Biological Trace Element Research

, Volume 187, Issue 2, pp 492–498 | Cite as

Requirement of TGFβ Signaling for Effect of Fluoride on Osteoblastic Differentiation

  • Jingmin Zhang
  • Ningning Jiang
  • Haolan Yu
  • Xiuhua Yu
  • Fengyang Guo
  • Zhitao Zhao
  • Hui XuEmail author


Research focused on transforming growth factor β (TGFβ) signaling in osteoblast is gradually increasing, whereas literature is rare in terms of fluorosis. This work aimed to investigate how TGFβ signaling participated in regulation of the osteoblast by different doses of fluoride treatment. Bone marrow stem cells (BMSCs) were developed into osteoblastic cells and exposed to 1, 4, and 16 mg/L F with and without 10 ng/mL of TGFβ. Cell viability and differentiation state of osteoblast under different settings were measured by means of cell counting kit and analysis of alkaline phosphatase (ALP) activity as well as formation of mineral nodules. Real-time PCR was utilized to test expression of ALP and Runt-related transcription factor 2 (Runx2) at gene level. The gene expression of TGFβ signaling effectors was also investigated, such as TGFβ receptors (TβRs), smad3, and mitogen-activated protein kinases (MAPK). Results demonstrated that fluoride treatment exhibited action on osteoblast viability and osteogenic differentiation and upregulated expression of TβR2, smad3, and MAPK in this process. Administration of TGFβ strengthened ALP activity but attenuated formation of mineral nodules. Co-treatment of TGFβ and low-dose fluoride increased ALP activity compared to same dose of single fluoride treatment, whereas it inhibited mineral nodule formation. Administration of TGFβ reversed the suppression of high-dose fluoride on osteogenic differentiation of BMSCs. Taken together, studies revealed that TβR2 acted as a target for fluoride and TGFβ treatment on BMSCs, and smad3 and MAPK were involved in the mechanism of fluoride regulating osteogenic differentiation. Together, our data indicated that TGFβ receptor-mediated signaling through smad3 and MAPK was required for modulation of fluoride on osteoblast viability and differentiation, and activating TβR2-smad3 signaling pathway reversed suppression of osteoblasts differentiation by high dose of fluoride treatment.


Transforming growth factor beta Osteoblast Alkaline phosphatase Transforming growth factor beta receptor Mitogen-activated protein kinase Smad3 



This study was funded by project (Study on role of osteocyte and PTH/TGF-beta signaling pathway in the mechanism of bone turnover occurred in skeletal fluorosis) supported by the National Natural Science Foundation of China (grant number 81673111). This study was supported by a grant from Technical Innovation Project of Health and Family Planning Commission in Jilin Province of China (2016J054). This study was supported by the Natural Science Foundation of Jilin Province of China (20180101151JC).

Compliance with Ethical Standards

All procedures performed in studies involving mice were in accordance with the ethical standards of the Animal Welfare Ethical Committee of Jilin University guidelines and regulations.

Conflict of Interest

The authors declare that they have no conflict of interest.


  1. 1.
    Ponikvar M (2008) Exposure of humans to fluoride and its assessment. In: Tressaud A, Haufe G (eds) Fluorine and health. Elsevier Science, pp 487–549Google Scholar
  2. 2.
    Dequeker J, Declerck K (1993) Fluor in the treatment of osteoporosis. An overview of thirty years clinical research. Schweiz Med Wochenschr 123(47):2228–2234PubMedGoogle Scholar
  3. 3.
    Murray TM, Harrison JE, Bayley TA et al (1990) Fluoride treatment of postmenopausal osteoporosis: age, renal function, and other clinical factors in the osteogenic response. J Bone Miner Res Suppl 1:S27–S35Google Scholar
  4. 4.
    Boivin G, Chavassieux P, Chapuy MC, Baud CA, Meunier PJ (1989) Skeletal fluorosis: histomorphometric analysis of bone changes and bone fluoride content in 29 patients. Bone 10(2):89–99CrossRefGoogle Scholar
  5. 5.
    Tang SY, Alliston T (2013) Regulation of postnatal bone homeostasis by TGFβ. Bonekey Rep 2:255. CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Chen G, Deng C, Li YP (2012a) TGF-β and BMP signaling in osteoblast differentiation and bone formation. Int J Biol Sci 8(2):272–288. CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Bonewald LF, Dallas SL (1994) Role of active and latent transforming growth factor beta in bone formation. J Cell Biochem 55:350–357CrossRefGoogle Scholar
  8. 8.
    Crane JL, Xian L, Cao X (2016) Role of TGF-β signaling in coupling bone remodeling. Methods Mol Biol 1344:287–300. CrossRefPubMedGoogle Scholar
  9. 9.
    Everett ET (2011) Fluoride's effects on the formation of teeth and bones, and the influence of genetics. J Dent Res 90(5):552–560. CrossRefPubMedPubMedCentralGoogle Scholar
  10. 10.
    Monjo M, Lamolle SF, Lyngstadaas SP, Rønold HJ, Ellingsen JE (2008) In vivo expression of osteogenic markers and bone mineral density at the surface of fluoride-modified titanium implants. Biomaterials 29:3771–3780. CrossRefPubMedGoogle Scholar
  11. 11.
    Okuda A, Kanehisa J, Heersche JN (1990) The effects of sodium fluoride on the resorptive activity of isolated osteoclasts. J Bone Miner Res 5(Suppl 1):115–120Google Scholar
  12. 12.
    Gazzano E, Bergandi L, Riganti C, Aldieri E, Doublier S, Costamagna C, Bosia A, Ghigo D (2010) Fluoride effects: the two faces of janus. Curr Med Chem 17(22):2431–2441CrossRefGoogle Scholar
  13. 13.
    Grafe I, Alexander S, Peterson JR (2017) TGF-β family signaling in mesenchymal differentiation. Cold Spring Harb Perspect Biol 10.
  14. 14.
    Wang CL, Xiao F, Wang CD, Zhu JF, Shen C, Zuo B, Wang H, Li D, Wang XY, Feng WJ, Li ZK, Hu GL, Zhang X, Chen XD (2017) Gremlin2 suppression increases the BMP-2-induced osteogenesis of human bone marrow-derived mesenchymal stem cells via the BMP-2/Smad/Runx2 signaling pathway. J Cell Biochem 118(2):286–297. CrossRefPubMedGoogle Scholar
  15. 15.
    de Gorter DJ, van Dinther M, Korchynskyi O (2011) Biphasic effects of transforming growth factor β on bone morphogenetic protein-induced osteoblast differentiation. J Bone Miner Res 26(6):1178–1187. CrossRefPubMedGoogle Scholar
  16. 16.
    Jordan GR, Loveridge N, Power J, Clarke MT, Parker M, Reeve J (2003) The ratio of osteocytic incorporation to bone matrix formation in femoral neck cancellous bone: an enhanced osteoblast work rate in the vicinity of hip osteoarthritis. Calcif Tissue Int 72(3):190–196CrossRefGoogle Scholar

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© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.School of Pharmaceutical SciencesJilin UniversityChangchunPeople’s Republic of China

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