Cobalt and chromium ions reduce human osteoblast‐like cell activity in vitro, reduce the OPG to RANKL ratio, and induce oxidative stress

Abstract

Metal-on-metal hip arthroplasty is associated with elevated levels of cobalt and chromium ions. The effects of cobalt and chromium ions on cell number, activity, expression of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa B ligand (RANKL) and oxidative stress on human osteoblast-like cells were addressed. Saos-2 cells were supplemented with Co 2þ , Cr 3þ , or Co 2þ þ Cr 3þ (1:2) at 0, 1, 10, and 100 mg/L and incubated for 24, 48, 72, and 96 h. Cell activity was assessed by MTT-assay and cell number by Crystal Violet staining. RNA levels of OPG and RANKL were evaluated using real-time quantitative polymerase chain reaction (qPCR). Compared to controls Co 2þ reduced cell numbers: at 10 mg/L by 17 AE 8% after 48 h and at 100 mg/L after 24 h by 35 AE 8%. Cr 3þ decreased cell numbers at 10 mg/L after 48 and 72 h. Co 2þ þ Cr 3þ combined at 1 mg/L lowered cell numbers after 24 and 96 h (17 AE 13, resp. 13 AE 4%). The 10 and 100 mg/L concentrations reduced cell numbers significantly after 24, 48, and 96 h. Cr 3þ reduced osteoblast activity at 1, 10, and 100 mg/L at all incubation times. The strongest reduction (11 AE 1%) was seen at 100 mg/L after 96 h. The OPG/RANKL ratio was reduced after 72 h with almost all Co 2þ and Cr 3þ concentrations. After 96 h, glutathione, superoxide dismutase, and catalase levels were indicative for an oxidative stress response in all samples. In conclusion, cobalt and chromium ions reduce human osteoblast activity, reduce OPG/RANKL ratio and lead to oxidative stress. Keywords: metal-on metal; osteoblasts; oxidative stress; OPG; RANKL Total hip arthroplasty (THA) is a well-accepted therapy for hip osteoarthritis, with long lasting good clinical results. Survival rates of most THA's are documented at more than 95% over 10-15 years. Wear, osteolysis, and loosening have emerged as the dominant concerns among orthopedic hip surgeons. Polyethylene wear particles trigger a foreign body granulomatous reaction, inflammation, and immune reactions leading to the secretion of various bone resorbing agents (cytokines, prostaglandins). This results in peri-prosthetic bone loss, contributing to loosening and failure of the prosthesis.1 One of the key factors in peri-prosthetic osteolysis is receptor activator of nuclear factor kappaB ligand (RANKL) and strong correlations have been found between its expression and the amount of polyethylene wear debris and the degree of osteolysis. 3 also related the presence of polyethylene to RANKL expression. In general, osteoclastogenesis is largely regulated by receptor activator of nuclear factor kappaB (RANK), RANKL, and osteoprotegerin (OPG). RANKL stimulates differentiation of osteoclast precursor cells into mature osteoclasts and is required for osteoclast activity; in fact, RANKLtreated mice show increased bone resorbtion. On the other hand, OPG decreases the number of osteoclasts and mice treated with OPG exhibit lowered osteoclast activity and increased bone volume. 4 In order to avoid polyethylene wear-related loosening and t...

Effects of thirty elements on bone metabolism

“…Reduced bone formation may be mediated via cytotoxic effects on osteoblasts, which reduce proliferation, inhibit osteocalcin release, and decrease alkaline phosphatase activity. These effects are associated with oxidative stress, which can be caused by imbalanced OPG/RANKL ratios, oxidation and nitration of proteins, and misregulation of antioxidant enzyme expression [85][86][87]. Chromium ions were also found to disturb the release of cytokines (TGF-␤1, TNF-␣, IL-␤1, TNF-G) from osteoblasts, promoting proliferation of preosteoclasts and maturation into active osteoclasts, ultimately, enhancing bone resorption [85,88].…”

“…In vitro experiments have also suggested that oxidative stress is an adverse effect of divalent cobalt ions. These effects may be mediated by the redox state in osteoblast-like cells, but the mechanism of action is not known, and contradictory results have been presented [86,87,91,92].…”

Fatal Cobalt Toxicity after a Non-Metal-on-Metal Total Hip Arthroplasty

“…Cobalt is a trace metal element which is essential for normal cellular metabolism but at high levels may lead to reduced human osteoblast activity, changes in osteoprotegerin (OPG)/receptor activator of nuclear factor kappa B ligand (RANKL) ratio leading to oxidative DNA damage, cellular apoptosis, necrosis, and oxidative DNA damage [1, 20]. Subsequently, elevated cobalt levels can elicit a multitude of symptoms including cardiomyopathy, hypothyroidism, polycythemia, cognitive dysfunction, neuropathy, and fatigue [1, 21].…”

Host Response to Orthopedic Implants (Metals and Plastics)

“…The cytotoxic effects of various metal ions might explain the extensive necrotic areas developing around MoM joint replacements; the intracellular danger signal molecules released during cell necrosis provide the danger signal necessary to initiate dendritic cell maturation and antigen presentation ultimately leading to activation of adaptive immunity. Other effects that have been attributed to metal ions include suppression of osteoblast function, with alteration of the osteoprotegerin (OPG)/RANKL ratio to favor osteoclastogenesis (Wang et al, 1997b;Fleury et al, 2006;Andrews et al, 2011;Zijlstra et al, 2012). Importantly, up-regulation of co-stimulatory molecules in macrophages by metal ions and particles have been described (Caicedo et al, 2010).…”