Metformin Combined with Alendronate Ameliorates Osteoarthritis by Attenuating RANKL-Induced Bone Resorption and Protecting Cartilage Against Degradation

Abstract BackgroundThe purpose of the present study was to investigate the effect of metformin combined with alendronate on subchondral bone and cartilage degeneration in a destabilization of the medial meniscus (DMM)-induced osteoarthritis (OA) mouse model.MethodThe effect of metformin on OA was assessed by H&E, safranin O/fast green, immunohistochemical and tartrate-resistant acid phosphatase (TRAP) staining. The Osteoarthritis Research Society International (OARSI) criteria were used to grade the degeneration of articular cartilage. ATDC5 cells were stimulated with 10 ng/mL IL-1β to establish an in vitro model of OA. Bone marrow macrophages were isolated. Cell proliferation was examined using CCK-8 assays. Cartilage extracellular matrix degradation was analyzed using Alcian Blue staining and high-density culture. Osteoclastogenesis was analyzed using TRAP staining. The mitochondrial membrane potential of ATDC5 cells and osteoclasts was determined by JC-1 fluorescent staining. Immunostaining, RT–PCR, and western blot analyses were carried out to examine protein and RNA expression levels.ResultThere were lower OARSI scores and fewer TRAP-positive cells in DMM mice after treatment with metformin combined with alendronate. In cartilage, metformin inhibited the expression of MMP3 and MMP13 in IL-1β-stimulated OA chondrocytes and increased the expression of Sox9 and aggrecan. In parallel, metformin also inhibited cartilage matrix degradation, as shown by Alcian blue staining and SO/FG staining in ATDC5 cells and cartilage tissues. Moreover, metformin and alendronate downregulated the levels of specific genes and proteins (NFATc1, c-Fos, DC-STAMP, TNFRSF11, CTSK), thereby inhibiting osteoclast differentiation in vitro. Mechanistically, metformin increased the expression of phospho-AMPK alpha1, attenuated the phosphorylation of NF-κB, and reduced reactive oxygen species (ROS) levels.ConclusionMetformin combined with alendronate may be a useful candidate for attenuating early-stage OA. This would be an ideal drug/drug combination for treating osteoarthritis in the presence of osteoporosis combined with T2D.