significant correlation between deterioration of subchondral bone and cartilage injury indicates that alterations in subchondral microstructure aggravate cartilage damage 10 .