Abstract
Magnesium alloys have been investigated by many researchers as a new absorbable biomaterial owing to their excellent degradability with non-maleficence or low-maleficence in living tissues. In the present work, the in vitro cytocompatibility of an Magnesium alloy was investigated by culturing cells directly on it. Investigations were carried out in terms of the cell viability along with the use of scanning electron microscopy to observe its morphology. The cell lines used were derived from fibroblast, endothelial, and smooth muscle cells. Pure magnesium and AZ31 alloy composed of magnesium (96 %), aluminum (3 %), and zinc (1 %) were adopted as models. The viability of cells on the metal samples and on the margin area of a multi-well plate was investigated. For direct culturing on metal, a depression in the viability and morphologically stressed cells were observed. In addition, the cell viability was also depressed for the margin area. To clarify the factors causing the negative effects, the amount of eluted metal ions and pH changes in the medium because of the erosion of the Magnesium samples were investigated, together with the cytotoxicity of sole metal ions corresponding to the composition of the metals. It was found that Mg2+, Zn2+, and Al3+ ions were less toxic at the investigated concentrations, and that these factors will not produce negative effects on cells. Consequently, these factors cannot fully explain the results.
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Stainger MP, Pietak AM, Huadmai J, Dias G. Magnesium and its alloys as orthopedic biomaterials: A review. Biomaterials 2006;27:1728–34.
Wolf FI, Cittadini A. Chemistry and biochemistry of magnesium. Mol Aspects Med. 2003;24:3–9.
Herring WB, Leavell BS, aixao LM, Yoe JH. Trace metal in human plasma and red blood cells. A study of magnesium, chromium, Nickel, Copper and Zinc I. Observation of normal subjects. Am J Clin Nutr. 1960;8:846–54.
Saris N-EL, Mervaala E, Kappanen H, Khawaja JA, Lewenstam A. Magnesium an update on physiological, clinical and analytical aspects. Clin Chim Acta. 2000;294:1–26.
Scheideler L, Füger C, Schille C, Rupp F, Wendel H-P, Hort N, Reichel HP, Geis-Gerstorfer J. Comparison of different in vitro tests for biocompatibility screening of Mg alloys. Acta Biometer. 2013;9:8740–45.
Zhu WR, Zheng YM, Leeflang MA, Zhou J. Mechanical property, biocorrosion and in vitro biocompatibility evaluation of Mg-Li-(Al)-(RE) alloys for future cardiovascular stent application. Acta Biomater. 2013;9:8488–98.
Gu XN, Li N, Zheng YF, Ruan L. In vitro degradation performance and biological response of a Mg-Zn Zr alloy. Mater Sci Eng B 2011;176:1778–84.
Fan J, Qiu X, Niu X, Tian Z, Sun W, Liu X, Li Y, Li W, Meng J. Microstructure, mechanical properties, in vitro degradation and cytotoxicity evaluations of Mg-1.5Y-1.2Zn-0.44Zr alloys for biodegradable metallic implants. Mater. Sci Eng C 2013;33:2345–52.
Zhou Y-L, Li Y, Luo DM, Ding Y, Hodgson P. Microstructures, mechanical and corrosion properties and biocompatibility of as extruded Mg–Mn–Zn–Nd alloys for biomedical applications. Mater Sci Eng C 2015;49:93–100.
Pompa L, Rahman ZU, Munoz E, Haider W. Surface characterization and cytotoxicity response of biodegradable magnesium alloys. Mater Sci Eng C 2015;49:761–8.
Weizbauer A, Seitz J-M, Werle P, Hegermann J, Willbold E, Eifler R, Windhagen H, Reifenrath J, Waizy. H. Novel magnesium alloy Mg–2La caused no cytotoxic effects on cells in physiological conditions. Mater Sci Eng C 2014;41:267–73.
Zhang S, hang X, Zhao C, Li J, Song Y, Xie C, Tao H, Zhang Y, He Y, Jiang Y, Bian Y. Research on an Mg–Zn alloy as a degradable biomaterial. Acta Biomater. 2010;6:626–40.
Li Z, Gu X, Lou S, Zheng Y. The development of binary Mg-Ca alloys for use as biodegradable materials within bone. Biomaterials. 2008;29:1329–44.
Hänzi AC, Gerber I, Schinhammer M, Löffler JF, Uggowitzer PJ. On the in vitro and in vivo degradation performance and biological response of new biodegradable Mg-Y-Zn alloys. Acta Biomater. 2010;6:1824–33.
Xu L, Yu G, Zhang E, Pan F, Yang K. In vivo corrosion behavior of Mg-Mn-Zn alloy for bone implant application. J Biomed Mater Res. 2007;83A:703–11.
Montoya R, Iglesias C, Escudero ML, García-Alonso MC. Modeling in vivo corrosion of AZ31 as temporary biodegradable implants. Experimental validation in rats. Mater Sci Eng C 2014;41:127–33.
Gu X, Zheng Y, Zhong S, Xi T, Wang J, Wang W. Corrosion of, and cellular responses to Mg–Zn–Ca bulk metallic glasses. Biomaterials. 2010;31:1093–103.
Jung O, Smeets R, Porchetta D, Kopp A, Ptock C, Müller U, Heiland M, Schwade M, Behr B, Kröger N, Kluwe L, Hanken H, Hartjen P. Optimized in vitro procedure for assessing the cytocompatibility of magnesium-based biomaterials. Acta Biomater. 2015;23:354–63.
Witte F, Feyerabend F, Maier P, Fischer J, Störmer M, Blawert C, Dietzel W, Hort N. Biodegradable magnesium–hydroxyapatite metal matrix composites. Biomaterials. 2007;28:2163–74.
Persaud-Sharma D, Budiansky N, McGoron AJ. Biocompatibility assessment of novel bioresorbable alloys Mg-Zn-Se and Mg-Zn-Cu for endovascular applications: in- vitro studies. J Biomim Biomater Tissue Eng. 2013;17:25–44.
Johnson I, Perchy D, Liu H. In vitro evaluation of the surface effects on magnesium-yttrium alloy degradation and mesenchymal stem cell adhesion. J Biomed Mater Res. 2012;100A:477–85.
Liu. H. The effects of surface and biomolecules on magnesium degradation and mesenchymal stem cell adhesion. J Biomed Mater Res. 2011;99A:249–60.
Keim S, Brunner JG, Fabry B, Virtanen S. Control of magnesium corrosion and biocompatibility with biomimetic coatings. J Biomed Mater Res. 2011;96B:84–90.
Lorenz C, Brunner JG, Kollmannsberger P, Jaafar L, Fabry B, Virtanen S. Effect of surface pre-treatments on biocompatibility of magnesium. Acta Biomater 2009;5:2783–89.
ASTM B275-05 standard practice for codification of certain nonferrous metals and alloys, cast and wrought. In: Annual book of ASTM standards. Philadelphia, PA: American Society for Testing and Materials; 2005.
Walter R, Kannano MB. Influence of surface roughness on the corrosion behaviour of magnesium alloy. Mater Des. 2011;32:2350–4.
Lincks J, Boyan BD, Blanchard CR, Lohmann CH, Liu Y, Cochran DL, Dean DD, Schwartz. Z. Response of MG63 osteoblast-like cells to titanium and titanium alloy is dependent on surface roughness and composition. Biomaterials. 1998;19:2219–32
Chunga T-W, Liub D-Z, Wangc S-Y, Wang S-S. Enhancement of the growth of human endothelial cells by surface roughness at nanometer scale. Biomaterials. 2003;24:4655–61.
Huan ZG, Leeflang MA, Zhou J, Fratila-Apachitei LE, Duszczyk. J. In vitro degradation behavior and cytocompatibility of Mg–Zn–Zr alloys. J Mater Sci. Mater Med. 2010;21:2623–35.
Yamamoto A, Hiromoto S. Effect of inorganic salts, amino acids and proteins on the degradation of pure magnesium in vitro. Mater Sci Eng C 2009;29:1559–68.
Xu L, Zhang E, Yin D, Zeng S, Yang K. In vitro corrosion behaviour of Mg alloys in a phosphate buffered solution for bone implant application. J Mater Sci Mater Med. 2008;19:1017–25.
Zhen Z, Liu X, Huang T, Xi T-F, Zheng. Y. Hemolysis and cytotoxicity mechanisms of biodegradable magnesium and its alloys. Mater Sci Eng C 2015;46:202–6.
Kita H, Kimoto M, Kudo M. Influence of Al content on corrosion resistance of Mg-Al-Zn alloys in chloride environments. J Japan Inst Metal. 2005;9:805–9.
Nordlien JH, Nisancioglu K, Ono S, Masuko NJ. Morphology and structure of oxide films formed on Mg Al alloys by exposure to air and water. Electrochem Soc. 1996;143:2564–72.
Beutler E. Anemia resulting from other nutritional deficiencies. In: Lichtma MA, Beutler E, Kipps TJ, Seligshon U, Kaushansky K, Prchal JT, editors. Williams hematology. 7th ed. New York: McGraw-Hill; 2006, Chapter 41, p. 555-60.
Dean JA. Lange’s handbook of chemistry, section 8, electrolytes, electromotive force, and chemical equilibrium. 8.2 equilibrium constants, table 8.6 solubility product constants. 15th ed. New York: McGraw Hill; 1998.
Hallab NJ, Messina CVC, Roebuck KA, Glant TT, Jacobs JJ. Concentration- and composition-dependent effects of metal ions on human MG-63 osteoblasts. J Biomed Mater Res. 2002;60:420–33.
Gu X, Zheng Y, Cheng Y, Zhong S, Xi T. In vitro corrosion and biocompatibility of binary magnesium alloys. Biomaterials. 2009;30:484–98.
Nguyen TY, Cipriano AF, Guan R-G, Zhao Z-Y, Liu H. In vitro interactions of blood, platelet, and fibroblast with biodegradable magnesium-zinc-strontium alloys. J Biomed Mater Res. 2015;103A:2974–86.
Zhao N, Watson N, Xu Z, Chen Y, Waterman J, Sankar J, Zhu D. In vitro biocompatibility and endothelialization of novel magnesium-rare earth alloys for improved stent applications. Plos One. 2014;9:e98674
Fischer J, Prosenc MH, Wolff M, Hort N, Willumeit R, Feyerabend F. Interference of magnesium corrosion with tetrazolium-based cytotoxicity assays. Acta Biomater. 2010;6:1813–23.
Acknowledgments
This work was sponsored by E.S.Q. Ltd., (Yabuki, Fukushima, Japan) in the year 2014 under the subsidiary enterprise “Research and Development Programs for Medical and Welfare Apparatus” by Fukushima prefectural government. We thank Support Center for Medical Research and Education, Tokai University for their technical support in SEM observation and Technology Joint Management Office for their measurement of surface roughness.
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Mochizuki, A., Yahata, C. & Takai, H. Cytocompatibility of magnesium and AZ31 alloy with three types of cell lines using a direct in vitro method. J Mater Sci: Mater Med 27, 145 (2016). https://doi.org/10.1007/s10856-016-5762-x
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DOI: https://doi.org/10.1007/s10856-016-5762-x