Isolation of Wear Debris from Periarticular Tissue and In Vivo Biocompatibility of Silicon Nitride Particles

Adverse biological reactions to orthopaedic wear particles have led to the investigation of silicon nitride (SiN) coatings for implants, which may improve the biocompatibility of hip replacements. However, the analysis of wear particles is impeded by suboptimal methods to isolate wear particles from tissue. The aim of this research was to develop an improved method to isolate wear particles from tissue samples and to evaluate the in vivo biocompatibility of SiN particles. The method involved digestion of tissue samples with papain (1.56 mg.ml-1) and proteinase K (1 mg.ml-1). Samples were then subjected to density gradient ultracentrifugation using sodium polytungstate (SPT) and washing through further rounds of ultracentrifugation. The isolation method was validated by subjecting tissue samples doped with 0.0025 mm3 of silicon nitride (SiN), or 0.025 mm3 of cobalt chromium (CoCr) or titanium particles to the isolation procedure. Particles were filtered and analysed using scanning electron microscopy (SEM), elemental analysis and image analysis software before and after isolation. The method had no significant effect on SiN, CoCr or titanium particle geometries. Volumes of 0.018 mm3 of SiN, CoCr or titanium particles were injected into rat stifle joints. After seven days animals were euthanised, and the stifle joints were formalin-fixed. Stifle joint tissues were harvested and subjected to particle isolation using the validated protocol. No significant changes to SiN or CoCr particle geometries occurred. Titanium particles were similar in size, aspect ratio and circularity before and after isolation, though particle numbers were insufficient for statistical analysis. Histology of stifle joint tissues demonstrated that all three particle types caused a degree of inflammation; in the SiN group, numbers of macrophages but not lymphocytes were significantly elevated. Necrosis and increases in synovial thickness were observed in the CoCr group, but were absent in the SiN and titanium groups. Rabbits were implanted with CoCr stifle joint prostheses, which were either uncoated or coated with SiN. After 12 weeks the rabbits were euthanised. Tissue samples from the joints were formalin-fixed and subjected to the particle isolation process. SiN coating particles, CoCr substrate particles, and zirconium dioxide particles from bone cement were isolated from the samples. Due to apparent coating failure, the coating particles were 1 – 100 μm in size and were columnar or shard-like in morphology. Histological analysis of rabbit samples from the coated group demonstrated a moderate macrophage infiltrate including giant cells, with few lymphocytes and no necrosis. Overall, SiN particles were relatively biocompatible, particularly in comparison to CoCr particles. However, analysis of a functional SiN coating must be carried out over a longer timeframe to enable chronic reactions to SiN coating particles to be fully evaluated.