The molecular mechanisms of action of antimicrobial ions released from orthopaedic implants on bone cell function and regeneration

Orthopaedic implant infection as a consequence of bacterial adhesion to an inert implant surface occurs in 1.7-40 % of open fractures treated with fixation devices. Prevention of bacterial adhesion that is the nidus for infection, through the addition of silver (Ag+) to the implant surface could reduce this incidence rate, but should not be detrimental to the biology of the fracture healing cascade. Mesenchymal stromal cells (MSCs) are the source of many progenitors involved in the fracture repair process and the effect of silver on in vitro MSC viability and function was studied. The application of Ag+ at concentrations and timeframes determined from a clinically relevant in vivo rat model of intramedullary implantation, did not affect MSC growth when cultured at high density. However, Ag+ induced a significant reduction in MSC clonogenicity compared to controls, which was abrogated through the co-application of Ag+ with conditioned media from untreated MSCs. Osteogenic and adipogenic differentiation of MSCs exposed to Ag+ during and prior to differentiation showed no statistical effect on osteogenesis. However adipogenic differentiation was significantly decreased in the presence of Ag+, but significantly elevated in its absence when using MSCs that had been expanded at clonal density with Ag+. The elevation of oxidative stress as a consequence of Ag+ treatment and the role of the antioxidant pathway in the neutralisation of reactive oxygen species was determined. Real-time PCR and proteomic analysis of MSCs generated under clonogenic conditions in the presence of Ag+ confirmed the upregulation of components of the peroxiredoxin/thioredoxin pathway, in addition to the mechanisms involved with glutathione (GSH) synthesis and reduction. Inhibition of GSH synthesis using BSO eliminated MSC clonogenicity in the presence of Ag+, while recovery was achieved through the concurrent addition of exogenous GSH, verifying the vital role of GSH in MSC tolerance to Ag+.