Aseptic loosening of total hip replacement - Macrophase expression of inducible nitric oxide synthase and cyclo-oxygenase-2, together with peroxynitrite formation, as a possible mechanism for early prosthesis failure

Aseptic loosening is a major cause of failure of total hip arthroplasty, The adverse tissue response to prosthetic wear particles, with activation of cytokine and prostanoid production, contributes to bone loss around the implants, We have investigated the possibility that inducible nitric oxide synthase (iNOS) and cyclo-oxygenase-2 (COX-2) are expressed in macrophages in the pseudomembrane at the bone-implant interface, thereby contributing to the periprosthetic bone resorption. We also assessed whether peroxynitrite, a nitric oxide (NO)-derived oxidant associated with cellular injury, is generated in the membrane, Enzymatic activity of iNOS was measured using the arginine-citrulline assay technique and prostaglandin E-2 (PGE(2)), as an indicator of COX-2 activity, was measured using an enzyme immunoassay. Cellular immunoreactivity for iNOS, nitrotyrosine (a marker of peroxynitrite-induced cellular injury) and COX-2 was assessed by quantitative peroxidase immunocytochemistry while immunofluorescence methods were used for subsequent co-localisation studies with CD68(+) macrophages. The presence of calcium-independent iNOS activity and PGE(2) production was confirmed in the homogenised interface membrane, Immunocytochemistry showed that periprosthetic CD68(+) wear-debris-laden macrophages were the most prominent cell type immunoreactive for iNOS, nitrotyrosine and COX-2, Other periprosthetic inflammatory and resident cell types were also found to immunolocalise nitrotyrosine thereby suggesting peroxynitrite-induced protein nitrosylation and cellular damage not only in NO-producing CD68(+) macrophages, but also in their neighbouring cells, These data indicate that both iNOS and COX-2 are expressed by CD68(+) macrophages in the interface membrane and peroxynitrite-induced cellular damage is evident in such tissue, If high-output NO and peroxynitrite generation were to cause macrophage cell death, this would result in the release of phagocytosed wear debris into the extracellular matrix, A detrimental cycle of events would then be established with further phagocytosis by newly-recruited inflammatory cells and subsequent NO, peroxynitrite and prostanoid synthesis, Since both NO and PGE(2) have been implicated in the induction and maintenance of chronic inflammation with resulting loss of bone, and peroxynitrite in the pathogenesis of disease states, they may be central to the pathogenesis of aseptic loosening.