International Orthopaedics

, Volume 31, Issue 2, pp 223–228 | Cite as

Gentamicin negatively influenced osteogenic function in vitro

  • Akif Ince
  • Norbert Schütze
  • Nadja Karl
  • Jochen F. Löhr
  • Jochen Eulert
Original Paper


Local delivery of gentamicin is an accepted method of infection prophylaxis in the surgery of open fractures. However, the few reports of studies into the effect of locally applied gentamicin on osteoblasts used inadequate methods. In our study, we used the well-characterised C2C12 cell line with reproducible differentiation pathway into the osteoblast lineage. We investigated the viability, cell number, alkaline phosphatase activity, and the expression of osteogenic genes of C2C12 cells after exposure to gentamicin at concentrations of 12.5–800 μg/ml for 48 h. Exposure of C2C12 cells to gentamicin (12.5–800 mg/ml) for 48 h showed no significant changes in the cell number, but cell viability was decreased by one-third at the tested concentrations of 200–800 μg/ml. The alkaline phosphatase activity was significantly decreased by one-third to one-half at any tested concentration (12.5–800 μg/ml) of gentamicin. Any tested concentration of gentamicin up to 800 μg/ml for 48 h did not inhibit or decrease the osteogenic gene expression of osterix and alkaline phosphatase of the C2C12 cells. In conclusion, gentamicin at high concentrations as achieved by local application reduced cellular viability and alkaline phosphatase activity in vitro and therefore may be detrimental for bone healing and repair in vivo.


La délivrance locale de gentamycine est une méthode usuelle de prophylaxie de l’infection dans la chirurgie des fractures ouvertes. Il y a peu d’études fiables sur les effets de la gentamycine sur les ostéoblastes. Dans cette étude est utilisée la lignée C2C12 avec reproductibilité des voies de différenciation dans la lignée ostéoblastique. Nous avons étudié la viabilité, le nombre de cellules, l’activité phosphatase alcaline et l’expression des gènes ostéogèniques des cellules C2C12 après exposition pendant 48 heures à des concentrations de gentamycine de 12,5 μg/ml à 800 μg/ml. Il n’y avait pas de modification significative du nombre de cellules mais la viabilité était diminuée d’un tiers pour les concentrations de 200 à 800 μg/ml. L’activité phosphatase alcaline était diminuée d’un tiers à la moitié pour toutes les concentrations étudiées. Dans aucun cas il n’y avait diminution ou inhibition de l’expression génique de l’osterix ou de la phosphatase alcaline des cellules C1C12. En conclusion, la gentamycine à haute concentration réduit la viabilité cellulaire et l’activité phosphatase alcaline in vitro, ce qui est peut-être néfaste pour la cicatrisation osseuse in vivo.


  1. 1.
    Beardmore AA, Brooks DE, Wenke JC, Thomas DB (2005) Effectiveness of local antibiotic delivery with an osteoinductive and osteoconductive bone-graft substitute. J Bone Joint Surg Am 87:107–112CrossRefPubMedGoogle Scholar
  2. 2.
    Blau HM, Chiu CP, Webster C (1983) Cytoplasmic activation of human nuclear genes in stable heterocaryons. Cell 32:1171–1180CrossRefPubMedGoogle Scholar
  3. 3.
    Boyan BD, Schwartz Z, Bonewald LF, Swain LD (1989) Localization of 1,25-(OH)2D3-responsive alkaline phosphatase in osteoblast-like cells (ROS 17/2.8, MG 63, and MC 3T3) and growth cartilage cells in culture. J Biol Chem 264:11879–11886PubMedGoogle Scholar
  4. 4.
    Campbell AA, Song L, Li XS, Nelson BJ, Bottoni C, Brooks DE, DeJong ES (2000) Development, characterization, and anti-microbial efficacy of hydroxyapatite-chlorhexidine coatings produced by surface-induced mineralization. J Biomed Mater Res 53:400–407CrossRefPubMedGoogle Scholar
  5. 5.
    Celil AB, Hollinger JO, Campbell PG (2005) Osx transcriptional regulation is mediated by additional pathways to BMP2/Smad signaling. J Cell Biochem 95:518–528CrossRefPubMedGoogle Scholar
  6. 6.
    Darouiche RO, Farmer J, Chaput C, Mansouri M, Saleh G, Landon GC (1998) Anti-infective efficacy of antiseptic-coated intramedullary nails. J Bone Joint Surg Am 80:1336–1340PubMedGoogle Scholar
  7. 7.
    DeJong ES, DeBerardino TM, Brooks DE, Nelson BJ, Campbell AA, Bottoni CR, Pusateri AE, Walton RS, Guymon CH, McManus AT (2001) Antimicrobial efficacy of external fixator pins coated with a lipid stabilized hydroxyapatite/chlorhexidine complex to prevent pin tract infection in a goat model. J Trauma 50:1008–1014PubMedCrossRefGoogle Scholar
  8. 8.
    Hendrich C, Noth U, Stahl U, Merklein F, Rader CP, Schutze N, Thull R, Tuan RS, Eulert J (2002) Testing of skeletal implant surfaces with human fetal osteoblasts. Clin Orthop 394:278–289PubMedCrossRefGoogle Scholar
  9. 9.
    Huddleston PM, Steckelberg JM, Hanssen AD, Rouse MS, Bolander ME, Patel R (2000) Ciprofloxacin inhibition of experimental fracture healing. J Bone Joint Surg Am 82:161–173PubMedGoogle Scholar
  10. 10.
    Ince A, Seemann K, Frommelt L, Katzer A, Lohr JF (2004) One-stage revision of shoulder arthroplasty in the case of periprosthetic infection (in German). Z Orthop Ihre Grenzgeb 142:611–617CrossRefPubMedGoogle Scholar
  11. 11.
    Isefuku S, Joyner CJ, Simpson AH (2003) Gentamicin may have an adverse effect on osteogenesis. J Orthop Trauma 17:212–216CrossRefPubMedGoogle Scholar
  12. 12.
    Ishiyama M, Tominaga H, Shiga M, Sasamoto K, Ohkura Y, Ueno K (1996) A combined assay of cell viability and in vitro cytotoxicity with a highly water-soluble tetrazolium salt, neutral red and crystal violet. Biol Pharm Bull 19:1518–1520PubMedGoogle Scholar
  13. 13.
    Jadlowiec J, Koch H, Zhang X, Campbell PG, Seyedain M, Sfeir C (2004) Phosphophoryn regulates the gene expression and differentiation of NIH3T3, MC3T3-E1, and human mesenchymal stem cells via the integrin/MAPK signaling pathway. J Biol Chem 279:53323–53330CrossRefPubMedGoogle Scholar
  14. 14.
    Katagiri T, Yamaguchi A, Komaki M, Abe E, Takahashi N, Ikeda T, Rosen V, Wozney JM, Fujisawa-Sehara A, Suda T (1994) Bone morphogenetic protein-2 converts the differentiation pathway of C2C12 myoblasts into the osteoblast lineage. J Cell Biol 127:1755–1766CrossRefPubMedGoogle Scholar
  15. 15.
    Keating JF, Blachut PA, O’Brien PJ, Meek RN, Broekhuyse H (1996) Reamed nailing of open tibial fractures: does the antibiotic bead pouch reduce the deep infection rate? J Orthop Trauma 10:298–303CrossRefPubMedGoogle Scholar
  16. 16.
    McKee MD, Wild LM, Schemitsch EH, Waddell JP (2002) The use of an antibiotic-impregnated, osteoconductive, bioabsorbable bone substitute in the treatment of infected long bone defects: early results of a prospective trial. J Orthop Trauma 16:622–627CrossRefPubMedGoogle Scholar
  17. 17.
    Miclau T, Edin ML, Lester GE, Lindsey RW, Dahners LE (1998) Effect of ciprofloxacin on the proliferation of osteoblast-like MG-63 human osteosarcoma cells in vitro. J Orthop Res 16:509–512CrossRefPubMedGoogle Scholar
  18. 18.
    Nakashima K, Zhou X, Kunkel G, Zhang Z, Deng JM, Behringer RR, de Crombrugghe B (2002) The novel zinc finger-containing transcription factor osterix is required for osteoblast differentiation and bone formation. Cell 108:17–29CrossRefPubMedGoogle Scholar
  19. 19.
    Pedersen JG, Lund B (1988) Effects of gentamicin and monomer on bone. An in vitro study. J Arthroplasty 3(Suppl):S63–S68PubMedCrossRefGoogle Scholar
  20. 20.
    Perry AC, Prpa B, Rouse MS, Piper KE, Hanssen AD, Steckelberg JM, Patel R (2003) Levofloxacin and trovafloxacin inhibition of experimental fracture-healing. Clin Orthop 414:95–100PubMedCrossRefGoogle Scholar
  21. 21.
    Sakano S, Murata Y, Miura T, Iwata H, Sato K, Matsui N, Seo H (1993) Collagen and alkaline phosphatase gene expression during bone morphogenetic protein (BMP)-induced cartilage and bone differentiation. Clin Orthop Relat Res 292:337–344PubMedGoogle Scholar
  22. 22.
    Steinbrink K, Frommelt L (1995) Treatment of periprosthetic infection of the hip using one-stage exchange surgery (in German). Orthopade 24:335–343PubMedGoogle Scholar
  23. 23.
    Sundin DP, Sandoval R, Molitoris BA (2001) Gentamicin inhibits renal protein and phospholipid metabolism in rats: implications involving intracellular trafficking. J Am Soc Nephrol 12:114–123CrossRefPubMedGoogle Scholar
  24. 24.
    Wahlig H, Dingeldein E, Buchholz HW, Buchholz M, Bachmann F (1984) Pharmacokinetic study of gentamicin-loaded cement in total hip replacements. Comparative effects of varying dosage. J Bone Joint Surg Br 66:175–179PubMedGoogle Scholar
  25. 25.
    Yaffe D, Saxel O (1977) Serial passaging and differentiation of myogenic cells isolated from dystrophic mouse muscle. Nature 270:725–727CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Akif Ince
    • 1
    • 2
    • 3
  • Norbert Schütze
    • 1
  • Nadja Karl
    • 1
  • Jochen F. Löhr
    • 2
  • Jochen Eulert
    • 1
  1. 1.Department of Orthopaedic SurgeryUniversity Hospital WürzburgWürzburgGermany
  2. 2.Department of Orthopaedic SurgeryENDO-KlinikHamburgGermany
  3. 3.HamburgGermany

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