Influence of Cu2+ on Osteoclast Formation and Activity In Vitro

Background: Copper-containing biomaterials are increasingly applied for bone regeneration due to their pro-angiogenetic, pro-osteogenetic and antimicrobial properties. Therefore, the effect of Cu2+ on osteoclasts, which play a major role in bone remodeling was studied in detail. Methods: Human primary osteoclasts, differentiated from human monocytes were differentiated or cultivated in the presence of Cu2+. Osteoclast formation and activity were analyzed by measurement of osteoclast-specific enzyme activities, gene expression analysis and resorption assays. Furthermore, the glutathione levels of the cells were checked to evaluate oxidative stress induced by Cu2+. Results: Up to 8 µM Cu2+ did not induce cytotoxic effects. Activity of tartrate-resistant acid phosphatase (TRAP) was significantly increased, while other osteoclast specific enzyme activities were not affected. However, gene expression of TRAP was not upregulated. Resorptive activity of osteoclasts towards dentin was not changed in the presence of 8 µM Cu2+ but decreased in the presence of extracellular bone matrix. When Cu2+ was added to mature osteoclasts TRAP activity was not increased and resorption decreased only moderately. The glutathione level of both differentiating and mature osteoclasts was significantly decreased in the presence of Cu2+. Conclusions: Differentiating and mature osteoclasts react differently to Cu2+. High TRAP activities are not necessarily related to high resorption.

[1]  C. Heinemann,et al.  How to Get Them off?—Assessment of Innovative Techniques for Generation and Detachment of Mature Osteoclasts for Biomaterial Resorption Studies , 2021, International journal of molecular sciences.

[2]  R. Bader,et al.  Comparison of Inflammatory Effects in THP-1 Monocytes and Macrophages after Exposure to Metal Ions , 2020, Materials.

[3]  M. Schlosser,et al.  Effects of copper-impregnated collagen implants on local pro- and anti-inflammatory and regenerative tissue reactions following implantation in rats. , 2019, Journal of biomedical materials research. Part A.

[4]  A. Boccaccini,et al.  Chitosan/hydroxyapatite composite bone tissue engineering scaffolds with dual and decoupled therapeutic ion delivery: copper and strontium. , 2019, Journal of materials chemistry. B.

[5]  J. Zhang,et al.  Concentration-dependent osteogenic and angiogenic biological performances of calcium phosphate cement modified with copper ions. , 2019, Materials science & engineering. C, Materials for biological applications.

[6]  Yanjin Lu,et al.  Copper-modified Ti6Al4V alloy fabricated by selective laser melting with pro-angiogenic and anti-inflammatory properties for potential guided bone regeneration applications. , 2018, Materials science & engineering. C, Materials for biological applications.

[7]  J. Weng,et al.  Preparation of Micro/Nano-Structure Copper-Substituted Hydroxyapatite Scaffolds with Improved Angiogenesis Capacity for Bone Regeneration , 2018, Materials.

[8]  P. Hartemann,et al.  Contact killing and antimicrobial properties of copper , 2018, Journal of applied microbiology.

[9]  J. Čapek,et al.  Comparison of glutathione levels measured using optimized monochlorobimane assay with those from ortho-phthalaldehyde assay in intact cells. , 2017, Journal of pharmacological and toxicological methods.

[10]  M. Gelinsky,et al.  Osteoclastic differentiation and resorption is modulated by bioactive metal ions Co2+, Cu2+ and Cr3+ incorporated into calcium phosphate bone cements , 2017, PloS one.

[11]  Livia Visai,et al.  POLITECNICO DI TORINO Repository ISTITUZIONALE Copper-containing mesoporous bioactive glass nanoparticles as multifunctional agent for bone regeneration / , 2022 .

[12]  A. Lode,et al.  Cu2+, Co2+ and Cr3+ doping of a calcium phosphate cement influences materials properties and response of human mesenchymal stromal cells. , 2017, Materials science & engineering. C, Materials for biological applications.

[13]  M. Gelinsky,et al.  Relevance of osteoclast-specific enzyme activities in cell-based in vitro resorption assays. , 2017, European cells & materials.

[14]  D. Medeiros.  Copper, iron, and selenium dietary deficiencies negatively impact skeletal integrity: A review , 2016, Experimental biology and medicine.

[15]  A. Boccaccini,et al.  Novel nanocomposite biomaterials with controlled copper/calcium release capability for bone tissue engineering multifunctional scaffolds , 2015, Journal of The Royal Society Interface.

[16]  A. Salem,et al.  Incorporation of copper into chitosan scaffolds promotes bone regeneration in rat calvarial defects. , 2015, Journal of biomedical materials research. Part B, Applied biomaterials.

[17]  Bernd Kreikemeyer,et al.  A dual function of copper in designing regenerative implants. , 2015, Biomaterials.

[18]  M. Gelinsky,et al.  Formation of osteoclasts on calcium phosphate bone cements and polystyrene depends on monocyte isolation conditions. , 2015, Tissue engineering. Part C, Methods.

[19]  Wenjie Zhang,et al.  Antibacterial property, angiogenic and osteogenic activity of Cu-incorporated TiO2 coating. , 2014, Journal of materials chemistry. B.

[20]  M. Valko,et al.  Redox active metal-induced oxidative stress in biological systems , 2012, Transition Metal Chemistry.

[21]  R. Jessberger,et al.  A novel resorption assay for osteoclast functionality based on an osteoblast‐derived native extracellular matrix , 2010, Journal of cellular biochemistry.

[22]  Huiqi Xie,et al.  Role of copper in angiogenesis and its medicinal implications. , 2009, Current medicinal chemistry.

[23]  Hongqiao Zhang,et al.  Glutathione: overview of its protective roles, measurement, and biosynthesis. , 2009, Molecular aspects of medicine.

[24]  Pamela Habibovic,et al.  Angiogenesis in calcium phosphate scaffolds by inorganic copper ion release. , 2009, Tissue engineering. Part A.

[25]  J. Rissanen,et al.  Secreted Tartrate-Resistant Acid Phosphatase 5b is a Marker of Osteoclast Number in Human Osteoclast Cultures and the Rat Ovariectomy Model , 2008, Calcified Tissue International.

[26]  C. Palacios.  The Role of Nutrients in Bone Health, from A to Z , 2006, Critical reviews in food science and nutrition.

[27]  Soo Young Lee,et al.  A crucial role for reactive oxygen species in RANKL-induced osteoclast differentiation. , 2005, Blood.

[28]  Hong-Hee Kim,et al.  Reactive oxygen species mediate RANK signaling in osteoclasts. , 2004, Experimental cell research.

[29]  C. Chow,et al.  Copper toxicity, oxidative stress, and antioxidant nutrients. , 2003, Toxicology.

[30]  Jinchao Zhang,et al.  Effects of Cu2+ and pH on osteoclastic bone resorption in vitro , 2003 .

[31]  J. Mönkkönen,et al.  Rapid screening method for osteoclast differentiation in vitro that measures tartrate-resistant acid phosphatase 5b activity secreted into the culture medium. , 2000, Clinical chemistry.

[32]  P. Lehenkari,et al.  Intracellular Fragmentation of Bone Resorption Products by Reactive Oxygen Species Generated by Osteoclastic Tartrate-resistant Acid Phosphatase* , 1999, The Journal of Biological Chemistry.

[33]  S. Rosalki,et al.  BIOCHEMICAL MARKERS OF BONE TURNOVER , 1998, International journal of clinical practice.

[34]  R. Burghardt,et al.  Kinetic analysis of glutathione in anchored cells with monochlorobimane. , 1995, Cytometry.

[35]  T. Cox,et al.  Purple acid phosphatase of the human macrophage and osteoclast. Characterization, molecular properties, and crystallization of the recombinant di-iron-oxo protein secreted by baculovirus-infected insect cells. , 1994, The Journal of biological chemistry.

[36]  I. Morita,et al.  Participation of oxidative stress in the process of osteoclast differentiation. , 1993, Biochimica et biophysica acta.

[37]  L. Bonewald,et al.  Oxygen-derived free radicals stimulate osteoclastic bone resorption in rodent bone in vitro and in vivo. , 1990, The Journal of clinical investigation.

[38]  T. Chambers,et al.  Purification and characterization of a tartrate-resistant acid phosphatase from human osteoclastomas. , 1989, The Biochemical journal.

[39]  P. Apostoli,et al.  Free copper in serum: An analytical challenge and its possible applications. , 2018, Journal of trace elements in medicine and biology : organ of the Society for Minerals and Trace Elements.

[40]  Lei Chen,et al.  Copper-containing mesoporous bioactive glass scaffolds with multifunctional properties of angiogenesis capacity, osteostimulation and antibacterial activity. , 2013, Biomaterials.

[41]  D. Gray,et al.  Inhibition of active bone resorption by copper , 2006, Calcified Tissue International.

[42]  Inorganic Additives,et al.  THE EFFECTS OF INORGANIC ADDITIVES TO CALCIUM PHOSPHATE ON IN VITRO BEHAVIOR OF OSTEOBLASTS AND OSTEOCLASTS , 2006 .

[43]  J. Rodríguez,et al.  Modulation of the proliferation and differentiation of human mesenchymal stem cells by copper , 2002, Journal of cellular biochemistry.