Effect Of Chromium(â…¢) Ions On Human HFOB Osteoblasts In Vitro: Safety And Gene Expression

Academic BackgroundMetal on metal hip prosthesis has more than 30 years of usage records. With the increasing number of young patients who used it, the metal ions cumulative in the body caused the biological effects, such as cell toxicity,genetic toxicity,carcinogenicity,metal allergy,etc, have been received attention and concern.ObjectiveTo explore the relationship between trivalent chromium concentration and osteoblasts activity cultured in vitro, to find out changes of osteoblasts gene expression which stimulated by the trivalent chromium in vitro,to explore the safety problem of trivalent chromium.MethodOsteoblasts hFOB 1.19 cultured 24, 48, 72 h by chromium chloride solution (according to the Cr³⁺ 0.01, 0.1, 1, 10, 100, 1000, 10000, 100000 mg/L gradient individually), then were detected the ability of MTT reduction and the expression of alkaline. Through gene-chip technology, the gene expression of osteoblasts cultured by chromium chloride (Cr³⁺ 10 mg/L) for 24 h was detected, and some of the differences in gene expression were verified through real-time PCR technology.ResultsThe effect of trivalent chromium on osteoblasts viability was dose-dependent and time-dependent. Toxicity of trivalent chromium was into the plateau period, after stimulated osteoblasts in vitro for 48h; trivalent chromium toxicity reached its peak, when concentration of trivalent chromium between 100 mg / L and 1000 mg / L ; trivalent chromium (≥0.01mg/L) could inhibit cell proliferation rates significantly. Microarray results showed up-regulated 1503 genes, down-regulated 1308 genes after trivalent chromium (10 mg/L) had stimulated 24h. Analysis revealed significant enrichments in some important GO terms often associated with cell growth and immune response, such as biological regulation, regulation of biological process, regulation of cellular process, cell communication, signal transduction, identical protein binding, membrane organization and biogenesis, cell-cell adhesion, hemophietic or lymphoid organ development, immune system development, hemopoiesis, myofibril, intramolecular oxidoreductase activity, GABA-B receptor activity. Real-time PCR results confirmed that card11, igsf6, tnfrsf17 and rora genes were significantly increased after exposure to trivalent chromium (10 mg/L) for 72 h. All 4 genes were expressed up-regulated after chromium-stimulated 72 h; their expression patterns were consistent with the microarray data.DiscussionMetal on metal prosthesis release of metal ions in the body through electrochemical process (corrosion) and enhanced machinery plus electrochemical process (wear - corrosion). It spreads to various body tissues by body fluid, raises a number of biological effects. Human reaction of metal ions in a dose-dependent manner, and metal ions serum level decides the size of biological effects. The metal ions biological effects include:â‘ inflammatory responseâ‘¡oxidative damageâ‘¢apoptosisâ‘£osteoporosis and extracellular matrix dissolution⑤Carcinogenesisâ‘¥host response. Its mechanism include corrosion, macrophage activation, reactive oxygen species generation, osteoblasts function inhibited osteoblasts activation, metal - protein complex formation, and other relevant. In this paper, the experiments in vitro proved that the proliferation inhibition effect of trivalent chromium on osteoblasts cultured in vitro was dose-dependent and time-dependent; Serum chromium concentration under normal circumstances indirectly proved to be safe to osteoblasts, at early postoperative. Osteoblasts cultured in vitro by stimulation of trivalent chromium showed most significantly differences in 43 genes, function of these genes was associated with inflammatory response, apoptosis. In addition, there were four genes (card11, igsf6, tnfrsf17 and rora) activated by trivalent chromium, they were involved in the biological effects of trivalent chromium through regulation of the immune response, apoptosis. Further investigation of the precise functional roles of these significantly changed genes will provide new insights in understanding the molecular mechanisms of metal toxic and pave the way for new therapeutic strategies and drug discovery.Conclusion Trivalent chromium (≥0.01mg/L) could inhibit osteoblasts proliferation rates significantly. Osteoblasts cultured in vitro by stimulation of trivalent chromium showed most significantly differences in function associated with inflammatory response, apoptosis (Cr³⁺ 10mg/L). In addition, there were four genes (card11, igsf6, tnfrsf17 and rora) activated by trivalent chromium (10mg/L), they were involved in the biological effects of trivalent chromium through regulation of the immune response, apoptosis. Further investigation of the precise functional roles of these significantly changed genes will provide new insights in understanding the molecular mechanisms of metal toxic.