In vitro macrophage response to nanometer‐size chromium oxide particles

Abstract

An increasing number of studies have reported adverse tissue reactions around metal-on-metal (MM) hip implants. However, the origin and mechanisms of these reactions remain unclear. Moreover, the biological effects of nanometer-size chromium oxide particles, the predominant type of wear particles produced by MM implants, remain mostly unknown. The purpose of this study was to analyze the cytotoxic effects of clinically relevant nanometer-size chromium oxide particles on macrophage response in vitro. J774.A1 macrophages were cultured with either 60 nm or 700 nm commercially available Cr2 O3 particles at different concentrations. Two different particle sizes were analyzed to evaluate potential volume effects. Cell mortality was analyzed by light microscopy, flow cytometry (annexin V-fluorescein isothiocyanate and propidium iodide assay), and using a cell death detection enzyme-linked immunosorbant assay (ELISA). Tumor necrosis factor alpha (TNF-α), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1 alpha (MIP-1α) release was measured by ELISA, and gene expression was analyzed by quantitative real-time PCR. Results showed that, at high concentrations, Cr2 O3 particles of both sizes can be cytotoxic, inducing significant decreases in total cell numbers and increases in necrosis. Results also suggested that these effects were dependent on particle volume. However, TNF-α, MCP-1, and MIP-1α cytokine release and gene expression remained low. Overall, this study demonstrates that nanometer-size particles of Cr2 O3 , a stable form of chromium oxide ceramic, have rather low cytotoxic effects on macrophages. Therefore, these particles may not be the main culprit in the initiation of the inflammatory reaction in MM periprosthetic tissues. However, other parameters (e.g., potential intracellular damage) remain to be investigated.

Effects of cobalt and chromium ions on lymphocyte migration

“…For each experimental condition, 0.5 × 10 6 macrophages were resuspended in 1 ml of DMEM‐based medium containing Co 2+ (2, 4, 8, 16, 24, 32, and 48 ppm) or Cr 3+ (25, 50, 100, 150, 250, 350, and 500 ppm). Macrophages incubated with neither Co 2+ nor Cr 3+ served as a negative control, and macrophages incubated with 1 μg/ml of lipopolysaccharide (LPS) from Escherichia coli O55:B5 (Sigma–Aldrich) were used as a positive control for TNF‐α and CC chemokine release (data not shown), as previously described …”

“…Macrophages incubated with neither Co 2þ nor Cr 3þ served as a negative control, and macrophages incubated with 1 mg/ml of lipopolysaccharide (LPS) from Escherichia coli O55:B5 (Sigma-Aldrich) were used as a positive control for TNF-a and CC chemokine release (data not shown), as previously described. 28 Triplicate samples in loosely capped 5-ml untreated polystyrene culture tubes (BD Biosciences) were incubated under cell culture conditions for 24 h. At the end of the incubation, the macrophage suspensions were centrifuged (150g for 6 min at room temperature), and the supernatants were frozen and stored in siliconized microcentrifuge tubes (Fisher Scientific) at À80˚C for cytokine measurements by enzyme-linked immunosorbent assays (ELISA; R&D Systems, Minneapolis, MN).…”

Macrophage Biocompatibility of CoCr Wear Particles Produced under Polarization in Physiological Hyaluronic Acid Solution

“…It is known that particles size is inversely proportional to the cytotoxicity degree [28,29], a data that could explain why particles from wear corrosion tests in PBS and in PBS supplemented with 0.3% HA in the absence of anodic polarization elicited a lower macrophages cell biocompatibility, as in these samples a higher proportion of small particles were detected (data not shown). The particles produced in these solutions and in these experimental conditions proceed from the breakdown of the passive film formed in the material surface where an enriched in chromium oxide was observed, a compound with high toxicity [30]. It is also necessary to consider that macrophages assays here reported were done taking in consideration the concentration of particles, that means that for the same total mass the proportion of particles with small size probably is higher in the assays performed in absence of polarization, either in PBS or PBS+0.3% HA, that could also give explanation for the lower macrophage biocompatibility of these particles.…”

In Vitro Analyses of the Toxicity, Immunological, and Gene Expression Effects of Cobalt-Chromium Alloy Wear Debris and Co Ions Derived from Metal-on-Metal Hip Implants

“…They found that CoCr-alloy particles were by far the most toxic and decreased viability and proliferation of human osteoblasts, fibroblasts, and macrophages. VanOs et al [90] used commercially available 60 nm and 700 nm round chromium oxide (Cr 2 O 3 ) particles to analyse the cytotoxic effects of chromium oxide particles on macrophage responses in vitro. With both particle sizes, cell mortality increased, resulting in a significant decrease in total cell numbers, as well as a significant increase in late apoptosis and necrosis.…”