Simvastatin and zinc synergistically enhance osteoblasts activity and decrease the acute response of inflammatory cells

  • Maryam Montazerolghaem
  • Yi Ning
  • Håkan Engqvist
  • Marjam Karlsson Ott
  • Maria Tenje
  • Gemma Mestres
Delivery Systems Original Research
Part of the following topical collections:
  1. Delivery Systems


Several ceramic biomaterials have been suggested as promising alternatives to autologous bone to replace or restore bone after trauma or disease. The osteoinductive potential of most scaffolds is often rather low by themselves and for this reason growth factors or drugs have been supplemented to these synthetic materials. Although some growth factors show good osteoinductive potential their drawback is their high cost and potential severe side effects. In this work the combination of the well-known drug simvastatin (SVA) and the inorganic element Zinc (Zn) is suggested as a potential additive to bone grafts in order to increase their bone regeneration/formation. MC3T3-E1 cells were cultured with Zn (10 and 25 µM) and SVA (0.25 and 0.4 µM) for 10 days to evaluate proliferation and differentiation, and for 22 days to evaluate secretion of calcium deposits. The combination of Zn (10 µM) and SVA (0.25 µM) significantly enhanced cell differentiation and mineralization in a synergetic manner. In addition, the release of reactive oxygen species (ROS) from primary human monocytes in contact with the same concentrations of Zn and SVA was evaluated by chemiluminescence. The combination of the additives decreased the release of ROS, although Zn and SVA separately caused opposite effects. This work shows that a new combination of additives can be used to increase the osteoinductive capacity of porous bioceramics.

Graphical Abstract


Reactive Oxygen Species Simvastatin Calcium Deposit Calcium Phosphate Cement Lower Cell Number 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



This work was co-funded by Marie Curie Actions FP7-PEOPLE-2011-COFUND (GROWTH 291795) via the VINNOVA programme Mobility for Growth (project n. 2013-01260) and Lars Hiertas Minne Foundation (project n. FO2014-0334). Financial support from Uppsala University is acknowledged by MT, HE and MM.


  1. 1.
    De Long WG, Einhorn TA, Koval K, McKee M, Smith W, Sanders R, et al. Bone grafts and bone graft substitutes in orthopaedic trauma surgery. A critical analysis. J Bone Joint Surg Am. 2007;89:649–58.CrossRefGoogle Scholar
  2. 2.
    Arrington ED, Smith WJ, Chambers HG, Bucknell AL, Davino NA. Complications of iliac crest bone graft harvesting. Clin Orthop Relat Res. 1996;329:300–9.CrossRefGoogle Scholar
  3. 3.
    Giannoudis PV, Dinopoulos H, Tsiridis E. Bone substitutes: an update. Injury. 2005;36(Suppl 3):S20–7.CrossRefGoogle Scholar
  4. 4.
    LeGeros RZ. Calcium phosphate-based osteoinductive materials. Chem Rev. 2008;108:4742–53.CrossRefGoogle Scholar
  5. 5.
    Ruhé PQ, Boerman OC, Russel FGM, Mikos AG, Spauwen PHM, Jansen JA. In vivo release of rhBMP-2 loaded porous calcium phosphate cement pretreated with albumin. J Mater Sci Mater Med. 2006;17:919–27.CrossRefGoogle Scholar
  6. 6.
    Wang H, Boerman OC, Sariibrahimoglu K, Li Y, Jansen JA. Leeuwenburgh SCG. Comparison of micro- versus nanostructured colloidal gelatin gels for sustained delivery of osteogenic proteins: bone morphogenetic protein-2 and alkaline phosphatase. Biomaterials. 2012;33:8695–703.CrossRefGoogle Scholar
  7. 7.
    Bessa P, Casal M, Reis R. Bone morphogenetic proteins in tissue engineering: the road from laboratory to clinic, part II (BMP delivery). J Tissue Eng Regen Med. 2008;2:81–96.CrossRefGoogle Scholar
  8. 8.
    Hari Reddi A. Role of morphogenetic proteins in skeletal tissue engineering and regeneration. Nature. 1998;16:291–4.Google Scholar
  9. 9.
    Poynton A, Lane J. Safety profile for the clinical use of bone morphogenetic proteins in the spine. Spine (Phila Pa 1976). 2002;27:S40–8.CrossRefGoogle Scholar
  10. 10.
    Germershausen JI, Hunt VM, Bostedor RG, Bailey PJ, Karkas JD, Alberts AW. Tissue selectivity of the cholesterol-lowering agents lovastatin, simvastatin and pravastatin in rats in vivo. Biochem Biophys Res Commun. 1989;158:667–75.CrossRefGoogle Scholar
  11. 11.
    Maeda T, Matsunuma A, Kawane T, Horiuchi N. Simvastatin promotes osteoblast differentiation and mineralization in MC3T3-E1 cells. Biochem Biophys Res Commun. 2001;280:874–7.CrossRefGoogle Scholar
  12. 12.
    Chen PY, Sun JS, Tsuang YH, Chen MH, Weng PW, Lin FH. Simvastatin promotes osteoblast viability and differentiation via Ras/Smad/Erk/BMP-2 signaling pathway. Nutr Res. 2010;30(3):191–9.CrossRefGoogle Scholar
  13. 13.
    Maeda T, Matsunuma A, Kurahashi I, Yanagawa T, Yoshida H, Horiuchi N. Induction of osteoblast differentiation indices by statins in MC3T3-E1 cells. J Cell Biochem. 2004;92:458–71.CrossRefGoogle Scholar
  14. 14.
    Mundy G. Stimulation of bone formation in vitro and in rodents by statins. Science. 1999;286:1946–9.CrossRefGoogle Scholar
  15. 15.
    Yamaguchi M. Role of nutritional zinc in the prevention of osteoporosis. Mol Cell Biochem. 2010;338:241–54.CrossRefGoogle Scholar
  16. 16.
    Gommans WM, Haisma HJ, Rots MG. Engineering zinc finger protein transcription factors: the therapeutic relevance of switching endogenous gene expression on or off at command. J Mol Biol. 2005;354:507–19.CrossRefGoogle Scholar
  17. 17.
    Kagami S, Kanari H, Suto A, Fujiwara M, Ikeda K, Hirose K, et al. HMG-CoA reductase inhibitor simvastatin inhibits proinflammatory cytokine production from murine mast cells. Int Arch Allergy Immunol. 2008;146(Suppl. 1):61–6.CrossRefGoogle Scholar
  18. 18.
    Funk JL, Chen J, Downey KJ, Clark RA. Bone protective effect of simvastatin in experimental arthritis. J Rheumatol. 2008;35:1083–91.Google Scholar
  19. 19.
    Moon HJ, Kim SE, Yun YP, Hwang YS, Bang JB, Park JH, Kwon IK. Simvastatin inhibits osteoclast differentiation by scavenging reactive oxygen species. Exp Mol Med. 2011;43(11):605–12.CrossRefGoogle Scholar
  20. 20.
    Zalewski PD, Truong-Tran AQ, Grosser D, Jayaram L, Murgia C, Ruffin RE. Zinc metabolism in airway epithelium and airway inflammation: basic mechanisms and clinical targets. A review. Pharmacol Ther. 2005;105:127–49.CrossRefGoogle Scholar
  21. 21.
    Prasad AS. Zinc: role in immunity, oxidative stress and chronic inflammation. Curr Opin Clin Nutr Metab Care. 2009;12:646–52.CrossRefGoogle Scholar
  22. 22.
    Anderson JM. Biological responses to materials. Annu Rev Mater Res. 2001;31(1):81–110.CrossRefGoogle Scholar
  23. 23.
    Kaesemeyer WH, Caldwell RB, Huang J, Caldwell RW. Pravastatin sodium activates endothelial nitric oxide synthase independent of its cholesterol-lowering actions. J Am Coll Cardiol. 1999;33(1):234–41.CrossRefGoogle Scholar
  24. 24.
    Wellinghausen N, Rink L. The significance of zinc for leukocyte biology. J Leukoc Biol. 1998;64:571–7.Google Scholar
  25. 25.
    Coelho MJ, Fernandes MH. Human bone cell cultures in biocompatibility testing. Part II: effect of ascorbic acid, beta-glycerophosphate and dexamethasone on osteoblastic differentiation. Biomaterials. 2000;21:1095–102.CrossRefGoogle Scholar
  26. 26.
    Aubin J, Liu F. The osteoblast lineage. In: Bilezikian J, Raisz L, Rodan G (Eds). Principles of Bone Biology. San Diego: Academic Press; 1996, p. 51–67.Google Scholar
  27. 27.
    Dahlgren C, Karlsson A. Respiratory burst in human neutrophils. J Immunol Methods. 1999;232:3–14.CrossRefGoogle Scholar
  28. 28.
    Sugiyama M, Kodama T, Konishi K, Abe K, Asami S, Oikawa S. Compactin and simvastatin, but not pravastatin, induce bone morphogenetic protein-2 in human osteosarcoma cells. Biochem Biophys Res Commun. 2000;271:688–92.CrossRefGoogle Scholar
  29. 29.
    Skoglund B, Forslund C, Aspenberg P. Simvastatin improves fracture healing in mice. J Bone Miner Res. 2002;17:2004–8.CrossRefGoogle Scholar
  30. 30.
    Yamaguchi M, Oishi H, Suketa Y. Stimulatory effect of zinc on bone formation in tissue culture. Biochem Pharmacol. 1987;36:4007–12.CrossRefGoogle Scholar
  31. 31.
    Stein G, Lian J. Molecular mechanisms mediating developmental and hormone-regulated expression of genes in osteoblasts: an integrated relationship of cell growth and differentiation. In: Noda M, editor. Cellular and Molecular Biology of Bone. Tokyo: Academic Press Inc; 1993. p. 47–95.Google Scholar
  32. 32.
    Owen T, Aronow M, Shalhoub V. Progressive development of the rat osteoblast phenotype in vitro: reciprocal relationships in expression of genes associated with osteoblast proliferation during formation of bone extracellular matrix. J Cell Phys. 1990;143:420–30.CrossRefGoogle Scholar
  33. 33.
    Vogel C, Marcotte EM. Insights into regulation of protein abundance from proteomics and transcriptomis analyses. Nat Rev Genet. 2013;13:227–32.CrossRefGoogle Scholar
  34. 34.
    Taylor RC, Webb Robertson B-JM, Markillie LM, Serres MH, Linggi BE, Aldrich JT, et al. Changes in translational efficiency is a dominant regulatory mechanism in the environmental response of bacteria. Integr Biol. 2013;5:1393.CrossRefGoogle Scholar
  35. 35.
    Fosmire GJ. Zinc toxicity. Am J Clin Nutr. 1990;51:225–7.Google Scholar
  36. 36.
    Law M, Rudnicka AR. Statin safety: a systematic review. Am J Cardiol. 2006;97:52C–60C.CrossRefGoogle Scholar
  37. 37.
    Mestres G, Santos CF, Engman L, Persson C, Ott KM. Scavenging effect of Trolox released from brushite cements. Acta Biomater. 2015;11:459–66.CrossRefGoogle Scholar
  38. 38.
    Canal C, Pastorino D, Mestres G, Schuler P, Ginebra MP. Relevance of microstructure for the early antibiotic release of fresh and pre-set calcium phosphate cements. Acta Biomater. 2013;9:8403–12.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Maryam Montazerolghaem
    • 1
  • Yi Ning
    • 1
  • Håkan Engqvist
    • 1
  • Marjam Karlsson Ott
    • 1
    • 2
  • Maria Tenje
    • 1
    • 2
    • 3
  • Gemma Mestres
    • 1
  1. 1.Department Engineering SciencesUppsala UniversityUppsalaSweden
  2. 2.Science for Life LaboratoryUppsalaSweden
  3. 3.Department Biomedical EngineeringLund UniversityLundSweden

Personalised recommendations