Genotoxic effects of particles of surgical cobalt chrome alloy on human cells of different age in vitro.

Humans are exposed to metals from industry, the environment and from wear debris from worn orthopaedic joint replacements. Patients exposed to worn cobalt chrome hip replacements show an increase of chromosome aberrations in the bone marrow adjacent to the implant and an increase of chromosome translocations and aneuploidy in the peripheral blood. This study has tested whether particles of surgical cobalt chrome alloy are able to induce similar DNA damage and chromosome aberrations in human cells in vitro. Because increasingly young patients are receiving hip replacements it has also tested whether the response is altered at different cellular age in vitro. Primary human fibroblasts, were tested at different pre senescent population doublings (PD10 (young) and PD35 (older)) to particles of cobalt chrome alloy for up to 15 days. As in patients there was an increase of aneuploidy, chromosome translocations and DNA damage after exposure to the cobalt chrome particles in vitro. The overall level of DNA damage and numerical and structural aberrations was approximately the same in young and older cells. However, the cellular reaction to the DNA damage was different. Older cells showed a greater loss of viability and induction of senescence and a lesser rate of mitosis and cell growth than young cells. They showed less change in transcription, particularly of p38 and caspase 10 mRNA levels, than young cells. They showed more complex aneuploidy in association with unseparated or prematurely separated chromatids. This study suggests that at least part of the chromosome changes in patients with worn implants may be due to direct effects of the metal wear particles from the implant. It would be of interest to test whether the altered reaction of the human cells at different in vitro age might correspond with a different incidence of chromosome aberrations in patients at different ages.

Interaction of micron and nano-sized particles with cells of the dura mater.

Intervertebral total disc replacements (TDR) are used in the treatment of degenerative spinal disc disease. There are, however, concerns that they may be subject to long-term failure due to wear. The adverse effects of TDR wear have the potential to manifest in the dura mater and surrounding tissues. The aim of this study was to investigate the physiological structure of the dura mater, isolate the resident dural epithelial and stromal cells and analyse the capacity of these cells to internalise model polymer particles. The porcine dura mater was a collagen-rich structure encompassing regularly arranged fibroblastic cells within an outermost epithelial cell layer. The isolated dural epithelial cells had endothelial cell characteristics (positive for von Willebrand factor, CD31, E-cadherin and desmoplakin) and barrier functionality whereas the fibroblastic cells were positive for collagen I and III, tenascin and actin. The capacity of the dural cells to take up model particles was dependent on particle size. Nanometer sized particles readily penetrated both types of cells. However, dural fibroblasts engulfed micron-sized particles at a much higher rate than dural epithelial cells. The study suggested that dural epithelial cells may offer some barrier to the penetration of micron-sized particles but not nanometer sized particles.

Biological Effects of Clinically Relevant CoCr Nanoparticles in the Dura Mater: An Organ Culture Study.

Medical interventions for the treatment of spinal disc degeneration include total disc replacement and fusion devices. There are, however, concerns regarding the generation of wear particles by these devices, the majority of which are in the nanometre sized range with the potential to cause adverse biological effects in the surrounding tissues. The aims of this study were to develop an organ culture model of the porcine dura mater and to investigate the biological effects of CoCr nanoparticles in this model. A range of histological techniques were used to analyse the structure of the tissue in the organ culture. The biological effects of the CoCr wear particles and the subsequent structural changes were assessed using tissue viability assays, cytokine assays, histology, immunohistochemistry, and TEM imaging. The physiological structure of the dura mater remained unchanged during the seven days of in vitro culture. There was no significant loss of cell viability. After exposure of the organ culture to CoCr nanoparticles, there was significant loosening of the epithelial layer, as well as the underlying collagen matrix. TEM imaging confirmed these structural alterations. These structural alterations were attributed to the production of MMP-1, -3, -9, -13, and TIMP-1. ELISA analysis revealed that there was significant release of cytokines including IL-8, IL-6, TNF-α, ECP and also the matrix protein, tenascin-C. This study suggested that CoCr nanoparticles did not cause cytotoxicity in the dura mater but they caused significant alterations to its structural integrity that could lead to significant secondary effects due to nanoparticle penetration, such as inflammation to the local neural tissue.

Biological effects of cobalt-chromium nanoparticles and ions on dural fibroblasts and dural epithelial cells.

The introduction of metal-on-metal total disc replacements motivated studies to evaluate the effects of cobalt-chromium (CoCr) nanoparticles on cells of the dura mater. Porcine fibroblasts and epithelial cells isolated from the dura mater were cultured with clinically-relevant CoCr nanoparticles and the ions, generated by the particles over 24 h, at doses up to 121 µm(3)per cell. Cell viability and production of proinflammatory cytokines was assessed over 4 days. The capacity of the particles to induce oxidative stress in the cells was evaluated at 24 h. The CoCr particles and their ions significantly reduced the viability of the dural epithelial cells in a dose-dependent manner but not the fibroblasts. Both cell types secreted IL-8 in response to particle exposure at doses of 60.5 µm(3) (epithelial cells) and 121 µm(3) (fibroblasts, epithelial cells) per cell. No significant release of IL-6 was observed in both cell types at any dose. Reactive oxygen species were induced in both cell types at 50 µm(3) per cell after 24 h exposure. The data suggested novel differences in the resistance of the dural epithelial cells and fibroblasts to CoCr nanoparticle/ion toxicity and demonstrated the inflammatory potential of the particles. The data contributes to a greater understanding of the potential biological consequences of the use of metal-on-metal total disc prostheses.