Methotrexate Prevents Wear Particle-induced Inflammatory Osteolysis in Mice via Activation of Adenosine A2A Receptor

Abstract

Objective. Adenosine, acting at the A2A receptor, mediates the antiinflammatory effects of methotrexate (MTX) in models of inflammation. We previously reported that A2A receptor ligation diminishes wear particle–driven osteolysis. The aim of this study was to investigate whether MTX treatment could prevent bone resorption caused by inflammatory osteolysis. Methods. C57BL/6 mice (6–8 weeks old) received intraperitoneal injections of 1 mg/kg MTX (n 10) or 0.9% saline (n 5), starting 2 weeks prior to surgical implantation of 3 mg of wear particles (ultrahigh molecular weight polyethylene [UHMWPE] particles). The MTX-treated mice received daily injections of vehicle or ZM241385 at the surgical site until they were killed, 14 days later. XenoLight RediJect Bone Probe was injected intravenously, and fluorescence analysis of the calvaria using an IVIS imaging system was performed to assess bone formation. Micro–computed tomography (micro- CT) and immunostaining for osteoclast and osteoblast markers were performed. Results. Implantation of wear particles induced bone pitting and thinning, as shown by micro-CT. MTX treatment markedly reduced osteolysis, and this effect was abrogated by treatment with the A2A receptor antagonist ZM241385. Implantation of UHMWPE reduced new bone formation, and MTX treatment restored new bone formation, an effect that was completely reversed by treatment with ZM241385. Histologic examination of particle-exposed calvarias demonstrated that MTX prevented accumulation of an inflammatory infiltrate at the site of particle implantation, increased the number of osteoblasts, and reduced the number of osteoclasts at the site of inflammation, an effect that was reversed by treatment with ZM241385. Conclusion. MTX reduces inflammatory osteolysis indirectly via stimulation of A2A receptor and may represent a novel approach to enhance orthopedic implant survival, delaying or eliminating the need for revision arthroplasty surgery.