Lasers in Medical Science

, Volume 32, Issue 1, pp 189–200 | Cite as

Comparison of the alendronate and irradiation with a light-emitting diode (LED) on murine osteoclastogenesis

  • Hong Moon Sohn
  • Youngjong Ko
  • Mineon Park
  • Bora Kim
  • Jung Eun Park
  • Donghwi Kim
  • Young Lae Moon
  • Wonbong LimEmail author
Original Article


Photomodulation therapy (PBMT) using light-emitting diode (LED) has been proposed as an alternative to conventional osteoporosis therapies. Our aim was to determine the effect of irradiation with a light-emitting diode on receptor activator of NF-κB ligand (RANKL)-mediated differentiation of mouse bone marrow macrophages into osteoclasts and compare it to alendronate treatment. The cells were irradiated with LED at 635±10 nm, 9-cm spot size, 5 mW/cm2, and 18 J for 60 min/day in a CO2 incubator. The differentiation of irradiated and untreated RANKL-stimulated bone marrow macrophages into osteoclasts was evaluated by tartrate-resistant acid phosphatase (TRAP) staining and by molecular methods. These included assessing messenger RNA (mRNA) expression of osteoclastic markers such as TRAP, c-Fos, Atp6v0d2, DC-STAMP, NFATc1, cathepsin K, MMP9 and OSCAR; phosphorylation of various MAPKs, including extracellular signal-regulated kinase ERK1/2, P38, and JNK; NF-κB translocation; and resorption pit formation. Results were compared to those obtained with sodium alendronate. Production of reactive oxygen species was measured by a 2’,7’-dihydrodichlorofluorescein diacetate assay. LED irradiation and alendronate inhibited mRNA expression of osteoclast-related genes, such as TRAP, c-Fos, and NFATc1, and reduced the osteoclast activity of RANKL-stimulated bone marrow macrophages. LED irradiation, but not alendronate, also inhibited the production of reactive oxygen species (ROS); phosphorylation of ERK, P38, and IκB; and NF-κB translocation. These findings suggest that LED irradiation downregulates osteoclastogenesis by ROS production; this effect could lead to reduced bone loss and may offer a new therapeutic tool for managing osteoporosis.


Osteoporosis LED ROS Osteoclastogenesis 



This study was supported by research funds from Chosun University (2014).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

All experimental procedures involving animals were compliant with institutional and governmental requirements and were approved by the Institutional Animal Care and Use Committee (CIACUC2014-A0023) of Chosun University, Gwangju, Korea.


  1. 1.
    Goltzman D (2002) Discoveries, drugs and skeletal disorders. Nat Rev Drug Discov 1:784–796CrossRefPubMedGoogle Scholar
  2. 2.
    Sims NA, Martin TJ (2014) Coupling the activities of bone formation and resorption: a multitude of signals within the basic multicellular unit. BoneKEy Rep 3:481PubMedPubMedCentralGoogle Scholar
  3. 3.
    Boyle WJ, Simonet WS, Lacey DL (2003) Osteoclast differentiation and activation. Nature 423:337–342CrossRefPubMedGoogle Scholar
  4. 4.
    Teitelbaum SL, Ross FP (2003) Genetic regulation of osteoclast development and function. Nat Rev Genet 4:638–649CrossRefPubMedGoogle Scholar
  5. 5.
    Isomura H, Fujie K, Shibata K, Inoue N, Iizuka T, Takebe G, Takahashi K, Nishihira J, Izumi H, Sakamoto W (2004) Bone metabolism and oxidative stress in postmenopausal rats with iron overload. Toxicology 197:93–100CrossRefPubMedGoogle Scholar
  6. 6.
    Bai XC, Lu D, Bai J, Zheng H, Ke ZY, Li XM, Luo SQ (2004) Oxidative stress inhibits osteoblastic differentiation of bone cells by ERK and NF-kappaB. Biochem Biophys Res Commun 314:197–207CrossRefPubMedGoogle Scholar
  7. 7.
    Garrett IR, Boyce BF, Oreffo RO, Bonewald L, Poser J, Mundy GR (1990) Oxygen-derived free radicals stimulate osteoclastic bone resorption in rodent bone in vitro and in vivo. J Clin Invest 85:632–639CrossRefPubMedPubMedCentralGoogle Scholar
  8. 8.
    Ha H, Kwak HB, Lee SW, Jin HM, Kim HM, Kim HH, Lee ZH (2004) Reactive oxygen species mediate RANK signaling in osteoclasts. Exp Cell Res 301:119–127CrossRefPubMedGoogle Scholar
  9. 9.
    Baron R, Ferrari S, Russell RG (2011) Denosumab and bisphosphonates: different mechanisms of action and effects. Bone 48:677–692CrossRefPubMedGoogle Scholar
  10. 10.
    Zhuo Y, Gauthier JY, Black WC, Percival MD, Duong LT (2014) Inhibition of bone resorption by the cathepsin K inhibitor odanacatib is fully reversible. Bone 67:269–280CrossRefPubMedGoogle Scholar
  11. 11.
    Ueda Y, Shimizu N (2003) Effects of pulse frequency of low-level laser therapy (LLLT) on bone nodule formation in rat calvarial cells. J Clin Laser Med Surg 21:271–277CrossRefPubMedGoogle Scholar
  12. 12.
    Hopkins JT, McLoda TA, Seegmiller JG, David Baxter G (2004) Low-level laser therapy facilitates superficial wound healing in humans: a triple-blind, sham-controlled study. J Athl Train 39:223–229PubMedPubMedCentralGoogle Scholar
  13. 13.
    Hussein AJ, Alfars AA, Falih MA, Hassan AN (2011) Effects of a low level laser on the acceleration of wound healing in rabbits. N Am J Med Sci 3:193–197CrossRefPubMedPubMedCentralGoogle Scholar
  14. 14.
    Amid R, Kadkhodazadeh M, Ahsaie MG, Hakakzadeh A (2014) Effect of low level laser therapy on proliferation and differentiation of the cells contributing in bone regeneration. J Lasers Med Sci 5:163–170PubMedPubMedCentralGoogle Scholar
  15. 15.
    Grassi FR, Ciccolella F, D’Apolito G, Papa F, Iuso A, Salzo AE, Trentadue R, Nardi GM, Scivetti M, De Matteo M, Silvestris F, Ballini A, Inchingolo F, Dipalma G, Scacco S, Tete S (2011) Effect of low-level laser irradiation on osteoblast proliferation and bone formation. J Biol Regul Homeost Agents 25:603–614PubMedGoogle Scholar
  16. 16.
    Lim W, Lee S, Kim I, Chung M, Kim M, Lim H, Park J, Kim O, Choi H (2007) The anti-inflammatory mechanism of 635 nm light-emitting-diode irradiation compared with existing COX inhibitors. Lasers Surg Med 39:614–621CrossRefPubMedGoogle Scholar
  17. 17.
    Bjordal JM, Couppe C, Chow RT, Tuner J, Ljunggren EA (2003) A systematic review of low level laser therapy with location-specific doses for pain from chronic joint disorders. Aust J Physiother 49:107–116CrossRefPubMedGoogle Scholar
  18. 18.
    Lim WB, Kim JS, Ko YJ, Kwon H, Kim SW, Min HK, Kim O, Choi HR, Kim OJ (2011) Effects of 635nm light-emitting diode irradiation on angiogenesis in CoCl(2) -exposed HUVECs. Lasers Surg Med 43:344–352CrossRefPubMedGoogle Scholar
  19. 19.
    Lubart R, Lavi R, Friedmann H, Rochkind S (2006) Photochemistry and photobiology of light absorption by living cells. Photomed Laser Surg 24:179–185CrossRefPubMedGoogle Scholar
  20. 20.
    Kanzaki H, Shinohara F, Kajiya M, Kodama T (2013) The Keap1/Nrf2 protein axis plays a role in osteoclast differentiation by regulating intracellular reactive oxygen species signaling. J Biol Chem 288:23009–23020CrossRefPubMedPubMedCentralGoogle Scholar
  21. 21.
    Pretel H, Lizarelli RF, Ramalho LT (2007) Effect of low-level laser therapy on bone repair: histological study in rats. Lasers Surg Med 39:788–796CrossRefPubMedGoogle Scholar
  22. 22.
    Kazem Shakouri S, Soleimanpour J, Salekzamani Y, Oskuie MR (2010) Effect of low-level laser therapy on the fracture healing process. Lasers Med Sci 25:73–77CrossRefPubMedGoogle Scholar
  23. 23.
    Pinheiro AL, Limeira Junior Fde A, Gerbi ME, Ramalho LM, Marzola C, Ponzi EA (2003) Effect of low level laser therapy on the repair of bone defects grafted with inorganic bovine bone. Braz Dent J 14:177–181CrossRefPubMedGoogle Scholar
  24. 24.
    Bartell SM, Kim HN, Ambrogini E, Han L, Iyer S, Serra Ucer S, Rabinovitch P, Jilka RL, Weinstein RS, Zhao H, O’Brien CA, Manolagas SC, Almeida M (2014) FoxO proteins restrain osteoclastogenesis and bone resorption by attenuating H2O2 accumulation. Nat Commun 5:3773CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Lim HJ, Bang MS, Jung HM, Shin JI, Chun GS, Oh CH (2014) A 635-nm light-emitting diode (LED) therapy inhibits bone resorptive osteoclast formation by regulating the actin cytoskeleton. Lasers Med Sci 29:659–670CrossRefPubMedGoogle Scholar
  26. 26.
    Bouvet-Gerbettaz S, Merigo E, Rocca JP, Carle GF, Rochet N (2009) Effects of low-level laser therapy on proliferation and differentiation of murine bone marrow cells into osteoblasts and osteoclasts. Lasers Surg Med 41:291–297CrossRefPubMedGoogle Scholar
  27. 27.
    Fisher JE, Rogers MJ, Halasy JM, Luckman SP, Hughes DE, Masarachia PJ, Wesolowski G, Russell RG, Rodan GA, Reszka AA (1999) Alendronate mechanism of action: geranylgeraniol, an intermediate in the mevalonate pathway, prevents inhibition of osteoclast formation, bone resorption, and kinase activation in vitro. Proc Natl Acad Sci U S A 96:133–138CrossRefPubMedPubMedCentralGoogle Scholar
  28. 28.
    Rao LG, Mackinnon ES, Josse RG, Murray TM, Strauss A, Rao AV (2007) Lycopene consumption decreases oxidative stress and bone resorption markers in postmenopausal women. Osteoporos Int 18:109–115CrossRefPubMedGoogle Scholar
  29. 29.
    Jiang F, Zhang Y, Dusting GJ (2011) NADPH oxidase-mediated redox signaling: roles in cellular stress response, stress tolerance, and tissue repair. Pharmacol Rev 63:218–242CrossRefPubMedGoogle Scholar
  30. 30.
    Ishizuka H, Garcia-Palacios V, Lu G, Subler MA, Zhang H, Boykin CS, Choi SJ, Zhao L, Patrene K, Galson DL, Blair HC, Hadi TM, Windle JJ, Kurihara N, Roodman GD (2011) ADAM8 enhances osteoclast precursor fusion and osteoclast formation in vitro and in vivo. J Bone Miner Res 26:169–181CrossRefPubMedGoogle Scholar
  31. 31.
    Takayanagi H, Kim S, Koga T, Nishina H, Isshiki M, Yoshida H, Saiura A, Isobe M, Yokochi T, Inoue J, Wagner EF, Mak TW, Kodama T, Taniguchi T (2002) Induction and activation of the transcription factor NFATc1 (NFAT2) integrate RANKL signaling in terminal differentiation of osteoclasts. Dev Cell 3:889–901CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag London 2016

Authors and Affiliations

  • Hong Moon Sohn
    • 1
  • Youngjong Ko
    • 1
  • Mineon Park
    • 1
  • Bora Kim
    • 1
  • Jung Eun Park
    • 2
  • Donghwi Kim
    • 1
  • Young Lae Moon
    • 1
  • Wonbong Lim
    • 1
    • 3
    Email author
  1. 1.Department of Orthopaedic SurgeryChosun University HospitalGwangjuKorea
  2. 2.Department of Biomedical Science, BK21-Plus Research Team for Bioactive ControlChosun UniversityGwangjuKorea
  3. 3.Department of Premedical SciencesCollege of Medicine, Chosun UniversityGwangjuRepublic of Korea

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